&

 Courtesy of Ramiro Martel Reyes
Chemical Approaches to Targeting Drug
Resistance in Cancer Stem Cells
COST Action CM1106
2nd Workshop
&
CIBICAN Conference on Molecular Pharmacology and
Mechanisms of New Anticancer Drugs
Puerto de la Cruz, Tenerife
October 14-15, 2014
http://www.cost.eu/
COST is an intergovernmental framework for European Cooperation in Science and
Technology, allowing the coordination of nationally-funded research on a European level. COST
has a very specific mission and goal. It contributes to reducing the fragmentation in European
research investments and opening the European Research Area to cooperation worldwide.
http://www.stemchem.org/
CMST COST Action CM1106 http://www.cost.eu/domains_actions/cmst/Actions/CM1106
From Memorandum of Understanding: This COST Action aims to unite researchers with
expertise in rational drug design and the medicinal chemistry of synthetic and natural
compounds with biomedical investigators dedicated to the understanding of the mechanisms
governing drug resistance in cancer stem cells. Through exchange of information, experience
and expertise, researcher mobility and fostering new collaboration between chemistry and
biology groups, the Action endeavours to develop new, effective methods for identifying novel
compounds and drug candidates that target drug-resistant cancer stem cells.
http://www.cibican.org/
The Centre for Biomedical Research of the Canary Islands (CIBICAN) is the result of the
collaboration between the University of La Laguna (ULL), the Insular Council of Tenerife and the
Canary Regional Government. It is aimed at further developing the international profile of its
Biomedical and Health Sciences research groups, as the core centre for biomedical and
biotechnological-based research of the ULL and the Canary Islands. Cibican has integrated the
research groups associated with the specialist ULL university institutes and the clinical research
units of the ULL associated hospitals bringing together the appropriate resources, scientific
equipment and knowledge base to accelerate biomedical discoveries and their application in the
promotion of health. Our mission is to combine interdisciplinary approaches from basic
biomedicine, medicinal chemistry and clinical research to develop new approaches towards the
transference of health knowledge to the industry and societal end users.
Joint COST CM1106 & CIBICAN Workshop
Scientific Committee
José M. Padrón University of La Laguna, Spain
Daniele Passarella University of Milan, Italy
Maurizio Botta University of Siena, Italy
Godefridus J. Peters VU University Medical Center, The Netherlands
Roger M. Phillips University of Bradford, United Kingdom
Organizing Committee
José M. Padrón University of La Laguna, Spain
Godefridus J. Peters VU University Medical Center, The Netherlands
Daniele Passarella University of Milan, Italy
Secretariat
Ioana Stupariu Secretary of the COST Action
Farah Cova Alonso Project Manager of IMBRAIN
Venue
Hotel Beatriz Atlantis and Spa, Avenida Venezuela 15, Puerto de la Cruz, Tenerife, Canarias
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 PROGRAMME
14 October, 2014
09:00
Registration
10:00
Introduction:
Section I
Chair:
10:15
L1 Roger Phillips
(University of Bradford, United Kingdom)
Should compounds with poor pharmacokinetic properties be
consigned to the archive or exploited for the loco-regional
treatment of cancer?
10:45
L2 Pierfausto Seneci – Invited Expert
(University of Milan, Italy)
Novel pro-apoptotic agents: chemistry-driven design, synthesis
and characterization
11:15
Authorities (University of La Laguna, Spain)
Rafael Alonso Solís (U. of La Laguna, Spain)
Daniele Passarella (University of Milan, Italy)
Frits Peters (VU University Medical Center, The
Netherlands)
Coffee break
Section II
Chair:
11:45
L3 Magdalena Krol
(Warsaw University of Life Sciences, Poland)
How tumor associated macrophages promote canine mammary
tumor metastasis?
12:05
L4 Ferenc Hudecz
(Eötvös Loránd University, Hungary)
Structure-activity relationship evaluation of ferrocene containing
antitumor compounds
12:25
L5 Ali O. Gure
(Bilkent University, Turkey)
Top-down and bottom-up approaches for identifying teranostic
biomarkers in cancer
12:45
Giovana Damia (Mario Negri Institute, Italy)
Lunch and Poster Session I
1
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 Section III
Chair:
15:00
L6 Frits Peters
(VU University Medical Center, The Netherlands)
Drug development at the EORTC-Drug Discovery Committee:
what can we learn from successful examples
15:30
L7 Wolfgang Link
(University of Algarve, Portugal)
Translocation in cancer therapy
15:55
L8 Miguel X. Fernandes
(University of La Laguna, Spain)
Coupling phenotypic drug discovery and computational methods
unravel anti-cancer therapeutic target
16:15
L9 Rengul Cetin-Atalay
(Bilkent University, Turkey)
Novel small molecule inhibitors against liver cancer
16:35
Bruno Botta (Sapienza University of Rome, Italy)
Coffee break and Poster Session I
17:00 – 18:00 MC meeting: Only for the members of the Management
Committee
2
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 15 October, 2014
Section IV
Chair:
09:30
L10 Andrea Menegon - Invited expert
(San Raffaele Scientific Institute, Italy)
Innovative optical approaches for ion channel functional studies
10:00
L11 Maurizio Botta
(University of Siena, Italy)
A possible strategy to combat HIV-associated cancers
10:20
L12 Cinzia Ingallina
(La Sapienza University, Rome)
Inhibitors of hedgehog pathway: new strategies in brain drug
delivery
10:40
Daniele Passarella (University of Milan, Italy)
Coffee break
Section V
Chair:
11:10
L13 Bernd Groner
(Institute for Tumor Biology and Experimental Therapy, Germany)
Induction of the miR-302/367 cluster in glioblastoma stem like
tumor initiating cells suppresses their tumorigenic gene
expression patterns and abolishes their transformation related
phenotypes
11:30
L14 Elena Petricci
(University of Siena, Italy)
Toward new antagonists of the DNA repair machinery and
hedgehog signaling pathway as anticancer agents
11:50
L15 Marcus Baumann
(University of Durham, United Kingdom)
Synthesis and biomedical evaluation of natural product inspired
chemical probes
12:10
L16 Mattia Mori
(Center for Life Nano Science, Italy)
Recent advances and opportunities in targeting cancer stem cells
12:35
L17 Jasna Banković
(University of Belgrade, Serbia)
Evaluating the mechanisms of new anticancer agents in multi-drug
resistant cancer cells
12:55
Andreanni Odysseos (EPOS-lasis, R&D, Cyprus)
Lunch break and Poster Session II
3
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 Section VI
Chair:
15:00
L18 Muriel Cuendet
(University of Geneva, Switzerland)
Natural products as cancer chemopreventive agents: a focus on
multiple myeloma cancer stem cells
15:20
L19 Richard Clarkson
(Cardiff University, United Kingdom)
Two novel cancer stem cell agents and a putative CSC platform
for drug evaluation
15:40
L20 Raimundo Freire
(Hospital Universitario de Canarias, Spain)
Ubiquitin hydrolases as novel regulators of DNA replication
16:00
L21 David Guillespie
(University of La Laguna, Spain)
Evaluation of Chk1 as a target for anti-cancer therapy in vivo
16:20
Panagiota Sotiropoulou (Université Libre de
Bruxelles, Belgium)
Coffee break and Poster Session II
16:50
Best poster competition
17:10
Best poster competition award
17:30
General discussion and closing remarks
4
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 POSTER SESSION
P1
Martins A, Csabi J, Amaral L, Molnar J, Hunyadi A. Postserone 2,3-dioxolanes
have improved activity sensitizing MCF7 and MDR mouse lymphoma cells
towards doxorubicin.
P2
Jovevska S, Mitanoska A, Bosnakovski D. Target cancer therapy by Notch and
WNT inhibitors-current approach and future prospective.
P3
Podolski-Renić A, Dinić J, Banković J, Milošević Z, Ríos-Luci C, Padrón JM,
Pešić M. Influence of MDR cancer phenotype on the efficacy of novel tubulin
destabilizing agent DTA0100.
P4
García-Caballero M, Medina MA, Quesada AR. Multilevel targeting for
leukemia therapy by the marine pyrrolidinedione AD0157.
P5
Monteiro A, Damia G, Ricci F, Passarella D, Santos MMM. Cytotoxic activity of
a new class of spirooxindoles in cancer stem cells.
P6
Maurizio B, Fallacara AL, Tintori C, Vignaroli G, Zamperini C, Crespan E, Maga
G, Angelucci A, Schenone S, Richters A, Iacono LD, Rauh D. Identification of
potent c-Src inhibitors affecting brain tumors in vivo.
P7
Hunyadi A, Martins A. An overview on our on-going research and collaboration
network against MDR cancer.
P8
Aikman B, de Graaf IAM, de Jager MH, Melgert BN, Groothuis GMM, Casini A.
Ex vivo precision cut tissue slices in the assessment of anticancer drug toxicity.
P9
Chadda R, Murphy PV. Novel macrocyclic glycuronides: Synthesis and
investigation of their potential as anti-tumour agents.
P10
Järvinen P, Muller CD. Importance of 3D culture in testing CSC lines for
apoptotic responses.
P11
Christodoulou MS, Fumagalli G, Sotiropoulou PA, Dosio F, Mazza D,
Passarella D. Self-assembled squalene-based fluorescent hetero-nanoparticles.
P12
Gozen D, Santos MMM, Monteiro A, Cetin-Atala R. Inhibition of MDM2-p53
interaction by novel compounds as therapeutics of hepatocellular carcinoma.
P13
Vilipić J, Stanojković T, Novaković I, Sladić D. Cytotoxic activity investigation of
twenty new amino acid tert-butylquinone and avarone derivatives.
P14
Hellmén E. Canine mammary osteosarcomas.
P15
Ghirga F, Mori M, Toscano S, Botta B, Ingallina C. Natural polyphenols and
derivatives as inhibitors of the hedgehog signalling pathway.
P16
Calogero F, Christodoulou MS, Bucci R, Passarella D. Convenient synthesis of
boehmeriasin A.
5
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P17
Mucha J, Majchrzak K, Taciak B, Hellmên E, Król M. MDSCs mediate
angiogenesis and predispose canine mammary tumor cells for metastasis via
IL-28/IL-28RA signaling.
P18
Dobiaš J, Hanquet G, Boháč A. An efficient identification of Mer TK validated
hits – exploitation of this methodology to Axl TK and Smo (a key mediator of the
Hedgehog pathway).
P19
Marucci C, Christodoulou MS, Bucci R, Passarella D. Pironetin-dumetorine
hybrids as new tubulin binders.
P20
García-Vilas JA, Cárdenas C, Quesada AR, Medina MA. Anti-angiogenic
natural compounds to evaluate their potential to target cancer stem cells.
P21
Stanković T, Balázs D, Dragoj M, Stojković S, Banković J, Martins A, Molnár J,
Amaral L, Hunyadi A, Pešić M. Lower antioxidant capacity of P-glycoprotein
overexpressing multi-drug resistant cancer cells as a mechanism for collateral
sensitivity to protoflavones.
P22
Dono R. Tumourless stem cells: uncoupling tumorigenicity from stemness by
acting on Glypican 4.
P23
Cincinelli R, Musso L, Dallavalle S. New heterocyclic scaffolds by
intramolecular reactions of 4-quinolone-2-carboxamides.
P24
Cincinelli R, Musso L, Dallavalle S, Podolski-Renić A, Pešić M. The effects of
new naphtoquinone and benzopyran derivatives studied in multi-drug resistant
cancer cells.
P25
Suleiman S, Schembri-Wismayer P, Cassar A, Aguis JC. Effects of biological
extracts on terminal differentiation of solid tumours and leukaemias.
P26
Borutinskaite V, Navakauskiene R. HDAC and HMT inhibitors in combination
with RA affect on gene methylation changes in leukemic cells.
P27
Filipović N, Pelliccia S, Bjelogrlić S, Todorović T, Schembri-Wismayere P,
Silvestri R. Novel Pd(II), Pt(II), Cu(II) and Ga(III) arylindole complexes as
potential leukaemia differentiation agents.
6
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 ORAL PRESENTATIONS
7
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L1. Should Compounds with Poor Pharmacokinetic Properties Be
Consigned to the Archive or Exploited for the Loco-Regional Treatment of
Cancer?
Roger Phillips
University of Bradford, United Kingdom
A key step in the development of anti-cancer drugs is the selection of
compounds with good pharmacokinetic (PK) properties. Whilst this is an essential
requirement for systemic based therapies targeting disseminated disease, it is of less
importance for loco-regional therapies where the aim is to deliver high concentrations
of drug to a localised tumour without systemic exposure. Indeed, compounds with poor
PK could paradoxically be advantageous in this setting and to exemplify this, the case
of the indolequinone bioreductive drug EO9 will be described.
Based on good pharmacodynamics (PD) properties in preclinical models, EO9
underwent clinical evaluation in the mid 1990’s but it failed to demonstrate efficacy in
phase II trials when administered intravenously. Rapid PK elimination and poor
penetration through avascular tissue were identified as the principle reason for the
failure of EO9. Instead of consigning EO9 to the archive, we argued that the ‘bad’ PK
properties of EO9 would in fact be advantageous in the treatment of superficial bladder
cancer. In this setting, intravesical administration of EO9 directly into the bladder would
circumvent the drug delivery problem and any drug that penetrated through the bladder
wall and into the systemic circulation would be rapidly eliminated thereby reducing the
risk of systemic toxicity. A phase I study was subsequently conducted and this
demonstrated that the intravesical administration of EO9 was not only safe (well
tolerated locally and no systemic toxicity) but it was efficacious with 8 out of 12
complete responses recorded. These results were reproduced in phase II studies and
phase III trials are being conducted in Europe and North America.
The example of EO9 demonstrates that compounds with good PD but poor
systemic PK can still be efficacious in the clinic if applied loco-regionally. As many
compounds are eliminated from further development because of undesirable PK
properties, it is very likely many compounds with similar characteristics reside on the
shelves of medicinal chemists laboratories. Rather than being consigned to the archive
and ignored, this presentation will argue that these compounds should be re-evaluated
as potential loco-regional therapies where poor systemic PK is actually an advantage
rather than a hindrance.
8
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L2. Novel Pro-Apoptotic Agents: Chemistry-Driven Design, Synthesis and
Characterization
Pierfausto Seneci
Invited expert
University of Milan, Italy
Apoptosis, or programmed cell death (PCD), is dysfunctional in a variety of
human pathologies, and has become the focus of extensive pharmaceutical research
[1]. Pathways leading to PCD include the extrinsic or death receptor-dependent path
and the intrinsic or mitochondrial path, both caspase-dependent. In a complex
scenario, hitting more than one putative significant target with a lead should maximize
the chances of therapeutic success.
The family of Inhibitor of Apoptosis Proteins (IAPs) [2] is characterized by one
or more Baculovirus IAP Repeat (BIR) domains. The most caspase-connected human
IAP is the X-Inhibitor of Apoptosis Protein (XIAP). XIAP is capable of binding caspase 9
(the initiator caspase) and both caspases 3 and 7 (the executioner caspases). XIAP
binding prevents activation of caspases and, consequently, prevents cells from
entering PCD. A protein released from mitochondria, Smac-DIABLO [3], binds XIAP as
a dimer on the same binding sites of caspase 9 (BIR3 domain). Smac interferes with
the binding site of caspases 3 and 7 (linker-BIR2 domain), promoting the extrinsic and
intrinsic PCD paths. Smac binding to other human IAP family members such as cIAP1
and cIAP2 through their BIR domains is also proven. Thus, Smac mimics may
modulate the pathological action of any IAP oncology target in both caspasedependent PCD paths.
We describe 4-substituted mono- and dimeric diazabicycloalkane-based Smac
mimics/IAP inhibitors [4-7] which establish novel, meaningful target-ligand interactions.
Their rational design via computational chemistry, their synthesis via original protocols
and their structural characterization via NMR and X-ray studies is described. Both
compound classes showed significant binding activity to BIR3 and linker-BIR2-BIR3
domains of XIAP and cIAPs, and a SAR is presented for them. Their cytotoxic activity
on tumor cell lines is reported, and key factors influencing cell penetration were
identified. A detailed characterization, including in vivo testing, is presented for the
selected lead Smac83, currently being licensed to a biotech company. Finally, the
design and synthesis of multi-targeted, “heterodimeric” Smac mimetics/caspase
activators based on the structure of recently reported PAC-1 [8] is presented in details.
References
[1]
[2]
[3]
[4]
[5]
[6]
[7]
[8]
Alberts K et al. Molecular Biology of the Cell (5th ed.). Garland Science, London, UK,
1115-1143 (2008).
Varfolomeev E, Vucic D. Future Oncol. 7, 633-648 (2011).
Chai J et al. Nature 406, 855-862 (2000).
