Extent Of Immune Infiltration Research Articles

A lactate‐responsive gene signature predicts the prognosis and immunotherapeutic response of patients with triple‐negative breast cancer

AbstractBackgroundIncreased glycolytic activity and lactate production are characteristic features of triple‐negative breast cancer (TNBC). The aim of this study was to determine whether a subset of lactate‐responsive genes (LRGs) could be used to classify TNBC subtypes and predict patient outcomes.MethodsLactate levels were initially measured in different breast cancer (BC) cell types. Subsequently, MDA‐MB‐231 cells treated with 2‐Deoxy‐d‐glucose or l‐lactate were subjected to RNA sequencing (RNA‐seq). The gene set variation analysis algorithm was utilized to calculate the lactate‐responsive score, conduct a differential analysis, and establish an association with the extent of immune infiltration. Consensus clustering was then employed to classify TNBC patients. Tumor immune dysfunction and exclusion, cibersort, single‐sample gene set enrichment analysis, and EPIC, were used to compare the tumor‐infiltrating immune cells between TNBC subtypes and predict the response to immunotherapy. Furthermore, a prognostic model was developed by combining 98 machine learning algorithms, to assess the predictive significance of the LRG signature. The predictive value of immune infiltration and the immunotherapy response was also assessed. Finally, the association between lactate and various anticancer drugs was examined based on expression profile similarity principles.ResultsWe found that the lactate levels of TNBC cells were significantly higher than those of other BC cell lines. Through RNA‐seq, we identified 14 differentially expressed LRGs in TNBC cells under varying lactate levels. Notably, this LRG signature was associated with interleukin‐17 signaling pathway dysregulation, suggesting a link between lactate metabolism and immune impairment. Furthermore, the LRG signature was used to categorize TNBC into two distinct subtypes, whereby Subtype A was characterized by immunosuppression, whereas Subtype B was characterized by immune activation.ConclusionWe identified an LRG signature in TNBC, which could be used to predict the prognosis of patients with TNBC and gauge their response to immunotherapy. Our findings may help guide the precision treatment of patients with TNBC.

Pan-cancer analysis reveals PDK family as potential indicators related to prognosis and immune infiltration

Pyruvate dehydrogenase kinases (PDKs) play a key role in glucose metabolism by exerting negative regulation over pyruvate dehyrogenase complex (PDC) activity through phosphorylation. Inhibition of PDKs holds the potential to enhance PDC activity, prompting cells to adopt a more aerobic metabolic profile. Consequently, PDKs emerge as promising targets for condition rooted in metabolic dysregulation, including malignance and diabetes. However, a comprehensive exploration of the distinct contribution of various PDK family members, particularly PDK3, across diverse tumor types remain incomplete. This study undertakes a systematic investigation of PDK family expression patterns, forging association with clinical parameters, using data from the TCGA and GTEx datasets. Survival analysis of PDKs is executed through both Kaplan–Meier analysis and COX regression analysis. Furthermore, the extent of immune infiltration is assessed by leveraging the CIBERSORT algorithm. Our study uncovers pronounced genetic heterogeneity among PDK family members, coupled with discernible clinical characteristic. Significantly, the study establishes the potential utility of PDK family genes as prognostic indicators and as predictors of therapeutic response. Additionally, our study sheds light on the immune infiltration profile of PDK family. The results showed the intimate involvement of these genes in immune-related metrics, including immune scoring, immune subtypes, tumor-infiltrating lymphocytes, and immune checkpoints expression. In sum, the findings of this study offer insightful strategies to guide the therapeutic direction, aiming at leveraging the impact of PDK family genes in cancer treatment.

