Transcriptomic miRNA and mRNA signatures in primary prostate cancer that are associated with lymph-node invasion

Abstract

Background: Nomograms or comparable techniques can be used to determine which patients with prostate cancer (PCa) will benefit from extended pelvic lymph node dissection (ePLND). While nomograms help guide clinical decisions, ∼80% of the patients undergo unnecessary ePLND. This pilot study aims to identify both transcriptomic mRNA and microRNA (miR) signatures in primary PCa tumours that are associated with the presence of lymph node metastasis (LNM).

Methods: Primary PCa tumours obtained from 88 patients (pathologically diagnosed as N0 [pN0, n = 44] or as N1 [pN1, n = 44]) were profiled using two different probe-based captured direct assays based on next-generation sequencing and targeting 19398 mRNA transcripts (human transcriptome panel [HTP] dataset) and 2083 miRs (miRs whole-transcriptome assay [WTA] dataset). The TCGA-PRAD (pN0 [n = 382] and pN1 [n = 70]) and GSE220095 (pN0 [n = 138] and pN1 [n = 17]) datasets were used for validation using bioinformatic analyses.

Results: A four-mRNA signature (CHRNA2, NPR3, VGLL3 and PAH) was found in primary tumour tissue samples from pN1 PCa patients, and then it was validated using the TCGA-PRAD and GSE220095 datasets. Adding serum prostate-specific antigen (PSA) values to the four-gene signature increased the performance to identify pN1 (HTP [AUC = .8487, p = 2.18e-09], TCGA-PRAD [AUC = .7150, p = 8.66e-08] and GSE220095 datasets [AUC = .8772, p = 4.09e-07]). Paired miR analyses showed that eight miRs were significantly upregulated in primary PCa that were pN1 (p < .01). The eight-miR signature performance increased when adding PSA (WTA dataset [AUC = .8626, p = 4.66e-10]) or Grade group (WTA dataset [AUC = .8689, p = 2e-10]). When combining the miR/mRNA signatures (miR-663b, CHRNA2 and PAH) with PSA levels, it showed the best performance to distinguish pN1 from pN0 PCa patients.

Conclusion: This study found miR/mRNA signatures in primary PCa tumours that in combination with serum PSA levels may complement nomograms for better detection of PCa patients with LNM and triage patients into better surgical decision-making.

Keywords

lymph node dissection / lymph node metastasis / mRNA-signature / prostate cancer

References

[1] Sandhu S, Moore CM, Chiong E, Beltran H, Bristow RG, Williams SG. Prostate cancer. Lancet. 2021; 398(10305): 1075-1090.

[2] Wei JT, Barocas D, Carlsson S, et al. Early detection of prostate cancer: AUA/SUO guideline part II: Considerations for a prostate biopsy. J Urol. 2023; 210(1): 54-63.

[3] Siegel RL, Miller KD, Wagle NS, Jemal A. Cancer statistics, 2023. CA: Cancer J Clin. 2023; 73(1): 17-48.

[4] Rebello RJ, Oing C, Knudsen KE, et al. Prostate cancer. Nat Rev Dis Primers. 2021; 7(1): 9.

[5] Sleeman JP. The lymph node pre-metastatic niche. J Mol Med. 2015; 93(11): 1173-1184.

[6] Gandaglia G, Ploussard G, Valerio M, et al. A novel nomogram to identify candidates for extended pelvic lymph node dissection among patients with clinically localized prostate cancer diagnosed with magnetic resonance imaging-targeted and systematic biopsies. Eur Urol. 2019; 75(3): 506-514.

[7] Lestingi JFP, Guglielmetti GB, Trinh QD, et al. Extended versus limited pelvic lymph node dissection during radical prostatectomy for intermediate- and high-risk prostate cancer: Early oncological outcomes from a randomized phase 3 trial. Eur Urol. 2021; 79(5): 595-604.

[8] Miura N, Sugihara N, Funaki K, et al. Extended robot-assisted laparoscopic prostatectomy and extended pelvic lymph node dissection as a monotherapy in patients with very high-risk prostate cancer Patients. Cancer Med. 2021; 10(22): 7968-7976.

[9] Di Pierro GB, Salciccia S, Frisenda M, et al. Comparison of four validated nomograms (Memorial Sloan Kettering Cancer Center, Briganti 2012, 2017, and 2019) predicting lymph node invasion in patients with high-risk prostate cancer candidates for radical prostatectomy and extended pelvic lymph node dissection: Clinical experience and review of the literature. Cancers. 2023; 15(6): 1683.