Seneci P et al. Bioorg. Med. Chem. 17, 5834-5856 (2009).
Cossu F et al. J. Mol. Biol. 392, 630-644 (2009).
Lecis D et al. Bioorg. Med. Chem. 20, 6709-6723 (2012).
Potenza D et al. Org. Biomol. Chem. 10, 3278-3287 (2012).
Putt, KS et al. Nat. Chem. Biol. 2, 543-550 (2006).
9
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L3. How Tumor Associated Macrophages Promote Canine Mammary
Tumor Metastasis?
Magdalena Król1, Joanna Mucha1, Kinga Majchrzak1,2, Agata Homa1, Małgorzata
Bulkowska1, Alicja Majewska1, Małgorzata Gajewska1, Marta Pietrzak1, Mikołaj
Perszko1, Karolina Romanowska1, Karol Pawłowski1,3, Elisabetta Manuali4, Eva
Hellmen5, Bartłomiej Taciak1, Tomasz Motyl1
1
Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of
Life Sciences Poland
2
Department of Animal Environment Biology, Faculty of Animal Sciences, Warsaw University of
Life Sciences Poland
3
Department of Large Animal Diseases with Clinic, Faculty of Veterinary Medicine, Warsaw
University of Life Sciences, Poland
4
Area Diagnostica Integrata Istologia e Microscopia Elettronica Istituto Zooprofilattico
Sperimentale dell'Umbria e delle Marche, Italy
5
Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural
Sciences, Sweden
Tumor-associated macrophages (TAMs) constitute a major component of tumor
microenvironments and influence cancer development. According to the current
hypothesis, these cells are “corrupted” by cancer cells and subsequently facilitate,
rather than inhibit, tumor progression and metastasis.
Using microarrays, we investigated miRNA expression in co-cultures of
macrophages and five canine mammary tumor cell lines and showed that the majority
of miRNA-targeted genes were involved in Wnt signaling. Furthermore, we showed that
co-culture with TAMs or treatment with macrophage-conditioned medium inhibited the
canonical Wnt pathway and activated the non-canonical Wnt pathway in tumor cells.
Subsequently, we demonstrated macrophage-induced invasive growth patterns and
epithelial–mesenchymal transition of tumor cells. Validation of these results ex vivo in
canine mammary carcinoma tissues (n = 50) and xenograft tumors indicated the
activation of non-canonical and canonical Wnt pathways in metastatic tumors and nonmetastatic malignancies, respectively.
We demonstrated that TAMs mediate a “switch” between canonical and noncanonical Wnt signaling pathways in canine mammary tumors, leading to increased
tumor invasion and metastasis.
Interestingly, similar changes in neoplastic cells were observed in the presence
of macrophage-conditioned medium or live macrophages. Hence, macrophages
secrete canonical Wnt inhibitors and non-canonical Wnt activators, even without
stimulation by neoplastic cells. However, co-culture with tumor cells enhanced this
effect. These observations indicate that rather than being “corrupted” by cancer cells,
TAMs constitutively secrete canonical Wnt inhibitors that decrease cancer proliferation
and development, but as a side effect, they induce the non-canonical Wnt pathway,
which leads to tumor activation and metastasis.
These data challenge the conventional understanding of TAM–cancer cell
interactions.
Acknowledgements: These studies have been supported by the grant no. UMO2013/09/D/NZ5/02496 from the National Science Centre.
10
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L4. Structure-Activity Relationship Evaluation of Ferrocene Containing
Antitumor Compounds
Szilvia Bősze1, Ildikó Szabó1, László Kocsis1, Antal Csámpai2, Ferenc Hudecz1,3
1
MTA-ELTE Research Group of Peptide Chemistry, Budapest, Hungary
Department of Inorganic Chemistry, Eötvös Loránd University (ELTE), Budapest, Hungary
3
Department of Organic Chemistry, Eötvös Loránd University (ELTE), Budapest, Hungary
2
Applying the concept of building block strategy a small library of ferrocenebased model compounds also incorporating quinchona-, chalchone- and [1,2,3]triazole
moieties as well as purely organic reference compounds were synthetized and
evaluated in in vitro antitumor tests. We envisaged that the combination of the
aforementioned structural elements encountered in a variety of molecules having
documented cytostatic effect [1-4] may give rise to a series of model derivatives of
enhanced level of activity. The contribution of the particular building blocks to the
antitumor effect was estimated on the basis of the results of the in vitro tests. Our
convergent synthetic strategy utilized the copper-catalysed [2+3] cycloaddition [5] of
dehydroquinine or dehydroquinidine with an azide component attached to the
previously constructed ferrocene-containing- or purely organic chalchone fragment to
afford 1,4-disubstituted triazoles as the targeted products. Definite correlations were
found between the structure and in vitro activity of the model compounds. While the
replacement of the ferrocene unit for phenyl group did not affect significantly the in vitro
antitumor activity, the presence of the quinchona moiety proved to be essential
irrespective to its relative configuration. Interestingly, significant activities were also
detected in the tests performed on azido-substituted chalchone derivatives. The
beneficial effects of many new and clinically used anticancer compounds are limited
because of their inability to reach the appropriate cellular targets. The cellular uptake
rate and the bioavailability of drug compounds can be enhanced by covalent
attachment to appropriate targeting or carrier peptide. Development of peptide
conjugates for drug targeting is one of the main topics of our research group. The
combination of drug candidates with specific targeting moiety for CSCs might lead to
highly selective conjugates [6].
Acknowledgements: This work was financially supported by the Hungarian Scientific Research
Fund (OTKA K104385, PD 83923, K83874, K104045 and PD104012). LK would like to thank
the MTA Postdoctoral Fellowship.
References
[1]
[2]
[3]
[4]
[5]
[6]
G. Jaouen, S. Top, A. Vessieres, P. Pigeon, G. Leclercq, I. Laios, Chem. Commun. 383
(2001).
B. I. Károlyi, Sz. Bősze, E. Orbán, P. Sohár, L. Drahos, E. Gál, A. Csámpai, Molecules
17, 2316 (2012).
Zsoldos-Mady V, Csampai A, Szabo R, Meszaros-Alapi E, Pasztor J, Hudecz F, Sohar P.
ChemMedChem 1, 1119 (2006).
L. V. Snegur, Y. S. Nekrasov, N. S. Sergeeva, Z. V. Zhilina, V. V. Gumenyuk, Z. A.
Starikova, A. A. Simenel, N. B. Morozova, I. K. Sviridova, V. N. Babin, Appl. Organomet.
Chem. 22, 139 (2008).
V. V. Rostovtsev, L. G. Green, V. V. Fokin, K. B. Sharpless, Angew Chemie Int Ed 41,
2596 (2002).
N. Mihala, F. Hudecz, Amino Acids, Pept. Proteins 37, 1 (2012).
11
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L5. Top-Down and Bottom-Up Approaches for Identifying Teranostic
Biomarkers in Cancer
Murat Isbilen, Waqas Akbar, Kerem Mert Senses, Anna Lucia Fallacara, Arthur
Machlenkin, Michal Lotem, Maurizio Botta, Ali Osmay Gure
Bilkent University, Ankara, Turkey
I will describe two approaches we use to delineate theranostic biomarkers in
cancer. The "top-down" approach is based on utilizing in silico data as starting material,
followed by in vitro validation, whereas the "bottom-up" approach is based on in vitro
data which is then expanded based on in silico secondary data, which is again
validated. The first approach was used to successfully identify candidate drugs that
could be used to treat a sub group of triple-negative breast cancer cell lines with stemcell like features. The second approach utilized melanoma primary cells with gene
expression data to identify biomarkers of chemosensitivity for novel Src inhibitors such
as 10a.
12
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L6. Drug Development at the EORTC-Drug Discovery Committee: What
Can We Learn from Successful Examples
Godefridus J Peters1, Hans R Hendriks2, Anne-Sophie Govaerts3, Andy Westwell4,
Iduna Fichtner5 on behalf of the EORTC-Drug Discovery Committee
1
Chair of the EORTC-Pharmacology and Molecular Mechanisms group, Brussels, Belgium, and
VU University Medical Center, Amsterdam, The Netherlands
2
Hendriks Pharmaceutical Consulting, Purmerend, The Netherlands;
3
EORTC Head Quarters, Brussels, Belgium
4
School of Pharmacy and Pharmaceutical Sciences, University of Cardiff, Wales
5
Max-Delbrück Center for Molecular Medicine, Berlin-Buch, Germany
Within the EORTC two main preclinical drug development programs are running, one
based on the initiative of individual investigators and the other based on a collaboration with the
NCI to identify new anticancer agents, using a pharmacological strategy. The first program was
initiated by the establishment of the Screening and Pharmacology Group in 1972 (in 2003
continued as a committee of the PAMM group: the Drug Discovery Committee), the second in
1993 when the EORTC Translational Research Division and Cancer Research UK established
a collaboration with the NCI in order to test compounds with a unique mechanism of action, and
new structures related to specific mechanism of actions which came from the NCI in-vitro 60tumor cell line screen. The latter compounds were awaiting further in-vivo evaluation and about
20% originated from Europe. It was decided that in the EORTC-NCI-compounds initiative these
would be further evaluated.
The first program was based on the interest of investigators in specific types of
compounds or mechanisms of action. Using classical and novel medicinal chemical approaches
a number of new compounds were synthesized, and initially tested in the SPG/DDC group for
activity (both in vitro and in vivo), often accompanied by mechanistic studies. Unusual paths
were sometimes followed, based on specific properties of a compound, such as temozolomide,
which was not active in several in vitro model systems but due to its unique properties (e.g.
crossing the blood-brain barrier) was further tested and is currently first-line treatment for
glioblastoma. More recent examples include Phortress, a potent aryl hydrocarbon receptor
(AhR) ligand (antagonist) which induces CYP1A1 and CYP1B1 transcription (mRNA and
expression). This compound is currently in Phase 1. Other compounds which were (initially)
developed within the SPG/DDC and went into the clinic include several prodrugs (e.g.
elacytarabine, CP4126) and SJG136, a novel DNA sequence selective minor groove crosslinking agent.
The NCI-compounds initiative included a review of available NCI data of compounds of
European origin by CRUK/EORTC members, focusing on novelty of chemical structure,
COMPARE-negative, mean-graph and growth curve profiles, potency and molecular target
data, and if available in-vivo data, later including hollow-fiber assay. Selected compounds were
tested in vivo with limited pharmacokinetics, drugs were formulated, and when sufficiently high
plasma concentrations were reached further tested for antitumor activity. Additional tests
included tubulin binding and anti-vascular effects. From the more than 1800 compounds that
have been reviewed about 150 compounds were selected. Criteria for dropping included
solubility, impurity, instability, insufficient bioavailability or insufficient activity in human tumour
xenograft models in vivo, poor pharmacology. The work resulted in fruitful multidisciplinary
interactions in Europe and increased the understanding of knowledge of structures and
mechanism of actions of drugs that made it to the clinic.
13
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L7. Translocation in Cancer Therapy
Richard Hill1,2, Patrícia Madureira1,2, Laura Colaço1, Selma Ugurel3, Murat Isbilen4, Ali
Gure4, Wolfgang Link1,2
1
Regenerative Medicine Program, Department of Medicine and Biomedical Sciences, University
of Algarve, 8005-139 Faro, Portugal
2
Centre for Molecular and Structural Biomedicine, CBME/IBB, University of Algarve, 8005-139
Faro, Portugal
3
Department of Dermatology, Julius-Maximilians University, Würzburg, Germany
4
Department of Molecular Biology and Genetics, Bilkent University, Ankara, Turkey
Intrinsic and acquired resistance to conventional and targeted chemotherapy is
the primary reason for treatment failure in many cancers. The identification of
molecular mechanisms that determine drug resistance and could be targeted has
enormous clinical importance. The identification of molecular mechanisms that
determine drug resistance and could be targeted has enormous clinical importance.
Crucial transcription factors such as forkhead box O (FOXO) proteins and p53 have
been shown to mediate the action of multiple anti-cancer drugs, including PI3K
pathway inhibitors. Here we reveal that tribbles homolog 2 (TRIB2) expression ablates
FOXO activation, disrupts the p53/MDM2 regulatory axis thus confers resistance to
anti-cancer drugs that include PI3K inhibitors. We show that TRIB2 expression is
significantly increased in melanoma, colon and pancreatic cancers that correlates with
extremely poor clinical prognosis. We report that TRIB2-mediated suppression of
FOXO and p53 activity is indirect and rather, is via the activation of AKT.
Mechanistically, the TRIB2 protein is stabilized by reduced proteasome-dependent
degradation and promotes AKT activation via its COP1 domain. Altogether, our study
reveals a novel regulatory mechanism underlying drug resistance in a range of
cancers, and suggests that TRIB2 functions as an important component within the AKT
signaling network in cancer cells.
14
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L8. Coupling Phenotypic Drug Discovery and Computational Methods
Unravel Anti-Cancer Therapeutic Target
Gastón Silveira-Dorta, Víctor S. Martín, José M. Padrón, Miguel X. Fernandes
Instituto Universitario de Bio-Organica "Antonio Gonzalez", Centro de Investigaciones
Biomédicas de Canarias (CIBICAN), Universidad de La Laguna, Avda. Astrofisico Francisco
Sanchez 2, 38206 La Laguna, Tenerife, Spain
Phenotypic drug discovery (PDD) studies the effect of small molecules, which
interact with unknown target(s), on some cellular parameters with no need to anticipate
mechanism(s) of action. We synthesized a series of lipidic aminoalcohols and studied
their effect on the growth of a representative panel of human solid tumor cell lines. The
cancer cell growth inhibition data were used as biological activity data for these
compounds. We performed molecular docking calculations of the lipidic aminoalcohols
against a panel of over 40 commonly used anti-tumoral therapeutic targets. We used
correlations between the docking scores for the aminoalcohols against a particular
target and the biological activity data to predict a possible therapeutic target for the
aminoalcohols. Experimental confirmation of computational prediction allowed us to
unravel a target for anti-cancer therapy. Structure-based efforts to design new
derivatives with improved biological activity are on course.
Acknowledgements: Cofinanced by the EU Research Potential (FP7-REGPOT-2012-CT201231637-IMBRAIN), the European Regional Development Fund (FEDER), and the Spanish
Instituto de Salud Carlos III (PI11/00840).
15
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L9. Novel Small Molecule Inhibitors against Liver Cancer
Irem Durmaz1, Tulin Ersahin1, Damla Gozen1,2, Deniz Cansen Yildirim1, Rengul CetinAtalay1,2
1
Department of Molecular Biology and Genetics, Faculty of Science, Bilkent University, Ankara,
Turkey
2
Informatics Institute, Middle East Technical University, 06531, Ankara, Turkey
Liver cancer is one of the leading causes of cancer-related deaths worldwide
and chemotherapy is the only treatment option for patients with Hepatocellular
Carcinoma (HCC). Yet, the only FDA-approved therapeutic agent, Sorafenib, which
targets RAF/MEK/ERK pathway, fails to prevent tumor recurrence due to persistent
signaling through alternative pathway activations.
Here, we present our studies on a various small molecules, which are cytotoxic
against liver cells.
First group of molecules are bioactive against PI3K/AKT/mTOR signaling. We
studied the synergistic effect of these molecules in combination with Sorafenib.
Inhibitors of PI3K or AKT kinases and Sorafenib leads to synergistic growth inhibition
and enhanced cell death. Suppression of cell cycle progression and migration, and
induction of apoptotic cell death were shown by flow cytometry, wound healing,
immunofluorescence and western blots experimentally. Computational analysis of
microarray and RNA-seq data identified the molecular mechanism of action of inhibitors
as single agents or in combination with Sorafenib, and determined specific gene sets.
Our transcriptomic analysis identified direct enzymatic targets and key downstream
effector proteins of each inhibitor. Additionally, we demonstrated synergistic anticancer
activities on mice xenografts.
Second group of molecules are cardiac glycosides. We focused on the
repositioning of these compounds to liver cancer treatment and to the chronic liver
diseases as chemopreventive agents for the first time. We initially showed that the
glycosides Lanatoside A, Lanatoside C and Glucogitoroside displayed significant timedependent cytotoxic activities on liver cancer cell lines in nano- and picomolar
concentrations. In this study, we have performed detailed biochemical analysis with
Lanatoside C, which is the natural precursor of the clinically preferred glycoside
digoxin. We showed that Lanatoside C induces oxidative stress dependent SAPK/JNK
protein activation in liver cancer cells. Cell death was characterized as a part of
extrinsic apoptotic pathway and G2/M cell cycle arrest. Furthermore in vivo tumor
experiments on nude mice with orally administered Lanatoside C significantly
demonstrated both chemopreventive and chemotherapeutic actions as followed by
tumor size and magnetic resonance imaging.
