Adoptive transfer of tumor infiltrating lymphocytes (TIL therapy) has proven highly effective for treating solid cancers, including non-small cell lung cancer (NSCLC). However, not all patients benefit from this therapy for yet unknown reasons. Defining markers that correlate with high tumor-reactivity of the autologous TIL products is thus key for achieving better tailored immunotherapies. We questioned whether the composition of immune cell infiltrates correlated with the tumor-reactivity of expanded TIL products. Unbiased flow cytometry analysis of immune cell infiltrates of 26 early-stage and 20 late-stage NSCLC tumor lesions was used for correlations with the T cell differentiation and activation status, and with the expansion rate and anti-tumor response of generated TIL products. The composition of tumor immune infiltrates was highly variable between patients. Spearman\'s Rank Correlation revealed that high B cell infiltration negatively correlated with the tumor-reactivity of the patient\'s expanded TIL products, as defined by cytokine production upon exposure to autologous tumor digest. In-depth analysis revealed that tumor lesions with high B cell infiltrates contained tertiary lymphoid structure (TLS)-related immune infiltrates, including BCL6+ antibody-secreting B cells, IgD+BCL6+ B cells and CXCR5+BLC6+ CD4+ T cells, and higher percentages of naïve CD8+ T cells. In conclusion, the composition of immune cell infiltrates in NSCLC tumors associates with the functionality of the expanded TIL product. Our findings may thus help improve patient selection for TIL therapy.

In breast cancer, triple negative (TN) breast cancer has most responses to immune checkpoint inhibitor (ICI) therapy. Lymphocyte infiltrate does not impact prognosis in Hormone receptor positive HER2 negative (HR + HER2-) breast tumors and few HR + HER2- tumors respond to ICI. We contrasted immune-associated gene expression between 119 TN and 475 HR + HER2- breast tumors from The Cancer Genome Atlas (TCGA) and confirmed our findings in 299 TN and 1369 HR + HER2- breast tumors in the METABRIC database. TN and HR+ HER2- tumors grouped into immune-high or -low tumors, both subtypes were represented in the immune-high group. The largest difference between the immune-high TN and HR + HER2- tumors was TN tumors had more abundant Th1 and Th2 CD4+ T cells while HR + HER2- tumors had more abundant fibroblasts (log2FC > 0.3; p < 10×10-10). This suggests an immune-high signature is not dictated by breast cancer subtype, but fibroblast subsets associated with worse outcome were higher in the immune-high HR + HER2- tumors.

Esophageal cancer is a highly lethal malignancy, representing 5% of all cancer-related deaths. The two main subtypes are esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC). While most research has focused on ESCC, few studies have analyzed EAC for transcriptional signatures linked to diagnosis or prognosis. In this study, we utilized single-cell RNA sequencing and bulk RNA sequencing to identify specific immune cell types that contribute to anti-tumor responses, as well as differentially expressed genes (DEGs). We have characterized transcriptional signatures, validated against a wide cohort of TCGA patients, that are capable of predicting clinical outcomes and the prognosis of EAC post-surgery with efficacy comparable to the currently accepted prognostic factors. In conclusion, our findings provide insights into the immune landscape and therapeutic targets of EAC, proposing novel immunological biomarkers for predicting prognosis, aiding in patient stratification for post-surgical outcomes, follow-up, and personalized adjuvant therapy decisions.

Pancreatic adenocarcinoma (PAAD) was characterized by dense fibrotic stroma and immunosuppressive tumor microenvironment (TME). TGFβ signaling pathways are highly activated in human cancers. However, the role of TGFβ2 in TME of PAAD remains to be elucidated. In this study, we showed that TGFβ2 was expressed at a relatively high level in PAAD tissues or cancer cells. Moreover, its high expression predicted unfavorable prognosis. In PAAD, gene set enrichment analysis showed that TGFβ2 correlated positively with leukocyte transendothelial migration, but negatively with aerobic metabolism, including oxidative phosphorylation. Results in Tumor and Immune System Interaction Database showed that TGFβ2 correlated with the infiltration of tumor-associated macrophages (TAMs), which could be attributed to that TGFβ2 promote CCL2 expression in PAAD. Moreover, correlation analysis showed that TGFβ2 could trigger cancer-associated fibroblasts (CAFs) activation in PAAD. The drug sensitivity analysis may indicate that patients with TGFβ2 high expression have higher sensitivity to chemotherapeutics, but the sensitivity to targeted drugs is still controversial. TGFβ2 could promote expansion of CAFs and infiltration of TAMs, thus participating in the construction of a fibrotic and immunosuppressive TME in PAAD. Targeting TGFβ2 could be a promising therapeutic approach, which needs to be elucidated by clinical and experimental evidences.

