Recurrent mutations in the spliceosome are observed in several human cancers, but their functional and therapeutic significance remains elusive. SF3B1, the most frequently mutated component of the spliceosome in cancer, is involved in the recognition of the branch point sequence (BPS) during selection of the 3' splice site (ss) in RNA splicing. Here, we report that common and tumor-specific splicing aberrations are induced by SF3B1 mutations and establish aberrant 3' ss selection as the most frequent splicing defect. Strikingly, mutant SF3B1 utilizes a BPS that differs from that used by wild-type SF3B1 and requires the canonical 3' ss to enable aberrant splicing during the second step. Approximately 50% of the aberrantly spliced mRNAs are subjected to nonsense-mediated decay resulting in downregulation of gene and protein expression. These findings ascribe functional significance to the consequences of SF3B1 mutations in cancer.

Genomic analyses of cancer have identified recurrent point mutations in the RNA splicing factor-encoding genes SF3B1, U2AF1, and SRSF2 that confer an alteration of function. Cancer cells bearing these mutations are preferentially dependent on wild-type (WT) spliceosome function, but clinically relevant means to therapeutically target the spliceosome do not currently exist. Here we describe an orally available modulator of the SF3b complex, H3B-8800, which potently and preferentially kills spliceosome-mutant epithelial and hematologic tumor cells. These killing effects of H3B-8800 are due to its direct interaction with the SF3b complex, as evidenced by loss of H3B-8800 activity in drug-resistant cells bearing mutations in genes encoding SF3b components. Although H3B-8800 modulates WT and mutant spliceosome activity, the preferential killing of spliceosome-mutant cells is due to retention of short, GC-rich introns, which are enriched for genes encoding spliceosome components. These data demonstrate the therapeutic potential of splicing modulation in spliceosome-mutant cancers.

A minimally invasive diagnostic assay for early detection of Alzheimer's disease (AD) is required to select optimal patient groups in clinical trials, monitor disease progression and response to treatment, and to better plan patient clinical care. Blood is an attractive source for biomarkers due to minimal discomfort to the patient, encouraging greater compliance in clinical trials and frequent testing. MiRNAs belong to the class of non-coding regulatory RNA molecules of ∼22 nt length and are now recognized to regulate ∼60% of all known genes through post-transcriptional gene silencing (RNAi). They have potential as useful biomarkers for clinical use because of their stability and ease of detection in many tissues, especially blood. Circulating profiles of miRNAs have been shown to discriminate different tumor types, indicate staging and progression of the disease and to be useful as prognostic markers. Recently their role in neurodegenerative diseases, both as diagnostic biomarkers as well as explaining basic disease etiology has come into focus. Here we report the discovery and validation of a unique circulating 7-miRNA signature (hsa-let-7d-5p, hsa-let-7g-5p, hsa-miR-15b-5p, hsa-miR-142-3p, hsa-miR-191-5p, hsa-miR-301a-3p and hsa-miR-545-3p) in plasma, which could distinguish AD patients from normal controls (NC) with >95% accuracy (AUC of 0.953). There was a >2 fold difference for all signature miRNAs between the AD and NC samples, with p-values<0.05. Pathway analysis, taking into account enriched target mRNAs for these signature miRNAs was also carried out, suggesting that the disturbance of multiple enzymatic pathways including lipid metabolism could play a role in AD etiology.

Activation of the fibroblast growth factor receptor FGFR4 by FGF19 drives hepatocellular carcinoma (HCC), a disease with few, if any, effective treatment options. While a number of pan-FGFR inhibitors are being clinically evaluated, their application to FGF19-driven HCC may be limited by dose-limiting toxicities mediated by FGFR1-3 receptors. To evade the potential limitations of pan-FGFR inhibitors, we generated H3B-6527, a highly selective covalent FGFR4 inhibitor, through structure-guided drug design. Studies in a panel of 40 HCC cell lines and 30 HCC PDX models showed that FGF19 expression is a predictive biomarker for H3B-6527 response. Moreover, coadministration of the CDK4/6 inhibitor palbociclib in combination with H3B-6527 could effectively trigger tumor regression in a xenograft model of HCC. Overall, our results offer preclinical proof of concept for H3B-6527 as a candidate therapeutic agent for HCC cases that exhibit increased expression of FGF19. .

