NUOVO-SOLDATI FOUNDATION FOR CANCER RESEARCH
Research grants in cancer research

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Laureates 2012 - 2013

Isabelle Tromme
Nicolò Riggi
Angela Di Giannatale
Krisztian Homicsko
Jean-Sebastien Frenel
François-Clément Bidard

Isabelle Tromme

Title of the project: Assessment of two methods to detect melanoma: dermoscopy and digital dermoscopy.

Place of the training course: Dermatology and epidemiology departments,
Catholic University of Louvain,
Brussels, Belgium.

Summary of the project: Melanoma is one of the first causes of death by cancer in the young Caucasian adult population. In light of the lack or limited efficiency of the treatments of the metastatic stages, early detection is of paramount importance since it is often a precondition to save a patient’s life by simple surgery. Dermoscopy allows the observation of the epidermis and the superficial dermis in order to significantly enhancing the diagnostic performances compared to the naked-eye examination, provided the practitioner is properly trained in this technique. Storage of the images and their comparison over time are made possible by digital dermoscopy. The present study aims at specifying the medical, psychological and socio-economic added value of dermoscopy and digital dermoscopy in terms of number of melanomas detected at a very early stage, avoidance of unnecessary surgeries and quality of life of high risk patients.

Nicolò Riggi

Title of the project:The role of microRNAs in Ewing’s sarcoma cancer stem cells.

Place of the training course: Research group of Prof. Andrew E. Rosenberg
Pathology Department at Harvard Medical School
Massachussets General Hospital (MGH)
55 Fruit Street
Boston, MA 02114
USA

Summary of the project: Ewing’s sarcoma family tumors (ESFT), the second most common bone malignancy in children and young adults, are characterized by unique chromosomal translocations believed to provide the key oncogenic event in ESFT pathogenesis. The most common fusion gene, EWS-FLI-1, is expressed in 85-90% of ESFT and believed to represent the initiating event in ESFT development by inducing and repressing target genes that lead to transformation of permissive primary mesenchymal stem cells (MSCs), currently considered to be the most likely cell of origin of these tumors.

We have recently identified ESFT cancer stem cells and showed that they express the CD133/Prominin-1 stem cell surface marker in addition to representing the tumor initiating population of ESFT. We have subsequently shown that under appropriate culture conditions, MSC expressing EWS-FLI-1 generate a cell subpopulation displaying ESFT CSC features in vitro, and demonstrated that induction of the ESFT CSC phenotype is the result of the combined effect of EWS-FLI-1 on its target gene expression and repression of microRNA-145 (miRNA145) promoter activity. MicroRNAs are now recognized as an abundant class of genome-encoded, small RNAs that repress gene expression at the post-transcriptional level via base pairing to complementary sites located in target messenger RNAs (mRNAs). In this way a particular miRNA sequence is thought to guide the regulation of several hundred different transcripts, and, additionally, an individual mRNA may be simultaneously targeted by multiple different miRNAs. Not surprisingly therefore, miRNAs have been implicated in the regulation of a variety of diverse biological processes, and their dysregulated expression is associated with several diseases, most notably cancer.

Based on our recent observations that miRNA-145 repression underlies the emergence of ESFT CSC, we plan to further dissect the mechanisms whereby EWS-FLI-1 expression and microRNAs dysregulation lead from CSC generation to Ewing’s sarcoma development. In order to achieve this goal, in a first set of experiments, we will compare the miRNA expression profile of three primary ESFT and six different ESFT cell lines to the one of wild type MSCs. In a second set of experiments we plan to isolate CSC from freshly surgical resected ESFT samples and perform miRNA expression profile analysis of transcripts from CD133 pos. (CSC) and neg. (not-CSC) cells.

These two approaches might allow us to identify a set of miRNA whose deregulation in ESFT might constitute a crucial oncogenic mechanism in ESFT pathogenesis and that could play a significant role in ESFT CSC biology, respectively. Each putative miRNA of potential interest found to be differentially expressed will subsequently be tested individually on ESFT cell lines and primary CSC cultures to assess the impact of its modulation on their biological behaviour in vitro as well as their tumorigenic potential and CSC properties in vivo. This study will therefore provide a comprehensive overview of the involvement of miRNA in CSC generation and ESFT development. More importantly, the intrinsic ability of a single miRNA to coordinately modulate the expression of dozens of genes involved in different molecular pathways and biological processes, suggests that miRNA might constitute central nodes in the genetic circuitry of cancer, rendering them a new very attractive and promising category of therapeutic targets.

Angela Di Giannatale, MD

Title of the project: Characterization and Functional Analysis of Pediatric Cancer-Derived Exosomes in Malignant Progression

Place of the training course: Department of Pediatric and Adolescent Oncology,
Institut Gustave Roussy,
Villejuif France