Finally, I will present our very recent data that we obtained with small molecules
coming from our COST collaborators: Potential p53-MDM2 inhibitors from Dr. Maria M.
Santos, (ULisboa), Src inhibitors from Dr. Maurizio Botta (Siena) and Large screening
with potential cytotoxic compounds from COST participants provided by Dr. Nadine
Martinet (Inserm).
16
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L10. Innovative Optical Approaches for Ion Channel Functional Studies
Andrea Menegon
Invited expert
Advanced Light and Electron Microscopy Bio-Imaging Centre, Experimental Imaging Centre,
San Raffaele Hospital Milan, Italy
Ion channels are involved in several socially relevant pathologies (such as
epilepsy, arrhythmia, hypertension, some psychological disorders, cancer etc.).
Although drug discovery has seen a rapid improvement in molecules, reagents,
technologies and cellular models, these advancements cannot be fully profited in ion
channel drug discovery. This is due to the intrinsic characteristics of ion channels (i.e.
channels open transiently and are modulated by different mechanisms) that make them
particularly difficult to be studied as pharmacological target. The quantitative study of
ion currents provides a direct measure of ion channel activity. However, the available
techniques (manual and automated patch clamping) have intrinsic limitations in drug
screening, even for the study of small libraries, because of time requirements and
complexity. Ion-luminescence or ion-fluorescence detection systems are compatible
with HTS (high throughput screening) studies but are intrinsically affected by low
temporal resolution. These approaches are virtually confined to the study of second
messenger variations as indirect readout for ion channel activation. A compromise
between the high precision of electrophysiology and the high processing capacity of
HTS is represented by the study of transmembrane potential variations by voltage
sensitive fluorescent dyes (VSD). In this talk, two applications of fast VSD, for direct
study of the membrane potential changes and the analysis of the functional state of ion
channels, will be shown. The possibility to apply these approaches to the screening of
molecules active on transient receptor potential channels (TRPCs), Acid sensing ion
channels (ASICs) as well as chloride channels (reported to be involved in cancer) will
be discussed.
17
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L11. A Possible Strategy to Combat HIV-Associated Cancers
Maurizio Botta1, Manfred Jung2, Canducci Filippo3, Rosa Martí-Centelles4, Eva
Falomir4, Juan Murga4, Miguel Carda4, J. Alberto Marco5, Cristina Tintori1, Giorgio
Maccari1, Marika Tiberi1, Giulia Iovenitti1, Sören Swyter2
1
Department of Biotechnology Chemistry and Pharmacy, University of Siena, Via A. Moro 2,
53100 Siena, Italy
2
Institute of Pharmaceutical Sciences, Albert-Ludwigs Universität, Albertstrasse 25, 79104
Freiburg, Germany
3
Department of Clinical and Experimental Medicine, Università degli Studi dell’Insubria, 21100
Varese, Italy
4
Departamento de Química Inorgánica y Orgánica, Univ. Jaume I, E-12071 Castellón, Spain
5
Departamento de Química Orgánica, Univ. de Valencia, E-46100 Burjassot, Valencia, Spain
Acquired immunodeficiency syndrome (AIDS), caused by the human
immunodeficiency virus (HIV), has become a major epidemic disease, infecting more
than 39 million people worldwide. The virus compromises the host immune system
increasing the risk of coinfections and development of opportunistic pathologies.
Among them, certain types of cancers are typical on HIV infected patients, such as the
Kaposi’s sarcoma and the Non-Hodgkin Lymphoma. The literature shows a doubled
risk of morbidity and mortality for Non-Hodgkin Lymphoma in HIV patients [1].
In previous works we have reported the synthesis and biological evaluation of
potent anti-viral compounds with a 5-(2-furfurylidene)-2-thioxo-thiazolidin-4-one or 2amino-5-(2-furfurylidene)-2-thiazolin-4-one structure. The biological target of these
compounds is not yet well defined, but the anti-viral activity spreads from micro- to the
nano-molar values.
Compounds with a similar 3D structure have been proposed by Maurer et al as
sirtuin 5 inhibitors [2]. Selection and testing of some own compounds have revealed
their activity against the isoform 2 of sirtuin at micro-molar concentration.
Consequently, we tested these compounds also against cancer cells finding good
activity and selectivity. The most promising compounds were further investigated
looking for the down-regulation of the expression of the oncogene hTERT and the
proto-oncogene cMYC. Remarkably, such compounds were also able to reduce the
expressions of the aforementioned genes.
We are still in a preliminary phase of our research but our findings could lead to
the design of new compounds active against two unrelated diseases. We have the
strong feeling that development and administration of a drug endowed with a similar
activity can increase the life expectancy of patients affected by HIV-associated
cancers.
References
[1]
[2]
Chao, C.; Xu, L.; Abrams, D.; Leyden, W.; Horberg, M.; Towner, W.; Klein, D.; Tang, B.;
Silverberg, M., Survival of non-Hodgkin lymphoma patients with and without HIV infection
in the era of combined antiretroviral therapy. AIDS 24, 1765-1770 (2010).
Maurer, B.; Rumpf, T.; Scharfe, M.; Stolfa, D. A.; Schmitt, M. L.; He, W. J.; Verdin, E.;
Sippl, W.; Jung, M., Inhibitors of the NAD(+)-Dependent Protein Desuccinylase and
Demalonylase Sirt5. Acs Medicinal Chemistry Letters 3, 1050-1053 (2012).
18
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L12. Inhibitors of Hedgehog Pathway: New Strategies in Brain Targeted
Drug Delivery
Cinzia Ingallina1,2, Mattia Mori2, Federica Rinaldi1,2, Maria Carafa1, Bruno Botta1,
Carlotta Marianecci1
1
Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Università di Roma, 00185
Roma, Italy
2
IIT Center for Life Nano Sciences, CLNS@Sapienza, 00161 Roma, Italy
Hedgehog (Hh) signalling pathway is essential for tissues development and
stemness and its deregulation leads to tumorigenesis. Hedgehog pathway aberrant
activation has been reported in many tumors, one is the medulloblastoma, the most
frequent malignant brain tumor of childhood, which belongs to the group of embryonal
neuroepithelial tumors and arise from stem cells or early progenitor cells in the
cerebellum [1]. Thus, inhibitors of Hedgehog pathway proved in recent years a widely
appreciated target for anticancer drugs [2].
The aim of this study is to discover and develop small organic molecules and/or
natural compounds capable of inhibiting the Hh signaling pathway and to prepare
niosomal formulations able to deliver the drugs through the blood-brain barrier (BBB).
To this purpose, we previously set up a robust computational protocol to screen a
library of bioactive natural products available in our laboratories. Some hits have been
selected in silico and submitted to biological evaluation (luciferase assays). Few of
them showed a very interesting activity against the Hh pathway, in particular those
belonging to the family of polyphenols. The most potent and less toxic compound
selected (whose structure is patented) [3] was encapsulated into niosomal vesicular
systems, with the aim of enhance its brain delivery. In fact, niosomes are versatile
carriers for drug delivery which have received growing attention in recent years [4], due
to the fact that they can entrap both lipophilic and hydrophilic drugs.
We prepared three niosomal formulations using polysorbates (Tweens) mixed
with cholesterol by film method, which were afterwards purified by both size exclusion
chromatography (SEC) and extruder, and fully characterized (for dimensions, zeta
potential, stability and bilayer fluidity, polarity and microviscosity). Fluorescent probes
loaded vesicles have been also prepared to simulate release studies in different media
(Hepes buffer pH 7.4, bovine serum and human serum), and to perform both cell
internalization and permeation studies trough a BBB model.
Entrapment efficiency of the lead compound into the prepared niosomes,
checked by HPLC, was found to be greater than 5%. Cytotoxicity of vesicle formulation
was tested on mouse fibroblast Balb/3T3, pointing out no significant toxicity for a time
of exposure of 24 hours. An in vitro BBB model was built by mouse endothelial bEnd3
co-cultured with astrocytes to investigate the mechanism of BBB permeation and
internalization of the selected formulation into the brain compartment.
References
[1]
[2]
[3]
[4]
Rudin C. M.; Hann, C. L.; Laterra, J.; Yauch, R. L.; Callahan, C. A.; Fu, L.; Holcomb, T.;
Stinson, J.; Gould, S. E.; Coleman, B.; LoRusso, P. M.; Von Hoff, D.D.; de Sauvage, F.
J.; Low, J. A. N. Engl. J. Med. 361, 1173 (2009).
Lin, T. L; Matsui, W. OncoTargets Ther. 5, 47 (2012).
B. Botta, B.; Gulino, A.; Botta, M.; Mori, M.; Di Marcotullio, L.; Infante, P.; Ghirga, F.;
Toscano, S.; Ingallina, C.; Alfonsi, R. PCT EP 2014063449 (2014).
Torchilin, V. P. Adv. Drug Del. Rev. 64, 302 (2012).
19
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L13. Induction of the miR-302/367 Cluster in Glioblastoma Stem Like
Tumor Initiating Cells Suppresses their Tumorigenic Gene Expression
Patterns and Abolishes their Transformation Related Phenotypes
Bernd Groner
Georg Speyer Haus, Institute for Tumor Biology and Experimental Therapy, Paul Ehrlich Str. 42,
D-60596 Frankfurt am Main, Germany
Cellular transformation is initiated by the activation of oncogenes and the
closely associated developmental reprogramming of the epigenetic landscape.
Transcription factors, regulators of chromatin states and microRNAs influence cell fates
in development and stabilize the phenotypes of normal, differentiated cells and of
cancer cells. The miR-302/367 cluster, e.g., is predominantly expressed in human
embryonal stem cells (hESs), it can promote the cellular reprogramming of human and
mouse cells and the generation of induced pluripotent stem cells, iPSC. We have
investigated if the epigenetic reprogramming potential of the miR-302/367 cluster can
be exploited to “de-program” tumor cells, i.e. shift their gene expression pattern
towards an alternative program associated with more benign cellular phenotypes.
Induction of the miR-302/367 cluster in U87MG glioblastoma cells drastically
suppressed the expression of transformation related proteins, e.g. the reprogramming
factors OCT3/4, SOX2, KLF4 and c-MYC, and the transcription factors POU3F2,
SALL2 and OLIG2, required for the maintenance of glioblastoma stem like tumor
propagating cells. It also diminished PI3K/AKT and STAT3 signaling, abolished the
features of epithelial to mesenchymal transition, altered the cytokine secretion patterns
and suppressed colony formation of the cells in soft agar. At the same time the miR302/367 cluster expression restored the expression of MAP2, a neuronal marker of
differentiation. Upon transplantation of the miR-302/367 cluster expressing cells into
mice, they had lost their ability to form tumors at the site of injection and to establish
liver metastasis.
20
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L14. Toward New Antagonists of the DNA Repair Machinery and hedgehog
Signaling Pathway as Anticancer Agents
Elena Petricci
Department of Biotechnology, Chemistry and Pharmacy, Università degli Studi di Siena, Via A.
Moro, 53100 Siena, Italy
Cancer stem cells (CSC) exhibit a survival advantage following chemo- and
radiotherapy, generating a real need for anticancer agents acting on new targets.
Between the different causes of tumor resistance the enhanced DNA repair
mechanisms and the higher expression of antiapoptotic factors (i.e. sonic hedgehog)
recently emerged as crucial [1].
MRE11 within the MRE11-RAD50-NBS1 (MRN) complex acts in DNA doublestrand break repair (DSBR), detection and signaling. MRN can be considered as an
ATM effector involved in hereditary breast and ovarian cancer syndrome, ataxiatelangiectasia, and Nijmegen breakage syndrome [2]. We recently employed structurebased design to project and synthesise a focused library of specific inhibitors of MRE11
endo- or exonuclease activity. PFM39 and PFM01 demonstrate to be selective
inhibitors of the exo- and endonuclease activity respectively and help on elucidating
MRE11 rule on DSBR [3]. Resection is an important step promoting translocations that
drive carcinogenesis. As resection is particularly active in CSCs generating resistance
to normal chemo- and radiotherapeutic agents, PFM derivatives have a great potential
biological and clinical value in cancer therapy.
Between the antiapoptotic factors over-expressed in CSCs, Sonic Hedgehog
(SHh) is a well known target for antineoplastic agents. Acylguanidine derivatives
recently developed by our group emerged as interesting Smo inhibitors [4] with an
optimal pharmacokinetic profile. Preliminary results on different cancer cell lines
indicate these compounds as very promising and effective tools to overcome
chemotherapy resistance and eradicate CSCs.
References
[1]
[2]
[3]
[4]
Blanpain, C.; Mohrin, M.; Sotiropoulou, P.A.; Passegue, E. Cell Stem Cell 8, 16-29
(2011).
Maugeri-Saccà, M.; Bartucci, M.; De Maria, R. Mol. Cancer Ther. 11, 1627-1636 (2012).
Shibata, A.; Moiani, D.; Arvai, A.S.; Perry, J.; Harding, S.M.; Genois, M.M.; Maity, R.;
Romoli, F.; Ismail, A.; Ismalaj, E.; Petricci, E.; Neale, M.J.; Bristow, R.G.; Masson, J.Y.;
Wyman, C.; Jeggo, P.A.; Tainer, J.A. Mol Cell. 53, 7-18 (2014).
Solinas, A.; Faure, H.; Roudaut, H.; Traiffort, E.; Schoenfelder, A.; Mann, A.; Manetti, F.;
Taddei, M.; Ruat, M. J. Med. Chem. 55, 1559-1571 (2012).
21
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L15. Synthesis and Biomedical Evaluation of Natural Product Inspired
Chemical Probes
Marcus Baumann
Department of Chemistry, University of Durham, United Kingdom
Natural products represent a unique class of bioactive compounds with
favourable pharmacological profiles, however, their high levels of cytotoxicity often
prevents their development into chemical probes or drug candidates. This talk will
present on the synthesis and biomedical evaluation of several natural product
analogues derived from boehmeriasin A and various epidithiodiketopiperazines (e.g.
chaetocine and gliocladine), which hold promise as novel epigenetic agents. The latter
series has been evaluated against a panel of human cancer cell lines showing high
activity in vitro and in vivo, whilst lacking any signs of cytotoxicity. Specifically, a distinct
histone methyltransferase (SUV39H1) was identified as molecular target allowing
further preclinical development as a new anticancer agent. Whilst this development is
ongoing further efforts towards studying these molecules in the context of cell
differentiation/reprogramming and cancer stem cells are being initiated and expected to
extend our knowledge of how to target related diseases.
22
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L16. Recent Advances and Opportunities in Targeting Cancer Stem Cells
Mattia Mori
Center for Life Nano Science@Sapienza, Istituto Italiano di Tecnologia, viale Regina Elena
291, 00161 Roma, Italy
Common cancer therapies target the bulky tumor and decrease it size, thus
allowing a more effective surgical removal or radiation therapy, but often lead to tumor
relapse. According to the “cancer stem cell hypothesis”, a small population of tumor
cells, thought to be cancer stem cells (CSCs), maintains the malignancy of the tumor
through self-renewal processes. CSCs proliferation and differentiation is largely
unaffected by conventional anticancer drugs, and CSCs are thought to be drugresistant. Therefore, targeting CSCs represents a new promising anticancer strategy
and small molecule targeting selectively the CSCs niche are in high demand.
A critical survey of the literature covering the year 2014 in the field of “targeting
CSCs” is presented. Overall, this is a growing and stimulating field of research
involving both biologists and chemists from academia and industry. While significant
progresses have been recorded especially from biological studies (i.e. role of CSCs in
tumor progression, characterization of CSCs niches in several types of tumors,
modulation of CSCs by small molecules), less advances have been provided from
chemical investigations. Indeed, very few preclinical candidates have been developed
and most researches are still carried out with old drugs or small molecules for which
the development up to drug candidates may be truly difficult. A big effort is required to
chemists and medicinal chemists for discovering new molecular entities targeting
selectively CSCs and develop them up to the drug market.
Besides this literature survey, recent successes achieved in targeting the
STAT3 transcription factor will be reported, and an overview of pitfalls and
opportunities for targeting the Hedgehog pathway will be given.
References
[1]
[2]
[3]
[4]
So JY et al, A Synthetic Triterpenoid CDDO-Im Inhibits Tumorsphere Formation by
Regulating Stem Cell Signaling Pathways in Triple-Negative Breast Cancer. PLoS ONE
9(9): e107616 (2014).
Tang SC et al, Novel therapeutic targets for pancreatic cancer. World J Gastroenterol
20(31): 10825-10844 (2014)
Arasada RR et al, EGFR Blockade Enriches for Lung Cancer Stem–like Cells through
Notch3-Dependent Signaling. Cancer Res: 74(19) (2014)
Shanshan Wan et al, Tumor-Associated Macrophages Produce Interleukin 6 and Signal
via STAT3 to Promote Expansion of Human Hepatocellular Carcinoma Stem Cells.
