Progress against GIST inches ahead

Advances in diagnosis, treatment reported by American Association for Cancer Research

Editor’s note: Jerry Call, science coordinator for the Life Raft Group, attended the 96th annual meeting of the American Association for Cancer Research held April 16-20 in Anaheim, Calif. This is his report on the handful of sessions that he was able to attend.

By Jerry Call Life Raft science coordinator

A potentially useful way for testing anti-GIST drugs aimed at KIT mutation that sparks cancer was unveiled by Dr. Brian Rubin, a pathologist at the University of Washington Medical Center in Seattle.

He presented a poster, abstract no. 1977, “A Mouse Model of Gastrointestinal Stromal Tumor.” This mouse model may be useful for testing new anti- KIT based therapies.

Rubin, Dr. Christina Antonescu of Memorial Sloan- Kettering Cancer Center, and their colleagues used a genetic “knock-in” strategy to replace one normal Kit allele with a mutate allele (for each characteristic or trait, organisms inherit two alternative forms of that gene, one from each parent. These alternative forms of a gene are called alleles). The KIT mutation introduced was K641E, which is equivalent to an exon 13 mutation found in a human familial GIST syndrome family. The mice developed hyperplasia in interstitial cells of cajal (ICC) and GIST tumors that are histologically, immunohistochemically, ultrastructurally and biochemically identical to human GISTs, and thus represent an excellent model for the study of ICC biology and the pathogenesis of GIST.

FLAVOPIRIDOL

Dr. Elliot Sambol and Dr. Samuel Singer, et al, from Memorial Sloan- Kettering presented a poster, abstract no. 1504, “Flavopiridol is an active agent in the treatment of gastrointestinal stromal tumor (GIST) cells with relative resistance to imatinib mesylate via both transcriptional suppression and inhibition of autophosphorylation of KIT.”

While flavopiridol is widely known as an inhibitor of cyclin- dependent kinase (CDK2 and CDK1), its effects may be mediated more by inhibition of transcription than anything else (Cell Cycle review, December 2004).

The Sloan-Kettering team used a GIST 882 cell line for their experiments. They found that total KIT protein was significantly diminished in flavopiridol-treated cells after 24 hours. This resulted in an indirect reduction in KIT autophosphorylation, comparable to Gleevec. They confirmed that transcriptional downregulation was the reason for reduced total KIT protein, by using quantitative real time RT-PCR.

Compared to control cells, KIT mRNA was reduced by 18.25 percent and 61.1T at 150nM and 300nM of flavopiridol, respectively, after 24 hours. They also found that flavopiridol, at clinically achievable doses, induces a high level of apoptosis (cell death) in GIST cells. They believe their findings justify a clinical evaluation of flavopiridol in treating patients with Gleevec-resistant GIST.

The way flavopiridol works against KIT is different than Gleevec. Flavopiridol reduces the total amount of KIT protein, where Gleevec inhibits the activation (phosyphorylation) of the KIT protein present without lowering the amount of KIT protein. One of the mechanisms of Gleevec-resistance is overexpression of KIT or PDGFRA (too much KIT or PDGFRA protein).

MICROARRAYS in DIAGNOSIS, TREATMENT

Memorial Sloan-Kettering teamed up with Affymetrix Inc. to present a poster, abstract LB-47, “Affymetrix Gene Expression Microarrays as a Tool for Detecting Copy Number Aberrations in Gastrointestinal Stromal Tumors.”

Microarrays are a tool for examining the expression of thousands of genes at the same time. By using microarrays and special software to study the expression of different proteins in cells, researchers hope to track down the genetic changes that promote cancer.

One of the changes that can occur in a cancer cell is the loss or gain of genetic material. This can be a whole chromosome, or a smaller segment like a gene. In this study, researchers were able to detect copy number aberrations such as deletions in Chromosome 1p, loss of Chromosome 22, and amplification in Chromosome 3q. Studies are in progress to determine the smallest change that can be reliably detected using this approach.

The potential for microarrays is vast, from diagnosis to personalized treatment. In what appears to be the first small step in moving microarray technology from the lab to the clinic, Affymetrix and Roche announced the availability of the world's first microarray instrument system for clinical diagnostics last September. The AmpliChip CYP450 Test will allow diagnostic laboratories to identify certain naturally occurring variations in the drug metabolism genes, CYP2D6 and CYP2C19 (note: CYP2D6 enzyme plays a role in Gleevec metabolism, where CYP3A4 is the major enzyme involved in Gleevec metabolism). These variations affect the rate at which people metabolize many drugs used to treat depression, schizophrenia, bi-polar disorder, cardiovascular disease and more.

Knowledge of these variations, considered with other factors, can help a physician select the best drug and set the right dose for a patient sooner, as well as avoid drugs that may cause the patient to suffer adverse reactions. This testing is currently approved only in the Europe.

ANTIANGIOGENESIS

Emerging concepts in translating antiangiogenesis therapy to the clinic was the topic of a session led by Dr. Rakesh K. Jain of Massachusetts General Hospital. One of the key hypotheses of this session is that antiangiogenic agents, when used judiciously, can transiently improve the function of tumor vessels and facilitate the delivery of oxygen (a radiation sensitizer) and drugs.