Mutations in estrogen receptor alpha (ER) that confer resistance to existing classes of endocrine therapies are detected in up to 30% of patients who have relapsed during endocrine treatments. Since a significant proportion of therapy-resistant breast cancer metastases continue to be dependent on ER signaling, there remains a critical need to develop the next generation of ER antagonists that can overcome aberrant ER activity. Through our drug discovery efforts, we identified H3B-5942 which covalently inactivates both wild-type and mutant ER by targeting Cys530 and enforcing a unique antagonist conformation. H3B-5942 belongs to a class of ER antagonist referred to as Selective Estrogen Receptor Covalent Antagonists (SERCAs).In vitro comparisons of H3B-5942 with standard of care (SoC) 10, 2018; DOI: 10.1158/2159-8290.CD-17-1229 3Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Author Manuscript Published OnlineFirst on July SignificanceNearly 30% of endocrine-therapy resistant breast cancer metastases harbor constitutively activating mutations in ER. Selective Estrogen Receptor Covalent Antagonist (SERCA) H3B-5942 engages C530 of both ER WT and ER MUT, promotes a unique antagonist conformation, and demonstrates improved in vitro and in vivo activity over standard of care (SoC) agents.Importantly, single agent efficacy can be further enhanced by combining with CDK4/6 or mTOR inhibitors.

A minimally invasive diagnostic assay for early detection of Alzheimer's disease (AD) is required to select optimal patient groups in clinical trials, monitor disease progression and response to treatment, and to better plan patient clinical care. Blood is an attractive source for biomarkers due to minimal discomfort to the patient, encouraging greater compliance in clinical trials and frequent testing. MiRNAs belong to the class of non-coding regulatory RNA molecules of ,22 nt length and are now recognized to regulate ,60% of all known genes through post-transcriptional gene silencing (RNAi). They have potential as useful biomarkers for clinical use because of their stability and ease of detection in many tissues, especially blood. Circulating profiles of miRNAs have been shown to discriminate different tumor types, indicate staging and progression of the disease and to be useful as prognostic markers. Recently their role in neurodegenerative diseases, both as diagnostic biomarkers as well as explaining basic disease etiology has come into focus. Here we report the discovery and validation of a unique circulating 7-miRNA signature (hsa-let-7d-5p, hsa-let-7g-5p, hsa-miR-15b-5p, hsa-miR-142-3p, hsamiR-191-5p, hsa-miR-301a-3p and hsa-miR-545-3p) in plasma, which could distinguish AD patients from normal controls (NC) with .95% accuracy (AUC of 0.953). There was a .2 fold difference for all signature miRNAs between the AD and NC samples, with p-values,0.05. Pathway analysis, taking into account enriched target mRNAs for these signature miRNAs was also carried out, suggesting that the disturbance of multiple enzymatic pathways including lipid metabolism could play a role in AD etiology.

4095 Background: FGF19 overexpression is hypothesized to hyperactivate FGFR4 and its downstream signaling pathway leading to enhanced tumor growth in HCC/ICC. Targeting FGFR4 may have therapeutic benefit in HCC/ICC with altered FGF19 signaling. A phase 1 study (NCT02834780) was initiated to assess H3B-6527, an investigational highly selective covalent FGFR4 inhibitor. Methods: Adult pts with advanced HCC or ICC, ECOG PS 0-1, well compensated liver function, and who progressed after at least one prior therapy, were administered H3B-6527 orally QD (once daily) on a 21-day cycle following a 3+3 design. Patients in the dose escalation phase were treated regardless of FGF19 status. Adverse events (AEs), pharmacokinetics (PK), and pharmacodynamics (PD) were assessed. Response was determined by RECIST 1.1 or modified RECIST every 6 weeks. Results: As of 06-Jan-2019, 37 pts have been treated with H3B-6527 at doses of 300 to 1400 mg QD (23 pts in escalation; 14 in expansion). In dose escalation, a total of 17 patients with HCC, Child-Pugh A received prior systemic therapy including 100% with prior TKI and 35% with prior IO. 12% had hepatitis B virus and 47% had hepatitis C virus. H3B-6527 plasma levels increased with dose from 300 to 1000 mg QD and plateaued. H3B-6527 was rapidly absorbed with a tmax of ~2-3 h and showed a terminal half-life of ~4-5 h, following administration of 1000 mg (fasted). No dose-limiting toxicities or ≥ Grade 3 treatment-related AEs (TRAE) have been observed in escalation. Most common TRAEs (≥ 10%) were diarrhea, nausea, and vomiting. Based on safety, PK, and PD, 1000 mg QD was the recommended phase 2 dose. Durable stable disease and partial responses (PR) have been observed on the once daily fasted schedule; 2 of 17 pts with HCC achieved PRs and an additional 7 with stable disease were on treatment for ≥ 5 months. Conclusions: H3B-6527 is well tolerated and demonstrates early signs of clinical activity. Dose expansion on QD schedule and exploration of BID (twice daily) schedule is ongoing. Clinical trial information: NCT02834780.

