Translational Research Program

1. Translational Research Program An Overview: 2000-2005 Annual Meeting 2005

2. Mission • The Translational Research Program was designed to provide the laboratorycomponent for achieving the overall INCTR mission of improving clinical outcome of cancer patients from developing countries. • Laboratory for generation of molecular correlates from clinical studies. • Conduct molecular epidemiological and educational programs.

3. Objectives • Major elements of the program driven by • Clinical studies conducted by INCTR (ALL in India) • Epidemiological studies of interest to INCTR • Differences in distribution of molecular subtypes of ALLs • Gene-environment interactions modulating oncogenic translocations • Epigenetic changes in bladder ca. modulated by infection • Enable capacity building • Training in molecular biology platforms that can provide clinically relevant tools • Pilot research studies for future fundable programs (ALL in India)

4. Specific ObjectivesBidirectional Interactions • Catalyze the translation of unique clinical observations from the patient setting in developing countries to a laboratory environment. • Is outcome of childhood ALL a function of the subtype distribution? • Rapidly move key and novel laboratory observations made in the West, to the patient setting in developing countries. • Are novel tumor types identified by molecular profiling clinically relevant in low resource settings?

5. Resources • Human • Scientists & technicians from KFNCCC&R • Trainees from developing countries • Space • KFNCCC&R, Riyadh, Saudi Arabia • Facilities • PCR, real time PCR, sequencer, tissue arrayer, microarrays, more. • Major Collaborators • 3 Indian Institutions (TMH, AIIMS, WIA) • NCI, Cairo, Egypt • Univ of Istanbul, Turkey

6. TUMOR SAMPLES Strategy

7. ALL Program 5 Projects: • Molecular subclassification • Gene expression profiling • Epigenetics • Pharmacogenetics • Proteomics of Apoptosis - target for treatment

8. ALL Program1- Molecular subclassification • Real-time RT-PCR for most common chromosomal translocations • Trained laboratory staff of INCTR centers in India • SOP for sample collection, storage, usage and analyses • Distribution of molecular subgroups TEL-AML1 are less frequent, BCR-ABLs are more frequent in India but not in Saudi Arabia • Clin Cancer Res, 2002 Leukemia, 2003 Am J Hematol, 2004 J Mol Diagnostics, 2005 • Clinical correlates - recent INCTR protocol

9. ALL Program2- Gene Expression Profiling • Real-time RT-PCR validation of data from the West • VEGF and its receptors are expressed in ALL • TEL/ETV6 expression can identify 12p del • Hox11L2 expression in T-ALL • Novel molecular subgroup identified in the West (Yeoh et al) is also found in India • Leuk Res, 2004 Int J Biol Markers, 2004 Cancer Lett (in press) • Affymetrix microarrays

10. ALL Program3- Epigenetics • MSP and COBRA assays for multiple genes and sequence analyses • Concurrent methylation pattern of ALL • E-cad is frequently methylated • B > T MLL > E2A-PBX1 • Differences with adult ALL • Less epigenetic lesions in cell cycle genes • Comparison with AML • Methylator phenotype: DAP-Kinase, p15, ER, SOCS1 • Peak of methylation in young adults • Leukemia 2003 Cancer Epidemiol Biomarkers Prev 2003 Am J Hematol 2004, 2005

11. ALL Program4- Pharmacogenetics • PCR-RFLP assays for multiple genes involved in metabolization of drugs and xenobiotics. • Population-based: SNPs influencing cancer risk • Middle-Eastern Arab population • The Pharmacogenomic J, 2004 • Patient-based: SNPs determining risk of toxicity and efficacy of treatment • Over 200 Indian patients • 100 pts in MCP943 • Clinical correlates: level of toxicity?

12. ALL Program5- Proteomics of Apoptosis • Standardized Western blot assay for proteins involved in the intrinsic and extrinsic pathways of apoptosis using cell lines. • Validated its application in clinical samples. • Proof of principle: Apoptosis as a target for therapy. • Haematologica 2003 Leuk & Lymphoma 2004 Genes, Chromosomes Cancer 2004

13. Additional ActivitiesManagement of ALL • Real time PCR assay for detection and quantification of fungal infections – can predict IFI? Preemptive therapy? • J Med Microbiol 2005 • Proof of principle of a novel marker of MRD and CNS involvement in ALL and AML (real time quantitative PCR for TdT) • Haematologica (2nd revision) • Real time PCR assay for EBV load (BMT).

14. Other Activities • Epigenetics of Bilharzial bladder cancer • SA > NSA • Modern Pathol 2004 • Geographic variation of oncogenic transl. in healthy individuals (Middle Eastern Arabs) • JH-BCL2 - 37% (> in older individuals) • BCR-ABL – 27% • TEL-AML1 – 4% • More than 1 translocation increased with age • Modulation by SNPs in DNA repair and synthesis genes (37 SNPs in 11 genes). • Manuscript in preparation

15. Future Directions1-Improvements • Provide the missing clinical links to prove the reliability and the usefulness of the assays and biomarkers developed. • Recent INCTR Protocol for ALL (India) • Uniform data collection and availability. • Establish a quality control system.

16. Future Directions2-Pathway • Comprehensively pursue the ALL translational program in India. • The pilot ALL project has already provided: • basic ground work to define feasibility. • technological training and capacity. • identified the team process required. • Now the Indo Leukemia Study Group should engage in writing a proposal to attract funding.

17. Future Directions3-Suggested Steps 1) Form a strategy group to prioritize the clinical and translational questions. Coordination and quality control. 2) Use data from the pilot project to write a grant application. 3) Identify partners (investigators from USA/ Europe) who can provide additional technological expertise and extend comparative studies. 4) Identify funding sources that will consider the proposal.