[10] Zhang X, Zhang G, Wang J, Bi J. Different lymph node dissection ranges during radical prostatectomy for patients with prostate cancer: a systematic review and network meta-analysis. World J Surg Oncol. 2023; 21(1): 80.

[11] Morton DL, Wen DR, Wong JH, et al. Technical details of intraoperative lymphatic mapping for early stage melanoma. Arch. Surg. 1992; 127(4): 392-399.

[12] Stultz J, Fong L. How to turn up the heat on the cold immune microenvironment of metastatic prostate cancer. Prostate Cancer Prostatic Dis. 2021; 24(3): 697-717.

[13] Hruby S, Englberger C, Lusuardi L, et al. Fluorescence guided targeted pelvic lymph node dissection for intermediate and high risk prostate cancer. J Urol. 2015; 194(2): 357-363.

[14] Jeschke S, Lusuardi L, Myatt A, Hruby S, Pirich C, Janetschek G. Visualisation of the lymph node pathway in real time by laparoscopic radioisotope- and fluorescence-guided sentinel lymph node dissection in prostate cancer staging. Urology. 2012; 80(5): 1080-1087.

[15] Miki J, Yanagisawa T, Tsuzuki S, et al. Anatomical localization and clinical impact of sentinel lymph nodes based on patterns of pelvic lymphatic drainage in clinically localized prostate cancer. Prostate. 2018; 78(6): 419-425.

[16] Chennamsetty A, Zhumkhawala A, Tobis SB, et al. Lymph node fluorescence during robot-assisted radical prostatectomy with indocyanine green: prospective dosing analysis. Clin Genitourin Cancer. 2017; 15(4): e529-e534.

[17] KleinJan GH, van den Berg NS, Brouwer OR, et al. Optimisation of fluorescence guidance during robot-assisted laparoscopic sentinel node biopsy for prostate cancer. Eur Urol. 2014; 66(6): 991-998.

[18] Manny TB, Patel M, Hemal AK. Fluorescence-enhanced robotic radical prostatectomy using real-time lymphangiography and tissue marking with percutaneous injection of unconjugated indocyanine green: the initial clinical experience in 50 patients. Eur Urol. 2014; 65(6): 1162-1168.

[19] Ramírez-Backhaus M, Mira Moreno A, Gómez Ferrer A, et al. Indocyanine green guided pelvic lymph node dissection: An efficient technique to classify the lymph node status of patients with prostate cancer who underwent radical prostatectomy. J Urol. 2016; 196(5): 1429-1435.

[20] van der Poel HG, Buckle T, Brouwer OR, Valdés Olmos RA, van Leeuwen FW. Intraoperative laparoscopic fluorescence guidance to the sentinel lymph node in prostate cancer patients: clinical proof of concept of an integrated functional imaging approach using a multimodal tracer. Eur Urol. 2011; 60(4): 826-833.

[21] Małkiewicz B, Kiełb P, Kobylański M, et al. Sentinel lymph node techniques in urologic oncology: Current knowledge and application. Cancers. 2023; 15(9): 2495.

[22] van den Berg NS, Buckle T, KleinJan GH, van der Poel HG, van Leeuwen FWB. Multispectral fluorescence imaging during robot-assisted laparoscopic sentinel node biopsy: A first step towards a fluorescence-based anatomic roadmap. Eur Urol. 2017; 72(1): 110-117.

[23] Fraser M, Livingstone J, Wrana JL, et al. Somatic driver mutation prevalence in 1844 prostate cancers identifies ZNRF3 loss as a predictor of metastatic relapse. Nat Commun. 2021; 12(1): 6248.

[24] Moreno CS, Winham CL, Alemozaffar M, et al. Integrated genomic analysis of primary prostate tumor foci and corresponding lymph node metastases identifies mutations and pathways associated with metastasis. Cancers. 2023; 15(23): 5671.

[25] Bustos MA, Tran KD, Rahimzadeh N, et al. Integrated assessment of circulating cell-free MicroRNA signatures in plasma of patients with melanoma brain metastasis. Cancers. 2020; 12(6): 1692.