Gastroenterology, doi: 10.1053/j.gastro.2014.08.039 (2014).
23
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L17. Evaluating the Mechanisms of New Anticancer Agents in Multi-Drug
Resistant Cancer Cells
Jasna Banković, Ana Podolski-Renić, Jelena Dinić, Tijana Stanković, Zorica
Milošević, Sonja Stojković, Miodrag Dragoj and Milica Pešić
University of Belgrade, Institute for Biological Research „Siniša Stanković“, Despota Stefana
142, 11060 Belgrade, Serbia
In vitro models of multi-drug resistance (MDR) are valuable tools for preclinical
testing of new anticancer drugs. We have developed three human MDR cancer cell
lines (non-small cell lung carcinoma NCI-H460/R, colorectal carcinoma DLD1-TxR, and
glioma U87-TxR) with unique molecular and chromosomal characteristics. These MDR
cancer cells showed chromosomal numerical and structural changes, differences in
tumor suppressors’ gene settings, overexpression of membrane efflux pumps and
changes in the expression of stemness transcription factors.
We have employed them to evaluate different approaches in overcoming MDR.
To that end, we tested P-glycoprotein (P-gp) inhibitors (jatrophane diterpenoids) and
prooxidant agents that are more active against P-gp overexpressing cells
(protoflavones). In addition, we used our MDR models to study the mechanisms of
microtubule destabilizing agent (DTA0100).
Besides considerably high P-gp inhibiting activity, jatrophane diterpenoids
showed significant potential to reverse the paclitaxel resistance in all MDR cancer cell
lines. The combinations of jatrophane diterpenoids with low paclitaxel concentrations
induced the cell cycle disturbance, which additionally contributes to their chemosensitizing activity.
We found that MDR cancer cells are more vulnerable to protoflavones
prooxidative activity. This is due to the low antioxidant capacity of P-gp overexpressing
cells. Therefore, protoflavones evade MDR and selectively act against P-gp
overexpressing cells by using their low antioxidant capacity as a collateral mechanism.
Microtubule interacting agent DTA0100 was studied in MDR models developed
by continuous treatment with paclitaxel. We found that differences in the expression
pattern of β tubulin isotypes might confer to DTA0100 sensitivity towards MDR cancer
cells.
Herein, we emphasize the importance of in vitro MDR cancer models as a first
step in the evaluation of new anticancer drugs. These models are essential for the
identification of new lead compounds that are able to preserve cytotoxic activity toward
MDR cancer cells or to restore the cytotoxicity of classic anticancer drugs.
24
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L18. Natural Products as Cancer Chemopreventive Agents: a Focus on
Multiple Myeloma Cancer Stem Cells
Muriel Cuendet
School of pharmaceutical sciences, University of Geneva, University of Lausanne, 30 Quai
Ernest-Ansermet, 1211 Geneva 4, Switzerland
As established by ample precedent, nature provides broad chemical diversity
and compounds isolated from plants are essential in cancer chemotherapy. Most
human cancers seem to be potentially preventable because of controllable or
removable causative exogenous factors (primary chemoprevention), but also by agents
interfering with carcinogenesis. These compounds can be divided into three categories:
blocking agents (anti-initiation), anti-promotion agents, and anti-progression agents.
Recent research has shown that stem cells may serve an important role in tumor
initiation and progression. Consistent with this notion, the removal of both therapysensitive tumor cells constituting the bulk of the tumor mass and cancer stem cells may
be necessary to achieve a chemopreventive response.
Despite advances in the development of novel therapies, multiple myeloma
(MM) remains an incurable malignancy, where the majority of patients relapses,
develops resistance and eventually dies from the disease. This may be attributed to the
fact that conventional therapies mainly target the bulk of tumor cells, but not the tumorinitiating cancer stem cells (CSCs). Thus, tumors composed of heterogeneous cell
populations, CSCs and bulk cells, may benefit from combination drug regimens that
target each population; however, compounds that target MM-CSCs remain largely
unknown. Such compounds are urgently needed as they may significantly improve the
prognosis of MM patients. We identified a compound which induced a significant
regression of tumors in a xenograft model using RPMI 8226 cells. In further studies,
this compound inhibited the growth and proliferation of MM-CSCs, induced apoptosis
and differentially regulated genes involved in cell maintenance in vitro.
25
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L19. Two Novel Cancer Stem Cell Agents and a Putative CSC Platform for
Drug Evaluation
Richard Clarkson
European Cancer Stem Cell Research Institute, Cardiff, United Kingdom
This presentation is divided into two parts. Initially I will describe our progress in
the development of a novel anti-metastatic agent targeting Bcl3, which has now been
verified to provide better than 90% long-term survival in mouse models of metastatic
breast cancer and the advent of a second pharmacological agent that both sensitises
cancer stem cells to the cytotoxic agent TRAIL, and furthermore appears to prevent the
acquisition of CSC-like properties from non-CSC tumour cells.
In the second half of the talk I will present ongoing collaborative work to develop
two CSC viability assay platforms that are based on the CSC properties of selfrenewal/asymmetric division and anoikis resistance of tumour initiation respectively.
The prospect of these assays for medium-scale drug screens will be discussed.
26
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L20. Ubiquitin Hydrolases as Novel Regulators of DNA Replication
Raimundo Freire
Unidad de Investigación, Hospital Universitario de Canarias, Instituto de Tecnologías
Biomédicas, Ofra s/n, La Laguna, Tenerife, Spain
The DNA damage checkpoint, as well as a correct regulation of DNA replication
during the cell cycle, is essential for the maintenance of genome integrity in
eukaryotes. Claspin plays a key role in the ATR-Chk1 branch of the DNA damage
checkpoint and is also required for accurate DNA replication. Cdt1 and Geminin are
essential players in the control of the initiation of DNA replication and Cdt1 is
additionally involved in delaying DNA replication after genotoxic stress. To achieve
these functions, Claspin, Cdt1 and Geminin are tightly regulated by ubiquitindependent proteasomal degradation. In our laboratory we have identified new
regulators of these three proteins among the ubiquitin hydrolases. In the case of
Claspin we found that the ubiquitin-specific peptidase 29, USP29 is able to interact with
Claspin. Downregulation of USP29 destabilizes Claspin, while its overexpression
increases Claspin protein levels. USP29 knockdown results in an impaired
phosphorylation of Chk1 after DNA damage and USP29-depleted cells show a big
defect in S phase progression. Equivalent results were obtained with new de-ubiquitin
enzymes that regulate Cdt1 and Geminin and will be presented here.
27
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 L21. Evaluation of Chk1 as a Target for Anti-Cancer Therapy in Vivo
David Guillespie
Institute of Biomedical Technologies, Centre for Biomedical Research of the Canary Islands,
Universidad de La Laguna, Tenerife
Chk1 is a key regulator of DNA damage checkpoint responses and genome
stability in eukaryotes. In recent years Chk1 inhibitors have been developed as
potential anti-cancer agents, however exactly how such agents should be deployed
remains unclear. To gain insight into this issue we investigated the effect of genetic
ablation of Chk1 in murine skin on tumor formation in response to chemical
carcinogens and activated oncogenes. We find that complete loss of Chk1 function in
keratinocytes strongly suppresses epithelial tumorigenesis in response to chemical
carcinogens without grossly disturbing tissue structure or function, most likely by
transiently purging the stem cells of origin from the skin as a result of spontaneous
DNA damage and cell death. We also find that Chk1 ablation in the melanocytic
lineage leads to the death of both normal melanocytes and melanoma cells due to
accumulation of spontaneous DNA damage during DNA replication. Taken together,
these data argue that Chk1 is likely to be inherently essential for the proliferation and
survival of at least some tumor types, such as melanoma, and for a subset of cells
within normal tissues that have the potential to undergo malignant transformation. An
important question now is whether these potent anti-cancer effects can be reproduced
using pharmacological inhibitors of Chk1.
28
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 POSTER SESSIONS
29
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P1. Postserone 2,3-Dioxolanes Have Improved Activity Sensitizing MCF7
and MDR Mouse Lymphoma Cells Towards Doxorubicin
Ana Martins1,2, J. Csabi3, L. Amaral4, J. Molnar1, Attila Hunyadi1
1
Department of Medical Microbiology and Immunobiology, Faculty of Medicine, University of
Szeged, 6720, Szeged, Hungary
2
Unidade de Parasitologia e Microbiologia Médica, Instituto de Higiene e Medicina Tropical,
Universidade Nova de Lisboa, Lisboa 1349-008, Portugal
3
Institute of Pharmacognosy, Faculty of Pharmacy, University of Szeged, 6720, Szeged,
Hungary
4
Center for Malaria and Other Tropical Diseases (CMDT), Instituto de Higiene e Medicina
Tropical, Universidade Nova de Lisboa, Lisboa 1349-008, Portugal
Ecdysteroids are plant-derived hydroxysteroids expressing a 7-en-6-one
moiety in their B-ring, that show some structural similarities to calcitriol, one of the
active forms of vitamin D, by means of their side-chain, typically trans C/D ring junction
and partly to their hydroxylation pattern [1]. The most abundant phytoecdysteroid is 20hydroxyecdysone (20E) [2]. The main difference between 20E and postserone is the
presence or absence of the side chain, respectively.
20E was shown to be non-toxic to mammals (oral LD50 in mice > 6g/kg) and to
exert a wide range of beneficial pharmacological effects (adaptogenic, anabolic,
antihyperglycemic, etc.) [3]. Due to their structural differences from vertebrate steroid
hormones, ecdysteroids do not interact with the vertebrate steroid-hormone system.
Their supposedly safe, non-hormonal anabolic activity lead to the worldwide marketing
of a large number of ecdysteroid-containing food supplements.
Previously we found that some ecdysteroids increased the activity of doxorubicin
against an MDR mouse lymphoma cell line over-expressing the human ABCB1 [4,5].
The observed structure activity relationships (SARs) highlighted the importance of
apolar groups at positions 2,3 [5]. Therefore, complementing the information obtained
for the 2,3 and 2,3;20,22-dioxolane substituted derivatives of 20E, new 2,3 derivatives
were synthesised from ponasterone. The new compounds were not cytotoxic against
the studied cell lines (IC50 > 150μM). However they highly improved the activity of
doxorubicin against both MDR mouse lymphoma cells (expresses the human ABCB1
transporter) and MCF7 cells (do not express the ABCB1 transporter) when used at 50
μM. These findings confirm our previous observations on the importance of apolar
substituents at position 2,3 and show that absence of the side chain can improve
activity as compared with the corresponding analogs derived from the 20E.
Acknowledgments: The authors acknowledge the Szeged Foundation for Cancer Research;
the European Union and the European Social Fund: TÁMOP 4.2.2.A-11/1/KONV-2012-0035;
and the Fundação para a Ciência e a Tecnologia: PEst-OE/SAU/UI0074/2011, PEstOE/SAU/UI0074/2014, SFRH/BPD/81118/2011.
References
[1]
[2]
[3]
[4]
[5]
Toth N, et al. Curr Med Chem 17(18): 1974-1994 (2010).
Lafont R and Dinan L. J Insect Sci 3(7): 1-30 (2003).
Dinan L and Lafont R. J Endocrinology 191(1): 1-8 (2006)
Martins A, et al. J Med Chem 55 (11): 5034–5043 (2012).
Martins A, et al. Molecules 18(12): 15255-15275 (2013).
30
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P2. Target Cancer Therapy by Notch and WNT Inhibitors - Current
Approach and Future Prospective
Sanja Jovevska1, Ana Mitanoska2, Darko Bosnakovski1
1
Faculty of Medical Sciences University, 2Faculty of Natural and Technical Sciences Goce
Delcev, Stip, 2000, R. Macedonia
Notch signaling pathway is important for communication between cells, which
includes gene regulatory mechanisms that control cellular processes during embryonic
life. It takes part in neuronal development and function, stabilization of arterial
endothelial and angiogenesis, communication between endocardium and myocardium
during ventricular development and differentiation. Notch signaling pathway is involved
in growth of some cancer cells, migration, invasion and angiogenesis. Its contribution to
carcinogenesis is through inhibition of differentiation, inhibition of apoptosis and
stimulation of proliferation.
WNT signaling plays a central role in embryonic development, tissue
regeneration, protein phosphorylation, osteoblast differentiation, signal transduction. Its
involvement in carcinogenesis results in increased growth of tumor cells, the creation of
metastasis and cell migration.
Inhibitors of Notch and WNT fall into these categories: small molecule drugs,
peptides and blocking antibodies. Here we will present currently exploited molecules
that target Notch and WNT signaling pathways and we will discuss future prospective.
31
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P3. Influence of MDR cancer phenotype on the efficacy of novel tubulin
destabilizing agent DTA0100
Ana Podolski-Renić1, Jelena Dinić1, Jasna Banković1, Zorica Milošević1, Carla RíosLuci2, José M. Padrón2, Milica Pešić1
1
Institute for Biological Research „Siniša Stanković“, University of Belgrade, Despota Stefana
142, 11060 Belgrade, Serbia
2
BioLab, Instituto Universitario de Bio-Organica "Antonio Gonzalez", Centro de Investigaciones
Biomédicas de Canarias (CIBICAN), Universidad de La Laguna, Avda. Astrofisico Francisco
Sanchez 2, 38206 La Laguna, Tenerife, Spain
Microtubule targeting agents that disrupt microtubule/tubulin dynamics are
widely used in cancer chemotherapy. However, development of drug resistance limits
their effectiveness in cancer treatment. Therefore, it is important to determine possible
drug resistance mechanism related to the application of anti-mitotic cancer therapy
(changes in microtubule dynamics and overexpression of efflux transporters).
In the present study we evaluated the effectiveness of propargylic enol ether
derivative DTA0100, a microtubule destabilizing agent on multi-drug resistant (MDR)
cancer cell lines. To that end, we employed two different MDR cancer cell lines with Pglycoprotein (P-gp) overexpression and their sensitive counterparts (colorectal
carcinoma DLD1-TxR and DLD1, glioblastoma U87-TxR and U87). Both MDR cancer
cell lines demonstrated resistance to various microtubule targeting agents including
paclitaxel, vinblastine and colchicine which are also substrates for P-gp. Tested
compound DTA0100 showed significant cytotoxicity against colon cancer cell lines
reaching IC50 values in nanomolar range. Importantly, DTA0100 was 2-fold more
efficient towards MDR colon cancer cells. However, its efficacy against U87-TxR cells
was reduced in comparison to their sensitive counterparts. DTA0100 treatment caused
microtubule disruption in all cell lines as verified by losing typical long and dense
microtubule network. AnnexinV/propidium iodide staining revealed that DTA0100
increased a portion of apoptotic cells after 72 h treatment in U87 and DLD1-TxR cells.
However, the changes were more prominent in glioblastoma cells. Cell cycle analysis
revealed that both glioblastoma cell lines undergo G2/M cell cycle block following
DTA0100 treatment. At the mRNA level, we have assessed the changes between
sensitive and MDR cell lines in the expression of I, II, III and IVb -  tubulin isotypes.
The expression of II, III and IVb -  tubulin was increased in U87-TxR cells, while the
class III was decreased in DLD-TxR.
The observed changes in the expression pattern of  tubulin isotypes between
MDR glioblastoma and MDR colon cancer cells may confer to their different sensitivity
to DTA0100. Further studies will be conducted in order to reveal whether mutational
status of class I  tubulin in MDR cancer cells may influence DTA0100 efficacy. In
addition, we will compare affinity of P-gp to colchicine and DTA0100 in given MDR in
vitro models.
Acknowledgements: Cofinanced by the EU Research Potential (FP7-REGPOT-2012-CT201231637-IMBRAIN), the European Regional Development Fund (FEDER), and the Spanish
Instituto de Salud Carlos III (PI11/00840).
32
COST CM1106 & CIBICAN Workshop P4. Multilevel Targeting
Pyrrolidinedione AD0157
October 14‐15, 2014 for
Leukemia
Therapy
by
the
Marine
Melisa García-Caballero, Miguel Ángel Medina, Ana R. Quesada
Department of Molecular Biology and Biochemistry, Faculty of Sciences, University of Málaga,
and Unit 741 of CIBER de Enfermedades Raras (CIBERER), Faculty of Sciences. University of
Málaga, Campus de Teatinos, E-29071 Málaga, Spain
AD0157, a natural pyrrolidinedione isolated from the fermentation broth of the
marine fungus Paraconiothyrium sp. HL-78-gCHSP3-B005, has been previously shown
to exhibit strong antiangiogenic activities. Although the mechanism of action of
AD0157 has not been fully characterized, a direct inhibition of the receptors of vascular
endothelial growth factors (VEGF) could contribute to the observed inhibition of
angiogenesis exerted by this compound. VEGF receptors are also expressed by tumor
and cancer stem cells, what explains their functional relevance not only for tumor
angiogenesis, but also for tumor initiation and growth and provides an important
opportunity for the development of new therapeutic approaches, especially for highly
aggressive tumours.
