Presence of Urinary Exosomes for Liquid Biopsy of Clear Cell Renal Cell Carcinoma: Protocol for a Pilot Feasibility Study

Guorong Li, Nora Mallouk, Pascale Flandrin, Arnauld Garcin, Claude Lambert, Sid Ali Berremila, Hocine Habchi, Nicolas Mottet, Guorong Li, Nora Mallouk, Pascale Flandrin, Arnauld Garcin, Claude Lambert, Sid Ali Berremila, Hocine Habchi, Nicolas Mottet

Abstract

Background: Approximately 70%-80% of kidney cancers are clear cell renal cell carcinomas (CCRCCs). Patient management is based on imaging (abdominal ultrasound and computerized tomography), surgical excision of the tumor, and pathological analysis. A tissue biopsy is therefore necessary to confirm the diagnosis and avoid unnecessary nephrectomy. For metastatic cancers, a tissue biopsy is essential for establishing the targeted therapy. This biopsy of tumor material is invasive and painful. Other techniques such as liquid biopsy would help reduce the need for tissue biopsy. The development of a simple biological test for diagnosis is essential. CA9 is a powerful marker for the diagnosis of CCRCC. Exosomes have become a major source of liquid biopsy because they carry tumor proteins, RNA, and lipids. Urine is the most convenient biological liquid for exosome sampling.

Objective: The aim of this study (PEP-C study) is mainly to determine whether it is possible to detect urinary exosomal CA9 for the molecular diagnosis of CCRCC.

Methods: This study will include 60 patients with CCRCC and 40 noncancer patients. Exosomes will be isolated from urine samples and exosomal CA9 will be detected by transmission electron microscopy, flow cytometry, and reverse transcription-quantitative polymerase chain reaction.

Results: This study is currently underway with funding support from the CHU Saint-Etienne of France.

Conclusions: We expect to demonstrate that urinary tumor exosomes could be a novel liquid biopsy to diagnose CCRCC and to guide clinicians in treatment decision-making.

Trial registration: ClinicalTrials.gov NCT04053855; https://ichgcp.net/clinical-trials-registry/NCT04053855.

International registered report identifier (irrid): DERR1-10.2196/24423.

Keywords: CA9; clear cell renal cell carcinoma; kidney cancer; liquid biopsy; urine exosome.

Conflict of interest statement

Conflicts of Interest: None declared.

©Guorong Li, Nora Mallouk, Pascale Flandrin, Arnauld Garcin, Claude Lambert, Sid Ali Berremila, Hocine Habchi, Nicolas Mottet. Originally published in JMIR Research Protocols (https://www.researchprotocols.org), 20.07.2021.

Figures

Figure 1
Figure 1
Outline of study. RT-qPCR: reverse transcription-quantitative polymerase chain reaction
Figure 2
Figure 2
Transmission electron microscopy observation of urinary exosomes.

