Tumor-associated macrophage enhances PD-L1-mediated immune escape of bladder cancer through PKM2 dimer/STAT3 complex nuclear translocation

Abstract Background Tumor-associated macrophages (TAMs) are important components of the tumor microenvironment (TME) and strongly associated with poor prognosis and drug resistance, including immune checkpoint blockade (ICB) in solid tumor patients. However, the mechanism by which TAM affects immune metabolism reprogramming and immune checkpoint signaling pathway in the bladder cancer (BLCA) TME remains elusive. Methods We analyzed the TCGA cohort of BLCA patients in relation to M2-TAMs signature genes as well as PD-L1 expression, and then explored the cellular and molecular roles by RNA sequencing on constructed BLCA cells with M2-TAMs co-culture systems. we used nucleoplasmic separation and protein cross-linking to search for mechanisms related to tumor metabolism and immune reprogramming following crosstalk effects of M2-TAMs, and constructed a mouse model of BLCA with a high occupancy rate of M2-TAMs and a group of organoids derived from human BLCA fresh tissues to validate therapeutic efficacy of the targets in vivo and in vitro. Results Transforming growth factor-beta (TGF-β) secreted by M2-TAMs increased the level of glycolysis and was involved in PD-L1-mediated immune escape through pyruvate kinase isoenzyme type M2 (PKM2) in the malignant BLCA microenvironment. TGF-β promoted the nuclear translocation of PKM2 dimers in conjunction with phosphorylated signal transducer and activator of transcription (p-STAT3), which exerts its kinase activity, and subsequently promotes PD-L1 expression through the transcription factor role of STAT3. Moreover, the high secretion level of TGF-β activated the JAK2/STAT3 signaling pathway to upregulate PD-L1 expression in BLCA. In vivo, SB-431542 (a TGF-β blocker) and shikonin (a PKM2 inhibitor) significantly reduced PD-L1 expression and inhibited BLCA growth by increasing immune infiltration. In addition, SB-431542 and shikonin inhibited the growth of bladder carcinoma organoids. Conclusions Our work sheds new light on that M2-TAM-derived TGF-β promotes PD-L1-mediated immune evasion in BLCA by increasing the PKM2/STAT3 complex nuclear translocation. Targeting TGF-β receptor blockade and PKM2 inhibition significantly reduces BLCA progression and immunosuppression, and their combination provides a promising target for the treatment of BLCA.