Protein and Peptide Pharmaceutical Laboratory, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.
University of Chinese Academy of Sciences, Beijing, 100049, China.
Breast Cancer Res. 2023 Apr 7;25(1):38. doi: 10.1186/s13058-023-01641-4.
Radiotherapy is widely applied in breast cancer treatment, while radiotherapy resistance is inevitable. TGF-β1 has been considered to be an endogenous factor for the development of radiotherapy resistance. As a large portion of TGF-β1 is secreted in an extracellular vesicles-associated form (TGF-β1), particularly in radiated tumors. Thus, the understanding of the regulation mechanisms and the immunosuppressive functions of TGF-β1 will pave a way for overcoming the radiotherapy resistance in cancer treatment.
The superoxide-Zinc-PKC-ζ-TGF-β1 pathway in breast cancer cells was identified through sequence alignments of different PKC isoforms, speculation and experimental confirmation. A series of functional and molecular studies were performed by quantitative real-time PCR, western blot and flow cytometry analysis. Mice survival and tumor growth were recorded. Student's t test or two-way ANOVA with correction was used for comparisons of groups.
The radiotherapy resulted in an increased expression of the intratumoral TGF-β1 and an enhanced infiltration of the Tregs in the breast cancer tissues. The intratumoral TGF-β1 was found mainly in the extracellular vesicles associated form both in the murine breast cancer model and in the human lung cancer tissues. Furthermore, radiation induced more TGF-β1 secretion and higher percentage of Tregs by promoting the expression and phosphorylation of protein kinase C zeta (PKC-ζ). Importantly, we found that naringenin rather than 1D11 significantly improved radiotherapy efficacy with less side effects. Distinct from TGF-β1 neutralizing antibody 1D11, the mechanism of naringenin was to downregulate the radiation-activated superoxide-Zinc-PKC-ζ-TGF-β1 pathway.
The superoxide-zinc-PKC-ζ-TGF-β1 release pathway was elucidated to induce the accumulation of Tregs, resulting in radiotherapy resistance in the TME. Therefore, targeting PKC-ζ to counteract TGF-β1 function could represent a novel strategy to overcome radiotherapy resistance in the treatment of breast cancer or other cancers.
The using of patient tissues with malignant Non-Small Cell Lung Cancer (NSCLC) was approved by the ethics committees at Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China (NCC2022C-702, from June 8th, 2022).
放射疗法广泛应用于乳腺癌治疗，但放射抵抗是不可避免的。TGF-β1 被认为是放射抵抗发展的内源性因素。由于大部分 TGF-β1 以细胞外囊泡相关形式（TGF-β1）分泌，特别是在放射肿瘤中。因此，了解 TGF-β1 的调控机制及其免疫抑制功能将为克服癌症治疗中的放射抵抗铺平道路。
通过对不同 PKC 同工型的序列比对、推测和实验验证，确定乳腺癌细胞中超氧化物-Zn-PKC-ζ-TGF-β1 途径。通过定量实时 PCR、Western blot 和流式细胞术分析进行了一系列功能和分子研究。记录小鼠的存活和肿瘤生长。使用学生 t 检验或带校正的双向 ANOVA 进行组间比较。
放射治疗导致肿瘤内 TGF-β1 表达增加，乳腺癌组织中 Tregs 浸润增强。在小鼠乳腺癌模型和人肺癌组织中，均发现肿瘤内 TGF-β1 主要以细胞外囊泡相关形式存在。此外，辐射通过促进蛋白激酶 C ζ（PKC-ζ）的表达和磷酸化，促进 TGF-β1 的分泌和 Tregs 比例升高。重要的是，我们发现，与 TGF-β1 中和抗体 1D11 相比，柚皮苷显著提高了放射治疗效果，且副作用较小。与 TGF-β1 中和抗体 1D11 不同，柚皮苷的作用机制是下调辐射激活的超氧化物-Zn-PKC-ζ-TGF-β1 途径。
阐明了超氧化物-Zn-PKC-ζ-TGF-β1 释放途径诱导 Tregs 积累，导致 TME 中的放射抵抗。因此，靶向 PKC-ζ 以拮抗 TGF-β1 功能可能代表克服乳腺癌或其他癌症放射治疗抵抗的一种新策略。
使用恶性非小细胞肺癌（NSCLC）患者组织得到中国医学科学院北京协和医学院伦理委员会的批准（NCC2022C-702，自 2022 年 6 月 8 日起）。
