Antiproliferative small-molecule inhibitors of transcription factor LSF reveal oncogene addiction to LSF in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide. Despite the prevalence of HCC, there is no effective, systemic treatment. The transcription factor LSF is a promising protein target for chemotherapy; it is highly expressed in HCC patient samples and cell lines, and promotes oncogenesis in rodent xenograft models of HCC. Here, we identify small molecules that effectively inhibit LSF cellular activity. The lead compound, factor quinolinone inhibitor 1 (FQI1), inhibits LSF DNA-binding activity both in vitro, as determined by electrophoretic mobility shift assays, and in cells, as determined by ChIP. Consistent with such inhibition, FQI1 eliminates transcriptional stimulation of LSF-dependent reporter constructs. FQI1 also exhibits antiproliferative activity in multiple cell lines. In LSF-overexpressing cells, including HCC cells, cell death is rapidly induced; however, primary or immortalized hepatocytes are unaffected by treatment with FQI1. The highly concordant structure-activity relationship of a panel of 23 quinolinones strongly suggests that the growth inhibitory activity is due to a single biological target or family. Coupled with the striking agreement between the concentrations required for antiproliferative activity (GI(50)s) and for inhibition of LSF transactivation (IC(50)s), we conclude that LSF is the specific biological target of FQIs. Based on these in vitro results, we tested the efficacy of FQI1 in inhibiting HCC tumor growth in a mouse xenograft model. As a single agent, tumor growth was dramatically inhibited with no observable general tissue cytotoxicity. These findings support the further development of LSF inhibitors for cancer chemotherapy.

STK38L kinase ablation promotes loss of cell viability in a subset of KRAS-dependent pancreatic cancer cell lines.

Pancreatic ductal adenocarcinomas (PDACs) are highly aggressive malignancies, associated with poor clinical prognosis and limited therapeutic options. Oncogenic KRAS mutations are found in over 90% of PDACs, playing a central role in tumor progression. Global gene expression profiling of PDAC reveals 3-4 major molecular subtypes with distinct phenotypic traits and pharmacological vulnerabilities, including variations in oncogenic KRAS pathway dependencies. PDAC cell lines of the aberrantly differentiated endocrine exocrine (ADEX) subtype are robustly KRAS-dependent for survival. The KRAS gene is located on chromosome 12p11-12p12, a region amplified in 5-10% of primary PDACs. Within this amplicon, we identified co-amplification of KRAS with the STK38L gene in a subset of primary human PDACs and PDAC cell lines. Therefore, we determined whether PDAC cell lines are dependent on STK38L expression for proliferation and viability. STK38L encodes a serine/threonine kinase, which shares homology with Hippo pathway kinases LATS1/2. We show that STK38L expression is elevated in a subset of primary PDACs and PDAC cell lines displaying ADEX subtype characteristics, including overexpression of mutant KRAS. RNAi-mediated depletion of STK38L in a subset of ADEX subtype cell lines inhibits cellular proliferation and induces apoptosis. Concomitant with these effects, STK38L depletion causes increased expression of the LATS2 kinase and the cell cycle regulator p21. LATS2 depletion partially rescues the cytostatic and cytotoxic effects of STK38L depletion. Lastly, high STK38L mRNA expression is associated with decreased overall patient survival in PDACs. Collectively, our findings implicate STK38L as a candidate targetable vulnerability in a subset of molecularly-defined PDACs.

MEK and TAK1 Regulate Apoptosis in Colon Cancer Cells with KRAS-Dependent Activation of Proinflammatory Signaling.

MEK inhibitors have limited efficacy in treating RAS-RAF-MEK pathway-dependent cancers due to feedback pathway compensation and dose-limiting toxicities. Combining MEK inhibitors with other targeted agents may enhance efficacy. Here, codependencies of MEK, TAK1, and KRAS in colon cancer were investigated. Combined inhibition of MEK and TAK1 potentiates apoptosis in KRAS-dependent cells. Pharmacologic studies and cell-cycle analyses on a large panel of colon cancer cell lines demonstrate that MEK/TAK1 inhibition induces cell death, as assessed by sub-G1 accumulation, in a distinct subset of cell lines. Furthermore, TAK1 inhibition causes G2-M cell-cycle blockade and polyploidy in many of the cell lines. MEK plus TAK1 inhibition causes reduced G2-M/polyploid cell numbers and additive cytotoxic effects in KRAS/TAK1-dependent cell lines as well as a subset of BRAF-mutant cells. Mechanistically, sensitivity to MEK/TAK1 inhibition can be conferred by KRAS and BMP receptor activation, which promote expression of NF-κB-dependent proinflammatory cytokines, driving tumor cell survival and proliferation. MEK/TAK1 inhibition causes reduced mTOR, Wnt, and NF-κB signaling in TAK1/MEK-dependent cell lines concomitant with apoptosis. A Wnt/NF-κB transcriptional signature was derived that stratifies primary tumors into three major subtypes: Wnt-high/NF-κB-low, Wnt-low/NF-κB-high and Wnt-high/NF-κB-high, designated W, N, and WN, respectively. These subtypes have distinct characteristics, including enrichment for BRAF mutations with serrated carcinoma histology in the N subtype. Both N and WN subtypes bear molecular hallmarks of MEK and TAK1 dependency seen in cell lines. Therefore, N and WN subtype signatures could be utilized to identify tumors that are most sensitive to anti-MEK/TAK1 therapeutics. IMPLICATIONS: This study describes a potential therapeutic strategy for a subset of colon cancers that are dependent on oncogenic KRAS signaling pathways, which are currently difficult to block with selective agents. Mol Cancer Res; 14(12); 1204-16. ©2016 AACR.