1. IGF1R targeting in pediatric and wild-type GIST.

Hypothesis: Gene expression profiling has identified high levels of OGF1R expression in pediatric GIST, compared to the adult counterpart. As various IGF1R antibodies drugs are now available and being tested in clinical trials, our goal is to identify the mechanism of upregulation of IGF1R in pediatric GIST. The following methods are being applied:

    1. Immunohistochemical screening for IGF1R protein expression, using a monoclonal antibody, clone G11, from Ventana-Roche, Tuscon, AZ. This monoclonal antibody recognizes the C-terminus of both un-phospho-rylated and phosphorylated forms of the IGF-1R beta chain and importantly does not cross-react with the insulin receptor

    2. Western Blot detection of IGF1R total and phosphorylated proteins

    3. IGF1R copy number changes by FISH (preliminary results showed no amplifications in 15 cases).

    4. IGF1R full cDNA sequencing (preliminarily 16 cases screened, with no gene mutations identified).

    5. Drug testing in GIST cell lines and short term culture of combination agents, such as Rapamycin and R1507 (Roche IGF1R antibody).

  1. Drug testing in short-term culture of WT GIST, using KIT and downstream targets inhibitors. As obtaining cell lines from pediatric or wild-type tumors is extremely challenging, we have perfected an alternative method to screen various TKI or downstream inhibitors in this rare GIST subset.

  1. SDH screening in pediatric and Wild-type GIST (collaborative effort with Su Kim and Constantine Stratakis, from NIH), including MSKCC samples as well as NIH pediatric clinic cases. An extensive effort in full length cDNA sequencing of the SDH complex gene (SDH-B, C, D) and SDH5 is being undertaken by both labs. Work from Katie Janeway’s lab has shown the SDHB protein expression is lost in most if not all pediatric GIST, in the absence of loss of function SDH mutations.

Future Directions: whole-transcriptome sequencing from 20 pediatric wild type GIST samples and 10 mutant adult cases with the SOLiD System 3 (Applied Biosystems, now Life Technologies). Sequencing mRNA will allow us to identify the majority of expressed mutations, rearrangements, and splice variants in individual tumors.

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