Lidia Sambucetti

Selective growth inhibition of tumor cells by a novel histone deacetylase inhibitor, NVP-LAQ824.

We have synthesized a histone deacetylase inhibitor, NVP-LAQ824, a cinnamic hydroxamic acid, that inhibited in vitro enzymatic activities and transcriptionally activated the p21 promoter in reporter gene assays. NVP-LAQ824 selectively inhibited growth of cancer cell lines at submicromolar levels after 48–72 h of exposure, whereas higher concentrations and longer exposure times were required to retard the growth of normal dermal human fibroblasts. Flow cytometry studies revealed that both tumor and normal cells arrested in the G2-M phase of the cell cycle after compound treatment. However, an increased sub-G1 population at 48 h (reminiscent of apoptotic cells) was observed only in the cancer cell line. Annexin V staining data supported our hypothesis that NVP-LAQ824 induced apoptosis in tumor and transformed cells but not in normal cells. Western blotting experiments showed an increased histone H3 and H4 acetylation level in NVP-LAQ824-treated cancer cells, suggesting that the likely in vivo target of NVP-LAQ824 was histone deacetylase(s). Finally, NVP-LAQ824 exhibited antitumor effects in a xenograft animal model. Together, our data indicated that the activity of NVP-LAQ824 was consistent with its intended mechanism of action. This novel histone deacetylase inhibitor is currently in clinical trials as an anticancer agent.

Abstract 4193: A triple exon-skipping luciferase reporter assay identifies a new CLK inhibitor scaffold

The splicing of pre-mRNA is a critical process in normal cells and is deregulated in cancer. Compounds that modulate this process have recently been shown to target a specific vulnerability in tumors. We have developed a novel cell-based assay that specifically activates luciferase in cells exposed to SF3B1 targeted compounds, such as sudemycin D6. This assay was used to screen a combined collection of approved drugs and bioactive compounds. This screening approach identified several active hits, the most potent of which were CGP-74514A and aminopurvalanol A, both have been reported to be cyclin-dependent kinases (CDKs) inhibitors. We found that these compounds, and their analogs, show significant cdc2-like kinase (CLK) inhibition and clear structure-activity relationships (SAR) at CLKs. We prepared a set of analogs and were able to ‘dial out’ the CDK activity and simultaneously developed CLK inhibitors with low nanomolar activity. Thus, we have demonstrated the utility of our exon-skipping assay and identified new molecules that exhibit potency and selectivity for CLK, as well as some structurally related dual CLK/CDK inhibitors. Citation Format: Yihui Shi, Jaehyeon Park, Chandraiah Lagisetti, Wei Zhou, Lidia C. Sambucetti, Thomas R. Webb. A triple exon-skipping luciferase reporter assay identifies a new CLK inhibitor scaffold [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4193. doi:10.1158/1538-7445.AM2017-4193

Abstract LB-135: New cancer prevention model using imaging for early detection of mesothelin-expressing cancers

A major challenge to the discovery and development of novel cancer prevention strategies is the availability of reliable animal models. Key elements for chemoprevention testing in animal include relevance to human cancers, with tumors of similar pathology and genetic abnormalities, the presence of intermediate lesions that resemble human cancer development both histologically and molecularly; and consistent tumor incidence of >60% within less than 6 months. To our knowledge no established animal model responds to these requirements for ovarian cancer prevention. We sought to adapt the Dicerflox/flox-Ptenflox/flox double knockout (DKO) ovarian cancer mouse model (Kim, Matzuk PNAS 2012) for cancer prevention studies, by synchronized induction of the disease with intra-ovary injection of an adenovirus encoding CRE protein (Ad-mCherry-Cre). Premalignant lesions in DKO AdCRE mice were visible by HE 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-135. doi:10.1158/1538-7445.AM2017-LB-135