A comprehensive analysis of essential meiotic endonuclease 1 to prognosis and immune infiltration in kidney renal clear cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common type of cancer, and its molecular pathogenesis is unclear. In this study, we investigated the prognostic value of essential meiotic endonuclease 1 (EME1) in kidney renal clear cell carcinoma (KIRC). We downloaded the RNA-Seq expression of 526 KIRC tissues and 72 normal tissues from the TCGA database and the corresponding clinical data. The gene expression profiles associated with four clear cell renal cell carcinomas were downloaded from the GEO database for analysis. The expression of EME1 in clear renal cell carcinoma and its correlation with the clinical baseline data were analyzed. Kaplan-Meier survival curve analysis was performed to assess the relationship between EME1 and patient survival. Enrichment analysis was performed to elucidate the possible functions of EME1. We also analyzed the relationship between the EME1 expression and immune infiltration through TIMER2.0 and TISIDB online databases as well as the relationship between EME1 and common immune checkpoints. EME1 was identified as a risk factor for overall survival in clear cell renal cell carcinoma with a hazard ratio of 3.201 (95% confidence interval: 2.430-4.215; p < 0.001). EME1 was highly expressed in KIRC compared to that in normal tissues (p < 0.001) and in the worse TNM stages and late stages (stage 3/4) (p < 0.001). High EME1 expression was strongly associated with the advanced T stage (p = 0.003), advanced N stage (p = 0.002), and advanced M stage (p = 0.006). Research data on KIRC were simultaneously collected and analyzed from the GEO database, including GSE40435, GSE53000, GSE68417, and GSE53757. EME1 predicted the survival status in KIRC patients (AUC = 0.62). We further established a nomogram including the correlation between the high and low EME1 expression, and EME1 was found to contribute to the prediction of the probability of patient survival with a c-index = 0.796. Kaplan-Meier analysis revealed a lower likelihood of survival with a high EME1 expression (p < 0.001). In addition, further bioinformatics analysis suggested that EME1 may be associated with the extent of immune infiltration in KIRC. An increased expression of EME1 in KIRC is thus associated with advanced clinicopathological features, possibly acting as a potential biomarker of poor prognosis in KIRC.

Prognostic Role of Mitochondrial Transcription Termination Factor 3 in Thyroid Carcinoma.

Background: Studies have shown that the Mitochondrial Transcription Termination Factor 3 (MTERF3) negatively regulates mitochondrial gene expression and energy metabolism, and plays a significant role in many cancer types. Nevertheless, the expression and prognostic role of MTERF3 in patients with thyroid carcinoma (THCA) is still unclear. Thus, we investigated the expression, clinicopathological significance, and prognostic value of MTERF3 in THCA. Methods: The protein and mRNA expression levels of MTERF3 were, respectively, analyzed using immunohistochemistry (IHC) from THCA tissues and RNA-Seq data downloaded from The Cancer Genome Atlas. In addition, the relationships among the expression of MTERF3, the stemness feature, the extent of immune infiltration, drug sensitivity, the expression of ferroptosis, and N6-methyladenosine (m6A) methylation regulators, were evaluated as prognostic indicators for patients with THCA using the Kaplan-Meier plotter database. Results: The IHC and RNAseq results showed that the protein and mRNA expression levels of MTERF3 in adjacent nontumor tissues were significantly higher than in THCA tissues. The survival analysis indicated that decreased expression of MTERF3 was associated with a poorer prognosis. Furthermore, the expression of MTERF3 not only negatively correlated with the enhancement of the stemness of THCA and the reduction of drug sensitivity but also was implicated in ferroptosis and m6A methylation. Conclusion: The data from this study support the hypothesis that decreased expression of MTERF3 in THCA is associated with a poor prognosis.

Creation of a Rat Takotsubo Syndrome Model and Utilization of Machine Learning Algorithms for Screening Diagnostic Biomarkers.

Ferroptosis, a crucial type of programmed cell death, is directly linked to various cardiac disorders. However, the contribution of ferroptosis-related genes (FRGs) to Takotsubo syndrome (TTS) has not been completely understood. The objective of this study was to investigate the relationship between the FRGs and TTS. TTS rat models were established by isoprenaline injection. Heart tissues were subsequently harvested for total RNA extraction and library construction. Transcriptome data wereobtained transcriptome data for TTS and FRGs from our laboratory, and sources such as the Ferroptosis Database (FerrDb) and the Gene Expression Omnibus Database (GEO). 57 differentially expressed FRGs (DE-FRGs) were discovered. The LASSO and SVM-RFE algorithms were employed to identify Enpp2, Pla2g6, Etv4, and Il1b as marker genes, and logistic regression was applied to construct a diagnostic model. The important genes were validated by real time PCR and the external dataset. Finally, the extent of immune infiltration was explored. Among the 57 genes, there were 36 up-regulated and 21 down-regulated genes that exhibited distinct expression patterns in the TTS and healthy control samples. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that the enriched pathways were primarily associated with pathways of neurodegeneration-multiple disease, while Gene Ontology (GO) analysis revealed that these genes were primarily linked to cellular response to external stimuli, outer membrane functions, and ubiquitin protein ligase binding. After the identification of four marker genes as potentially effective biomarkers for TTS diagnosis, subsequent logistic regression modeling revealed a receiver operating characteristic curve (ROC) with an AUC of 1.0. The examination of immune cell infiltration showed significantly higher prevalence of activated CD4+ T cells, mast cells, etc., in TTS. Our findings support the theoretical importance of ferroptosis in TTS, highlighting Enpp2, Pla2g6, Etv4, and Il1b as potential diagnostic and therapeutic biomarkers for TTS.

CD8+ T cell/cancer-associated fibroblast ratio stratifies prognostic and predictive responses to immunotherapy across multiple cancer types.

