Identification of Fangjihuangqi Decoction as a late-stage autophagy inhibitor with an adjuvant anti-tumor effect against non-small cell lung cancer.

BACKGROUND: Clinically, although chemotherapy is one of the most commonly used methods of treating tumors, chemotherapeutic drugs can induce autophagic flux and increase tumor cell resistance, leading to drug tolerance. Therefore, theoretically, inhibiting autophagy may improve the efficacy of chemotherapy. The discovery of autophagy regulators and their potential application as adjuvant anti-cancer drugs is of substantial importance. In this study, we clarified that Fangjihuangqi Decoction (FJHQ, traditional Chinese medicine) is an autophagy inhibitor, which can synergistically enhance the effect of cisplatin and paclitaxel on non-small cell lung cancer (NSCLC) cells. METHODS: We observed the changes of autophagy level in NSCLC cells under the effect of FJHQ, and verified the level of the autophagy marker protein and cathepsin. Apoptosis was detected after the combination of FJHQ with cisplatin or paclitaxel, and NAC (ROS scavenger) was further used to verify the activation of ROS-MAPK pathway by FJHQ. RESULTS: We observed that FJHQ induced autophagosomes in NSCLC cells and increased the levels of P62 and LC3-II protein expression in a concentration- and time-gradient-dependent manner, indicating that autophagic flux was inhibited. Co-localization experiments further showed that while FJHQ did not inhibit autophagosome and lysosome fusion, it affected the maturation of cathepsin and thus inhibited the autophagic pathway. Finally, we found that the combination of FJHQ with cisplatin or paclitaxel increased the apoptosis rate of NSCLC cells, due to increased ROS accumulation and further activation of the ROS-MAPK pathway. This synergistic effect could be reversed by NAC. CONCLUSION: Collectively, these results demonstrate that FJHQ is a novel late-stage autophagy inhibitor that can amplify the anti-tumor effect of cisplatin and paclitaxel against NSCLC cells.

Berbamine Hydrochloride inhibits lysosomal acidification by activating Nox2 to potentiate chemotherapy-induced apoptosis via the ROS-MAPK pathway in human lung carcinoma cells.

Autophagy is typically activated in cancer cells as a rescue strategy in response to cellular stress (e.g., chemotherapy). Herein, we found that Berbamine Hydrochloride (Ber) can act as an effective inhibitor of the late stage of autophagic flux, thereby potentiating the killing effect of chemotherapy agents. Lung carcinoma cells exposed to Ber exhibited increased autophagosomes, marked by LC3-II upregulation. The increased level of p62 after Ber treatment indicated that the autophagic flux was blocked at the late stage. The lysosome staining assay and cathepsin maturation detection indicated impaired lysosomal acidification. We found that Nox2 exhibited intensified co-localization with lysosomes in Ber-treated cells. Nox2 is a key enzyme for superoxide anion production capable of transferring electrons into the lysosomal lumen, thereby neutralizing the inner protons; this might explain the aberrant acidification. This hypothesis is further supported by the observed reversal of lysosomal cathepsin maturation by Nox2 inhibitors. Finally, Ber combined with cisplatin exhibited a synergistic killing effect on lung carcinoma cells. Further data suggested that lung carcinoma cells co-treated with Ber and cisplatin accumulated excessive reactive oxygen species (ROS), which typically activated MAPK-mediated mitochondria-dependent apoptosis. The enhanced anti-cancer effect of Ber combined with cisplatin was also confirmed in an in vivo xenograft mouse model. These findings indicate that Ber might be a promising adjuvant for enhancing the cancer cell killing effect of chemotherapy via the inhibition of autophagy. In this process, Nox2 might be a significant mediator of Ber-induced aberrant lysosomal acidification.

Integrated analysis reveals the pivotal interactions between immune cells in the melanoma tumor microenvironment.

Melanoma is the most lethal type of skin cancer. Despite the breakthroughs in the clinical treatment of melanoma using tumor immunotherapy, many patients do not benefit from these immunotherapies because of multiple immunosuppressive mechanisms. Therefore, there is an urgent need to determine the mechanisms of tumor-immune system interactions and their molecular determinants to improve cancer immunotherapy. In this study, combined analysis of microarray data and single-cell RNA sequencing data revealed the key interactions between immune cells in the melanoma microenvironment. First, differentially expressed genes (DEGs) between normal and malignant tissues were obtained using GEO2R. The DEGs were then subjected to downstream analyses, including enrichment analysis and protein-protein interaction analysis, indicating that these genes were associated with the immune response of melanoma. Then, the GEPIA and TIMER databases were used to verify the differential expression and prognostic significance of hub genes, and the relationship between the hub genes and immune infiltration. In addition, we combined single cell analysis from GSE123139 to identify immune cell types, and validated the expression of the hub genes in these immune cells. Finally, cell-to-cell communication analysis of the proteins encoded by the hub genes and their interactions was performed using CellChat. We found that the CCL5-CCR1, SELPLG-SELL, CXCL10-CXCR3, and CXCL9-CXCR3 pathways might play important roles in the communication between the immune cells in tumor microenvironment. This discovery may reveal the communication basis of immune cells in the tumor microenvironment and provide a new idea for melanoma immunotherapy.