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KIF5B-RET fusion kinase promotes cell growth by multilevel activation of STAT3 in lung cancer.

ABSTRACT: BACKGROUND:Lung cancer in nonsmokers tends to be driven by a single somatic mutation or a gene fusion. KIF5B-RET fusion is an oncogene identified in non-small cell lung cancers. In this study, we verified the oncogenic activity of KIF5B-RET fusion and investigated how KIF5B-RET activates the specific signaling pathways for cellular transformation. We aimed to provide a basis for the further development of the therapy for KIF5B-RET positive lung cancer patients. METHODS:RT-PCR was used to screen for KIF5B-RET fusions in Chinese lung cancer patients. To verify the oncogenic activity of KIF5B-RET kinase in lung cancer cells, we manipulated its expression genetically followed by colony formation and tumor formation assays. The mechanism by which KIF5B-RET kinase induces proliferation was investigated by western blot, coimmunoprecipitation, and administration of RET, MAPK and STAT3 inhibitors. RESULTS:Our study identified a KIF5B-RET fusion in Chinese NSCLC patients and demonstrated that KIF5B-RET transfected cells showed a significantly increased proliferation rate and colony-forming ability. Furthermore, we found that KIF5B-RET fusion kinase induced multilevel activation of STAT3 at both Tyr705 and Ser727, and KIF5B-RET-STAT3 signaling related inhibitors repressed the proliferation and tumorigenicity of lung cancer cells significantly. CONCLUSIONS:Our data suggest that KIF5B-RET promotes the cell growth and tumorigenicity of non-small cell lung cancers through multilevel activation of STAT3 signaling, providing possible strategies for the treatment of KIF5B-RET positive lung cancers.

SUBMITTER: Qian Y

PROVIDER: S-EPMC4114102 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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KIF5B-RET fusion kinase promotes cell growth by multilevel activation of STAT3 in lung cancer.

Qian YingYing Y Chai Shoujie S Liang Zuyu Z Wang Yongfang Y Zhou You Y Xu Xia X Zhang Chenchen C Zhang Min M Si Jingxing J Huang Feiteng F Huang Zhangdan Z Hong Wei W Wang Kai K

Molecular cancer 20140721

<h4>Background</h4>Lung cancer in nonsmokers tends to be driven by a single somatic mutation or a gene fusion. KIF5B-RET fusion is an oncogene identified in non-small cell lung cancers. In this study, we verified the oncogenic activity of KIF5B-RET fusion and investigated how KIF5B-RET activates the specific signaling pathways for cellular transformation. We aimed to provide a basis for the further development of the therapy for KIF5B-RET positive lung cancer patients.<h4>Methods</h4>RT-PCR was ...[more]

PMID: 25047660

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Project description:KIF5B-RET gene rearrangement occurs in ~1% of lung adenocarcinomas. Recently, targeted agents that inhibit RET phosphorylation have been evaluated in several clinical studies; however, little is known about the role of this gene fusion in driving lung cancer. Immunohistochemistry was used to evaluate the expression of the FOXA2 protein in tumor tissues of patients with lung adenocarcinoma. KIF5B-RET fusion cells proliferated in a cohesive form and grew tightly packed with variable-sized colonies. The expression of RET and its downstream signaling molecules, including p-BRAF, p-ERK, and p-AKT, increased. In KIF5B-RET fusion cells, the intracellular expression of p-ERK was higher in the cytoplasm than in the nucleus. Two transcription factors, STAT5A and FOXA2, exhibiting significantly different expressions at the mRNA level, were finally selected. p-STAT5A was highly expressed in the nucleus and cytoplasm, whereas the expression of the FOXA2 protein was lower; however, it was much higher in the nucleus than in the cytoplasm. Compared with the expression of FOXA2 in the RET rearrangement-wild NSCLC (45.0%), high expression (3+) were observed in most RET rearrangement NSCLCs (94.4%). Meanwhile, KIF5B-RET fusion cells began to increase belatedly from day 7 and only doubled on day 9 in 2D cell culture. However, tumors in mice injected with KIF5B-RET fusion cells began to rapidly increase from day 26. In cell cycle analyses, the KIF5B-RET fusion cells in G0/G1 were increased on day 4 (50.3 ± 2.6%) compared with the empty cells (39.3 ± 5.2%; P = 0.096). Cyclin D1 and E2 expressions were reduced, whereas CDK2 expression slightly increased. pRb and p21 expression was diminished compared with the empty cells, TGF-β1 mRNA was highly expressed, and the proteins were accumulated mostly in the nucleus. Twist mRNA and protein expression was increased, whereas Snail mRNA and protein expression was decreased. Particularly, in KIF5B-RET fusion cells treated with FOXA2 siRNA, the expression of TGF-β 1 mRNA was remarkably reduced but Twist1 and Snail mRNA were increased. Our data suggest that cell proliferation and invasiveness in KIF5B-RET fusion cells are regulated by the upregulation of STAT5A and FOXA2 through the continuous activation of multiple RET downstream signal cascades, including the ERK and AKT signaling pathways. We found that TGF-β1 mRNA, where significant increments were observed in KIF5B-RET fusion cells, is regulated at the transcriptional level by FOXA2.

Project description:BackgroundRearranged during transfection (RET) gene fusions are a validated target in non-small-cell lung cancer (NSCLC). RET-selective inhibitors selpercatinib (LOXO-292) and pralsetinib (BLU-667) recently demonstrated favorable antitumor activity and safety profiles in advanced RET fusion-positive NSCLC, and both have received approval by the US Food and Drug Administration for this indication. Insights into mechanisms of resistance to selective RET inhibitors remain limited.Patients and methodsThis study was performed at five institutions. Tissue and/or cell-free DNA was obtained from patients with RET fusion-positive NSCLC after treatment with selpercatinib or pralsetinib and assessed by next-generation sequencing (NGS) or MET FISH.ResultsWe analyzed a total of 23 post-treatment tissue and/or plasma biopsies from 18 RET fusion-positive patients who received an RET-selective inhibitor (selpercatinib, n = 10; pralsetinib, n = 7; pralsetinib followed by selpercatinib, n = 1, with biopsy after each inhibitor). Three cases had paired tissue and plasma samples, of which one also had two serial resistant tissue specimens. The median progression-free survival on RET inhibitors was 6.3 months [95% confidence interval 3.6-10.8 months]. Acquired RET mutations were identified in two cases (10%), both affecting the RET G810 residue in the kinase solvent front. Three resistant cases (15%) harbored acquired MET amplification without concurrent RET resistance mutations, and one specimen had acquired KRAS amplification. No other canonical driver alterations were identified by NGS. Among 16 resistant tumor specimens, none had evidence of squamous or small-cell histologic transformation.ConclusionsRET solvent front mutations are a recurrent mechanism of RET inhibitor resistance, although they occurred at a relatively low frequency. The majority of resistance to selective RET inhibition may be driven by RET-independent resistance such as acquired MET or KRAS amplification. Next-generation RET inhibitors with potency against RET resistance mutations and combination strategies are needed to effectively overcome resistance in these patients.

Dataset Information

KIF5B-RET fusion kinase promotes cell growth by multilevel activation of STAT3 in lung cancer.

Publications

KIF5B-RET fusion kinase promotes cell growth by multilevel activation of STAT3 in lung cancer.

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