Enhanced Complement Expression in the Tumor Microenvironment Following Neoadjuvant Therapy: Implications for Immunomodulation and Survival in Pancreatic Ductal Adenocarcinoma.

Background: Neoadjuvant therapy (NAT) is increasingly being used for pancreatic ductal adenocarcinoma (PDAC) treatment. However, its specific effects on carcinoma cells and the tumor microenvironment (TME) are not fully understood. This study aims to investigate how NAT differentially impacts PDAC's carcinoma cells and TME. Methods: Spatial transcriptomics was used to compare gene expression profiles in carcinoma cells and the TME between 23 NAT-treated and 13 NAT-naïve PDAC patients, correlating with their clinicopathologic features. Analysis of an online single-nucleus RNA sequencing (snRNA-seq) dataset was performed for validation of the specific cell types responsible for NAT-induced gene expression alterations. Results: NAT not only induces apoptosis and inhibits proliferation in carcinoma cells but also significantly remodels the TME. Notably, NAT induces a coordinated upregulation of multiple key complement genes (C3, C1S, C1R, C4B and C7) in the TME, making the complement pathway one of the most significantly affected pathways by NAT. Patients with higher TME complement expression following NAT exhibit improved overall survival. These patients also exhibit increased immunomodulatory and neurotrophic cancer-associated fibroblasts (CAFs); more CD4+ T cells, monocytes, and mast cells; and reduced immune exhaustion gene expression. snRNA-seq analysis demonstrates C3 complement was specifically upregulated in CAFs but not in other stroma cell types. Conclusions: NAT can enhance complement production and signaling within the TME, which is associated with reduced immunosuppression in PDAC. These findings suggest that local complement dynamics could serve as a novel biomarker for prognosis, evaluating treatment response and resistance, and guiding therapeutic strategies in NAT-treated PDAC patients.

RNA sequencing analysis of hepatocellular carcinoma identified oxidative phosphorylation as a major pathologic feature.

Dysregulation of expression of functional genes and pathways plays critical roles in the etiology and progression of hepatocellular carcinoma (HCC). Next generation-based RNA sequencing (RNA-seq) offers unparalleled power to comprehensively characterize HCC at the whole transcriptome level. In this study, 17 fresh-frozen HCC samples with paired non-neoplastic liver tissue from Caucasian patients undergoing liver resection or transplantation were used for RNA-seq analysis. Pairwise differential expression analysis of the RNA-seq data was performed to identify genes, pathways, and functional terms differentially regulated in HCC versus normal tissues. At a false discovery rate (FDR) of 0.10, 13% (n = 4335) of transcripts were up-regulated and 19% (n = 6454) of transcripts were down-regulated in HCC versus non-neoplastic tissue. Eighty-five Kyoto Encyclopedia of Genes and Genomes pathways were differentially regulated (FDR, <0.10), with almost all pathways (n = 83) being up-regulated in HCC versus non-neoplastic tissue. Among the top up-regulated pathways was oxidative phosphorylation (hsa00190; FDR, 1.12E-15), which was confirmed by Database for Annotation, Visualization, and Integrated Discovery (DAVID) gene set enrichment analysis. Consistent with potential oxidative stress due to activated oxidative phosphorylation, DNA damage-related signals (e.g., the up-regulated hsa03420 nucleotide excision repair [FDR, 1.14E-04] and hsa03410 base excision repair [FDR, 2.71E-04] pathways) were observed. Among down-regulated genes (FDR, <0.10), functional terms related to cellular structures (e.g., cell membrane [FDR, 3.05E-21] and cell junction [FDR, 2.41E-07], were highly enriched, suggesting compromised formation of cellular structure in HCC at the transcriptome level. Interestingly, the olfactory transduction (hsa04740; FDR, 1.53E-07) pathway was observed to be down-regulated in HCC versus non-neoplastic tissue, suggesting impaired liver chemosensory functions in HCC. Our findings suggest oxidative phosphorylation and the associated DNA damage may be the major driving pathologic feature in HCC.

Dcx expression defines a subpopulation of Gdf5 + cells with chondrogenic potentials in E12.5 mouse embryonic limbs.

Growth differentiation factor 5 (Gdf5) and doublecortin (Dcx) genes are both expressed in joint interzone cells during synovial joint development. In this study, we re-analyzed the single cell RNA-sequencing data (Gene Expression Omnibus GSE151985) generated from Gdf5 + cells of mouse knee joints at embryonic stages of E12.5, E13.5, E14.5, and E15.5, with a new focus on Dcx. We found that Dcx expression was enriched in clusters of Gdf5 + cells, with high expression levels of pro-chondrogenic genes including sex determining region Y-box transcription factor 5 (Sox5), Sox6, Sox9, Gdf5, versican, matrilin 4, collagen type II α 1 chain (Col2a1), Col9a1, Col9a2, and Col9a3 at E12.5. Dcx + and Dcx - cells had differential gene expression profiles. The up-regulated genes in Dcx + vs. Dcx - cells at E12.5 and E13.5 were enriched in chondrocyte differentiation and cartilage development, whereas those genes up-regulated at E14.5 and E15.5 were enriched in RNA splicing, protein stability, cell proliferation, and cell growth. Gene expression profiles in Dcx + cells showed rapid daily changes from E12.5 to E15.5, with limited number of genes shared across the time period. Expression of Gdf5, Sox5, Sox6, melanoma inhibitory activity, noggin, odd-skipped related transcription factor 2, matrilin 4, and versican was positively correlated with Dcx expression. Our results demonstrate that Dcx expression defines a subpopulation of Gdf5 + cells with chondrogenic potentials in E12.5 mouse embryonic limbs.

Genome and transcriptome profiling of FBXW family in human prostate cancer.

