Anti-Angiogenic Effects of Sorafenib in Relapsed Chronic Lymphocytic Leukemia: Correlative Studies of a Phase 2 Clinical Trial.

Chronic lymphocytic leukemia (CLL) is a malignant B-cell disorder that remains a largely incurable disease. We and others have previously shown that angiogenesis is important in the pathogenesis of CLL. Sorafenib is a small molecule signal transduction inhibitor that exhibits anti-tumor activity through a number of mechanisms including blocking of the Raf/MEK/ERK pathway and inhibition of receptor tyrosine kinases involved in angiogenic signaling such as the VEGF receptors. The data presented here are correlative studies performed on a Phase 2 clinical trial designed to measure the overall response rate to oral doses of sorafenib in CLL patients who have failed previous nucleoside analog therapy (alone or in combination with alkylating agents) or monoclonal antibody therapy. Each cycle consisted of 400 mg of sorafenib administered orally twice a day for 28 days. Treatment continued until any of the withdrawal criteria (unacceptable toxicity, disease progression, patient’s decision to withdraw) were reached. Peripheral blood samples for correlative studies were obtained before initiation of therapy, after 12 weeks (beginning of cycle 4) and after 24 weeks (beginning of cycle 7), or when the patient went off-study for any indication. In these studies, we specifically focused on examining VEGF and the VEGF receptors VEGFR-1 (flt-1), VEGFR-2 (KDR), VEGFR-3 (flt-4), neuropilin-1 and neuropilin-2. These molecules were analyzed at the transcript level by RT-PCR and at the protein level by western blotting or ELISA. In addition we examined plasma levels of IL-8 by ELISA, since IL-8 has been recently shown to be an important mediator of cell survival and angiogenesis in CLL. Specimens were analyzed from a total of five patients. Plasma IL-8 levels were markedly decreased in all three patients who underwent therapy beyond 12 weeks. The two patients went off study at or before 12 weeks did not show any decrease in IL-8 levels. No decrease was noted between the pre-treatment and post-treatment plasma concentrations of VEGF for any of the patients. No consistent change in cellular expression was observed between the pre-treatment and post-treatment samples for VEGF, VEGFR-1, VEGFR-2 or VEGFR-3. Neuropilin-1 expression was decreased or absent in the post therapy samples in patients who received therapy beyond 12 weeks, while no change in expression was seen in the two patients who went off study at or before 12 weeks. Neuropilin-2 expression was not detected in any of the samples. These results suggest that sorafenib suppresses angiogenic pathways by inhibiting secretion of IL-8 and expression of neuropilin-1. Additional studies and samples are needed to confirm these interesting findings, and to further delineate the anti-angiogenic effects of sorafenib in CLL.