ViroLogic, Inc., has presented the data demonstrating the ability of the company's eTag Assay System to provide a direct measure of activated Vascular Endothelial Growth Factor (VEGF) receptor type 2 (VEGFR2, also known as KDR), and its effects on downstream signal transduction pathways in an in vitro model system at the 2005 American Society of Clinical Oncology (ASCO) Annual Meeting.
Currently, there is no direct method of detecting activated VEGFR2, and an assay able to measure this, and potentially indicate its contribution to tumour blood vessel development in individual patients, could aid in the prescription of anti-angiogenic therapies. Angiogenesis is the process by which new blood vessels are generated in the body and are directed to tissues undergoing rapid growth or in need of additional nutrients. It is a critical factor in tumour growth and metastasis because tumour cells divide and grow very rapidly, and typically secrete angiogenic factors to ensure an adequate blood supply, the company says in a release.
ViroLogic's eTag assays enable detailed analysis of protein drug targets and signalling pathways in cancer cells, including samples that are formalin-fixed, paraffin-embedded, which is the standard format in most pathology labs.
There are number of peptide and protein factors that stimulate angiogenesis and a key one is VEGF, which acts through the VEGFR2 receptor. Much effort is being directed toward blocking VEGF, VEGFR2 and other angiogenic signalling pathways in different types of cancer with targeted therapeutics such as monoclonal antibodies like Avastin (bevacizumab), approved for use in colon cancer and exhibiting clinical activity in studies in lung and breast cancer, as well as small molecule VEGFR2 kinase inhibitors that are still in development.
"Our data in these cell lysate studies show that the eTag assay gives a direct measure of the activation of VEGFR2 and the signalling pathway it modulates," said Sharat Singh, ViroLogic's Chief Technical Officer, Oncology, and co-author of the study. "We are currently utilizing very similar eTag assays for activated EGFR receptors with human clinical samples. We feel that the VEGFR2 assay could also be used in this setting, first as a selection tool to determine whether a VEGF pathway inhibitor, like Avastin, is an appropriate therapy for a patient, and then potentially to measure modulation of activated VEGFR2 as a marker for efficacy of certain angiogenesis inhibitors," he added.
"Our work on clinical tumour samples to date with eTag assays has focused on activated EGFR and HER receptors, which we expect to be the focus of our first commercial oncology product," said William D. Young, Chairman and CEO of ViroLogic.
Researchers at ViroLogic stimulated Human Umbilical Vein Endothelial Cells with increasing concentrations of VEGF for different periods of time. The eTag assays were used to detect VEGFR2 homodimers, VEGFR2 phosphorylation, and downstream pathway activation in whole cells lysates. The downstream proteins assayed in the signal transduction pathway stimulated by VEGF included Erk, BAD, RSK, and Akt.
The level of VEGFR2 homodimers was found to increase with increasing doses of VEGF. Phosphorylation of VEGFR2 was transient, peaking at one minute of VEGF treatment and falling to background level after five minutes. One minute dosing of VEGF showed increasing VEGFR2 phosphorylation with increasing levels of VEGF. Downstream pathway proteins Erk, RSK, and BAD were activated with increasing levels of VEGF.
The assays can provide information on a drug's mechanism of action, selectivity and potency in a biological setting in pre-clinical research, and enable enrichment or selection of clinical trial populations later in a drug's development. In addition, ViroLogic believes these assays may ultimately be used to help physicians better determine whether certain therapies are more appropriate for individual cancer patients, and whether to combine therapies with different mechanisms or properties.