Molecular and transcriptional basis of bidirectional CD4+ T cell exhaustion in oropharyngeal squamous cell carcinoma.

Tumor-infiltrating CD4+ T cells orchestrate the adaptive immune response through remarkable plasticity, and the expression patterns of exhaustion-related inhibitory receptors in these cells differ significantly from those of CD8+ T cells. Thus, a better understanding of the molecular basis of CD4+ T cell exhaustion and their responses to immune checkpoint blockade (ICB) is required. Here, we integrated multiomics approaches to define the phenotypic and molecular profiles of exhausted CD4+ T cells in oropharyngeal squamous cell carcinoma (OPSCC). Two distinct immune-promoting (Module 1) and immunosuppressive (Module 2) functional modules in tumor-infiltrating CD4+ T cells were identified, and both the immune-promoting function of Module 1 cells and immunosuppressive function of Module 2 cells were positively associated with their corresponding exhaustion states. Furthermore, the application of ICBs targeting effector CD4+ T cells in Module 1 (αPD-1) and Treg cells in Module 2 (αCTLA-4) in mouse models could help reinvigorate the effector function of Module 1-exhausted CD4+ T cells and reduce the immunosuppressive function of Module 2-exhausted CD4+ T cells, ultimately promoting OPSCC tumor regression. Taken together, our study provides a crucial cellular basis for the selection of optimal ICB in treating OPSCC.

The diversity of inhibitory receptor co-expression patterns of exhausted CD8+ T cells in oropharyngeal carcinoma.

Exhausted CD8+ T cells (Texs) are characterized by the expression of various inhibitory receptors (IRs), whereas the functional attributes of these co-expressed IRs remain limited. Here, we systematically characterized the diversity of IR co-expression patterns in Texs from both human oropharyngeal squamous cell carcinoma (OPSCC) tissues and syngeneic OPSCC model. Nearly 60% of the Texs population co-expressed two or more IRs, and the number of co-expressed IRs was positively associated with superior exhaustion and cytotoxicity phenotypes. In OPSCC patients, programmed cell death-1 (PD-1) blockade significantly enhanced PDCD1-based co-expression with other IR genes, whereas dual blockades of PD-1 and cytotoxic T lymphocyte-associated protein 4 (CTLA-4) significantly upregulated CTLA4-based co-expression with other IR genes. Collectively, our findings demonstrate that highly diverse IR co-expression is a leading feature of Texs and represents their functional states, which might provide essential clues for the rational selection of immune checkpoint inhibitors in treating OPSCC.

mRNA-LNP vaccination-based immunotherapy augments CD8+ T cell responses against HPV-positive oropharyngeal cancer.

Although mRNA vaccines are known as potent activators of antigen-specific immune responses against infectious diseases, limited understanding of how they drive the functional commitment of CD8+ T cells in tumor microenvironment (TME) and secondary lymphoid organs hinders their broader application in cancer immunotherapy. Here, we systematically evaluated the immunological effects of a lipid nanoparticle (LNP)-encapsulated mRNA vaccine that encodes human papillomavirus E7 protein (HPV mRNA-LNP), a tumor-specific antigen of HPV-positive oropharyngeal squamous cell carcinoma (OPSCC). HPV mRNA-LNP vaccination activated overall and HPV-specific CD8+ T cells, as well as differentially drove the functional commitment of CD8+ T cells through distinct IFN-response and exhaustion trajectories in the spleen and TME, respectively. Combination therapies of HPV mRNA-LNP vaccination with immune checkpoint blockades boosted HPV-specific CD8+ T cells while maintaining their anti-tumor function, thus further promoting tumor regression. Our results showed that the HPV mRNA-LNP vaccination combined with immune checkpoint blockade is a promising approach for immunotherapy of HPV-positive OPSCC.

Is postauricular injection a systemic or a topical route for inner ear drug delivery?

The challenges associated with the blood-labyrinth barrier and unwanted systemic adverse effects severely impede the development of systemic drug administration for inner ear therapy. Previous studies have demonstrated that postauricular injection might be an alternative route for the systemic administration for inner ear therapy because it is minimally invasive and can achieve a high drug concentration in the inner ear. However, the in vivo effects remain unclear. This study aimed to compare differences in cisplatin-induced ototoxicity and systemic adverse effects between postauricular and systemic administration. Here, we compared the differences in hair cell loss, body weight change, and renal dysfunction (indicated by serum levels of blood urea nitrogen and creatinine) between these two routes of administration. The results showed that although postauricular injection of cisplatin might induce greater hair cell damage in the ipsilateral cochleae than that after intraperitoneal injection, significant hair cell damage in the contralateral cochleae as well as systemic adverse effects were also observed. Our results indicated that: (1) topical administration effects might occur in the ipsilateral inner ear following postauricular injection, and (2) systemic circulation might also be an underestimated route of drug delivery to the inner ear following postauricular injection. Overall, postauricular injection is a hybrid administration route, which simultaneously shares the features of topical and systemic administration for inner ear drug delivery.

N-Glycosylation Engineering to Improve the Constitutive Expression of Rhizopus oryzae Lipase in Komagataella phaffii.

Our previous studies demonstrated that the N-glycans in Rhizopus chinensis lipase (RCL) was important for its secretion. In order to improve the secretion of Rhizopus oryzae lipase (ROL) under the control of the GAP promoter in Komagataella phaffii, two extra N-glycosylation sites were introduced in ROL according to the position of the N-glycosylation sites of RCL by sequence alignment. The results indicated that the secretion level of ROL was strongly improved by N-glycosylation engineering, and the highest value of extracellular enzyme activity was increased from 0.4 ± 0.2 U/mL to 207 ± 6 U/mL in a shake flask. In the 7-L fermenter, the extracellular enzyme activity of the mutant (2600 ± 43 U/mL) and the total protein concentration (2.5 ± 0.2 g/L) were 218- and 6.25-fold higher than these of the parent, respectively. This study presents a strategy for constitutive recombinant expression of ROL using the GAP promoter combined with N-glycosylation engineering, providing a potential enzyme for application in the food industry.