A recent report found that rare predicted loss-of-function (pLOF) variants across 13 candidate genes in TLR3- and IRF7-dependent type I IFN pathways explain up to 3.5% of severe COVID-19 cases. We performed whole-exome or whole-genome sequencing of 1,864 COVID-19 cases (713 with severe and 1,151 with mild disease) and 15,033 ancestry-matched population controls across 4 independent COVID-19 biobanks. We tested whether rare pLOF variants in these 13 genes were associated with severe COVID-19. We identified only 1 rare pLOF mutation across these genes among 713 cases with severe COVID-19 and observed no enrichment of pLOFs in severe cases compared to population controls or mild COVID-19 cases. We found no evidence of association of rare LOF variants in the 13 candidate genes with severe COVID-19 outcomes.

Various forms of oncogenic ALK proteins have been identified in various types of human cancers. While Crizotinib, an ALK inhibitor, has been found to be therapeutically useful against a subset of ALK⁺ tumours, clinical resistance to this drug has been well recognized and the mechanism of this phenomenon is incompletely understood. Using the cellular thermal shift assay (CETSA), we measured the Crizotinib—ALK binding in a panel of ALK⁺ cell lines, and correlated the findings with the ALK structure and its interactions with specific binding proteins. The Crizotinib IC₅₀ significantly correlated with Crizotinib—ALK binding. The suboptimal Crizotinib—ALK binding in Crizotinib-resistant cells is not due to the cell-specific environment, since transfection of NPM-ALK into these cells revealed substantial Crizotinib—NPM-ALK binding. Interestingly, we found that the resistant cells expressed higher protein level of β-catenin and siRNA knockdown restored Crizotinib—ALK binding (correlated with a significant lowering of IC₅₀). Computational analysis of the crystal structures suggests that β-catenin exerts steric hindrance to the Crizotinib—ALK binding. In conclusion, the Crizotinib—ALK binding measurable by CETSA is useful in predicting Crizotinib sensitivity, and Crizotinib—ALK binding is in turn dictated by the structure of ALK and some of its binding partners.

Purpose: STAT3 is known to have both oncogenic and tumor suppressive effects, but the regulation of these opposing effects is elusive. We hypothesized that STAT3β, one of the two STAT3 isoforms, is the key determinant in this context. Experimental Design: The prognostic significance of STAT3β and phospho-STAT3α^Y705 (pSTAT3α^Y705) was evaluated in 286 cases of patients with esophageal squamous cell carcinoma (ESCC). STAT3β-induced changes in the chemosensitivity to cisplatin and 5-fluorouracil were assessed both in vitro and in vivo. STAT3β-induced changes in the frequency of cancer stem cells were evaluated using Hoechst and CD44 staining. How STAT3β regulates STAT3α was determined using immunoprecipitation, confocal microscopy, DNA-binding, and chromatin immunoprecipitation-PCR. Results: STAT3β expression is an independent protective prognostic marker in patients with ESCC, which strongly correlated with longer overall survival (P = 0.0009) and recurrence-free survival (P = 0.0001). STAT3β significantly decreased the cancer stem cell population, and sensitized ESCC cells to cisplatin and 5-fluorouracil in tumor xenografts. Mechanistically, STAT3β markedly attenuated the transcription activity of STAT3α via inducing STAT3α:STAT3β heterodimers. However, the heterodimer formation decreased the binding between STAT3α and PTPN9 (better known as PTP-MEG2), a protein tyrosine phosphatase, thereby promoting the phosphorylation of STAT3α^Y705 and enhancing its nuclear translocation and DNA binding. Correlating with this, high STAT3β expression converts the prognostic value of pSTAT3α^Y705 from unfavorable to favorable in patients with ESCC. Conclusions: STAT3β suppresses chemoresistance and cancer stemness by blocking the transcriptional activity of STAT3α. The paradoxical increase in pSTAT3α^Y705 induced by STAT3β carries important implications as to how the biologic and prognostic significance of STAT3 in cancers should be interpreted. Clin Cancer Res; 22(3); 691–703. ©2015 AACR.

Loss of miR-200b promotes invasion via activating the Kindlin-2/integrin 1/AKT pathway in esophageal squamous cell carcinoma: An E-cadherin-independent mechanism Loss of miR-200b promotes invasion via activating the Kindlin-2/integrin β1/AKT pathway in esophageal squamous cell carcinoma: An E-cadherin-independent mechanism

MiR-200b is a pleiotropically acting microRNA in cancer progression, representing an attractive therapeutic target. We previously identified miR-200b as an invasiveness repressor in esophageal squamous cell carcinoma (ESCC), whereas further understanding is warranted to establish it as a therapeutic target. Here, we show that miR-200b mitigates ESCC cell growth by inducing G2-phase cell cycle arrest and apoptosis. The expression/activation of multiple key cell cycle regulators such as CDK1, CDK2, CDK4 and Cyclin B, and the Wnt/β-Catenin signaling are modulated by miR-200b. We identified CDK2 and PAF (PCNA-associated factor), two important tumor-promoting factors, as direct miR-200b targets in ESCC. Correlating with the frequent loss of miR-200b in ESCC, both CDK2 and PAF levels are significantly increased in ESCC tumors compared to case-matched normal tissues (n=119, both P<0.0001), and correlate with markedly reduced survival (P=0.007 and P=0.041, respectively). Furthermore, CDK2 and PAF are also associated with poor prognosis in certain subtypes of breast cancer (n=1802) and gastric cancer (n=233). Although CDK2 could not significantly mediate the biological function of miR-200b, PAF siRNA knockdown phenocopied while restored expression of PAF abrogated the biological effects of miR-200b on ESCC cells. Moreover, PAF was revealed to mediate the inhibitory effects of miR-200b on Wnt/β-Catenin signaling. Collectively, the pleiotropic effects of miR-200b in ESCC highlight its potential for therapeutic intervention in this aggressive disease.