Validation of hyperthermia as an enhancer of chemotherapeutic efficacy: insights from a bladder cancer organoid model

This study aimed to evaluate the combined efficacy of hyperthermia and chemotherapy using a bladder cancer organoid model and to explore hyperthermia-related molecular pathways. Tumor organoids were generated by embedding RT4 bladder cancer cells into Matrigel. The resulting organoids were treated with pirarubicin or gemcitabine at 37 °C or 42 °C. Proliferation was determined by Ki67 immunofluorescence staining, and apoptosis was assessed using a TdT-mediated dUTP nick end labeling (TUNEL) assay. RNA sequencing was used to identify the differentially expressed genes. Bladder cancer organoids were successfully established and exhibited robust proliferative abilities. Treatment with gemcitabine or pirarubicin under hyperthermic conditions caused pronounced structural damage to the organoids and increased cell death compared to that in the normothermically treated group. Furthermore, Ki67 labeling and TUNEL assays showed that the hyperthermia chemotherapy group showed a significantly reduced proliferation rate and high level of apoptosis. Finally, RNA sequencing revealed the IFN-γ signaling pathway to be associated with hyperthermia. Overall, hyperthermia combined with chemotherapy exerted better therapeutic effects than those of normothermic chemotherapy in grade 1-2 non-muscle-invasive bladder cancer, potentially through activation of the IFN-γ-JAK-STAT pathway.

本研究旨在利用膀胱癌类器官模型评估热疗联合化疗的协同疗效，并探索热疗相关的分子通路。本研究通过将RT4膀胱癌细胞接种于Matrigel（基质胶）中构建肿瘤类器官，将所得类器官分别置于37℃与42℃环境下，采用吡柔比星或吉西他滨进行处理。通过Ki67免疫荧光染色检测细胞增殖水平，利用TdT介导的dUTP缺口末端标记（TUNEL）实验评估细胞凋亡情况，同时采用RNA测序技术鉴定差异表达基因。本研究成功构建的膀胱癌类器官具备较强的增殖能力。相较于常温处理组，热疗条件下联合吉西他滨或吡柔比星处理可对类器官造成显著结构损伤，并提升细胞死亡比例。此外，Ki67标记与TUNEL实验结果显示，热疗联合化疗组的细胞增殖率显著降低，细胞凋亡水平显著升高。最终，RNA测序分析发现，IFN-γ（干扰素γ）信号通路与热疗密切相关。总体而言，针对1-2级非肌层浸润性膀胱癌，热疗联合化疗的治疗效果优于单纯常温化疗，其潜在机制可能与激活IFN-γ-JAK-STAT信号通路有关。