Cancer-associated fibroblasts (CAFs) within the tumor microenvironment (TME) are critical for immune suppression by restricting immune cell infiltration in the tumor stromal zones from penetrating tumor islands and changing their function status, particularly for CD8+ T cells. However, assessing and quantifying the impact of CAFs on immune cells and investigating how this impact is related to clinical outcomes, especially the efficacy of immunotherapy, remain unclear. The TME was characterized using immunohistochemical (IHC) analysis using a large-scale sample size of gene expression profiles. The CD8+ T cell/CAF ratio (CFR) association with survival was investigated in The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) lung cancer cohorts. The correlation between CFR and immunotherapeutic efficacy was computed in five independent cohorts. The correlation between CFR and objective response rates (ORRs) following pembrolizumab monotherapy was investigated in 20 solid tumor types. To facilitate clinical translation, the IHC-detected CD8/α-SMA ratio was applied as an immunotherapeutic predictive biomarker in a real-worldlung cancer cohort. Compared with normal tissue, CAFs were enriched in cancer tissue, and the amount of CAFs was overwhelmingly higher than that in other immune cells. CAFs are positively correlated with the extent of immune infiltration. A higher CFR was strongly associated with improved survival in lung cancer, melanoma, and urothelial cancer immunotherapy cohorts. Within most cohorts, there was no clear evidence for an association between CFR and programmed death-ligand 1 (PD-L1) or tumor mutational burden (TMB). Compared with TMB and PD-L1, a higher correlation coefficient was observed between CFR and the ORR following pembrolizumab monotherapy in 20 solid tumor types (Spearman's r = 0.69 vs. 0.44 and 0.21). In a real-world cohort, patients with a high CFR detected by IHC benefited considerably from immunotherapy as compared with those with a low CFR (hazard ratio, 0.37; 95% confidence interval, 0.19-0.75; p < 0.001). CFR is a newly found and simple parameter that can be used for identifying patients unlikely to benefit from immunotherapy. Future studies are needed to confirm this finding.

PD-L1 Dependent Immunogenic Landscape in Hot Lung Adenocarcinomas Identified by Transcriptome Analysis.

Simple SummaryLung cancer, with non-small-cell lung cancer as its most common form, is the leading cause of cancer-related mortality and shows a poor prognosis. Despite recent advantages in the field of immunotherapy, there is still a great need for an improved understanding of PD-1/PD-L1 checkpoint blockade-responsive biology. Since immune cell infiltration is regarded as an important parameter in this field, we aimed to identify the immunogenic landscape in primary lung adenocarcinoma on the transcriptomic level in context with tumoral PD-L1 expression (positive vs. negative) and extent of immune infiltration (“hot” vs. “cold” phenotype). Our results reveal that genes that are related to the tumor microenvironment are differentially expressed based on tumoral PD-L1 expression indicating novel aspects of PD-L1 regulation, with potential biological relevance, as well as relevance for immunotherapy response stratification.Background: Lung cancer is the most frequent cause of cancer-related deaths worldwide. The clinical development of immune checkpoint blockade has dramatically changed the treatment paradigm for patients with lung cancer. Yet, an improved understanding of PD-1/PD-L1 checkpoint blockade-responsive biology is warranted. Methods: We aimed to identify the landscape of immune cell infiltration in primary lung adenocarcinoma (LUAD) in the context of tumoral PD-L1 expression and the extent of immune infiltration (“hot” vs. “cold” phenotype). The study comprises LUAD cases (n = 138) with “hot” (≥150 lymphocytes/HPF) and “cold” (<150 lymphocytes/HPF) tumor immune phenotype and positive (>50%) and negative (<1%) tumor PD-L1 expression, respectively. Tumor samples were immunohistochemically analyzed for expression of PD-L1, CD4, and CD8, and further investigated by transcriptome analysis. Results: Gene set enrichment analysis defined complement, IL-JAK-STAT signaling, KRAS signaling, inflammatory response, TNF-alpha signaling, interferon-gamma response, interferon-alpha response, and allograft rejection as significantly upregulated pathways in the PD-L1-positive hot subgroup. Additionally, we demonstrated that STAT1 is upregulated in the PD-L1-positive hot subgroup and KIT in the PD-L1-negative hot subgroup. Conclusion: The presented study illustrates novel aspects of PD-L1 regulation, with potential biological relevance, as well as relevance for immunotherapy response stratification.

Analysis of the immune landscape of small bowel neuroendocrine tumors.

