Cancer immunotherapy is increasingly used in clinical practice, but its success rate is reduced by tumor escape from the immune system. This may be due to the genetic instability of tumor cells, which allows them to adapt to the immune response and leads to intratumoral immune heterogeneity. The study investigated spatial immune heterogeneity in the tumor microenvironment and its possible drivers in a mouse model of tumors induced by human papillomaviruses (HPV) following immunotherapy. Gene expression was determined by RNA sequencing and mutations by whole exome sequencing. A comparison of different tumor areas revealed heterogeneity in immune cell infiltration, gene expression, and mutation composition. While the mean numbers of mutations with every impact on gene expression or protein function were comparable in treated and control tumors, mutations with high or moderate impact were increased after immunotherapy. The genes mutated in treated tumors were significantly enriched in genes associated with ECM metabolism, degradation, and interactions, HPV infection and carcinogenesis, and immune processes such as antigen processing and presentation, Toll-like receptor signaling, and cytokine production. Gene expression analysis of DNA damage and repair factors revealed that immunotherapy upregulated Apobec1 and Apobec3 genes and downregulated genes related to homologous recombination and translesion synthesis. In conclusion, this study describes the intratumoral immune heterogeneity, that could lead to tumor immune escape, and suggests the potential mechanisms involved.

• MeSH
• imunoterapie * metody   MeSH
• infekce papilomavirem imunologie   virologie   MeSH
• lidé MeSH
• modely nemocí na zvířatech * MeSH
• mutace * MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádorové mikroprostředí * imunologie   MeSH
• regulace genové exprese u nádorů MeSH
• sekvenování exomu MeSH
• únik nádoru z imunitní kontroly genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Human papillomaviruses (HPVs) induce a subset of head and neck squamous cell carcinomas (HNSCC) and anogenital cancers, particularly cervical cancer (CC). The major viral proteins that contribute to tumorigenesis are the E6 and E7 oncoproteins, whose expression is usually enhanced after the integration of viral DNA into the host genome. Recently, an alternative tumorigenesis pathway has been suggested in approximately half of HNSCC and CC cases associated with HPV infection. This pathway is characterized by extrachromosomal HPV persistence and increased expression of the viral E2, E4, and E5 genes. The E6, E7, E5, and E2 proteins have been shown to modify the expression of numerous cellular immune-related genes. The antitumor immune response is a critical factor in the prognosis of HPV-driven cancers, and its characterization may contribute to the prediction and personalization of the increasingly used cancer immunotherapy. METHODS: We analyzed the immune characteristics of HPV-dependent tumors and their association with carcinogenesis types. Transcriptomic HNSCC and CC datasets from The Cancer Genome Atlas were used for this analysis. RESULTS: Clustering with immune-related genes resulted in two clusters of HPV16-positive squamous cell carcinomas in both tumor types: cluster 1 had higher activation of immune responses, including stimulation of the antigen processing and presentation pathway, which was associated with higher immune cell infiltration and better overall survival, and cluster 2 was characterized by keratinization. In CC, the distribution of tumor samples into clusters 1 and 2 did not depend on the level of E2/E5 expression, but in HNSCC, most E2/E5-high tumors were localized in cluster 1 and E2/E5-low tumors in cluster 2. Further analysis did not reveal any association between the E2/E5 levels and the expression of immune-related genes. CONCLUSIONS: Our results suggest that while the detection of immune responses associated with preserved expression of genes encoding components of antigen processing and presentation machinery in HPV-driven tumors may be markers of better prognosis and an important factor in therapy selection, the type of carcinogenesis does not seem to play a decisive role in the induction of antitumor immunity.

• MeSH
• dlaždicobuněčné karcinomy hlavy a krku komplikace   MeSH
• infekce papilomavirem * komplikace   MeSH
• karcinogeneze genetika   MeSH
• lidé MeSH
• lidské papilomaviry MeSH
• nádory děložního čípku * MeSH
• nádory hlavy a krku * genetika   komplikace   MeSH
• onkogenní proteiny virové * genetika   metabolismus   MeSH
• Papillomavirus E7 - proteiny genetika   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Tumor-associated macrophages (TAMs) plentifully infiltrate the tumor microenvironment (TME), but their role in anti-tumor immunity is controversial. Depending on the acquired polarization, they can either support tumor growth or participate in the elimination of neoplastic cells. In this study, we analyzed the TME by RNA-seq and flow cytometry and examined TAMs after ex vivo activation. Tumors with normal and either reversibly or irreversibly decreased expression of major histocompatibility complex class I (MHC-I) molecules were induced with TC-1, TC-1/A9, and TC-1/dB2m cells, respectively. We found that combined immunotherapy (IT), composed of DNA immunization and the CpG oligodeoxynucleotide (ODN) ODN1826, evoked immune reactions in the TME of TC-1- and TC-1/A9-induced tumors, while the TME of TC-1/dB2m tumors was mostly immunologically unresponsive. TAMs infiltrated both tumor types with MHC-I downregulation, but only TAMs from TC-1/A9 tumors acquired the M1 phenotype upon IT and were cytotoxic in in vitro assay. The anti-tumor effect of combined IT was markedly enhanced by a blockade of the colony-stimulating factor-1 receptor (CSF-1R), but only against TC-1/A9 tumors. Overall, TAMs from tumors with irreversible MHC-I downregulation were resistant to the stimulation of cytotoxic activity. These data suggest the dissimilarity of TAMs from different tumor types, which should be considered when utilizing TAMs in cancer IT.

• Publikační typ
• časopisecké články MeSH

Cluster of differentiation (CD) 80 is mainly expressed in immune cells but can also be found in several types of cancer cells. This molecule may either activate or inhibit immune reactions. Here, we determined the immunosuppressive role of CD80 in the tumor microenvironment by CRISPR/Cas9-mediated deactivation of the corresponding gene in the mouse oncogenic TC-1 cell line. The tumor cells with deactivated CD80 (TC-1/dCD80-1) were more immunogenic than parental cells and induced tumors that gained sensitivity to cytotoxic T-lymphocyte antigen 4 (CTLA-4) blockade, as compared with the TC-1 cells. In vivo depletion experiments showed that the deactivation of CD80 switched the pro-tumorigenic effect of macrophages observed in TC-1-induced tumors into an anti-tumorigenic effect in TC-1/dCD80-1 tumors and induced the pro-tumorigenic activity of CD4+ cells. Moreover, the frequency of lymphoid and myeloid cells and the CTLA-4 expression by T helper (Th)17 cells were increased in TC-1/dCD80-1- compared with that in the TC-1-induced tumors. CTLA-4 blockade downregulated the frequencies of most immune cell types and upregulated the frequency of M2 macrophages in the TC-1 tumors, while it increased the frequency of lymphoid cells in TC-1/dCD80-1-induced tumors. Furthermore, the anti-CTLA-4 therapy enhanced the frequency of CD8+ T cells as well as CD4+ T cells, especially for a Th1 subset. Regulatory T cells (Treg) formed the most abundant CD4+ T cell subset in untreated tumors. The anti-CTLA-4 treatment downregulated the frequency of Treg cells with limited immunosuppressive potential in the TC-1 tumors, whereas it enriched this type of Treg cells and decreased the Treg cells with high immunosuppressive potential in TC-1/dCD80-1-induced tumors. The immunosuppressive role of tumor-cell-expressed CD80 should be considered in research into biomarkers for the prediction of cancer patients' sensitivity to immune checkpoint inhibitors and for the development of a tumor-cell-specific CD80 blockade.

• Publikační typ
• časopisecké články MeSH