In this communication, evidence of the growth inhibitory activity of AD0157 on
human HL60 promyelocytic leukemia cells, U937 histiocytic lymphoma cells and in
KU812F basophilic leukemia cells will be presented. Antileukemic activity of this
compound is exerted through the induction of apoptosis, reflected in an induction of
chromatin condensation, DNA fragmentation, and an increase in the subG1 cell
population. AD0157 promotes the translocation of phosphatidylserine from the inner
leaflet of the phospholipid bilayer to the cell surface and decreases the mitochondrial
membrane potential. Our data indicate that AD0157 interferes with the leukemia cell
survival and proliferation through the PI3K/Akt and MAPK/ERK pathways, inducing
apoptosis in human leukemic cells through both the extrinsic and the intrinsic pathway.
These data reinforce the potential pharmacological interest of AD0157 as a
drug candidate for the treatment of leukemia, based on a multilevel targeting on several
cells of the leukemia microenvironment.
Acknowledgements: Supported by grant PIE P12-CTS-1507 (Andalusian Government and
“Fondo Europeo de Desarrollo Regional” (FEDER)). The “Centros de Investigación Biomédica
en Red” or CIBER de Enfermedades Raras” is an initiative from the “Instituto de Salud Carlos
III” (ISCIII) (Spain).
33
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P5. Cytotoxic Activity of a New Class of Spirooxindoles in Cancer Stem
Cells
Ângelo Monteiro1, Giovanna Damia2, Francesca Ricci2, Daniele Passarella3, Maria M.
M. Santos1
1
The Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of
Lisbon. Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal
2
Department of Chemistry, University of Milan. Via Golgi 19, 20133 Milan, Italy
3
Department of Oncology, Mario Negri Institute for Pharmacological Research. Via Giuseppe
La Masa 19, 20156 Milan, Italy
Cancer remains one of the greatest medical concerns and its burden is set to
increase with the increase in an aging population. Despite enormous advances in the
area, World Health Organization projects a rise in deaths from cancer to 13.1 million in
2030 [1]. As such, cancer continues to pose a major threat to human health and further
research regarding new therapeutic strategies that more effectively combat cancer are
needed.
In the last 20 years, has re-emerged the cancer stem cells (CSCs) hypothesis
as a valid concept in fighting cancer [2]. The CSCs are a rare cell population known as
immortal tumor-initiating cells, since they can self-renew and generate tumor cells with
different phenotypes, a pluripotent capacity. Due to their astonishing characteristics, it’s
though that CSCs are the basis of tumor initiation, development, metastases and
recurrence [3].
Recently, Santos’s research group reported the potential use of a new class of
spirooxindoles as potential anticancer agents. The compounds were evaluated for their
cytotoxic activity in two breast cancer cell lines. Most of them showed good activities
against MCF-7 cell line (GI50 up to 7 µM), and had low cytotoxic activity against HEK
293T non tumor derived cell line. In addition, two compounds were more selective for
MCF-7 cell line (ER-positive) than for MDA-MB-231 tumor cell line (ER-negative) [4].
Here, we report the evaluation of the most promising compounds (GI50 30 µM)
in two ovarian enriched cancer stem cell lines (#83 and #110) [5] and one ovarian
cancer cell line (OVCAR5). In a total of ten compounds, eight of them had good activity
against the two cancer stem enriched cell lines (GI50 < 20 µM). Three compounds had
around 2-fold selectivity for stem cell lines over MCF-7 cell line. Six of eight active
compounds were also evaluated in differentiated cells derived from #83 and #110 cell
lines in order to compare with the results obtained in the stem cell enriched cell lines.
Acknowledgements: This study was supported by FCT (Fundação para a Ciência e a
Tecnologia, Portugal) through grants PTDC/QUI-QUI/111664/2009, PEst-OE/SAU/UI4013/2014
and Cost action CM1106.
References
[1]
[2]
[3]
[4]
[5]
Siegel R., Naishadham D., Jemal A., CA Cancer J. Clin., 63, 11–30 (2013).
O’Connor M. L., et al., Cancer Letters, 344, 180-187 (2014).
Chen K., Huang Y., Chen J., Acta Pharm. Sin., 34, 732–740 (2013).
Monteiro A., Gonçalves L., Santos M.M.M., Eur. J. Med. Chem., 79, 266-272 (2014).
Ricci F., Bernasconi S., Perego P., Ganzinelli M., Russo G., Bono F., Mangioni C.,
Fruscio R., Signorelli M., Broggini M., Damia G., Cell Cycle, 11, 1966-1976 (2012).
34
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P6. Identification of Potent c-Src Inhibitors Affecting Brain Tumors in Vivo
Botta Maurizio1, Anna Lucia Fallacara1, Cristina Tintori1, Giulia Vignaroli1, Claudio
Zamperini1, Emmanuele Crespan2, Giovanni Maga2, Adriano Angelucci3, Silvia
Schenone4, André Richters5, Lucia Dello Iacono1, Daniel Rauh5
1
Università degli studi di Siena, Italy
2
IGM-CNR Pavia, Italy
3
Università dell’Aquila, Italy
4
Università degli Studi di Genova, Italy
5
TU Dortmund, Germany
Neuroblastoma (NB) and Glioblastoma Multiforme (GMB) are the most frequent
solid malignancies of nervous system. In particular, NB is the most common extracranial solid cancer in childhood and the most common cancer in infancy, while GMB is
an aggressive primary brain tumor, with an extremely poor prognosis (Wen and Kesari,
2008).
The tyrosine kinase c-Src has been reported to play an important role in the
differentiation, cell adhesion and survival of NB cells. Accordingly, our patented c-Src
inhibitors characterized by a pyrazolo[3,4-d]pyrimidine (Pyr-Pyr) scaffold have been
studied as anticancer agents against NB.
Pyr-Pyr against neuroblastoma. Few years ago our team started a research
in the field of TKs inhibitors which led to the synthesis of new Pyr-Pyr derivatives,
structurally related to the c-Src inhibitors PP1 and PP2, but bearing a different
substitution pattern on the heterocyclic scaffold. Selected members of this family
displayed antiproliferative activity, led to cell cycle arrest, induced apoptosis, decreased
adhesion and invasiveness and reduced Src phosphorylation in SH-SY5Y cell cultures
of human neuroblastoma with IC50 ranging from 0.8 to 0.08 µM.
Neuroblastoma. These compounds showed in vivo activity reducing the weight
of tumor mass in mouse models. Treatment with the most active compound 1 (50
mg/Kg for 60 days) led to a 50% growth reduction of the tumor weight.
Glioblastoma. Compound 1 was administered in vivo to nude mice inoculated
subcutaneously with U87 cells. Mice received 50 mg/kg of 1 for 60 days. The
antitumoral effect of the compound was also evaluated in combination with a single
radiotherapic treatment (4Gy). At the end point mice that have received the
combination therapy showed the smallest tumors respect to other experimental groups
(< 80% respect to untreated group).
References
[1]
[2]
[3]
Radi M, Brullo C, Crespan E et al. Bioorg Med Chem Lett. 21(19):5928-33 (2011).
Navarra M, Celano M, Maiuolo J et al. BMC Cancer. 10:602 (2010).
Schenone, S; Bondavalli, F.; Bruno, O.; Botta, M. et al WO 2009034547 (2009).
35
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P7. An Overview on our On-Going Research and Collaboration Network
Against MDR Cancer
Attila Hunyadi1, Ana Martins2,3
1
Institute of Pharmacognosy, Faculty of Pharmacy, University of Szeged, 6720, Szeged,
Hungary
2
Department of Medical Microbiology and Immunobiology, Faculty of Medicine, University of
Szeged, 6720, Szeged, Hungary
3
Unidade de Parasitologia e Microbiologia Médica, Instituto de Higiene e Medicina Tropical,
Universidade Nova de Lisboa, Lisboa 1349-008, Portugal
Phytoecdysteroids, herbal analogs of the insect molting hormone, as well as
protoflavonoids, rare natural flavonoids bearing a non-aromatic B-ring, have been identified by
our group as potentially useful chemical tools for targeting efflux-mediated multi-drug resistance
(MDR) in cancer [1,2]. Briefly, certain natural and semi-synthetic ecdysteroids were found to be
able to sensitize MDR cancer cells to a variety of chemotherapeutics, and this activity does not
appear to be a result of direct ABC transporter inhibition [1] Protoflavonoids, on the other hand,
exert selective cytotoxic activity [2] on several MDR cancer cell lines, which, as a result of their
adaptation process towards chemotherapeutics, have apparently become more sensitive to
oxidative stress as compared to their parental cells.
The presentation aims to give a brief overview on our current research directions on
these two promising compound groups, including our chemical approaches to obtain further
derivatives for bioactivity testing. The on-going collaborations and key findings, within or outside
of the framework of COST Action CM1106, are also presented, in order to facilitate discussion
towards possible new initiatives.
Acknowledgements: The authors acknowledge the Szeged Foundation for Cancer Research;
the European Union and the European Social Fund: TÁMOP 4.2.2.A-11/1/KONV-2012-0035;
and the Fundação para a Ciência e a Tecnologia: PEst-OE/SAU/UI0074/2011, PEstOE/SAU/UI0074/2014, SFRH/BPD/81118/2011.
References
[1]
[2]
Martins A, et al. J Med Chem 55 (11): 5034–5043 (2012).
Dankó B et al. Anticancer Res 32: 2863-2870 (2012).
36
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P8. Ex Vivo Precision Cut Tissue Slices in the Assessment of Anticancer
Drug Toxicity
Brech Aikman, I.A.M de Graaf, M.H. de Jager, B.N. Melgert, G. M.M. Groothuis,
Angela Casini
Division of Pharmacokinetics, Toxicology and Targeting, Groningen Research Institute of
Pharmacy, University of Groningen, A. Deusinglaan 1, 9713 AV, The Netherlands
Traditional pre-clinical in vitro models used in studies regarding metabolism,
targeting or toxicity are often limited in representing the complex and dynamic
homeostasis/interplay that exists in vivo. Static models fail to take the heterogeneity
that exists in pathogens/cancer in account, making predictions concerning potential
novel drugs possibly inaccurate. Recent potential new anticancer drugs selected only
based on classical in vitro screening have shown to be lacking in efficacy, thus
emphasizing the need for different methods [1].
Within this frame, the so called precision-cut tissue slice (PCTS) is an
investigational model that closely resembles the organ of origin in both configuration
and homeostasis [2]. These tissue samples can be obtained from various organs such
as the liver, kidney, etc. PCTS are already applied as ex vivo models to assess drug
activity, toxicity, metabolism and transport. Maintaining the original tissue configuration
preserves the organ specific interactions between different tissues and cell types,
enabling more accurate metabolism and function studies of both endogenous and
exogenous substrates in healthy and pathogenic tissue.2
The aim of our study was the investigation of the mechanisms of toxicity of a
new experimental anticancer metallodrug in rat precision cut liver slices. Thus, we
present here the obtained results that are also aimed at optimizing the experimental
conditions for anticancer drug testing. Overall, our results show that PCTS is a valuable
tool not only in the discovery and development of novel drugs, but also in research
aimed at contributing to our understanding of already registered drugs, focussing on
anticancer metallodrugs. For this purpose, we also used the PCTS technology in the
toxicity evaluation of cytotoxic metal compounds such as cisplatin [3] in healthy rat liver
samples to predict possible side effects.
Acknowledgements: COST Action CM1106 is gratefully acknowledged for financial support
and useful discussion.
References
[1]
[2]
[3]
Hickman JA, Graeser R, De Hoogt R. Three-dimensional models of cancer for
pharmacology and cancer cell biology: Capturing tumor complexity in vitro/ex vivo.
Biotechnol. J. 9: 1115-1128 (2014).
De Graaf IA, Olinga P, de Jager MH, et al. Preparation and incubation of precision-cut
liver and intestinal slices for application in drug metabolism and toxicity studies. Nature
Protocols 5(9):1540-1551 (2010).
Wheate, N J, Walker, S, Craig, G E, et al. The status of platinum anticancer drugs in the
clinic and in clinical trials. Dalton transactions 39(35): 8113-8127 (2010).
37
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P9. Novel Macrocyclic Glycuronides: Synthesis and Investigation of Their
Potential as Anti-Tumour Agents
Rekha Chadda, Paul V. Murphy
School of Chemistry, National University of Ireland
Tumour metastasis is responsible the majority of cancer deaths and
development of inhibitors is considered important [1]. In addition cancer stem cells are
highly metastatic.
Preliminary results have shown macrolactam glucuronide 2 is an inhibitor of
angiogenesis in a zebrafish model and an inhibitor of breast tumour cell migration. This
compound could be considered to be structurally related to isomigrastatin 3 and quinic
acid derivative 1 prepared at the NIH, both of which showed inhibition of tumour cell
migration. For this reason a range of simpler analogues based on glucuronic acid have
been prepared for testing as inhibitors of tumour metastasis. In some cases chelation
induced anomerisation was utilised to generate the macrocyclic compounds. The
syntheses will be presented.
References
[1]
Weigelt, B.; Peterse, J. L.; Veer, J. van’t, Nature Rev. Cancer, 5, 591-602 (2005).
38
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P10. Importance of 3D Culture in Testing CSC Lines for Apoptotic
Responses
Päivi Järvinen, Christian D Muller
University of Strasbourg, UMR 7200 CNRS, Illkirch, France
Introduction. Drug discovery models for studying anti-cancer properties of
compounds have traditionally been conducted on two-dimensional (2D) cell cultures,
which present significant limitations in reproducing the complexity and pathophysiology
of in vivo tumor tissue. Growing on artificial plastic surfaces, cells lose their abilities to
differentiate, polarize, and interact with each other (cell-cell interactions) but also with
the surrounding extra cellular matrix (ECM). Cells proliferate fast and form a
homogeneous population, and thus fail to mimic the in vivo conditions. Limitations in
methodologies have slowed down the process of finding new drugs. New techniques
using three-dimensional (3D) cell cultures more accurately reflect the complex in vivo
organotypic growth of cancer cells and the microenvironment, which is shown, e.g., in
gene expression profiles, signaling pathway activities, and drug sensitivities.
Methods. In 3D cell culture set-ups, cancer cells and cancer stem cells were
grown on 96-well plates with ultra-low binding surface and U-shape, which promote the
formation of a single cell aggregate i.e. spheroid per well. The spheroids were grown
for four days, then, half of the medium was replaced with sample dissolved in medium.
In this project, natural extracts were studied. On day four the diameter of the spheroids
was around 400 µm. On day seven the spheroids were dissociated using TrypsinEDTA solution and the apoptotic responses were measured by capillary flow cytometer
(Guava, Millipore) using Annexin V and Propidium iodide as the fluorescent markers.
Large spheroids have differences on cell types depending on the location. The core of
the spheroid starts to form hypoxic conditions when the diameter exceeds 500 µm, but
also availability of nutrients decreases. These gradients affect cells, especially their
proliferation rates, which also affect the drug activities. During the treatment the
spheroid diameter exceeded 500 µm. In comparison, the apoptotic effects were also
studied in traditional 2D cell cultures.
Results. The apoptotic responses were studied using four cell lines, NTERA-2
and MCF-7 grown on 2D and 3D, and THP-1 and BXPC-3 grown on 2D conditions.
NTERA-2, MCF-7 and BXPC-3 are considered as CSC as they express the OCT-4
factor. THP-1 is a non-adherent monocytic cell line. As pancreatic BXPC-3 cells
formed spheroids in 3D with irregular forms and non-linear growth rates, only in 2D
cultures were used for screening. Six fungi excretion natural extracts were evaluated
for their potential to induce apoptosis. Two of the extracts were active on all cell lines in
both 2D and 3D conditions, although, the EC50 values varied from 27 to 160 µg/ml.
Three of the extracts were active on 2D culture only. Interestingly, one of the 6 extracts
was more active in 3D culture conditions than 2D. In drug discovery pipeline, the
primary screening of compounds is most often evaluated in 2D cultures only. Our
results reveal that one of the active extracts in 3D conditions might have been
interpreted as false negative if discrimination of samples was done according to results
obtained in 2D. Further, three of the extracts would have been interpreted as false
positives, because they did not induce apoptosis in 3D culture conditions. Also, there
were differences between cell lines of different organ origin. In conclusion, our results
shows that primary screening of anti-cancer compounds should be conducted not only
using a set of cell lines of different origins but as well using 2D and 3D culture
conditions to improve the predictive value of the screening.