Summary of the project: Tumor-derived factors stimulate the formation of ‘pre-metastatic niches’ favoring subsequent tumor cells colonization in distant tissues. However, how a solid tumor exerts such systemic influences is unknown. Exosomes are a potential candidate for such exchanges. These membrane-derived microvesicles cells contain RNAs and proteins, are shed prolifically by tumor cells, and act to stimulate angiogenesis, activate fibroblasts and m odulate immune function. Exosomes are present in the circulation of cancer patients and m ay carry tumor-derived signals that promote the progression of both primary tumor growth and metastases. Our hypothesis is that the proteomic profiles of exosomes isolated from the plasma of pediatric cancer could provide predictive diagnostic information on the primary tumors metastatic capacity. The goal of this work is thus to analyze the biological and clinical profiles of pediatric-tumor derived exosomes. Isolate and characterize exosomes from the plasma of pediatric cancers and control subjects; conduct proteomic analysis to com pare exosome content changes as a function of tumor subtype and stage; determine how the proteomic content of 3D cultured pediatric cancer cells (PCC) exosomes changes with progression; develop m ore complex co-cultures to test the uptake of these exosomes by resident cells in the tumor microenvironment, determine the functional consequence of such uptake; examine whether circulating PCC-exosomes help establish a metastatic niche and/or disrupt Bone Marrow (BM) niches; develop 3D co-culture models to mimic the complexity of the BM microenvironment and examine the mechanism by which exosomes could disturb the equilibrium ; target Itgb1, Fibronectina or other suspects on/in-these exosomes to inhibit their harmful actions. Any molecule that plays a direct or indirect role in mediating exosome uptake by target cells would represent an attractive target for therapies that block the pro-metastatic effects of tumor exosomes.

Dr Krisztian Homicsko, MD-PhD

Title of the project: Assessing angiogenesis during tumor development and in response to antiangiogenic and B-Raf inhibitors in a genetically engineered mouse model of melanoma

Place of the training course:Centre Pluridisciplinaire d’Oncologie, CHUV, Lausanne Switzerland
The Hanahan Laboratory, Ecole Polytechnique Federale de Lausanne, EPFL, Lausanne, Switzerland

Summary of the project: Melanoma is a challenging disease for which the standards of care have transitory benefit, leading to aggressive and often lethal progression. Even the recently FDA approved B-Raf inhibitor (vemurafenib, Zelboraf©)1, which produces in some patients dramatic initial responses and therefore will become standard-of-care for patients with B-Raf mutations, typically fails with disease recurrence after a few months. Angiogenesis is a hallmark of melanoma progression at the transition from horizontal to vertical growth phase2. Late stage melanoma also shows extensive angiogenesis but targeting angiogenesis either with small molecular tyrosine kinase inhibitors or by monoclonal antibodies have failed to show clinical benefit. This apparent discordance in melanoma is not well understood. A number of genetically engineered mouse models (GEMM) of melanoma have been developed, and the Hanahan laboratory is studying one such model, called iBIP, that develops melanomas in rather synchronous fashion amenable to pre-clinical trials (L. Kwong and L. Chin, unpublished). This project will focus on three questions concerning angiogenesis in melanoma.
First, the angiogenesis phenotype of the iBIP mouse model of melanoma will be studied. Second, the response and mechanism of resistance to anti-angiogenic treatments and the effect of B-Raf inhibition on angiogenesis will be evaluated. Finally, to improve control of melanoma, the combinations of B-Raf inhibition with anti-angiogenic therapy and with a modified version of the chemo-switch metronomic therapy will be studied.

Dr Jean-Sebastien Frenel

Title of the project: Evaluating plasma cell free DNA for multi-purpose biomarker studies for the treatment of cancer patients with PI3K inhibitors.

Place of the training course: Drug Development Unit,
The Institute of Cancer Research and The Royal Marsden Hospital,
Sutton, UK,

Summary of the project: The existence of circulating cell-free DNA (cfDNA) is a common and relatively early phenomenon in various cancer types. CfDNA is tumor-derived, has been found in higher concentrations in cancer patients than healthy controls and has been proven to share similar genetic features to the primary tumour. As it can easily be isolated from human plasma, cfNA may represent a ‘liquid biopsy’ enabling the real-time analyses of tumour DNA for the evaluation of tumor-related genetic and epigenetic alterations relevant to carcinogenesis and resistance to therapy.
The phosphoinositide-3 kinase (PI3K) pathway has been identified as an important target in cancer and is activated by several different mechanisms, including somatic mutation and amplification of genes encoding key components. Consequently, therapeutics targeting the PI3K pathway are being developed at a rapid pace.
Our main goal is to assess the utility of circulating cell free plasma DNA as a multi-purpose biomarker for patients treated with PI3K inhinitors. We hypothesize that plasma cfDNA profile analyses can be utilized A)to detect PI3K mutations/amplification and the deletion of tumor suppressors that activate PI3K signaling in comparison with the tumor B) Monitor disease response to treatment; C) Identify novel genomic changes associated with treatment resistance.

François-Clément Bidard

Title of the project: Applications to the detection and characterization of circulating tumours cells

Place of the training course: ChELSI Institute (Chemical Engineering at the Life Science Interface)
University of Sheffield, UK

Summary of the project: Circulating tumor cells (CTC) can be isolated and counted by several techniques in the blood of patients diagnosed with localized or metastatic cancers. CTC counts became over the past few year a new prognostic factor in several cancer types. Beyond this quantitative approach, the molecular characterization of CTC may help us to better understand the biology of these cancer cells and of the metastatic process. New therapeutic targets might be also discovered on these tumor cells. The project is about the characterization of the CTC surface membrane proteins, by a bottom-up proteomic strategy. This project will use liquid chromatography coupled with tandem mass spectrometry (Lc-MS/MS) of peptide sequences which can be isolated from the CTC surface. A comparative quantative analysis may be also performed by isobaric labelling (iTRAQ).