肺癌是全球癌症相关死亡的主要原因，其临床结果和预后仍不理想。了解潜在的分子靶点对于精密诊断和/或治疗目的非常必要。组蛋白去乙酰化酶6（HDAC6）是一种重要的去乙酰化酶酶，是癌症治疗的有前途的靶点；然而，调节癌症发病机制的分子机制尚不清楚。基于TCGA和GEO数据库，分析了HDAC6表达水平的临床相关性及其与总生存率的相关性。使用qRT-PCR和Western blot分析评估了肺癌组织和患者来源的原发性肺癌细胞的HDAC6表达。通过蛋白质组分析确定了HDAC6的潜在调节机制，并使用特异性HDAC6抑制剂三环霉素A（TSA）和HDAC6 RNA干扰（siHDAC6）进行了验证，包括免疫泳膜法、免疫荧光法、微管沉淀和免疫光谱法（IP-MS）。使用来自患者的肺癌细胞进行2D和3D肿瘤球体形成评估了肺癌细胞生长。结果表明，HDAC6在肺癌标本中上调且与预后不良显著相关。通过TSA和siHDAC6抑制HDAC6，导致磷酸化的ERK解除微管上的依赖状态，并降低p-ERK的水平。通过TSA和siHDAC6诱导微管醋酸化介导GRP78-p-ERK从微管上解离，从而抑制癌细胞生长。抑制HDAC6可导致上调微管醋酸化，进而导致GRP78-p-ERK从微管上解离。结果，p-ERK水平降低，肺癌细胞生长被压制。本研究揭示了HDAC6作为肿瘤促进因子的有趣作用和分子机制，其抑制代表着一种有前途的抗癌疗法。 © 2023. The Author(s).

The leading cause of cancer-related mortality worldwide is lung cancer, and its clinical outcome and prognosis are still unsatisfactory. The understanding of potential molecular targets is necessary for clinical implications in precision diagnostic and/or therapeutic purposes. Histone deacetylase 6 (HDAC6), a major deacetylase enzyme, is a promising target for cancer therapy; however, the molecular mechanism regulating cancer pathogenesis is largely unknown.The clinical relevance of HDAC6 expression levels and their correlation with the overall survival rate were analyzed based on the TCGA and GEO databases. HDAC6 expression in clinical samples obtained from lung cancer tissues and patient-derived primary lung cancer cells was evaluated using qRT-PCR and Western blot analysis. The potential regulatory mechanism of HDAC6 was identified by proteomic analysis and validated by immunoblotting, immunofluorescence, microtubule sedimentation, and immunoprecipitation-mass spectrometry (IP-MS) assays using a specific inhibitor of HDAC6, trichostatin A (TSA) and RNA interference to HDAC6 (siHDAC6). Lung cancer cell growth was assessed by an in vitro 2-dimensional (2D) cell proliferation assay and 3D tumor spheroid formation using patient-derived lung cancer cells.HDAC6 was upregulated in lung cancer specimens and significantly correlated with poor prognosis. Inhibition of HDAC6 by TSA and siHDAC6 caused downregulation of phosphorylated extracellular signal-regulated kinase (p-ERK), which was dependent on the tubulin acetylation status. Tubulin acetylation induced by TSA and siHDAC6 mediated the dissociation of p-ERK on microtubules, causing p-ERK destabilization. The proteomic analysis demonstrated that the molecular chaperone glucose-regulated protein 78 (GRP78) was an important scaffolder required for p-ERK localization on microtubules, and this phenomenon was significantly inhibited by either TSA, siHDAC6, or siGRP78. In addition, suppression of HDAC6 strongly attenuated an in vitro 2D lung cancer cell growth and an in vitro 3D patient derived-lung cancer spheroid growth.HDAC6 inhibition led to upregulate tubulin acetylation, causing GRP78-p-ERK dissociation from microtubules. As a result, p-ERK levels were decreased, and lung cancer cell growth was subsequently suppressed. This study reveals the intriguing role and molecular mechanism of HDAC6 as a tumor promoter, and its inhibition represents a promising approach for anticancer therapy.© 2023. The Author(s).