39
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P11. Self-Assembled Squalene-Based Fluorescent Hetero-Nanoparticles
Michael S. Christodoulou1, Gaia Fumagalli1, Panagiota A. Sotiropoulou2, Franco
Dosio3, Davide Mazza4, Daniele Passarella1
1
Dipartimento di Chimica, Università degli Studi di Milano, Via Golgi 19, 20133 Milano, Italy
2
IRIBHM, Université Libre de Bruxelles, Brussels, Belgium
3
Dipartimento di Scienza e Tecnologia del Farmaco, Via Giuria 9, 10125 Torino, Italy
4
Centro di Imaging Sperimentale Ospedale San Raffaele Scientific Institute, Via Olgettina 58,
20133 Milano, Italy
Cancer stem cells (CSCs) are a subpopulation of cancer cells with high
clonogenic capacity and ability to reform parental tumours upon transplantation.
Resistance to therapy has been shown for several types of CSC and, therefore, they
have been proposed as the cause of tumour relapse. Consequently, much effort has
been made to design molecules that can target CSCs specifically and sensitize them to
therapy [1].
The recent advances in nanotechnology and nanomaterials have been
integrated into analytical chemistry for the design of large numbers of fluorescent
chemical and biological probes. In particular, nanoparticles have some advantages
because they are much brighter than the single dyes since one particle contains
several dyes molecules and their molecular size minimizes physical perturbation of
living cells. Our continuous interest in the field of chemical approaches to target cancer
cells moved us to study the preparation of a novel class of squalene conjugates with
paclitaxel, podophyllotoxin, camptothecin and epothilone A. All of them were
characterized by a squalene tail that makes them able to self-assemble in water, and to
secure the release inside the cells by a disulfide-containing linker [2]. The need to trace
the delivery of the nanoassemblies and to demonstrate the internalization of the drugs
pushed us toward the formation of heterogeneous fluorescent nanoassemblies by
mixing a paclitaxel-squalene conjugate and fluorescein-squalene conjugate.
The following application of hetero-nanoparticles is in the combined therapy.
Mixing a paclitaxel-squalene conjugate with a squalene derivative of an inhibitor of the
hedgehog pathway such as cyclopamine [3] we were able to obtain again heteronanoparticles. The preparation of fluorescent hetero-nanoparticles containing a
paclitaxel-squalene
conjugate,
a
cyclopamine-squalene
conjugate
and
tetramethylrhodamine-sqalene conjugate serves to demonstrate the internalization of
the nanoassemblies.
References
[1]
[2]
[3]
P. A. Sotiropoulou, M. S. Christodoulou, A. Silvani, C. Herold-Mende, D. Passarella Drug
Discov. Today, DOI: 10.1016/j.drudis.2014.05.002 (2014).
S. Borrelli, M. S. Christodoulou, I. Ficarra, A. Silvani, G. Cappelletti, D. Cartelli, G. Damia,
F. Ricci, M. Zucchetti, F. Dosio, D. Passarella Eur. J. Med. Chem. 85, 179-190 (2014).
P. Heretsch, L. Tzagkaroulaki, A. Giannis Angew. Chem. Int. Ed. 49, 3418-3427 (2010).
40
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P12. Inhibition of MDM2-p53 Interaction by Novel Compounds as
Therapeutics of Hepatocellular Carcinoma
Damla Gozen1, Maria M. M. Santos1,2, Ângelo Monteiro2, Rengul Cetin-Atala1,3
1
Bilkent University, Department of Molecular Biology and Genetics, Ankara, Turkey
2
Research Institute for Medicines, Universidade de Lisboa, Lisbon, Portugal
3
Informatics Institute, Middle East Technical University, Ankara, Turkey
Hepatocellular carcinoma (HCC) is the sixth most commonly diagnosed cancer
and second most common cause of cancer related death worldwide [1]. Although there
are several treatment strategies, among which chemotherapy is the major one, their
efficacies are low due to drug resistance. Therefore, more studies focusing on the
development of novel therapeutics of HCC with increased effectiveness and decreased
side effects are needed. The tumor suppressor protein, p53 is one of the major
regulators of cell cycle; mutations of which are involved frequently in the development
of HCC [2]. One of the most well known regulators of p53 activity is Mdm2, which is
known to be overexpressed in many cancer cells leading to the protosomal degradation
of p53 [3]. Thus, inhibition of p53-Mdm2 interaction might be a promising therapeutic
strategy for HCC. The aim of this study is to determine anticancer activities of newly
synthesized potential inhibitors of p53-Mdm2 interaction on HCC cell lines and define
the underlying mechanisms. The cytotoxic effects of the inhibitors were initially tested
by SRB assay on HCC cell lines carrying various p53 mutations. In order to determine
the underlying mechanisms behind these cytotoxic effects, cell cycle analysis and cell
staining methods were used. Among 17 different inhibitors, 3 of them were further
studied since they were found to have significant IC50 values in HepG2 cell line, which
has wild type p53 protein expression when compared to other HCC cell lines with
various p53 mutations. They were shown to cause apoptosis by Hoechst staining and
FACS experiments, while the immunofluorescence results showed the localization of
p53 to nucleus after treatment in some cell lines.
Keywords: Hepatocellular carcinoma, p53, mdm2
References
[1]
[2]
[3]
Ferlay J, Soerjomataram I EM. IARC Cancer Base No. 11 [Internet]. Lyon: IARC.
GLOBOCAN. 2012;1.0
S P Hussain, J Schwank, F Staib, X W Wang and C C Harris. TP53 mutations and
hepatocellular carcinoma: insights into the etiology and pathogenesis of liver cancer.
Oncogene. 26: 2166-2176 (2007).
Endo K, Ueda T, Ohta T, Terada T. Protein expression of MDM2 and its
clinicopathological relationships in human hepatocellular carcinoma. Liver.; 20(3): 209215 (2000).
41
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P13. Cytotoxic Activity Investigation of Twenty New Amino Acid tertButylquinone and Avarone Derivatives
Jovana Vilipić1, Tatjana Stanojković2, Irena Novaković3, Dušan Sladić4
1
Innovation Center of the Faculty of Chemistry, University of Belgrade, Studentski trg 12-16,
P.O.Box 158, 11001 Belgrade Serbia
2
Institute of Oncology and Radiology of Serbia, Pasterova 14, 11001 Belgrade, Serbia
3
Center for Chemistry, Institute of Chemistry, Technology and Metallurgy, University of
Belgrade, Studentski trg 12-16, P.O.Box. 473, 11001 Belgrade, Serbia
4
Faculty of Chemistry, University of Belgrade, Studentski trg 12-16, P.O.Box 158, 11001
Belgrade, Serbia
A series of derivatives of marine sesquiterpene quinoneavarone (A) with amino
acids has been synthesized by nucleophilic addition of amino acids to the quinone and
their cytotoxic activity against a panel of tumor cell lines has been investigated.
Although avarone is the major constituent of the sponge Dysidea avara, and D. avara
has been a subject of cultivation and cell culture projects, it is very important to
investigate whether a simplification of the structure would lead to satisfactory biological
activity. Therefore, a very simple model, tert-butylquinone (T) has been selected as
target for modification. In this work, twenty new amino acid derivatives of avarone and
tert-butylquinone have been synthesized.
Cytotoxic activities of investigated compounds against five cancer cell lines –
human cervix adenocarcinoma cell line (HeLa), non-small cell lung carcinoma (A549),
human melanoma cell (Fem-X), chronic myelogenous leukemia (K562), human breast
cancer (MDA-MB-453) and a non-cancerous cell line, human embryonic lung fibroblast
(MRC-5), were determined by MTT assay. The obtained results (Table 1) were
expressed as IC50 values (µM) determined from cell survival diagrams and compared
with a widely used anticancer drug cisplatin as positive control. Given that compounds
A3, A4, A5, A6 and A8 showed the highest cytotoxicity towards HeLa cells, these
compounds were selected for examination of the mechanism of action by
cytofluorimetric analysis, using propidium iodide to label DNA. The present study
suggested that the cell cycle arrest and induction of apoptosis might be one possible
mechanism of action of these compounds in human cancer cells. The greatest effect,
both in cytotoxicity and in cell cycle perturbation and induction of apoptosis was
achieved with avarone derivatives with branched amino acid chains.
42
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P14. Canine Mammary Osteosarcomas
Eva Hellmén
Swedish University of Agricultural Sciences, Department of Anatomy, Physiology and
Biochemistry, Box 7011, 75007 Uppsala, Sweden
Bone forming mammary tumours appear in dogs and humans and are poorly
understood. This presentation describes five representative cases of canine mammary
osteosarcomas and induced tumours by a cloned canine mammary osteosarcoma cell
line in nude mice. All five primary tumours were combined mammary osteosarcomas
i.e. composed of both cartilage and bone tissues. In four of the five cases the
metastases were also combined osteosarcomas. However, for the metastases in one
dog and in the nude mice, only bone forming and spindle cell tumours were seen. The
metastases were spread directly to the lungs by the blood in three dogs, and via the
lymph nodes in two dogs. The metastases differed in morphology, both within and
between different metastatic sites. Some of the metastases had an even lower grade
than the corresponding primary tumour.
The interest in the COST CM1106 Action is to assay compounds relevant for
(canine) mammary tumour stem cells and collaboration would be appreciated.
43
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P15. Natural Polyphenols and Derivatives as Inhibitors of the Hedgehog
Signalling Pathway
Francesca Ghirga1,2, Mattia Mori2, Sara Toscano1,2, Bruno Botta1, Cinzia Ingallina1,2
1
Dipartimento di Chimica e Tecnologie del Farmaco, Università "La Sapienza" di Roma, 00185
Roma
2
IIT Center for Life Nano Sciences, CLNS@Sapienza, 00161 Roma
Hedgehog (Hh) signaling pathway is essential for tissues development and
stemness and its deregulation leads to tumorigenesis. Hedgehog pathway aberrant
activation has been reported in many tumors, one is the medulloblastoma, the most
frequent malignant brain tumor of childhood, which belongs to the group of embryonal
neuroepithelial tumors and arise from stem cells or early progenitor cells in the
cerebellum [1].
The aim of this study is to discover and develop small organic molecules and/or
natural compounds capable of inhibiting the Hh signaling pathway by antagonizing the
transmembrane transducer Smoothened (Smo) receptor and/or the Gli1 protein (the
pathway downstream effector for binding to DNA), thereby providing anticancer activity.
A virtual screening of a library of natural products allowed us to select a family of
promising compounds which afterwards was submitted to activity tests. One of the
most interesting active hit was a chalcone isolated from the flowers of Coreopsis
stillmanii (namely, stillopsidine Fig. 1), which has been optimized through rational
design and synthesis. The first modification of the stillopsidine nucleus was the
synthesis of the pentamethoxylated PCM-1 (Fig. 2) derivative by a Claisen-Schmidt
condensation [2].
Later, the activity of such compound was evaluated by measuring its ability to inhibit
the signal transduction of Hedgehog pathway in a cellular context characterized by
hyper-activation of the pathway. The excellent results obtained in vitro prompted us to
synthesize structural analogues, suggested by molecular modeling studies at three
steps: ring A, ring B and the double bond. (Tables 1 and 2). Biological assays of the
analogues synthesized are in progress, which will allow to refine the computational
model and to investigate the structure-activity relationships.
References
[1]
[2]
Gulino, A.; Arcella, A.; Giangaspero, F. Current Opinion Oncol. 20, 668 (2008).
Bukhari, S.N.A.; Jasamai, M.; Jantan, I.; Ahmad, W. Mini-Reviews Org. Chem., 10, 73
(2013).
44
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P16. Convenient Synthesis of Boehmeriasin A
Francesco Calogero, Michael S. Christodoulou, Raffaella Bucci, Daniele Passarella
Università degli Studi di Milano, Dipartimento di Chimica, via Golgi 19, Milano
Boehmeriasin A (1) is a potent cytotoxic natural alkaloid, it is found to be active
over many tumor cell lines (lung, breast, kidney, colon and leukemia) with an IG50
between 0.2 and 100 ng/mL. However the molecular target and the mechanism of
action are still unknown.
First asymmetric synthesis of Boehmeriasin A had been disclosed in 2010, but it
took several steps to obtain the enantiopure compound.
Like most alkaloids in nature, this compound holds piperidine moiety in is
structure; Passarella’s group developed and optimised an enzymatic process to obtain
piperidine ethanol with high enantiomeric excess (more than 95%) [1]. Taking
advantage of this methodology a new synthesis of Boehmeriasin A was achieved.
Piperidine ethanol was protected as tert-butyl carbamate, then kinetic resolution
was accomplished by sequential transesterification mediated by two different enzymes,
Lipase PS and Porcine Pancreatic Lipase. Enantiopure compound 3 was oxidised to
aldehyde and reacted with p-methoxyphenylmagnesium bromide to form alcohol 4,that
was suddenly oxidised to ketone, deprotected and reacted with activated phenylacetic
acid to form amide 5. Compound 5 under basic condition underwent intramolecular
Claisen condensation to form cyclised compound 6.
Phenanthrene ring was built performing an intramolecular Pd catalysed cross
coupling via C-H activation, finally enantiopure Boehmeriasin A 1 was obtained after
amide reduction into amine using lithium aluminium hydride.
This synthetic route is very versatile and could be used to prepare various
analogues.
References
[1]
M. Angoli, A. Barilli, G. Lesma, D. Passarella, S. Riva, A. Silvani, B. Danieli, J. Org.
Chem. 68, 9525-9527 (2003).
45
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P17. MDSCs Mediate Angiogenesis and Predispose Canine Mammary
Tumor Cells for Metastasis via IL-28/IL-28RA Signaling
Joanna Mucha1, Kinga Majchrzak2, Bartłomiej Taciak1, Eva Hellmên3, Magdalena
Król1
1
Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of
Life Sciences – WULS, Nowoursynowska 159, 02-776 Warsaw, Poland
2
Department of Animal Environment Biology, Faculty of Animal Sciences, Warsaw University of
Life Sciences – WULS, Ciszewskiego 8, 02-786 Warsaw, Poland
3
Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural
Sciences, Box 7011, SE-750 07 Uppsala, Sweden
Myeloid-derived
suppressor
cells
(MDSCs)
are
responsible
for
immunosuppression and tumor development by induction of angiogenesis in a STAT3dependend manner. Unfortunately, knowledge of MDSCs is mainly based on mice
studies and more clinical investigations using spontaneous tumor models are required.
Present paper includes both in vitro and clinical data obtained from canine patients.
Due to the current efforts to introduce the dog in a mainstream of cancer research and
clinical trials, these results may be interesting not only for veterinarians, but also for
broader audience.
We showed that in dogs with mammary cancer the number of circulating
MDSCs increases with tumor development. Using microarrays we showed that MDSCs
significantly alter molecular pathways within tumor cells in vitro. Especially important is
increased activation of IL-28/IL-28RA signaling. IL-28 secreted by MDSCs (the highest
expression observed in stage III/IV patients) stimulates STAT3 in tumor cells which
results in induction of 3D vessel formation by HUVECs, epithelial-mesenchymal
transition (EMT) and increased migration of tumor cells in vitro. Knock-down of IL-28
receptor decreases tumor cell invasion and migration in Boyden chambers.
As far as we realize this was the first microarray examination of molecular
interactions between MDSCs and tumor cells. We showed, for the first time, that
MDSCs secrete IL-28 which drives STAT3 activation to promote angiogenesis, and
EMT, invasion and migration of tumor cells. Thus, IL-28 may constitute an interesting
target for further therapies.
Moreover, similarity in linear increase of circulating MDSCs levels between
canine and human patients indicates dog as a good model for clinical trials of drugs
targeting MDSCs.
46
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P18. An Efficient Identification of Mer TK Validated Hits – Exploitation of
this Methodology to Axl TK and Smo (a Key Mediator of the HEDGEHOG
Pathway)
Juraj Dobiaš1, Gilles Hanquet2, Andrej Boháč1,3
1
Comenius University, Faculty of Natural Sciences, Department of Organic Chemistry, Mlynská
dolina, 842 15 Bratislava, Slovakia
2
Université de Strasbourg, Ecole européenne de Chimie, Polymères et Matériaux (ECPM)
Laboratoire de stéréochimie (UMR CNRS 7509), 25, rue Becquerel, F-67087 Strasbourg,
France
3
Biomagi, Ltd., Mamateyova 26, 851 04 Bratislava, Slovakia
Axl TK is a downstream effector of epithelial-to-mesenchymal transition (EMT)
signalling involved in cancer invasion and metastatic process [1]. Axl plays a pivotal
role in resistance to chemotherapy regimens [2]. X-ray structure of Axl TK was not yet
published. Recently, one homology model of Axl kinase domain was described [3]. We
plan to compose Axl TK model(s) for development of Axl inhibitors. At first we decided
to prove efficiency of kinase inhibitor identification based on an available X-ray
structure of Axl relative Mer TK and VHTS (virtual high throughput screening). By this
approach 800 Mer TK drug-like inhibitor candidates were selected from 8 million
purchasable compounds from the ZINC database. The first 11 selected compounds
were purchased and screened by Mer TK enzymatic assay. Seven from them were
active (2.75, 8.01, 9.70, 23.7, 58, 643 and 985 uM). We are performing a lead
optimization with the best Mer TK verified hit.
