Margit Hagel

The integrin family of cell adhesion receptors are important for a diverse set of biological responses during development. Although many integrins have been shown to engage a similar set of cytoplasmic effector proteins in vitro, the importance of these proteins in the biological events mediated by different integrin receptors and ligands is uncertain. We have examined the role of one of the best-characterized integrin effectors, the focal adhesion protein paxillin, by disruption of the paxillin gene in mice. Paxillin was found to be critically involved in regulating the development of mesodermally derived structures such as heart and somites. The phenotype of the paxillin(−/−) mice closely resembles that of fibronectin(−/−) mice, suggesting that paxillin is a critical transducer of signals from fibronectin receptors during early development. Paxillin was also found to play a critical role in fibronectin receptor biology ex vivo since cultured paxillin-null fibroblasts display abnormal focal adhesions, reduced cell migration, inefficient localization of focal adhesion kinase (FAK), and reduced fibronectin-induced phosphorylation of FAK, Cas, and mitogen-activated protein kinase. In addition, we found that paxillin-null fibroblasts show some defects in the cortical cytoskeleton and cell spreading on fibronectin, raising the possibility that paxillin could play a role in structures distinct from focal adhesions. Thus, paxillin and fibronectin regulate some common embryonic developmental events, possibly due to paxillin modulation of fibronectin-regulated focal adhesion dynamics and organization of the membrane cytoskeletal structures that regulate cell migration and spreading.

UNLABELLED: Aberrant signaling through the fibroblast growth factor 19 (FGF19)/fibroblast growth factor receptor 4 (FGFR 4) signaling complex has been shown to cause hepatocellular carcinoma (HCC) in mice and has been implicated to play a similar role in humans. We have developed BLU9931, a potent and irreversible small-molecule inhibitor of FGFR4, as a targeted therapy to treat patients with HCC whose tumors have an activated FGFR4 signaling pathway. BLU9931 is exquisitely selective for FGFR4 versus other FGFR family members and all other kinases. BLU9931 shows remarkable antitumor activity in mice bearing an HCC tumor xenograft that overexpresses FGF19 due to amplification as well as a liver tumor xenograft that overexpresses FGF19 mRNA but lacks FGF19 amplification. Approximately one third of patients with HCC whose tumors express FGF19 together with FGFR4 and its coreceptor klotho β (KLB) could potentially respond to treatment with an FGFR4 inhibitor. These findings are the first demonstration of a therapeutic strategy that targets a subset of patients with HCC. SIGNIFICANCE: This article documents the discovery of BLU9931, a novel irreversible kinase inhibitor that specifically targets FGFR4 while sparing all other FGFR paralogs and demonstrates exquisite kinome selectivity. BLU9931 is efficacious in tumors with an intact FGFR4 signaling pathway that includes FGF19, FGFR4, and KLB. BLU9931 is the first FGFR4-selective molecule for the treatment of patients with HCC with aberrant FGFR4 signaling.

Recent evidence suggests that blocking aberrant hedgehog pathway signaling may be a promising therapeutic strategy for the treatment of several types of cancer. Cyclopamine, a plant Veratrum alkaloid, is a natural product antagonist of the hedgehog pathway. In a previous report, a seven-membered D-ring semisynthetic analogue of cyclopamine, IPI-269609 (2), was shown to have greater acid stability and better aqueous solubility compared to cyclopamine. Further modifications of the A-ring system generated three series of analogues with improved potency and/or solubility. Lead compounds from each series were characterized in vitro and evaluated in vivo for biological activity and pharmacokinetic properties. These studies led to the discovery of IPI-926 (compound 28), a novel semisynthetic cyclopamine analogue with substantially improved pharmaceutical properties and potency and a favorable pharmacokinetic profile relative to cyclopamine and compound 2. As a result, complete tumor regression was observed in a Hh-dependent medulloblastoma allograft model after daily oral administration of 40 mg/kg of compound 28.

Abstract Outcomes for patients with advanced hepatocellular carcinoma (HCC) remain poor despite recent progress in drug development. Emerging data implicate FGF19 as a potential HCC driver, suggesting its receptor, FGFR4, as a novel therapeutic target. We evaluated fisogatinib (BLU-554), a highly potent and selective oral FGFR4 inhibitor, in a phase I dose-escalation/dose-expansion study in advanced HCC using FGF19 expression measured by IHC as a biomarker for pathway activation. For dose escalation, 25 patients received 140 to 900 mg fisogatinib once daily; the maximum tolerated dose (600 mg once daily) was expanded in 81 patients. Fisogatinib was well tolerated; most adverse events were manageable, grade 1/2 gastrointestinal events, primarily diarrhea, nausea, and vomiting. Across doses, the overall response rate was 17% in FGF19-positive patients [median duration of response: 5.3 months (95% CI, 3.7–not reached)] and 0% in FGF19-negative patients. These results validate FGFR4 as a targetable driver in FGF19-positive advanced HCC. Significance: Fisogatinib elicited clinical responses in patients with tumor FGF19 overexpression in advanced HCC. These results validate the oncogenic driver role of the FGFR4 pathway in HCC and the use of FGF19 as a biomarker for patient selection. See related commentary by Subbiah and Pal, p. 1646. This article is highlighted in the In This Issue feature, p. 1631

Paxillin is a focal adhesion scaffolding protein which was originally identified as a substrate of the oncogenic tyrosine kinase, v-src. Paxillin has been proposed to be involved in regulation of focal adhesion dynamics. Two alternatively spliced mouse paxillin cDNAs were cloned and in the process, a paxillin-related protein, Hic-5, was also identified. Cloning and characterization of Hic-5 indicates that this protein shares extensive homology with paxillin. Although Hic-5 was originally characterized as a TGF-beta-inducible gene and proposed to be a transcription factor involved in senescence, the studies here demonstrate that Hic-5 is localized to focal adhesion in REF52 cells and can interact with the focal adhesion proteins, Fak, Frnk, and vinculin. In addition, like paxillin, Hic-5 can bind to a negative regulator of Src PTKs, csk but does not bind to the adaptor protein Crk. Like paxillin, localization of this protein to focal adhesions is mediated primarily by the LIM domains; however, sequences outside the LIM domains also play a minor role in focal adhesion targeting. These results suggest that Hic-5 like paxillin could be involved in regulation of focal adhesion dynamics and raise the possibility that Hic-5 and paxillin could have overlapping or opposing functions in the overall regulation of cell growth and differentiation.