No abstract

Vaccines against SARS-CoV-2 have alleviated infection rates, hospitalization and deaths associated with COVID-19. In order to monitor humoral immunity, several serology tests have been developed, but the recent emergence of variants of concern has revealed the need for assays that predict the neutralizing capacity of antibodies in a fast and adaptable manner. Sensitive and fast neutralization assays would allow a timely evaluation of immunity against emerging variants and support drug and vaccine discovery efforts. Here we describe a simple, fast, and cell-free multiplexed flow cytometry assay to interrogate the ability of antibodies to prevent the interaction of Angiotensin-converting enzyme 2 (ACE2) and the receptor binding domain (RBD) of the original Wuhan-1 SARS-CoV-2 strain and emerging variants simultaneously, as a surrogate neutralization assay. Using this method, we demonstrate that serum antibodies collected from representative individuals at different time-points during the pandemic present variable neutralizing activity against emerging variants, such as Omicron BA.1 and South African B.1.351. Importantly, antibodies present in samples collected during 2021, before the third dose of the vaccine was administered, do not confer complete neutralization against Omicron BA.1, as opposed to samples collected in 2022 which show significant neutralizing activity. The proposed approach has a comparable performance to other established surrogate methods such as cell-based assays using pseudotyped lentiviral particles expressing the spike of SARS-CoV-2, as demonstrated by the assessment of the blocking activity of therapeutic antibodies (i.e. Imdevimab) and serum samples. This method offers a scalable, cost effective and adaptable platform for the dynamic evaluation of antibody protection in affected populations against variants of SARS-CoV-2.

Tumor infiltrating lymphocytes (TILs) exert their antitumor activity in the tumor microenvironment (TME). Chronic TCR stimulation, together with features of the tumor microenvironment such as hypoxia, result in T cell exhaustion. Exhausted T cells (TEX) are characterized by oxidative stress that leads to cellular accumulation of oxidized proteins. Because the proteasome complex clears oxidized proteins, we hypothesized that its capacity might be overloaded in TEX, limiting their fitness. Analysis of single-cell transcriptomics data from 19 tumor types indicated an increased proteasome activity in TEX compared to non-exhausted TILs. Higher levels of ROS-induced protein oxidation, expression of proteasome genes and proteasome activity were confirmed in human TEX, suggesting that increased proteasome activity might be a compensatory mechanism to exhaustion-associated oxidative stress. We show that pharmacological activation of the proteasome prevented T cell exhaustion, and lead to superior anti-tumor control in the context of adoptive cell therapy in mouse models. Thus, modulation of the proteasome activity is an attractive strategy to prevent T cell dysfunction promoted by the TME, and to potentially overcome this major obstacle in cancer immunotherapies against solid tumors. Citation Format: Ander de Blas, Massimo Andreatta, Asier Antoñana-Vildosola, Paloma Velasco, Leire Egia-Mendikute, Borja Jimenez-Lasheras, Samanta Romina Zanetti, Santiago J. Carmona, Endika Prieto-Fernández, Asis Palazon. Proteasome activation prevents T cell exhaustion and improves antitumor activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1354.