Hedgehog (Hh) pathway is involved in stem cell maintenance, tissue repair and
oncogenesis. Smo receptor is an important molecular target to antagonize the Hh
pathway [4]. Five X-ray complexes of hu-Smo were deposited in PDB database. We
applying the above SBDD methodology on two different hu-Smo variants with the aim
to get variable Smo hits for further development.
We are open for collaboration on development of Mer, Axl, Smo or other target
modulators.
Acknowledgements: We are grateful for financial and other support to Biomagi, Slovakia,
VEGA 1/0634/13. The COST action CM1106 (StemChem) for research networking is also
acknowledged.
References
[1]
[2]
[3]
[4]
K. Vuoriluoto, H. Haugen, S. Kiviluoto, J.-P. Mpindi, J. Nevo, C. Gjerdrum, C. Tiron, J.B.
Lorens, J. Ivaska Vimentin regulates EMT induction by Slug and oncogenic H-Ras and
migration by governing Axl expression in breast cancer Oncogene 2011 30 1436-1448.
J.D. Paccez, M. Vogelsang, M.I. Parker, L.F. Zerbini The receptor tyrosine kinase Axl in
cancer: Biological functions and therapeutic implications Int J Cancer 2014 134 1024-33.
A. Mollard., S.L. Warner, L.T. Call, M.L. Wade, J.J. Bearss, A. Verma, S. Sharma, H.
Vankayalapati, D.J. Bearss Design, Synthesis, and Biological Evaluation of a Series of
Novel AXL Kinase Inhibitors ACS Med Chem Lett 2011, 2, 907-912.
M. Ruat, L. Hoch, H. Faure, D. Rognan Targeting of Smoothened for therapeutic gain
Trends in Pharmacological Sciences 2014 35 237-246.
47
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P19. Pironetin-Dumetorine Hybrids as New Tubulin Binders
Cristina Marucci, Michael S. Christodoulou, Raffaella Bucci, Daniele Passarella
Università degli Studi di Milano, Dipartimento di Chimica, via Golgi 19, Milano
Microtubules are dynamic polymers which play a central role in a number of
cellular process, most particularly cell division, as they are the key constituents of the
mitotic spindle [1]. They are constituted of a heterodimeric protein named tubulin. It is
composed by two polipeptide called α-tubulin and β-tubulin, which through
polymerization process assembly to form microtubules. Thus, any molecule which
exhibits some interaction with microtubules dynamics will be able to influence the cell
division process [2]. Most of these antimitotic agents interact with β-tubulin. In contrast,
the number of products that bind to α–tubulin is very small. One of these compounds
that bind -tubulin is the Pironetin 1.
We designed a library of compounds characterized by the presence of the 5,6dihydro pyran-2-one ring and by exploiting the synthetic plan previously studied for the
preparation of dumetorine [3].
References
[1]
[2]
[3]
T. Fojo, The role of Microtubules in Cell Biology, Neurobiology and Oncology, Humana
Press, Totowa, New Jersey, 2008
J.G. Pla, M. Carda, J. Murga, E. Falomir, C. Trigili, S. Notararigo, F. J. Dìaz, I. Barasoain,
Eur J Med Chem 46 1630 (2011).
a) E. Riva, A. Rencurosi, S. Gagliardini, D. Passarella, M Martinelli, Chem Eur J 17 62216226 (2011); b) D. Passarella, S. Riva, G Grieco, F. Cavallo, B. Checa, F. Arioli, E Riva,
D. Comi, B. Danieli, Tetrahedron: Asymmetry 20 192 (2009).
48
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P20. Anti-Angiogenic Natural Compounds to Evaluate Their Potential to
Target Cancer Stem Cells
Javier A. García-Vilas1, Casimiro Cárdenas1, Ana R. Quesada1,2, Miguel Ángel
Medina1,2
1
Universidad de Málaga, Andalucía Tech, Departamento de Biología Molecular y Bioquímica,
Facultad de Ciencias, e IBIMA (instituto de Biomedicina de Málaga), Málaga
2
Unidad 741, CIBER de Enfermedades Raras (CIBERER)
Tight relationships of cancer stem cells (CSCs) with the cancer vascular niche
are being experimentally studied and could have important therapeutic consequences.
In fact, there are several angiogenic markers expressed by CSCs, including VEGF,
VEGF-C, VEGFR-2, Tie2, and Ang1/2, among others. On the other hand, CSCs
features include lower ROS concentrations and increased expression of anti-apoptotic
proteins. In our lab we have characterized a number of new natural anti-angiogenic
compounds that could be potentially useful to target CSCs in combined antitumor
therapies. This communication will show data regarding the anti-angiogenic effects of
some of these natural compounds and will discuss their potential to target CSCs.
Acknowledgements: Our experimental work is supported by grant P12-CTS-1507 (Andalusian
Government and FEDER) and funds from group BIO-267 (Andalusian Government). The
"CIBER de Enfermedades Raras" is an initiative from the ISCIII (Spain).
49
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P21. Lower Antioxidant Capacity of P-Glycoprotein Overexpressing MultiDrug Resistant Cancer Cells as a Mechanism for Collateral Sensitivity to
Protoflavones
Tijana Stanković1, Dankó Balázs2, Miodrag Dragoj1, Sonja Stojković1, Jasna Banković1,
Ana Martins3,4, Joseph Molnár3, Leonard Amaral5, Attila Hunyadi2, Milica Pešić1
1
Institute for Biological Research „Siniša Stanković“, University of Belgrade, Despota Stefana
142, 11060 Belgrade, Serbia
2
Institute of Pharmacognosy, University of Szeged, Eötvös u. 6, 6720 Szeged, Hungary
3
Department of Medical Microbiology and Immunobiology, University of Szeged, Dóm tér 9,
Szeged 6720, Hungary
4
Unidade de Parasitologia e Microbiologia Médica, Institute of Hygiene and Tropical Medicine,
Universidade Nova de Lisboa, Rua da Junqueira 100, Lisbon 1349-008, Portugal
5
Center for Malaria and Other Tropical Diseases (CMDT), Institute of Hygiene and Tropical
Medicine, Universidade Nova de Lisboa, Rua da Junqueira 100, Lisbon 1349-008, Portugal
Protoflavones are oxidized flavonoid derivatives with an unusual non-aromatic
B-ring. These compounds act as pro-oxidants. It has been shown previously that
protoflavones are not substrates for P-glycoprotein (P-gp), an efflux membrane pump
involved in classic mechanism of multi-drug resistance (MDR) in cancer. Moreover,
some of their derivatives were found to act selectively towards certain P-gp
overexpressing cancer cells.
To evaluate the effects of protoflavones, we employed three different human
MDR cancer cell lines with high P-gp expression and a rat MDR cancer cell line with
different mechanism of resistance (changes in apoptotic machinery). Tested
compounds showed significant cytotoxicity even acting in nanomolar range. Their
efficacy and selectivity varied among different cancer cell lines. The compounds were
more active against human P-gp overexpressing cancer cells compared to their
sensitive counterparts but less active against the rat MDR cancer cells.
In order to analyze the preference of tested compounds for MDR cells, we
compared the level of reactive oxygen species (ROS) between pairs of MDR and
corresponding non-MDR cells. Interestingly, we found that ROS level in all human P-gp
overexpressing cells was lower in comparison with parallel sensitive cells while rat
MDR cells were adapted to higher ROS level. Generally, overloading of ROS inside the
cell generates cell damage and death. To prevent such irreversible cell damage, the
increase of ROS induces a compensatory upregulation of antioxidant systems as an
adaptive response especially in cancer cells. This mechanism can be applied to rat
MDR cells that we have examined. In case of tested human P-gp overexpressing cells,
lower level of ROS indicate inactivity of antioxidant systems. To prove this concept, we
assessed the level of glutathione and the expression of antioxidant enzymes. In
addition, we examined the mechanism of the most selective protoflavone derivative.
We found that MDR cells are more vulnerable to protoflavone pro-oxidative
activity. This is due to their low antioxidant capacity probably caused as a collateral
mechanism inside the P-gp overexpressing cells.
Acknowledgements: This research was supported by the Ministry of Education, Science and
Technological Development of the Republic of Serbia (Grant No III41031). The authors
acknowledge the support from the European Union co-funded by the European Social Fund
(TÁMOP 4.2.2.A-11/1/KONV-2012-0035), the Szeged Foundation for Cancer Research and the
Fundação para a Ciência e a Tecnologia (FCT), Portugal (PEsT-OE/SAU/UI0074/2011 and
PEsT-OE/SAU/UI0074/2014). A. Martins was supported by the grant SFRH/BPD/81118/2011,
FCT, Portugal.
50
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P22. Tumourless Stem Cells: Uncoupling Tumorigenicity from Stemness
by Acting on Glypican 4
Rosanna Dono
Aix-Marseille University – Developmental Biology Institute of Marseille (IBDM) – CNRS
UMR7288, Campus de Luminy, case 907, 13288 Marseille, France
Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) –
defined by their capacities for self-renewal and for differentiation into all body cell types
– have opened new avenues in biomedical sciences as these cells can be suitable for
understanding disease mechanisms, for drug development/drug toxicity tests and for
cell replacement therapies. Interestingly, iPSCs can be generated not only from normal
tissue cells but also from malignant cells thus providing a direct human cell model to
study oncogenic events.
It has been also shown that gene expression networks that are responsible for
the maintenance of stemness versus cell lineage entry in these SCs are interconnected
and in many cases share components with networks implicated in oncogenesis and in
cancer SCs. Moreover, SCs generate tumours when transplanted in animal models
and the molecular basis of their tumorigenicity appears linked their cancer-resembling
properties. Tumorigenicity of SCs has garnered significant attention and interest in the
fields of regenerative medicine and tumour biology for issues related to safety and
tumour prevention. In this perspective, uncovering the signalling network crosstalkunderlying stemness and tumorigenicity of pluripotent SCs could permit the
development of new therapeutic strategies targeting cancer SCs.
We found that it is possible to uncouple self-renewal and differentiation of
pluripotent SCs versus tumorigenicity by manipulating the activity levels of Glypican 4
(Gpc4), the homologue of the drosophila dally-like morphogen regulator. In particular,
pluripotent cells lacking Gpc4 lose their intrinsic tumorigenic properties after
implantation into nude mice and in rat brains, while maintain the pluripotent
differentiation program both in vitro and in vivo (Stem Cells 2012; J. Neurosci 2014). As
Gpc4 is involved in the regulation of cell responses to extracellular signals, our
outcomes show that it is possible to uncouple tumorigenicity from stemness of
pluripotent SCs by acting on mechanisms regulating perception of instructing
environmental cues. We define the biological context of Gpc4 mutant pluripotent SCs
as a “tumourless state”.
We have begun addressing whether there is a molecular signature underlying the
“tumourless state”. Comparison of the transcriptome profile of mouse Gpc4 mutant
versus control ESCs revealed that the “tumourless state” corresponds to distinct
thresholds of molecular components relevant to self-renewal/differentiation and
tumorigenicity. These results suggest that there is a molecular signature that: 1)
ensures an efficient switch from self-renewal to lineage commitment, 2) permits
depletion of self-renewing cells, thus avoiding tumorigenesis. We have established
human iPSCs lacking Gpc4 and currently using them to define the molecular signature
of the “tumourless state” by combining large-scale gene expression profile screens with
bioinformatics.
Yet, Glypican4 is over-expressed in human tumours such as colon and breast
cancers where cancer SCs have been described. We are currently evaluating Gpc4
expression profile and function in different cancer cell types. We anticipate that Gpc4
may be a potential cell surface marker of cancer SCs and that agents targeting Gpc4
could elicit cytotoxic and/or differentiation effects on cancer cells.
51
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P23. New heterocyclic scaffolds by intramolecular reactions of 4quinolone-2-carboxamides
Raffaella Cincinelli, Loana Musso, Sabrina Dallavalle
Department of Food, Environmental and Nutritional Sciences, University of Milan, via Celoria 2,
I-20133, Milano, Italy
The quinolone moiety is an important structural unit in medicinal chemistry and
many compounds with this scaffold have shown a broad range of biological properties
including anticancer, antimicrobial, antiviral and antimalarial activity.
In pursuance of our research on the development of new antitumor compounds,
we became interested in accessing structurally diverse heterocyclic rings containing
the quinolone moiety.
O
TFA
O
HO
R
N
H
CONH
R
O
N
n=1
NH
1
n
O
TFA
n = 3,4
R
O
N
N
2
m = 1,2
m
We have devised a reliable synthetic route to 4-quinolone-based fused systems
starting from 4-quinolone-2-carboxylic acid oxoamides. The acid-catalyzed
intramolecular reaction of N-unsubstituted quinolones gives structurally diverse
compounds, depending on the length of the chain. Acid treatment of β-oxoamides
furnishes 3H-pyrazino[1,2-a]quinoline-4,6-diones, due to the nucleophilic attack of N-1
to the carbonyl group, whereas acid treatment of δ- and ε-oxoamides leads to the
formation of tetracyclic compounds by a tandem heteroannulation reaction.
As no examples of such heterocyclic structures have been reported in the
literature so far, the sequence represents a versatile approach to new scaffolds and
specifically provides a method for the rapid preparation of differently substituted
derivatives.
The results of a preliminary test on compound 2 (m = 1) (IC50 = 10 µM on H460
tumor cell lines) suggest that these classes of compounds could be worth of further
investigation.
52
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P24. The Effects of New Naphtoquinone and Benzopyran Derivatives
Studied in Multi-Drug Resistant Cancer Cells
Raffaella Cincinelli1, Loana Musso1, Sabrina Dallavalle1, Ana Podolski-Renić2, Milica
Pešić2
1
Department of Food, Environmental and Nutritional Sciences, University of Milan, via Celoria
2, I-20133, Milano, Italy
2
University of Belgrade, Institute for Biological Research „Siniša Stanković“, Despota Stefana
142, 11060 Belgrade, Serbia
Multi-drug resistance (MDR) is a significant obstacle to the efficient treatment of
cancer. Overexpression of membrane transporters P-glycoprotein (P-gp) and Breast
Cancer Resistance Protein (BCRP) is responsible for the classic mechanism of MDR –
the extrusion of drugs from cancer cells.
The search for anti-cancer agents able to overcome or evade MDR has become
an imperative in the field of drug design and discovery.
Structure modification of natural products may still deliver new hope for the
discovery of new scaffolds able to preserve cytotoxic activity toward MDR cancer cells.
As a part of a research program aimed at studying new natural product–inspired
compounds we synthesized some representative scaffolds containing a benzopyrane
or a naphtoquinone core.
Benzopyrans are privileged medicinal pharmacophores which appear as
important structural components in natural compounds and generated great attention
because of their interesting biological activity. Quinones are another class of important
natural products showing remarkable anti-cancer activity, mostly connected with their
redox properties.
Based on our previous work, a series of naphtoquinones and benzopyrans were
designed and synthesized.
The compounds were tested in in vitro models of MDR (pairs of sensitive and
MDR human cancer cell lines with different origin: non-small cell lung carcinoma,
colorectal carcinoma and glioblastoma). Importantly, the characterization of MDR
cancer cells revealed that they possess overexpression of P-gp or both P-gp and
BCRP.
The most potent compound, belonging to the class of naphtoquinones, reached
IC50 in nanomolar range of concentrations in all the cell lines.
53
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P25. Effects of Biological Extracts on Terminal Differentiation of Solid
Tumours and Leukaemias
Sherif Suleiman, Pierre Schembri-Wismayer, Analisse Cassar, Jean Calleja Aguis
University of Malta, Malta
Cancer is the second leading cause of death (22.8%) worldwide following heart
disease (26.6%). The main approach in the management of patients suffering from
cancer is the use of chemotherapy and/or radiation therapy, once resection is no longer
an option. Despite the fact that these therapies generally succeed in reducing the
overall tumour size, relapse is still the primary cause of poor survival rates with
recurrence occurring in many patients with metastatic cancer.
Unlike normal cells, cancer cells show a block in differentiation leading to
uncontrolled proliferation, eventually invading surrounding tissues and organs. This can
lead to metastases as the cancer cells spread to other parts of the body through the
haemopoietic and lymphatic systems.
