The Acrylonitrile Analog, VJ-289 Ablates Acute Myelogenous Leukemia Blast, Progenitor and Stem Cell Populations by Inducing Tubulin Acetylation and Caspase Activation
Authors: Zong Hongliang , Penthala Narsimha Reddy , Sen Siddhartha , Brennan Sarah , Sonar Vijayakumar , Crooks Peter A. , Guzman Monica L. 
Published: October 2019

Abstract 2496

Combretastatin A-4, a derivative of combretastatin, a natural product of the South African tree Combretum caffrum, has been reported to have anti-angiogenic and anti-tubulin effects in different cancer cell lines. We synthesized 48 novel combretastatin analogs to assess anti-leukemia activity in a panel of 12 leukemia cell lines. We identified an analog, VJ-289 [(Z)-3-(1H-indol-2-yl)-2-(3,4,5-trimethoxyphenyl)acrylonitrile] with robust anti-leukemic activity. VJ-289 showed a dose-dependent toxicity to most of the leukemic cell lines tested. The average LD50 for the 12 different leukemia cell lines was 132 nM (95% CI, 91.8–170.5). Specifically, MV4-11 cells demonstrated the most sensitivity to VJ-289 (LD50 = 66 nM), whereas THP-1 was the most resistant (LD50 = 227 nM). Furthermore, when the activity of VJ-289 was tested, five out of 14 primary AML samples demonstrated resistance to VJ-289 with an LD50 > 300nM. The average LD50 for the sensitive primary AML samples was 64.06 nM (95% CI, 35.36–92.76; N=9). Most importantly, normal CD34+ cord blood cells were significantly less affected by VJ-289 (LD50 > 500 nM).

Furthermore, VJ-289 was capable of eliminating AML progenitor/stem cells as determined by phenotypic analysis in 15 primary AML samples, colonies forming ability (N=6) and xenotransplant assays (N=6). Overall, we observed a 90.3% decrease in colony formation after treatment with 150 nM VJ-289 relative to untreated control. In contrast, VJ-289 had less impact on colony forming ability of normal hematopoietic stem/progenitor cells from cord blood cells (66.1% decrease relative to untreated; p=0.013).

To investigate the role of VJ-289 in leukemic cell apoptosis, various cell survival signaling pathways were examined. Western blotting and intracellular staining/flow cytometry data showed that caspases, including caspase 3 and 8, were activated alongside the cleavage of PARP in a dose-dependent manner. Caspase activation was observed as early as 4 h after treatment with 100 nM VJ-289. PI3K/AKT, MAPK and NF-κB were decreased upon VJ-289 treatment. Moreover, the degradation of MCL1 and the cleavage of Bcl2, two anti-apoptotic Bcl2 family members, were decreased by VJ-289 in a dose- and time-dependent manner. Interestingly, the acetylation of α-tubulin, which is critical for microtubule stabilization, and is involved in multiple cellular functions, ranging from cell motility, cell cycle progression or cell differentiation to intracellular trafficking and signaling, was transiently induced by VJ-289 within 2 hours, and was inhibited dramatically after 4 hours.

In summary, we have identified a combrestastatin A-4 analog, VJ-289, as a new anti-leukemia agent with the ability to ablate blast, progenitor and stem cell populations via induction of caspase activation and α-tubulin acetylation. Studies are underway to determine what modulates sensitivity to VJ-289 across AML specimens.

Disclosures:

No relevant conflicts of interest to declare.