This “vessel normalization” hypothesis underscores the importance of dose and schedule in combining antiangiogenic therapies with cytotoxic therapies. The hypothesis was that normal tissues have normal blood vessels. Blood vessels that feed tumors are abnormal. As antiangiogenic treatment progresses, tumor blood vessels change from abnormal to normal before they start to shrink and deteriorate. The period of transition, when vessels are changing from abnormal to normal, may represent a window of time when both radiation and delivery of cytotoxic drugs to the tumor may be improved.

The presenters hypothesized that after beginning VEGF blockage (VEGF is one of the most common/important antiangiogenic targets), tumors might be more sensitive to radiation three to six after starting VEGF blockage. They noted that this might be dependent on tumor type and, in the case of chemotherapy, the window might also be drug and dose dependent. The presenters offered a few other opinions:

That proper dose of antiangiogenic drugs is important for tumor blood vessel normalization.

That antiangiogenic therapies can have different goals, i.e., tumor starvation, improved drug delivery, sensitizer to radiation.

TARGETED CHEMOTHERAPIES

Presentations were given on some newer types of chemotherapy, with specific targets but perhaps rather general or wide-ranging effects. These included heat shock protein 90 (HSP- 90) inhibitors, proteasome inhibitors, and histone deacetylase inhibitors (HDAC inhibitors). These classes of drugs were reviewed in our report of the 2004 AACR meeting (see the April 2004 edition of the LRG newsletter).

Another class of drugs, mTOR inhibitors, with a specific target also is similar in some ways to these other drugs, in that it may treat a rather wide range of cancers due to the importance of the target to cancer cells generally. Most notable among these drugs is the addition of at least two drugs with better solubility/better formulation in the HSP-90 inhibitor class. These two drugs are IPI-504 from Infinity Pharmaceuticals, and 17-DMAG from Kosan Biosciences. 17- DMAG is in phase I clinical trials, and IPI- 504 should be entering phase I trials very soon. Impressions from the presentations are that these drugs are likely to work better when combined with other drugs (including between the 4 classes listed here) and that they may have not only synergistic benefits, but perhaps synergistic toxicities as well.

Jeffrey Tong from Infinity Pharmaceuticals gave a presentation on Infinity’s HSP-90 inhibitor, IPI-504. HSP- 90 is a protein that helps fold other proteins into their final shape. Inhibition of HSP-90 prevents proteins (including KIT) from folding into their three-dimensional shape. This indirectly inhibits KIT signaling (presumably by reducing the amount of active KIT protein). Tong described three paths for unfolded proteins.
— They can be refolded by HSP-90 protein.
— They can be degraded by a “cellular machine” called the proteasome.
— They can accumulate in the cell, triggering cell death (apoptosis). Because the first two items can both be inhibited by different drugs, there exists the possibility that combining the two classes of drugs would lead to more cell death as the unfolded proteins accumulate.

It should be noted that Dr. Julian Adams, formerly of Millennium Pharmaceuticals, developed the first drug approved in the proteasome inhibitor class, Velcade, and has since joined Infinity Pharmaceuticals and was instrumental in developing IPI-504.

THE FUTURE

Dr. Charles Sawyers was the chair of the session “The future of oncology drugs in the new era of molecularly targeted drugs.” Part of this discussion reviewed the history of Gleevec. Researchers initially had wondered if the early results of Gleevec in CML could be duplicated. They wondered whether CML, which is a genetically simple type of cancer, was in fact “real cancer.” They got their answer in the Gleevec for GIST trials. Gleevec worked very well in GIST, and GIST was indeed a “real cancer.” A PET scan of the first GIST patient that responded to Gleevec was a part of the presentation. This scan was described as “One of the most famous PET scans in oncology” by the presenter.

Some of the challenges highlighted in this session were:
— Selecting the right targets
— Discovering the right biomarkers to enable selection of the right patients
— Smarter trials

Some of the speculation is that biomarkers will evolve to pathway markers, and many of the next generation targets will not have to be activated to be crucial targets.

Presenters also discussed the concept of “synthetic lethality.” Researchers are looking for synthetic lethal interactions (such as targeting the most important, and second most important genes/targets).

Several new KIT and/or PDGFRA inhibitors had poster displays. Many of these are just starting or almost ready to start phase I trials. Since there is no indication that they were specifically designed to overcome Gleevecresistant secondary mutations in GIST, it is unknown whether any of these will have applications in GIST.

In addition to AMG706, BMS- 354825 and AMN107, which have ongoing or planned GIST trials, inhibitors included: OSI-930 from OSI Pharmaceuticals, SU14813 from Pfizer, BAY 43-9006 from Bayer Pharmaceuticals, ABT-869 from Abbott, JNJ- 10198409 (RWJ-540473) from Johnson & Johnson, AZD2171 from Astra- Zeneca, and CHIR-258 from Chiron Oncology.

Montigen Pharmaceuticals and the University of Arizona also had a poster that found that in two patients and one GIST cell line, KIT was downregulated and another protein, AXL, was upregulated. A lead compound that inhibits AXL was presented. This work requires further validation.

One group (abstract 4956) found that glucose uptake in GIST cells (such as in PET scans) is dependent on AKT, but that GIST cell survival isn’t dependent on AKT. AKT is a protein downstream in GIST.