[26] Bustos MA, Yokoe T, Shoji Y, et al. MiR-181a targets STING to drive PARP inhibitor resistance in BRCA- mutated triple-negative breast cancer and ovarian cancer. Cell Biosci. 2023; 13(1): 200.

[27] Dragomir MP, Knutsen E, Calin GA. Classical and noncanonical functions of miRNAs in cancers. Trends Genet. 2022; 38(4): 379-394.

[28] Hayashi Y, Millen JC, Ramos RI, et al. Cell-free and extracellular vesicle microRNAs with clinical utility for solid tumors. Mol Oncol. 2024.

[29] Bustos MA, Gottlieb J, Choe J, et al. Diagnostic miRNA signatures in paired tumor, plasma, and urine specimens from renal cell carcinoma patients. Clin Chem. 2023; 70(1): 261-272.

[30] Koh Y, Bustos MA, Moon J, et al. Urine cell-free MicroRNAs in localized prostate cancer patients. Cancers. 2022; 14(10): 2388.

[31] Bustos MA, Rahimzadeh N, Ryu S, et al. Cell-free plasma microRNAs that identify patients with glioblastoma. Lab Investig. 2022; 102(7): 711-721.

[32] Finotello F, Mayer C, Plattner C, et al. Molecular and pharmacological modulators of the tumor immune contexture revealed by deconvolution of RNA-seq data. Genome Med. 2019; 11(1): 34.

[33] Egevad L. Handling of radical prostatectomy specimens. Histopathology. 2012; 60(1): 118-124.

[34] Lindh C, Delahunt B, Samaratunga H, et al. A novel technique for biobanking of large sections of radical prostatectomy specimens. Histopathology. 2018; 72(3): 481-489.

[35] Borchert S, Herold T, Kalbourtzis S, et al. Transcriptome-wide gene expression profiles from FFPE materials based on a nuclease protection assay reveals significantly different patterns between synovial sarcomas and morphologic mimickers. Cancers. 2022; 14(19): 4737.

[36] Hurwitz SN, Lockhart B, Önder Ö, et al. Proteogenomic profiling of high-grade B-Cell lymphoma with 11q aberrations and burkitt lymphoma reveals lymphoid enhancer binding factor 1 as a novel biomarker. Modern pathology: an official journal of the United States and Canadian Academy of Pathology, Inc. 2023; 36(7): 100170.

[37] Koll FJ, Metzger E, Hamann J, et al. Overexpression of KMT9α Is associated with aggressive basal-like muscle-invasive bladder cancer. Cells. 2023; 12(4): 589.

[38] Qiu L, Wang SA, Tang G, et al. Blastoid B-Cell neoplasms: Diagnostic challenges and solutions. Cancers. 2023; 15(3): 848.

[39] Rade M, Kreuz M, Borkowetz A, et al. A reliable transcriptomic risk-score applicable to formalin-fixed paraffin-embedded biopsies improves outcome prediction in localized prostate cancer. Mol Med. 2024; 30(1): 19.

[40] Yin Y, Liu Q, Shao Y, et al. Regulatory mechanism of androgen receptor on NCAPD3 gene expression in prostate cancer. The Prostate. 2022; 82(1): 26-40.

[41] Jing Z, Liu Q, Xie W, et al. NCAPD3 promotes prostate cancer progression by up-regulating EZH2 and MALAT1 through STAT3 and E2F1. Cell Signal. 2022; 92: 110265.

[42] Zhang Y, Shao Y, Ren J, et al. NCAPD3 exerts tumor-promoting effects in prostatic cancer via dual impact on miR-30a-5p by STAT3-MALAT1 and MYC. Cell Death Discov. 2024; 10(1): 159.

[43] Wang I, Song L, Wang BY, Rezazadeh Kalebasty A, Uchio E, Zi X. Prostate cancer immunotherapy: a review of recent advancements with novel treatment methods and efficacy. Am J Clin Exp Urol. 2022; 10(4): 210-233.

[44] Strasner A, Karin M. Immune infiltration and prostate cancer. Front Oncol. 2015; 5: 128.

[45] Ayers M, Lunceford J, Nebozhyn M, et al. IFN-γ-related mRNA profile predicts clinical response to PD-1 blockade. J Clin Invest. 2017; 127(8): 2930-2940.

[46] Koll FJ, Schwarz A, Köllermann J, et al. CK5/6 and GATA3 defined phenotypes of muscle-invasive bladder cancer: Impact in adjuvant chemotherapy and molecular subtyping of negative cases. Front Med. 2022; 9: 875142.