Hepatocellular carcinoma (HCC) has limited treatment options and generally poor prognosis. Recent genomic studies have identified FGF19 as a driver oncogene in HCC. FGF19 is a gut secreted hormone that acts in the liver through FGFR4 to regulate bile acid synthesis. Consistent with the notion that FGF19 is a driver oncogene in HCC, transgenic mice overexpressing FGF19 form liver tumors and genetic ablation of FGFR4 prevented tumor formation. These data suggest targeting FGFR4 would have therapeutic benefit in HCC with altered FGF19 signaling. While a number of Pan-FGFR inhibitors are being clinically evaluated, their application to FGF19-driven HCC may be limited by their FGFR1-3 related dose limiting toxicities. Using structure guided drug design, we have generated a highly selective covalent FGFR4 inhibitor, H3B-6527. Biochemical and cellular selectivity assays showed that H3B-6527 is &gt;300 fold selective towards FGFR4 compared to other FGFR isoforms. Addition of H3B-6527 to FGF19 amplified HCC cell lines led to dose dependent inhibition of FGF19/FGFR4 signaling and concomitant reduction in cell viability. In a panel of 40 HCC cell lines, H3B-6527 selectively reduced the viability of cells that harbor FGF19 amplification and showed no effect in FGF19 non-amplified HCC cell line models. Oral dosing of H3B-6527 to mice led to dose-dependent pharmacodynamic modulation of FGFR4 signaling and tumor regression in FGF19 altered HCC cell line derived xenograft models. H3B-6527 demonstrated inhibition of tumor growth in an orthotopic liver xenograft model of FGF19 altered HCC grown in nude mice. Importantly, the inhibition of tumor growth occurred at doses that were well tolerated in mice and no evidence of FGFR1-3 related toxicities were observed at efficacious doses. In a panel of 30 HCC patient-derived xenograft (PDX) models, H3B-6527 demonstrated tumor regressions in the context of FGF19-amplified tumors. In addition, H3B-6527 showed antitumor activity and tumor regressions in PDX models with high FGF19 expression but no FGF19 amplification. The mechanism for FGF19 overexpression in the absence of gene amplification is under investigation. In conclusion, our preclinical studies demonstrate that FGF19 expression is a predictive biomarker for response to FGFR4 inhibitor therapy. Genomic analysis of public and proprietary data sets indicates that at least approximately 30% of HCC patients exhibit altered FGF19 expression and could potentially benefit from H3B-6527 monotherapy treatment. Citation Format: Anand Selvaraj, Erik Corcoran, Heather Coffey, Sudeep Prajapati, Ming-Hong Hao, Nicholas Larsen, Jennifer Tsai, Takashi Satoh, Kana Ichikawa, Julie Jaya Joshi, Raelene Hurley, Jeremy Wu, Chia-Ling Huang, Suzanna Bailey, Craig Karr, Pavan Kumar, Victoria Rimkunas, Crystal Mackenzie, Nathalie Rioux, Amy Kim, Sandeep Akare, George Lai, Lihua Yu, Peter Fekkes, John Wang, Markus Warmuth, Peter Smith, Dominic Reynolds. H3B6527, a selective and potent FGFR4 inhibitor for FGF19-driven hepatocellular carcinoma [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 3126. doi:10.1158/1538-7445.AM2017-3126