OBJECTIVE: B7 molecules (B7s) are crucial synergistic signals for effective immune surveillance against tumor cells. While previous studies have explored the association between the B7 family and cancer, most have been limited to specific genes or cancer subtypes.
METHODS: Our study utilized multi-omics data to investigate potential correlations between B7s expression (B7s exp.) and prognosis, clinicopathological features, somatic mutations (SMs), copy number variations (CNVs), immune characteristics, tumor microenvironment (TME), microsatellite instability, tumor mutation burden, immune checkpoint gene (ICG), and drug responsiveness in TCGA tumors. Furthermore, the connection between B7s exp. and immunotherapy (IT) performance assessed in various validated datasets. Following this, immune infiltration analysis (IIA) was conducted based on B7s exp., CNVs, or SMs in bladder cancer (BLCA), complemented by real-time PCR (RT-PCR) and protein confirmation of B7-H3.
RESULTS: Across most cancer types, B7s exp. was related to prognosis, clinicopathological characteristics, mutations, CNVs, ICG, TMB, TME. The examination of sensitivity to anticancer drugs unveiled correlations between B7 molecules and different drug sensitivities. Specific B7s exp. patterns were linked to the clinical effectiveness of IT. Using GSEA, several enriched immune-related functions and pathways were identified. Particularly in BLCA, IIA revealed significant connections between B7 CNVs, mutation status, and various immune cell infiltrates. RT-PCR confirmed elevated B7-H3 gene levels in BLCA tumor tissues.
CONCLUSIONS: This study confirmed the significance of B7s exp. and genomic changes in predicting outcomes and treatment across different cancer types. Moreover, they indicate a critical function of B7s in BLCA and their potential as IT biomarkers.

OBJECTIVE: Triple-Negative Breast Cancer (TNBC) is known for its aggressiveness and treatment challenges due to the absence of ER, PR, and HER2 receptors. Our work emphasizes the prognostic value of LCP1 (Lymphocyte cytosolic protein 1), which plays a crucial role in cell processes and immune cell activity, to predict outcomes and guide treatments in TNBC.
METHODS: We explored LCP1 as a potential biomarker in TNBC and investigated the mRNA and protein expression levels of LCP1. We investigated different databases, including GTEX, TCGA, GEO, cBioPortal and Kaplan-Meier Plotter. Immunohistochemistry on TNBC and benign tumor samples was performed to examine LCP1\'s relationship with patient clinical characteristics and macrophage markers. We also assessed survival rates, immune cell infiltration, and drug sensitivity related to LCP1 using various bioinformatics tools.
RESULTS: The results indicated that LCP1 expression was higher in TNBC tissues compared to adjacent normal tissues. However, high expression of LCP1 was significantly associated with favorable survival outcomes in patients with TNBC. Enrichment analysis revealed that genes co-expressed with LCP1 were significantly enriched in various immune processes. LCP1 showed a positive correlation with the infiltration of resting dendritic cells, M1 macrophages, and memory CD4 T cells, and a negative correlation with M2 macrophages. Further analysis suggested a link between high levels of LCP1 and increased survival outcomes in cancer patients receiving immunotherapy.
CONCLUSIONS: LCP1 may serve as a potential diagnostic and prognostic biomarker for TNBC, which was closely associated with immune cell infiltration, particularly M1 and M2 macrophages. Our findings may provide valuable insights into immunotherapeutic strategies for TNBC patients.