The aim of this research is to cause terminal differentiation of cancer cells using
biological extracts from insects and other organisms with regenerating and
differentiating cellular systems such as planarians. Fractions of insect conditioned
medium have already been proven to increase significantly the degree of differentiation
of the leukaemic cell line - HL60. (Cassar A., 2009) Chemical analysis and
identification will be carried out to identify the active ingredients within the conditioned
medium and extracts. In the future, this can potentially lead to the development of
improved treatments for patients suffering of cancer.
Work has already started using Planaria (Schmidtea Mediterranea) conditioned
media and this was tested on Osteosarcoma Cell line SaOS2 and leukaemia cell line
KG1a. Cell proliferation was determined using the MTT assay and differentiation was
determined using alkaline phosphatase staining for SaOS2 and NBT for KG1a. Further
biological extracts are being prepared from other organisms with regenerating potential
and these will be tested for their ability to cause differentiation in different cancer cell
lines.
54
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P26. HDAC and HMT Inhibitors in Combination with RA Affect on Gene
Methylation Changes in Leukemic Cells
Veronika Borutinskaite, Ruta Navakauskiene
Department of Molecular Cell Biology, Institute of Biochemistry, Vilnius University, Lithuania
Acute promyelocytic leukemia (APL) is one of the most common subtypes of
acute myeloid leukemia (AML). It is characterized by the arrest of myeloid
differentiation in the promyelocytic stage, uncontrolled cell division and growth. The cell
line NB4 has the characteristic reciprocal translocation t(15;17). The product of this
aberrant gene is the fusion protein and differentiation suppressor PML-RARα. HL-60
cells do not possess this translocation but have the amplification of c-MYC and lack the
product of TP53 (p53).
The aim of this investigation was to evaluate the effect of HDACI and MHTI
together with RA on leukemic cell differentiation, as well as, the specific gene
expression and protein quantity changes in treated APL cells. We used HDACI −
Belinostat (PXD 101) and HMTI − 3-Deazaneplanocin A (DZNep). Using cell staining,
counting, flow cytometric analysis, Western blotting and MS-PCR methods some
findings were made. The biggest effect on depression of cell viability and induction of
apoptosis was observed when APL cells were treated with 1 μM RA + 0.2 μM Bel + 0.5
μM DZNep. What is more, we observed some changes in the gene expression level of
myeloid differentiation specific genes and histone modifications associated genes.
Our study showed that combined RA, HDACI and HMTI treatments were more
effective on APL cell differentiation than these chemicals alone. This was proved by the
assessement of cell viability, growth inhibition, apoptosis, examined gene and protein
expression profiles. These combined RA, HDACI and HMTI treatments could be further
investigated and used in in vivo experiments.
55
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 P27. Novel Pd(II), Pt(II), Cu(II) and Ga(III) Arylindole Complexes as
Potential Leukaemia Differentiation Agents
Nenad Filipović1, Sveva Pelliccia2, Snežana Bjelogrlić3, Tamara Todorović4, Pierre
Schembri-Wismayer5, Romano Silvestri2
1
University of Belgrade – Faculty of Agriculture, Nemanjina 6, 11000 Belgrade, Serbia
Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Università di Roma, I-00185
Roma, Italy
3
National Cancer Research Center of Serbia, University of Belgrade, 11000 Belgrade, Serbia
4
University of Belgrade – Faculty of Chemistry, Studentski trg 12-16, 11000 Belgrade, Serbia
5
Department of Anatomy, Faculty of Medicine and Surgery, University of Malta, Msida MSD
2080, Malta
2
3-Arylithio/aroylndoles are a class of potent inhibitors of tubulin polymerization
and cancer cell growth. They inhibit tubulin polymerization by binding to the colchicine
site, thereby inhibiting the binding of [3H]colchicine to tubulin. After analysis of more
than 70 drug candidates [1], (2-(pyridin-2-yl)-1H-indol-3-yl)(3,4,5-trimethoxyphenyl)methanone (HL) was selected as a compound capable to bind bidentately to metal ions
by forming 5-membered chelate ring. HL was used as a ligand for the synthesis of
Pd(II), Pt(II), Cu(II) and Ga(III) complexes. The complexes were characterized by
elemental analysis, IR and heteronuclear-multidimensional NMR spectroscopy, as well
as a single crystal X-ray analysis (palladium and copper complexes).
The ligand and the complexes have been evaluated as growth inhibitors of
HL60 cells which are an in vitro model for acute myeloid leukaemia. Due to lack of the
t15:17 translocation these cells are resistant to retinoic acid differentiation treatment.
The study was divided into two limbs to establish the range of concentrations in which
the compounds are cytotoxic to HL60 cells, and to evaluate whether the compounds
are capable to induce differentiation of HL60 cells toward monocytes and/or
granulocytes. Preliminary results have shown that all the compounds act as cytotoxic
agents at nanomolar concentrations indicating on their ability to accumulate in treated
cells, while Ga(III) complex appears to be a potent inducer of differentiation of HL60
leukaemia cells.
References
[1]
G. La Regina et al., J. Med. Chem. 56, 123-149 (2013).
56
COST CM1106 & CIBICAN Workshop October 14‐15, 2014 PARTICIPANTS
57
COST Participants
Name
Affiliation
e-mail address
Activity in the
Workshop
Aikman, Brech
University of Groningen
B.Aikman@student.rug.nl
Poster
Almeida, Gabriela
Institute of Molecular Pathology
and Immunology UP
galmeida@ipatimup.pt
STSM
coordinator, MC
member
Athanassopoulos,
Constantinos
University of Patras
kath@chemistry.upatras.gr
Bayir, Ece
Ege University
ece.bayir@biyomuhendis.com
Banković, Jasna
Institute for Biological Research
"Sinisa Stankovic", Belgrade
jasnam@ibiss.bg.ac.rs
MC substitute,
Talk
Baumann, Markus
University of Durham
marcus.baumann@durham.ac.uk
Talk
Boháč, Andrej
Comenius University, Bratislava
andrej.bohac@fns.uniba.sk
Borutinskaite, Veronika
Vilnius University, Vilnius
veronika.borutinskaite@bchi.vu.lt
Bosch, Joan
Bosnakovski, Darko
Botta, Bruno
University of Barcelona
"Goce Delcev" Stip University
Sapienza University, Rome
joanbosch@ub.edu
darko.bosnakovski@ugd.edu.mk
bruno.botta@uniroma1.it
MC member
MC substitute,
Poster
MC member
MC member
Botta, Maurizio
University of Siena
botta.maurizio@gmail.com
Cetin-Atalay, Rengul
Bilkent University, Ankara
rengul@bilkent.edu.tr
Chadda, Rekha
National University of Ireland
r.chadda1@nuigalway.ie
MC member,
Talk
MC substitute,
Talk
Poster
Christodoulou, Michalis
University of Milan
michalis.christ@gmail.com
Poster
Working
Group
Group
responsible
Country
Angela Casini
The
Netherlands
WG-1
Portugal
WG-3
Greece
Aylin Sendemir
Urkmez
Turkey
WG-3
Serbia
WG-3
United
Kingdom
Slovakia
WG-1
Lithuania
WG-3
WG-1
WG-3
Spain
FYROM
Italy
WG-2
Italy
Turkey
Paul Murphy
Daniele
Passarella
Ireland
Italy
Clarkson, Richard
Cardiff University
clarksonr@cf.ac.uk
Cuendet Licea, Muriel
University of Geneva
muriel.cuendet@unige.ch
Dallavalle, Sabrina
Damia, Giovanna
Dobiaš, Juraj
Dono, Rosanna
Fallacara, Anna-Lucia
Filipović, Nenad
Ghirga, Francesca
University of Milan
Mario Negri Institute, Milan
Comenius University, Bratislava
Aix-Marseille University, Marseille
University of Siena
University of Belgrade
Sapienza University, Rome
sabrina.dallavalle@unimi.it
giovanna.damia@marionegri.it
jur.dobias@gmail.com
rosanna.dono@ibdml.univmed.fr
al.fallacara@gmail.com
nenadf@chem.bg.ac.rs
francesca.ghirga@uniroma1.it
MC member,
Talk
Poster
Chair
Poster
Poster
Poster
Poster
Poster
Gozen, Damla
Bilkent University, Ankara
damla.gozen@bilkent.edu.tr
Poster
Groner, Berndt
Institute for Tumor Biology and
Experimental Therapy, Frankfurt
am Main
groner@em.uni-frankfurt.de
Talk
Gure, Ali O.
Bilkent University, Ankara
agure@bilkent.edu.tr
Hanquet, Gilles
Strasbourg University
ghanquet@unistra.fr
Hellmen, Eva
Swedish University of Agricultural
Sciences
eva.hellmen@slu.se
Herold-Mende, Christel
University of Heidelberg
Christel.Herold-Mende@med.uniheidelberg.de
Hunyadi, Attila
Eötvös Loránd University,
Budapest
University of Szeged
hunyadi.a@pharm.u-szeged.hu
Kikelj, Danijel
University of Ljubljana
Danijel.Kikelj@ffa.uni-lj.si
Król, Magdalena
Warsaw University of Life Sciences
magdalena_krol@sggw.pl
Hudecz, Ferenc
fhudecz@elte.hu
Talk
MC member,
Talk
MC member,
Poster
MC member,
Poster
MC substitute
MC member,
Talk
Poster
MC member,
Poster
MC member,
Talk
WG-1
United
Kingdom
WG-1
Switzerland
WG-3
WG-1
Italy
Italy
Slovakia
France
Italy
Serbia
Italy
Andrej Boháč
WG-1
Maurizio Botta
WG-1
Bruno Botta
Rengul CetinAtalay
Turkey
WG-1
Germany
WG-1
Turkey
WG-3
France
WG-1
Sweden
WG-1
Germany
WG-3
Hungary
WG-3
Hungary
WG-3
Slovenia
WG-1
Poland
Link, Wolfgang
University of Algarve, Faro
walink@ualg.pt
Martins, Ana
Medina, Miguel Angel
Menegon, Andrea
Mitanoska, Ana
Monteiro, Angelo
Mori, Mattia
Mucha, Joanna
Muller, Christian D.
University of Szeged
University of Málaga
San Raffaele Scientific Institute
"Goce Delcev" Stip University
University of Lisbon
Center for Life Nano Science, Italy
Warsaw University of Life Sciences
University of Strasbourg
martins.a@pharm.u-szeged.hu
MEDINA@uma.es
menegon.andrea@hsr.it
ana.mitanoska@ugd.edu.mk
afmonteiro@ff.ulisboa.pt
m.mattia79@gmail.com
doro60@poczta.onet.pl
cdmuller@unistra.fr
Navakauskiene, Ruta
Vilnius University
ruta.navakauskiene@bchi.vu.lt
Odysseos, Andreani
EPOS-lasis, R&D, Nicosia
andreani@epos-iasis.com
Ofir, Rivka
Ben Gurion University of the
Negev, Beer Sheva
rivir@bgu.ac.il
MC Substitute,
Talk
Poster
Poster
Invited speaker
Poster
Poster
Talk
Poster
Poster
MC member,
Poster
MC member,
Session Chair
MC member
Local organizer,
MC substitute,
DDC Member
MC Chair
MC member,
Poster
WG-1
Portugal
Attila Hunyadi
Ana R. Quesada
WG-1
Hungary
Spain
Italy
FYROM
Portugal
Italy
Poland
France
WG-1
Lithuania
WG-1
Cyprus
WG-1
Israel
WG-3
Spain
WG-3
Italy
WG-1
Serbia
WG-1
Maria M. Santos
Magdalena Król
Padrón, José M.
IUBO-AG, University of La Laguna
jmpadron@ull.es
Passarella, Daniele
University of Milan
Institute for Biological Research
"Sinisa Stankovic", Belgrade
National Centre for Scientific
Research “DEMOKRITOS”, Aghia
Paraskevi
daniele.passarella@unimi.it
pitsinos@chem.demokritos.gr
Poster
WG-3
Greece
Quesada, Ana R.
University of Málaga
quesada@uma.es
MC member,
Poster
WG-1
Spain
Ricci, Francesca
Santos, Maria M. M.
Mario Negri Institute, Milan
University of Lisbon
francesca.ricci@marionegri.it
mariasantos@ff.ul.pt
Pešić, Milica
Pitsinos, Emmanuel
camala@ibiss.bg.ac.rs
Giovanna Damia
WG-3
Italy
Portugal
Schembri-Wismayer,
Pierre
Sendemir Urkmez, Aylin
Seneci, Pierfausto
Ege University, Izmir
University of Milan, Italy
pierre.schembriwismayer@um.edu.mt
sendemir@gmail.com
pierfausto.seneci@unimi.it
Sladić, Dušan
University of Belgrade
dsladic@chem.bg.ac.rs
Sotiropoulou, Panagiota
Suleiman, Sherif
Vizirianakis, Ioannis
Université Libre de Bruxelles
University of Malta
Aristotle University of Thessaloniki
Panagiota.Sotiropoulou@ulb.ac.be
sherif.s.suleiman@um.edu.mt
ivizir@pharm.auth.gr
University of Malta
MC member,
Poster
Poster
Invited expert
MC member,
Poster
MC member
Poster
MC member
WG-1
Malta
WG-2
Turkey
Italy
WG-3
Serbia
WG-1
Belgium
Pierre Wismayer
WG-3
Greece
Participants without reimbursement
Name
Affiliation
e-mail address
Callogero, Francesco
University of Milan
francesco.calogero@unimi.it
Activity in the
Workshop
Poster
Dinić, Jelena
Institute for Biological Research
“Siniša Stanković”, Belgrade
jelena.dinic@ibiss.bg.ac.rs
MC Substitute
Serbia
Ingallina, Cinzia
Sapienza University, Rome
cinzia.ingallina@uniroma1.it
Talk
Italy
Podolski-Renić, Ana
Institute for Biological Research
“Siniša Stanković”, Belgrade
ana.podolski@ibiss.bg.ac.rs
Poster
Petricci, Elena
University of Siena
elena.petricci@unisi.it
Talk
Marucci, Cristina
University of Milan
cristina.marucci@gmail.com
Poster
Stupariu, Ioana
Secretariat CM1106
stemchem@gmail.com
Secretary CM1106
Group responsible
Country
Daniele Passarella
Italy
Milica Pešić
Serbia
Italy
Daniele Passarella
Italy
Italy
DDC & CIBICAN Participants
Activity in the
Workshop
DDC Chair
Name
Affiliation
e-mail address
Godefridus J. Peters
VU University Medical Center
g.j.peters@vumc.nl
Roger Phillips
University of Bradford
R.M.Phillips@bradford.ac.uk
Rafael Alonso Solís
ITB, University of La Laguna
ralonsosolis@gmail.com
Víctor S. Martín
IUBO-AG, University of La Laguna
vmartin@ull.es
Miguel X. Fernandes
IUBO-AG, University of La Laguna
mfernand@ull.edu.es
Talk
Talk
DDC Member
ITB Director, IMBRAIN
Coordinator
Country
The Netherlands
United Kingdom
Spain
Spain
Spain
David Gillespie
ITB, University of La Laguna
dgillesp@ull.es
Romen Carrillo Fumero
IUBO-AG, University of La Laguna
rocarril@ull.es
Spain
Ana R. Díaz Marrero
IUBO-AG, University of La Laguna
adiazmar@ull.edu.es
Spain
Isabel López Bazzocchi
IUBO-AG, University of La Laguna
ilopez@ull.es
Spain
Ignacio A. Jiménez Díaz
IUBO-AG, University of La Laguna
ignadiaz@ull.es
Spain
Oliver Callies
IUBO-AG, University of La Laguna
callies.oliver@gmail.com
Spain
Leticia González León
University of La Laguna
gl.leticia@gmail.com
Spain
Raimundo Freire
Hospital Universitario de Canarias
rfreire@ull.edu.es
Veronique Smits
Hospital Universitario de Canarias
vsmits@ull.es
Ignacio Alonso
Hospital Universitario de Canarias
Spain
Santiago Hernandez
Hospital Universitario de Canarias
Spain
Elisa Cabrera
Hospital Universitario de Canarias
Spain
Rocío Delgado Díaz
Hospital Universitario de Canarias
Spain
Fernando García Tellado
IPNA, CSIC
fgarcia@ipna.csic.es
Spain
Felix Machín Concepción
Hospital Universitario NS de Candelaria
fmacconw@gmail.com
Spain
Sebastián Jiménez Reyes
University of La Laguna
University of La Laguna, Secretariat
IMBRAIN
sebastianjimenez@cibican.org
Innovation manager
Spain
fpcova@ull.es
Project manager
Spain
Farah Cova Alonso
Talk
Spain
Spain
Spain