Dual-loss of PBRM1 and RAD51 identifies hyper-sensitive subset patients to immunotherapy in clear cell renal cell carcinoma.

BACKGROUND: Homologous recombination deficiency (HRD), though largely uncharacterized in clear cell renal cell carcinoma (ccRCC), was found associated with RAD51 loss of expression. PBRM1 is the second most common mutated genes in ccRCC. Here, we introduce a HRD function-based PBRM1-RAD51 ccRCC classification endowed with diverse immune checkpoint blockade (ICB) responses. METHODS: Totally 1542 patients from four independent cohorts were enrolled, including our localized Zhongshan hospital (ZSHS) cohort and Zhongshan hospital metastatic RCC (ZSHS-mRCC) cohort, The Cancer Genome Atlas (TCGA) cohort and CheckMate cohort. The genomic profile and immune microenvironment were depicted by genomic, transcriptome data and immunohistochemistry. RESULTS: We observed that PBRM1-loss ccRCC harbored enriched HRD-associated mutational signature 3 and loss of RAD51. Dual-loss of PBRM1 and RAD51 identified patients hyper-sensitive to immunotherapy. This dual-loss subtype was featured by M1 macrophage infiltration. Dual-loss was, albeit homologous recombination defective, with high chromosomal stability. CONCLUSIONS: PBRM1 and RAD51 dual-loss ccRCC indicates superior responses to immunotherapy. Dual-loss ccRCC harbors an immune-desert microenvironment but enriched with M1 macrophages. Dual-loss ccRCC is susceptible to defective homologous recombination but possesses high chromosomal stability.

NKG2A+CD8+ T cells infiltration determines immunosuppressive contexture and inferior response to immunotherapy in clear cell renal cell carcinoma.

BACKGROUND: Immunotherapy is gaining momentum, but current treatments have limitations in terms of beneficiaries. Clear cell renal cell carcinoma (ccRCC) harbors the highest expression of human leukocyte antigen E (HLA-E), ligand of NKG2A, among all solid tumors. In this study, we aim to investigate the role of NKG2A+CD8+ T cells in tumor microenvironment and its potential as a novel target in ccRCC. METHODS: This study included four independent cohorts, including 234 patients from Zhongshan cohort (ZSHC) who underwent partial or radical nephrectomy at Zhongshan Hospital, and 117 metastatic patients from metastatic Zhongshan cohort (ZSHC-metastatic renal cell carcinoma) who were treated with immune checkpoint inhibitor or tyrosine kinase inhibitor alone. We also incorporated a cohort of 530 patients diagnosed with ccRCC from The Cancer Genome Atlas (referred to as TCGA-kidney renal clear cell carcinoma) and 311 patients from CheckMate cohort for bioinformatics exploration and hypothesis validation. Fresh surgical specimens from 15 patients who underwent ccRCC surgery at Zhongshan Hospital were collected for flow cytometry analysis. Another 10 fresh surgical specimens were used to investigate the therapeutic potential of NKG2A blockade after in vitro intervention. The infiltration of NKG2A+CD8+ T cells was assessed using immunohistochemical staining, flow cytometry, and immunofluorescence staining in ZSHC cohort. RESULTS: Patients with higher infiltration of NKG2A+CD8+ T cells in ccRCC exhibited shorter overall survival and resistance to immunotherapy. NKG2A+CD8+ T cells expressed upregulated checkpoint molecules and displayed impaired effector functions, along with tissue-residency characteristics. Combination of programmed cell death protein-1 (PD-1) blockade and NKG2A blockade demonstrated an enhanced capability in reactivating CD8+ T cells effector functions. CONCLUSION: Intense infiltration of NKG2A+CD8+ T cells were associated with poorer prognosis and response to immunotherapy. NKG2A blockade combined with current immunotherapy exhibited a robust ability to reactivate CD8+ T cells effector functions.

Connexin 36 Mediates Orofacial Pain Hypersensitivity Through GluK2 and TRPA1.

Trigeminal neuralgia is a debilitating condition, and the pain easily spreads to other parts of the face. Here, we established a mouse model of partial transection of the infraorbital nerve (pT-ION) and found that the Connexin 36 (Cx36) inhibitor mefloquine caused greater alleviation of pT-ION-induced cold allodynia compared to the reduction of mechanical allodynia. Mefloquine reversed the pT-ION-induced upregulation of Cx36, glutamate receptor ionotropic kainate 2 (GluK2), transient receptor potential ankyrin 1 (TRPA1), and phosphorylated extracellular signal regulated kinase (p-ERK) in the trigeminal ganglion. Cold allodynia but not mechanical allodynia induced by pT-ION or by virus-mediated overexpression of Cx36 in the trigeminal ganglion was reversed by the GluK2 antagonist NS102, and knocking down Cx36 expression in Nav1.8-expressing nociceptors by injecting virus into the orofacial skin area of Nav1.8-Cre mice attenuated cold allodynia but not mechanical allodynia. In conclusion, we show that Cx36 contributes greatly to the development of orofacial pain hypersensitivity through GluK2, TRPA1, and p-ERK signaling.