OBJECTIVE: This study aimed to examine the associations of FTO expression with prognosis, tumor microenvironment (TME), immune cell infiltration, immune checkpoint genes, and relevant signaling pathways in GC. Furthermore, the relationship between FTO and TGF-β was studied in GC.
METHODS: The mRNA expression and clinical survival data of GC samples were obtained from The Cancer Genome Atlas Stomach Adenocarcinoma (TCGA-STAD). TIMER2, TNM plot, and GEPIA database were used to analyze FTO expression. The associations of FTO with prognosis and clinicopathologic features were assessed using the Kaplan-Meier plotter and UALCAN database, respectively. The R software was employed to analyze its related signaling pathways and the associations with TME, immune cell infiltration, and immune checkpoint genes. GEPIA and ENCORI were used to examine the association of FTO with TGF-β expression. The SRAMP website was utilized to predict m6A modification of TGF-β. IHC, Western blot, and qPCR were used to analyze the expression levels of FTO and TGF-β in clinical gastric cancer tissue samples or gastric cancer cell lines. In addition, a m6A RNA methylation assay kit was used to determine m6A levels in gastric cancer cells.
RESULTS: FTO mRNA and protein levels were significantly elevated in GC compared to normal gastric tissues. Kaplan-Meier survival analysis suggested that upregulated FTO was associated with a worse prognosis in GC. Upregulated FTO was markedly correlated with differentiation degree, lymph node metastasis, and clinical TNM stage. GO and KEGG pathway analyses revealed that FTO-associated molecules were enriched in neuroactive ligand-receptor interaction, calcium signaling, PI3k-Akt signaling, cAMP signaling pathways, and TGF-β signaling pathways, among others. The TME score was remarkably higher in the high-FTO group than in the low-FTO group. Furthermore, FTO expression had positive correlations with different types of immune cells and immune checkpoint genes. Moreover, FTO may regulate TGF-β in an m6A RNA modification manner in GC.
CONCLUSIONS: FTO may become an independent predictive prognostic biomarker correlating with TME, immune cell infiltration, and immune checkpoint genes in gastric cancer and might influence GC progression by regulating TGF-β expression.

UNASSIGNED: Cyclin B2 (CCNB2) is associated with cell cycle progression, acting as a cell cycle checkpoint in progression of G2/M transition. In many cancer patients, it has been observed that overexpression of CCNB2 enhances tumor invasiveness and leads to adverse prognosis. However, the association of CCNB2 with the tumor microenvironment remains unclear. Therefore, it is necessary to clarify the associations of CCNB2 with the immune status and prognosis of breast carcinoma (BRCA).
UNASSIGNED: Gene expression and clinical data for BRCA were obtained from The Cancer Genome Atlas and Gene Expression Omnibus databases, followed by association analyses of CCNB2 expression with prognosis, immune cell infiltration, and immune checkpoints. This study further performed drug sensitivity analysis and constructed a prognostic nomogram for CCNB2.
UNASSIGNED: 3619 differentially expressed genes were identified in BRCA, including CCNB2 that emerged as a key gene in the network. High CCNB2 expression correlated with poor prognosis. Functional analysis demonstrated enrichment of CCNB2 co-expressed genes with the cell cycle, cancer progression, cell energy, and immune pathways. Microsatellite instability and tumor mutation burden analyses indicated CCNB2 as a candidate immunotherapy target. Tumor-infiltrating myeloid-derived suppressor cells, regulatory T cells, and T helper 2 cells were associated with CCNB2-related tumor progression and metastasis. CCNB2 expression positively correlated with immune checkpoints, indicating that high CCNB2 expression might facilitate tumor immune escape. Tumors with high CCNB2 expression showed sensitivity to phosphoinositide 3-kinase-protein kinase B-mammalian target of rapamycin and cyclin-dependent kinase (CDK) 4/6 inhibitors, and the nomogram had good prognostic predictive ability for patients with BRCA.
UNASSIGNED: CCNB2 may play a crucial role in tumorigenesis and serve as an independent prognostic biomarker associated with tumor microenvironment, tumor immune infiltration and immunotherapy in BRCA.