F-box and WD repeat domain containing (FBXW) family of E3 ligases has 10 members that ubiquitinate substrate proteins for proteasome-mediated degradation. Publicly archived datasets from The Cancer Genome Atlas (TCGA), Prostate Cancer Transcriptome Atlas (PCTA), and cBioPortal were analyzed for mRNA expression and genetic alterations of 10 FBXW genes. We found that FBXW7 mRNA expression was significantly decreased in primary prostate cancers compared to normal prostate tissues, whereas mRNA expression of FBXW8-10 was significantly increased in primary prostate cancers compared to normal prostate tissues. FBXW7 mRNA expression was also significantly decreased in breast invasive carcinoma, glioblastoma multiforme, head and neck squamous cell carcinoma, lung squamous cell carcinoma, and uterine corpus endometrial carcinoma. In contrast, FBXW7 mRNA expression was significantly increased in cholangiocarcinoma, colon adenocarcinoma, kidney renal clear cell carcinoma, kidney renal papillary cell carcinoma, liver hepatocellular carcinoma, lung adenocarcinoma, pheochromocytoma and paraganglioma, and thyroid carcinoma. Compared to normal tissues, FBXW5 mRNA expression was significantly increased in breast invasive carcinoma, cholangiocarcinoma, kidney chromophobe, kidney renal clear cell carcinoma, liver hepatocellular carcinoma, lung adenocarcinoma, lung squamous cell carcinoma, prostate adenocarcinoma, thyroid carcinoma, and uterine corpus endometrial carcinoma, whereas FBXW5 mRNA expression was only significantly decreased in colon adenocarcinoma. There were not any significant differences in gene copy number gains, losses, or gene simple somatic mutations between primary prostate cancers and normal prostate tissues. The mRNA expression levels of FBXW5, 7, 8, 9, and 12 were significantly higher in metastatic castration-resistant prostate cancers (mCRPCs) than primary prostate cancers, whereas mRNA expression levels of FBXW1 and 4 were significantly lower in mCRPCs than primary prostate cancers. All 10 FBXW genes had significantly more overall gene alterations including gene amplifications in mCRPCs than primary prostate cancers. FBXW5 and 7 had significantly more gene deep deletions in mCRPCs than primary prostate cancers and FBXW7 had significantly more gene missense mutations in mCRPCs than primary prostate cancers. Our findings suggest that different FBXW genes have differential mRNA expression in prostate cancer and other cancer types and their gene amplifications are significantly more in mCRPCs than primary prostate cancers. FBXW7 mRNA expression is consistently decreased in primary prostate cancers compared to normal prostate tissues.

Th17 cells promote tumor growth in an immunocompetent orthotopic mouse model of prostate cancer.

Interleukin-17 (IL-17) has been demonstrated to promote development of a variety of cancers including prostate cancer in genetically modified mouse models. IL-17 is the main product secreted by T helper 17 (Th17) cells. A recent study has shown that Th17 cells and related genes are upregulated in human prostate cancers. However, there is no direct experimental evidence to demonstrate Th17's role in prostate cancer. In the present study, we co-implanted mouse prostate cancer MPC3-luc cells with Th17-polarized mouse splenocytes in the prostate of immunocompetent C57BL/6J male mice. We found that Th17-polarized splenocytes promoted orthotopic allograft prostate tumor growth compared to the control splenocytes. The numbers of IL-17-positive lymphocytes and macrophages were higher in the prostate tumors grown from co-implantation of MPC3-luc cells and Th17-polarized splenocytes, compared to the prostate tumors grown from co-implantation of MPC3-luc cells and control splenocytes. Our findings provide the first direct experimental evidence that Th17 cells may promote prostate cancer growth.

Genetic alterations of interleukin-17 and related genes in human prostate cancer.

Interleukin-17 (IL-17) has been shown to promote development of hormone-naïve prostate cancer (HNPC) and castration-resistant prostate cancer (CRPC) as well as lymph node metastasis in mouse models. Gene alterations of IL-17 family of cytokines and their downstream genes in human prostate cancer have not been investigated. We studied 7 datasets archived in cBioPortal and queried gene alterations in a total of 1303 cases of human prostate cancers. 35 genes were examined, including IL-17 family of cytokines and receptors, IL-17-downstream genes, and genes related to IL-17-downstream genes. We found that 34/35 (97%) genes had significantly more alterations in metastatic prostate cancer (with alteration rates ranging from 3.42% to 13.01%) than primary prostate cancer (with alteration rates ranging from 0.40% to 2.96%). 15/35 (43%) genes had significantly more alterations in primary CRPC than primary HNPC. 34/35 (97%) genes had significantly more alterations in metastatic CRPC than primary HNPC. Only three genes (S100A7, S100A8, and S100A9) had significantly more alterations in metastatic CRPC than primary CRPC. The gene alterations were mostly gene amplifications (97%), while gene deep deletions, missense mutations, and truncating mutations were very rare. 7/35 (20%) genes had significantly more alterations in primary neuroendocrine prostate cancer (NEPC) than primary adenocarcinoma (AC). 23/35 (66%) genes had significantly more alterations in metastatic NEPC than metastatic AC. Only three genes (S100A7, S100A8, and S100A9) had significantly more alterations in metastatic NEPC than metastatic AC with neuroendocrine features. Most of the gene alterations in metastatic NEPC were gene amplifications (80%), while gene deep deletions, missense mutations, and truncating mutations were very rare. Our findings suggest that gene amplifications of IL-17 and related genes are more frequently found in metastatic CRPC and NEPC than primary hormone-naïve prostate adenocarcinomas, implying that IL-17 and related genes may play important roles in the progression from HNPC to CRPC and from primary location to metastasis as well as in development of metastatic NEPC.