Chen Guodong, Kong Peizhong, Yang Miaomiao, Hu Wanglai, Prise Kevin M, Yu K N, Cui Shujun, Qin Feng, Meng Gang, Almahi Waleed Abdelbagi, Nie Lili, Han Wei
Anhui Province Key Laboratory of Medical Physics and Technology, Institute of Health and Medical Technology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei, PR China; Hefei Cancer Hospital, Chinese Academy of Sciences, Hefei, PR China.
Anhui Province Key Laboratory of Medical Physics and Technology, Institute of Health and Medical Technology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei, PR China; University of Science and Technology of China, Hefei, PR China; Clinical Pathology Center, The Fourth Affiliated Hospital of Anhui Medical University, Hefei, PR China.
Int J Radiat Oncol Biol Phys. 2022 Apr 1;112(5):1216-1228. doi: 10.1016/j.ijrobp.2021.11.023. Epub 2021 Nov 26.
Radioresistance is a major cause of treatment failure in tumor radiation therapy, and the underlying mechanisms of radioresistance are still elusive. Golgi phosphoprotein 3 (GOLPH3) has been reported to associate tightly with cancer progression and chemoresistance. Herein, we explored whether GOLPH3 mediated radioresistance of lung adenocarcinoma (LUAD) and whether targeted suppression of GOLPH3 sensitized LUAD to radiation therapy.
The aberrant expression of GOLPH3 was evaluated by immunohistochemistry in LUAD clinical samples. To evaluate the association between GOLPH3 and radioresistance, colony formation and apoptosis were assessed in control and GOLPH3 knockdown cells. γ-H2AX foci and level determination and micronucleus test were used to analyze DNA damage production and repair. The rescue of GOLPH3 knockdown was then performed by exogenous expression of small interfering RNA-resistant mutant GOLPH3 to confirm the role of GOLPH3 in DNA damage repair. Mechanistically, the effect of GOLPH3 on regulating stability and nuclear accumulation of epidermal growth factor receptor (EGFR) and the activation of DNA-dependent protein kinase (DNA-PK) were investigated by quantitative real-time polymerase chain reaction, western blot, immunofluorescence, and coimmunoprecipitation. The role of GOLPH3 in vivo in radioresistance was determined in a xenograft model.
In tumor tissues of 33 patients with LUAD, the expression of GOLPH3 showed significant increases compared with those in matched normal tissues. Knocking down GOLPH3 reduced the clonogenic capacity, impaired double-strand break (DSB) repair, and enhanced apoptosis after irradiation. In contrast, reversal of GOLPH3 depletion rescued the impaired repair of radiation-induced DSBs. Mechanistically, loss of GOLPH3 accelerated the degradation of EGFR in lysosome, causing the reduction in EGFR levels, thereby weakening nuclear accumulation of EGFR and attenuating the activation of DNA-PK. Furthermore, adenovirus-mediated GOLPH3 knockdown could enhance the ionizing radiation response in the LUAD xenograft model.
GOLPH3 conferred resistance of LUAD to ionizing radiation via stabilizing EGFR, and targeted suppression of GOLPH3 might be considered as a potential therapeutic strategy for sensitizing LUAD to radiation therapy.
放射抗性是肿瘤放射治疗中治疗失败的主要原因，而放射抗性的潜在机制仍不清楚。据报道，高尔基体磷蛋白3（GOLPH3）与癌症进展和化学抗性密切相关。在此，我们探讨了GOLPH3是否介导肺腺癌（LUAD）的放射抗性，以及靶向抑制GOLPH3是否能使LUAD对放射治疗敏感。
通过免疫组织化学评估LUAD临床样本中GOLPH3的异常表达。为了评估GOLPH3与放射抗性之间的关联，在对照细胞和GOLPH3敲低细胞中评估集落形成和凋亡。使用γ-H2AX焦点和水平测定以及微核试验来分析DNA损伤的产生和修复。然后通过外源性表达小干扰RNA抗性突变体GOLPH3来挽救GOLPH3敲低，以确认GOLPH3在DNA损伤修复中的作用。从机制上讲，通过定量实时聚合酶链反应、蛋白质免疫印迹、免疫荧光和免疫共沉淀研究了GOLPH3对调节表皮生长因子受体（EGFR）稳定性和核积累以及DNA依赖性蛋白激酶（DNA-PK）激活的影响。在异种移植模型中确定了GOLPH3在体内对放射抗性的作用。
在33例LUAD患者的肿瘤组织中，GOLPH3的表达与配对的正常组织相比显著增加。敲低GOLPH3可降低克隆形成能力，损害双链断裂（DSB）修复，并增强照射后的凋亡。相反，GOLPH3缺失的逆转挽救了辐射诱导的DSB修复受损。从机制上讲，GOLPH3的缺失加速了EGFR在溶酶体中的降解，导致EGFR水平降低，从而减弱EGFR的核积累并减弱DNA-PK的激活。此外，腺病毒介导的GOLPH3敲低可增强LUAD异种移植模型中的电离辐射反应。
GOLPH3通过稳定EGFR赋予LUAD对电离辐射的抗性，靶向抑制GOLPH3可能被视为使LUAD对放射治疗敏感的潜在治疗策略。
