One of the most encouraging developments in oncology has been the success of BRAF inhibitors in -mutant melanoma. However, in contrast to its striking efficacy in-mutant melanomas, BRAF inhibitor monotherapy is ineffective in -mutant colorectal cancer. Although many studies on BRAF inhibitor resistance in colorectal cancer have focused on mechanisms underlying the reactivation of the EGFR/RAS/RAF/MEK/ERK pathway, the current study focuses on identifying novel adaptive signaling mechanisms, a fresh angle on colorectal cancer resistance to BRAF inhibition. We found that treatment with BRAF inhibitors (both current and next-generation BRAF inhibitors) upregulated the Wnt/β-catenin pathway in-mutant colorectal cancer cell lines through activating the cytoplasmic tyrosine kinase focal adhesion kinase (FAK). The results showed that FAK activation upon BRAF inhibitor treatment did not require EGFR or ERK1/2 activation, implying that BRAF inhibitor treatment-induced hyperactivation of Wnt signaling is "pathway reactivation"-independent. BRAF inhibition-induced Wnt pathway activation was further validated in preclinical models of -mutant colorectal cancer, including cell line xenograft model and a patient-derived xenograft model. Combined inhibition of BRAF/Wnt pathways or BRAF/FAK pathways exerted strong synergistic antitumor effects in cell culture model and mouse xenograft model. Overall, the current study has identified activation of the Wnt/β-catenin pathway as a novel fundamental cause of colon cancer resistance to BRAF inhibition. Our results suggest that although complete vertical pathway blockade is pivotal for effective and durable control of-mutant colorectal cancer, cotargeting parallel adaptive signaling-the Wnt/β-catenin pathway-is also essential. .

About 76% of patients with lung adenocarcinoma harbor activating mutations in the receptor tyrosine kinase (RTK)/RAS/RAF pathways, leading to aberrant activation of the mitogen‐activated protein kinase (MAPK) pathways particularly the MAPK/ERK pathway. However, many lung adenocarcinomas lacking these genomic mutations also display significant MAPK pathway activation, suggesting that additional MAPK pathway alterations remain undetected. This study has identified serine/threonine kinase mitogen‐activated protein 4 kinase 4 (MAP4K4) as a novel positive regulator of MAPK/ERK signaling in lung adenocarcinoma. The results showed that MAP4K4 was drastically elevated in lung adenocarcinoma independently of KRAS or EGFR mutation status. Knockdown of MAP4K4 inhibited proliferation, anchorage‐independent growth and migration of lung adenocarcinoma cells, and also inhibited human lung adenocarcinoma xenograft growth and metastasis. Mechanistically, we found that MAP4K4 activated ERK through inhibiting protein phosphatase 2 activity. Our results further showed that downregulation of MAP4K4 prevented ERK reactivation in EGFR inhibitor erlotinib‐treated lung adenocarcinoma cells. Together, our findings identify MAP4K4 as a novel MAPK/ERK pathway regulator in lung adenocarcinoma that is required for lung adenocarcinoma maintenance.

Background & AimsIdentification and validation of new functionally relevant and pharmacologically actionable targets for pancreatic ductal adenocarcinoma (PDAC) remains a great challenge. Premalignant acinar cell reprogramming (acinar-to-ductal metaplasia [ADM]) is a precursor of pancreatic intraepithelial neoplasia (PanIN) lesions that can progress to PDAC. This study investigated the role of proline-rich tyrosine kinase 2 (PYK2) in mutant Kras-induced and pancreatitis-associated ADM and PanIN formation, as well as in PDAC maintenance.MethodsGenetically engineered mouse models of mutant Kras (glycine 12 to aspartic acid) and Pyk2 deletion were used for investigating the role of PYK2 in PDAC genesis in mice. In vitro ADM assays were conducted using primary pancreatic acinar cells isolated from mice. Immunohistochemistry, immunofluorescence, and a series of biochemical experiments were used to investigate upstream regulators/downstream targets of PYK2 in pancreatic carcinogenesis. PDAC cell line xenograft experiments were performed to study the role of PYK2 and its downstream target in PDAC maintenance.ResultsPYK2 was increased substantially in ADM lesions induced by mutant Kras or inflammatory injury. Pyk2 deletion remarkably suppressed ADM and PanIN formation in a mutant Kras-driven and pancreatitis-associated PDAC model, whereas PYK2 knockdown substantially inhibited PDAC cell growth in vitro and in nude mice. This study uncovered a novel yes-associated protein 1/transcriptional co-activator with PDZ binding motif/signal transducer and activator of transcription 3/PYK2/β-catenin regulation axis in PDAC. Our results suggest that PYK2 contributes to PDAC genesis and maintenance by activating the Wnt/β-catenin pathway through directly phosphorylating β-cateninY654.ConclusionsThe current study uncovers PYK2 as a novel downstream effector of mutant KRAS signaling, a previously unrecognized mediator of pancreatitis-induced ADM and a novel intervention target for PDAC.

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<p>Supplementary Figure 1: BRAF inhibitor increase nuclear β-catenin in BRAF-mutant CRC cells; Supplementary Figure 2: Wnt/β-catenin pathway activation induced by BRAF inhibitor is dependent on FAK; Supplementary Figure 3: FAK inhibitor synergizes with BRAF inhibitor in the treatment of BRAF-mutant CRC; Supplementary Figure 4: BRAF inhibitor activates FAK and Wnt/β-catenin pathway in patient-derived xenografts (PDX) model; Supplementary Figure 5: Cyclin D1 is induced by BRAF inhibitor through FAK/β-catenin axis; Supplementary Figure 6: BRAF inhibitor combined with FAK inhibitor or β-catenin inhibitor strongly inhibited colony formation; Supplementary Figure 7: Next generation BRAF inhibitors activate FAK and increase cellular level of β-catenin</p>