Osteosarcoma is a primary malignant bone tumor that affects children and young adults. Understanding the molecular mechanisms underlying osteosarcoma is critical to develop effective treatments. This study aimed to identify core genes and explore the role of intercellular communication in osteosarcoma. We used GSE87437 and GSE152048 dataset to conduct a weighted correlation network analysis (WGCNA) and identify co-expression modules. The enriched biological processes and cellular components of the genes in the steelblue module were analyzed. Next, we explored the expression, diagnostic value, correlation, and association with immune infiltrate of CCSER1 and LOC101929154. Finally, we utilized CIBERSORT algorithm to predict the infiltrated immune cells in osteosarcoma tissues. Our results identified 44 co-expression modules, and the steelblue module was mainly associated with axon development, axonogenesis, and innervation. CCSER1 and LOC101929154 were significantly upregulated in osteosarcoma tissues with poor response to preoperative chemotherapy. Moreover, the expressions of CCSER1 and LOC101929154 were positively correlated. The area under the receiver operating characteristic curve of CCSER1 and LOC101929154 was 0.800 and 0.773, respectively. The expression of CCSER1 was negatively correlated with follicular helper T cells and positively correlated with M0 macrophages, while LOC101929154 was negatively correlated with activated mast cells. Besides, CD4 memory-activated T cells were observed at lower levels in patients who responded well to chemotherapy. Our study identified core genes CCSER1 and LOC101929154 and provided insight into the intercellular communication profile in osteosarcoma. Our results suggested that targeting CCSER1, LOC101929154, and CD4 memory-activated T cells may be a promising strategy for the treatment of osteosarcoma.

OBJECTIVE: To uncover the mechanisms underlying the development of colorectal cancer (CRC), we applied bioinformatic analyses to identify key genes and experimentally validated their possible roles in CRC onset and progression.
METHODS: We performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis on differentially expressed genes (DEGs), constructed a protein-protein interaction (PPI) network to find the top 10 hub genes, and analyzed their expression in colon adenocarcinoma (COAD) and rectum adenocarcinoma (READ). We also studied the correlation between these genes and immune cell infiltration and prognosis and validated the expression of SLC9A2 in CRC tissues and cell lines using qRT-PCR and Western blotting. Functional experiments were conducted in vitro to investigate the effects of SLC9A2 on tumor growth and metastasis.
RESULTS: We found 130 DEGs, with 45 up-regulated and 85 down-regulated in CRC. GO analysis indicated that these DEGs were primarily enriched in functions related to the regulation of cellular pH, zymogen granules, and transmembrane transporter activity. KEGG pathway analysis revealed that the DEGs played pivotal roles in pancreatic secretion, rheumatoid arthritis, and the IL-17 signaling pathway. We identified 10 hub genes: CXCL1, SLC26A3, CXCL2, MMP7, MMP1, SLC9A2, SLC4A4, CLCA1, CLCA4, and ZG16. GO enrichment analysis showed that these hub genes were predominantly involved in the positive regulation of transcription. Gene expression analysis revealed that CXCL1, CXCL2, MMP1, and MMP7 were highly expressed in CRC, whereas CLCA1, CLCA4, SLC4A4, SLC9A2, SLC26A3, and ZG16 were expressed at lower levels. Survival analysis revealed that 5 key genes were significantly associated with the prognosis of CRC. Both mRNA and protein expression levels of SLC9A2 were markedly reduced in CRC tissues and cell lines. Importantly, SLC9A2 overexpression in SW480 cells led to a notable inhibition of cell proliferation, migration, and invasion. Western blotting analysis revealed that the expression levels of phosphorylated ERK (p-ERK) and phosphorylated JNK (p-JNK) proteins were significantly increased, whereas there were no significant changes in the expression levels of ERK and JNK following SLC9A2 overexpression. Correlation analysis indicated a potential link between SLC9A2 expression and the MAPK signaling pathway.
CONCLUSIONS: Our study suggests that SLC9A2 acts as a tumor suppressor through the MAPK pathway and could be a potential target for CRC diagnosis and therapy.