[47] Horny K, Sproll C, Peiffer L, et al. Mesenchymal-epithelial transition in lymph node metastases of oral squamous cell carcinoma is accompanied by ZEB1 expression. J Transl Med. 2023; 21(1): 267.

[48] Wood S, Schertzer M, Yaremko ML. Identification of the human neuronal nicotinic cholinergic alpha 2 receptor locus, (CHRNA2), within an 8p21 mapped locus, by sequence homology with rat DNA. Somat Cell Mol Genet. 1995; 21(2): 147-150.

[49] Fan A, Zhang Y, Cheng J, Li Y, Chen W. A novel prognostic model for prostate cancer based on androgen biosynthetic and catabolic pathways. Front Oncol. 2022; 12: 950094.

[50] Chen H, Libertini SJ, George M, et al. Genome-wide analysis of androgen receptor binding and gene regulation in two CWR22-derived prostate cancer cell lines. ERC. 2010; 17(4): 857-873.

[51] Pudova EA, Kobelyatskaya AA, Katunina IV, et al. Lymphatic dissemination in prostate cancer: Features of the transcriptomic profile and prognostic models. Int J Mol Sci. 2023; 24(3): 2418.

[52] Alles J, Fehlmann T, Fischer U, et al. An estimate of the total number of true human miRNAs. Nucleic Acids Res. 2019; 47(7): 3353-3364.

[53] Kilikevicius A, Meister G, Corey DR. Reexamining assumptions about miRNA-guided gene silencing. Nucleic Acids Res. 2021; 50(2): 617-634.

[54] Tinel C, Lamarthée B, Anglicheau D. MicroRNAs: small molecules, big effects. Curr Opin Organ Transplant. 2021; 26(1): 10-16.

[55] Pudova EA, Krasnov GS, Nyushko KM, et al. miRNAs expression signature potentially associated with lymphatic dissemination in locally advanced prostate cancer. BMC medical genomics. 2020; 13(suppl 8): 129.

[56] Godínez-Rubí M, Ortuño-Sahagún D. miR-615 fine-tunes growth and development and has a role in cancer and in neural repair. Cells. 2020; 9(7): 1566.

[57] Laursen EB, Fredsøe J, Schmidt L, et al. Elevated miR-615-3p expression predicts adverse clinical outcome and promotes proliferation and migration of prostate cancer cells. Am J Pathol. 2019; 189(12): 2377-2388.

[58] Lichner Z, Fendler A, Saleh C, et al. MicroRNA signature helps distinguish early from late biochemical failure in prostate cancer. Clin Chem. 2013; 59(11): 1595-1603.

[59] Zhang Y, Campbell BK, Stylli SS, Corcoran NM, Hovens CM. The prostate cancer immune microenvironment, biomarkers and therapeutic intervention. Uro. 2022; 2(2): 74-92.

[60] McArdle PA, Canna K, McMillan DC, McNicol AM, Campbell R, Underwood MA. The relationship between T-lymphocyte subset infiltration and survival in patients with prostate cancer. Br J Cancer. 2004; 91(3): 541-543.

[61] Gollapudi K, Galet C, Grogan T, et al. Association between tumor-associated macrophage infiltration, high grade prostate cancer, and biochemical recurrence after radical prostatectomy. Am J Cancer Res. 2013; 3(5): 523-529.

[62] Ness N, Andersen S, Valkov A, et al. Infiltration of CD8+ lymphocytes is an independent prognostic factor of biochemical failure-free survival in prostate cancer. The Prostate. 2014; 74(14): 1452-1461.

[63] Zorko NA, Makovec A, Elliott A, et al. Natural killer cell infiltration in prostate cancers predict improved patient outcomes. Prostate Cancer Prostatic Dis. 2024; 28: 129-137.

[64] Xin S, Liu X, Li Z, et al. ScRNA-seq revealed an immunosuppression state and tumor microenvironment heterogeneity related to lymph node metastasis in prostate cancer. Exp Hematol Oncol. 2023; 12(1): 49.

[65] Ntala C, Salji M, Salmond J, et al. Analysis of prostate cancer tumor microenvironment identifies reduced stromal CD4 effector T-cell infiltration in tumors with pelvic nodal metastasis. Eur Urol Open Sci. 2021; 29: 19-29.

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2025 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.