Immune checkpoint inhibitors have shown promising results in different cancers, and correlation between immune infiltration, expression of programmed death-ligand 1 (PD-L1) by tumor cells and response to immunotherapy has been reported. There is limited knowledge regarding the immune microenvironment of small bowel (SB) neuroendocrine tumors (NETs). This work was aimed at characterizing the immune landscape of SB NETs. Expression of PD-L1 and programmed death-1 (PD-1) was evaluated by immunohistochemistry in 102 surgically resected, primary NETs of the duodenum, jejunum and ileum. Extent and characteristics of the tumor-associated immune infiltrate were also assessed and investigated in their prognostic potential. We detected the expression of PD-L1 in ≥1 and ≥50% of tumor cells in 40/102 (39%; 95% CI, 30-49%) and 14/102 (14%; 95% CI, 8-22%) cases respectively. Intratumor host immune response was apparently absent in 35/102 cases (34%; 95% CI, 25-44%), mild to moderate in 46/102 samples (45%, 95% CI, 35-55%), intense in 21/102 tumors (21%, 95% CI, 13-30%). Expression of PD-L1 and extent of immune infiltration were significantly higher in duodenal NETs as compared with jejunal/ileal NETs. A marked peritumoral host response was organized as ectopic lymph node-like structures in 18/102 cases (18%; 95% CI, 11-26%). Neither PD-L1 expression nor the degree of immune infiltration showed any prognostic significance. Overall, the immune landscape of SB NETs is heterogeneous, with adaptive immune resistance mechanisms prevailing in duodenal NETs. Clinical trials of immune checkpoint inhibitors should take into account the immune heterogeneity of SB NETs.

1587P - Tumor infiltrating lymphocytes and tertiary lymphoid structures in paired primary tumors and metastases from breast cancer patients

In primary tumors (P), infiltrating lymphocytes (TIL) and tertiary lymphoid structures (TLS) have been associated with better clinical outcomes, in HER2+ and triple negative breast cancer (BC). The temporal and spatial heterogeneity of TIL and TLS between P and their matched metastases (M) is currently unknown. We analyzed formalin fixed paraffin embedded tissue samples from a retrospective cohort of BC patients who underwent adjuvant surgery for the P and metastasectomy or biopsy for their metachronous M at a distant site(s) (n = 51). Immunohistochemistry stained sections using anti-CD3 (T cells) and anti-CD20 (B cells) antibodies in a double stain were scored by two trained pathologists blinded to the clinical data. TIL were scored as the percent (%) of stroma plus tumor surface area infiltrated with CD3+ plus CD20+ cells. The number of TLS were normalized to the tumor area. The extent of TIL in P (range 1-35%) was significantly higher with respect to the corresponding M (p = 0.04). Interestingly, when the extent of TIL in P was ≥10% (=TIL+) there was a significant decrease in the %TIL detected in M (p < 0.0001, n = 20). Alternatively, when TIL infiltration in the P was <10% (=TIL-) the %TIL was significantly higher at the distant M site (p = 0.002, n = 31). In secondary M lesions, TIL varied between 0-20% and TLS were essentially absent. Analysis of TIL in M from different organs revealed a significantly higher level in soft tissues (n = 4) compared to skin (n = 24), brain (n = 2) or breast relapses (n = 14) (p = 0.04). These preliminary data suggest that tumors can alter their immunogenicity during disease progression by their production of neoantigens and/or immunosuppressive factors, which potentially accounts for our observed differences between P and M. In metastatic disease, the extent of immune infiltration is globally lower than in early disease with almost no TLS found in M, signifying that the composition and organization of the immune infiltrate varies between disease stages. The extent of the immune infiltrate may also depend upon the organ site of relapse, further complicating the heterogeneity of both the tumor and the anti-tumor immune response.

CNS-targeted LIF Expression Improves Therapeutic Efficacy and Limits Autoimmune-mediated Demyelination in a Model of Multiple Sclerosis

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system (CNS) with an inflammatory and a neurodegenerative component. The neuropoietic cytokine leukemia inhibitory factor (LIF) is expressed in MS lesions, but its effect on lesion development is far from understood. LIF is an interesting candidate for MS therapy, as it has neuroprotective properties and may also promote the survival of myelinating oligodendrocytes (OLGs). However, therapeutic administration of LIF is complicated by its limited ability to cross the blood-brain barrier and its pleiotropic actions outside the CNS. In this study, lentiviral vectors (LVs) were used to achieve stable expression and secretion of LIF in the CNS of adult mice. CNS-targeted expression of LIF significantly reduced demyelination in a murine model of MS. In addition, local expression of LIF ameliorated clinical symptoms with enhanced efficacy compared to systemic treatment with recombinant protein. These findings demonstrate that gene therapeutic administration of LIF is a promising approach to limit lesion burden and clinical symptoms in neuroinflammatory disease.