ACAP1 Deficiency Predicts Inferior Immunotherapy Response in Solid Tumors

SIMPLE SUMMARY: ACAP1 plays a key role in endocytic recycling, which is essential for the normal function of lymphocytes. This study aimed to assess the expression and function of ACAP1 in lymphocytes. In this study, we revealed that ACAP1 is expressed primarily in lymphocytes and is necessary for the normal function of lymphocytes. Its expression is regulated by promoter DNA methylation and the transcription factor SPI1. ACAP1 levels positively correlate with the infiltrating levels of tumor-infiltrating lymphocytes (TILs) across a broad range of solid cancer types. Moreover, ACAP1 deficiency is associated with poor prognosis and immunotherapeutic response in multiple cancer types treated with immunotherapy. Thus, this study demonstrates that ACAP1 is a novel lymphocyte marker. We propose a widely applicable indicator to predict response to immunotherapy that may guide cancer patient stratification for appropriate therapy. ABSTRACT: Background: ACAP1 plays a key role in endocytic recycling, which is essential for the normal function of lymphocytes. However, the expression and function of ACAP1 in lymphocytes have rarely been studied. Methods: Large-scale genomic data, including multiple bulk RNA-sequencing datasets, single-cell sequencing datasets, and immunotherapy cohorts, were exploited to comprehensively characterize ACAP1 expression, regulation, and function. Gene set enrichment analysis (GSEA) was used to uncover the pathways associated with ACAP1 expression. Eight algorithms, including TIMER, CIBERSORT, CIBERSORT-ABS, QUANTISEQ, xCELL, MCPCOUNTER, EPIC, and TIDE, were applied to estimate the infiltrating level of immune cells. Western blotting, qPCR, and ChIP-PCR were used to validate the findings from bioinformatic analyses. A T-cell co-culture killing assay was used to investigate the function of ACAP1 in lymphocytes. Results: ACAP1 was highly expressed in immune-related tissues and cells and minimally in other tissues. Moreover, single-cell sequencing analysis in tumor samples revealed that ACAP1 is expressed primarily in tumor-infiltrating lymphocytes (TILs), including T, B, and NK cells. ACAP1 expression is negatively regulated by promoter DNA methylation, with its promoter hypo-methylated in immune cells but hyper-methylated in other cells. Furthermore, SPI1 binds to the ACAP1 promoter and positively regulates its expression in immune cells. ACAP1 levels positively correlate with the infiltrating levels of TILs, especially CD8+ T cells, across a broad range of solid cancer types. ACAP1 deficiency is associated with poor prognosis and immunotherapeutic response in multiple cancer types treated with checkpoint blockade therapy (ICT). Functionally, the depletion of ACAP1 by RNA interference significantly impairs the T cell-mediated killing of tumor cells. Conclusions: Our study demonstrates that ACAP1 is essential for the normal function of TILs, and its deficiency indicates an immunologically “cold” status of tumors that are resistant to ICT.