Neuroendocrine gene subsets are uniquely dysregulated in prostate adenocarcinoma

Abstract

Prostate cancer has heterogeneous growth patterns, and its prognosis is the poorest when it progresses to a neuroendocrine phenotype. Using bioinformatic analysis, we evaluated RNA expression of neuroendocrine genes in a panel of five different cancer types: prostate adenocarcinoma, breast cancer, kidney chromophobe, kidney renal clear cell carcinoma and kidney renal papillary cell carcinoma. Our results show that specific neuroendocrine genes are significantly dysregulated in these tumors, suggesting that they play an active role in cancer progression. Among others, synaptophysin (SYP), a conventional neuroendocrine marker, is upregulated in prostate adenocarcinoma (PRAD) and breast cancer (BRCA). Our analysis shows that SYP is enriched in small extracellular vesicles (sEVs) derived from plasma of PRAD patients, but it is absent in sEVs derived from plasma of healthy donors. Similarly, classical sEV markers are enriched in sEVs derived from plasma of prostate cancer patients, but weakly detectable in sEVs derived from plasma of healthy donors. Overall, our results pave the way to explore new strategies to diagnose these diseases based on the neuroendocrine gene expression in patient tumors or plasma sEVs.

Keywords: Prostate cancer; exosomes; extracellular vesicles; neuroendocrine genes; patient plasma; synaptophysin.

Figure 1.

Differential expression patterns of known neuroendocrine cancer genes across TCGA cancer types. (a) Venn diagram depicting the neuroendocrine genes reported in TCGA (The Cancer Genome Atlas) that are significantly upregulated (FDR: False Discovery Rate <0.05) in one or more indicated cancer types [Prostate adenocarcinoma (PRAD), Breast cancer (BRCA), Kidney Chromophobe (KICH), Kidney renal clear cell (KIRC) carcinoma, and Kidney renal papillary cell (KIRP) carcinoma] compared to normal samples. The table below indicates the genes that are significantly upregulated according to the cancer type. (b) Volcano plots of log2 fold-change (Log FC) in expression of the known neuroendocrine genes (X-axis) with statistical significance (Y-axis) in five cancer types. Significantly downregulated and upregulated genes are represented in blue and red colors, respectively. Grey color represents not significant (NS). The genes with 1.5 fold-change are shown. Y-axis represents mRNA expression (Normalized expression). Significantly (FDR <0.05) differentially expressed genes are shown. P-value < 0.05 are considered significant.

Figure 2.

Differential expression patterns of known downregulated neuroendocrine cancer genes across cancer types reported in TCGA. Venn diagram depicting the neuroendocrine genes that are significantly downregulated (FDR <0.05) in different TCGA cancer types (PRAD, BRCA, KICH, KIRC, and KIRP) compared to normal samples. The table (right panel) indicates the genes that are significantly downregulated according to the cancer type.

Figure 3.

Neuroendocrine gene expression levels in NEPC compared to CRPC. Violin plots represent mRNA expression levels (Y-axis) of neuroendocrine genes (X-axis) that are significantly dysregulated (FDR <0.05) in NEPC compared to CRPC as reported in TCGA. NEPC: neuroendocrine prostate cancer, CRPC: castrate-resistant prostate cancer. P-value  < 0.05 are considered significant.

Figure 4.

PRAD patient plasma-derived sEVs are enriched in SYP and sEV classical markers. (a) Immunoblotting (IB) analysis of pooled plasma sEV fractions (Fractions 6–8) from healthy donor (4–13) or PRAD patients (M-Z); 20 μg of protein were loaded per sEV sample. Expression of synaptophysin (SYP), CD63, Syntenin, Alix, and CD9 was analyzed. (b) Nanoparticle tracking analysis (NTA) of size and concentration of pooled sEVs (Fractions 6–8) from healthy donor plasma (7 + 8 + 9) or PRAD patient plasma (R+S+T).