Background: Immunotherapy has shown promising results in enhancing response rates for patients with triple-negative breast cancer (TNBC). The success of immunotherapy is affected by poor tumor antigen presentation. This immune evasion is facilitated by genetic and epigenetic alterations, including aberrant RNA splicing (AS). Here we examined the role of PTBP1, a key RNA splicing factor related to immune evasion in TNBC. Methods: Clinical and gene expression data from 3,614 breast cancer patients included in the METABRIC and TCGA projects were evaluated to identify the impact of PTBP1 on TNBC. Univariate and multivariate statistical modeling was performed to test the association of PTBP1 expression with relapse-free survival (RFS), disease-free survival (DFS), and overall survival (OS). CRISPR-Cas9 technology, followed by RNA sequencing (RNA-Seq) was utilized to identify differentially activated pathways in a TNBC cell model (MDA-MB-231). A portrait of infiltrating immune cells was compiled using the xCELL algorithm. Immunofluorescence (IF) and immunohistochemistry (IHC) were employed to validate our findings in TNBC cell lines, tissue microarrays, and FFPE specimens (n = 110). Results: We found that PTBP1 is significantly upregulated in patients with TNBCs (P< 0.001). TNBC patients with high PTBP1 presented significantly shorter RFS (P= 0.018; HR = 1.66, 95%CI 1.1-2.5; n = 255), DFS (P= 0.017; HR = 3.2, 95%CI 1.16-8.3; n = 151), and OS (P= 0.037; HR = 1.48, 95%CI 1.02-2.15; n = 162). To explore potential mechanisms linking PTBP1 expression and poor survival, CRISPR-guided knockout of PTBP1, followed by RNA-seq identified a significant enhancement on antigen presentation pathways, confirmed by IF staining. xCELL analysis showed that TNBC tumors with high PTBP1 present an immune infiltration profile compatible with immune evasion (low CD4+ naïve and memory T cells, low CD8+ T cells, low tumor-associated macrophages, and high suppressor T cells). These observations were further confirmed by IHC evaluation of TNBC clinical specimens. Conclusions: This study suggests an important and relatively unexplored role of AS in immune evasion, identifying new prognostic and potentially therapeutic targets for patients with aggressive TNBC.