References

    1. Ljungberg B, Bensalah K, Canfield S, Dabestani S, Hofmann F, Hora M, Kuczyk MA, Lam T, Marconi L, Merseburger AS, Mulders P, Powles T, Staehler M, Volpe A, Bex A. EAU guidelines on renal cell carcinoma: 2014 update. Eur Urol. 2015 May;67(5):913–924. doi: 10.1016/j.eururo.2015.01.005.
    1. Li G, Cuilleron M, Gentil-Perret A, Tostain J. Characteristics of image-detected solid renal masses: implication for optimal treatment. Int J Urol. 2004 Feb;11(2):63–67. doi: 10.1111/j.1442-2042.2004.00750.x.
    1. Palmirotta R, Lovero D, Cafforio P, Felici C, Mannavola F, Pellè E, Quaresmini D, Tucci M, Silvestris F. Liquid biopsy of cancer: a multimodal diagnostic tool in clinical oncology. Ther Adv Med Oncol. 2018 Aug 29;10:1758835918794630. doi: 10.1177/1758835918794630.
    1. Keller S, Ridinger J, Rupp A, Janssen JW, Altevogt P. Body fluid derived exosomes as a novel template for clinical diagnostics. J Transl Med. 2011 Jun 08;9(1):86. doi: 10.1186/1479-5876-9-86.
    1. Yang J, Wei F, Schafer C, Wong DTW. Detection of tumor cell-specific mRNA and protein in exosome-like microvesicles from blood and saliva. PLoS One. 2014 Nov 14;9(11):e110641. doi: 10.1371/journal.pone.0110641.
    1. Zhao A, Péoc'h M, Cottier M, Genin C, Mottet N, Li G. Cell-free RNA content in urine as a possible molecular diagnostic tool for clear cell renal cell carcinoma. Int J Cancer. 2015 Jun 01;136(11):2610–2615. doi: 10.1002/ijc.29313.
    1. Melo SA, Luecke LB, Kahlert C, Fernandez AF, Gammon ST, Kaye J, LeBleu VS, Mittendorf EA, Weitz J, Rahbari N, Reissfelder C, Pilarsky C, Fraga MF, Piwnica-Worms D, Kalluri R. Glypican-1 identifies cancer exosomes and detects early pancreatic cancer. Nature. 2015 Jul 09;523(7559):177–182. doi: 10.1038/nature14581.
    1. Miranda KC, Bond DT, McKee M, Skog J, Păunescu TG, Da Silva N, Brown D, Russo LM. Nucleic acids within urinary exosomes/microvesicles are potential biomarkers for renal disease. Kidney Int. 2010 Jul;78(2):191–199. doi: 10.1038/ki.2010.106.
    1. Murakami T, Oakes M, Ogura M, Tovar V, Yamamoto C, Mitsuhashi M. Development of glomerulus-, tubule-, and collecting duct-specific mRNA assay in human urinary exosomes and microvesicles. PLoS One. 2014 Oct 2;9(9):e109074. doi: 10.1371/journal.pone.0109074.
    1. Spanu S, van Roeyen CRC, Denecke B, Floege J, Mühlfeld AS. Urinary exosomes: a novel means to non-invasively assess changes in renal gene and protein expression. PLoS One. 2014 Oct 13;9(10):e109631. doi: 10.1371/journal.pone.0109631.
    1. Street J, Koritzinsky E, Glispie D, Star R, Yuen P. Urine exosomes: an emerging trove of biomarkers. Adv Clin Chem. 2017;78:103–122. doi: 10.1016/bs.acc.2016.07.003.
    1. Li G, Cuilleron M, Cottier M, Gentil-Perret A, Lambert C, Genin C, Tostain J. The use of MN/CA9 gene expression in identifying malignant solid renal tumors. Eur Urol. 2006 Feb;49(2):401–405. doi: 10.1016/j.eururo.2005.10.025.
    1. Song SH, Jeong IG, You D, Hong JH, Hong BS, Song C, Joung JY, Moon KH, Cho YM, Ahn H, Kim C. VEGF/VEGFR2 or PDGF-β/PDGFR-β expression in non-metastatic, renal cell carcinoma: a prospective study with 1,091 consecutive cases. J Urol. 2013 Apr;189(4S):7681–7689. doi: 10.1016/j.juro.2013.02.160.
    1. Jung MK, Mun JY. Sample preparation and imaging of exosomes by transmission electron microscopy. J Vis Exp. 2018 Jan 04;(131):1–5. doi: 10.3791/56482.
    1. Pospichalova V, Svoboda J, Dave Z, Kotrbova A, Kaiser K, Klemova D, Ilkovics L, Hampl A, Crha I, Jandakova E, Minar L, Weinberger V, Bryja V. Simplified protocol for flow cytometry analysis of fluorescently labeled exosomes and microvesicles using dedicated flow cytometer. J Extracell Vesicles. 2015 Mar 31;4(1):25530. doi: 10.3402/jev.v4.25530.
    1. Peterson MF, Otoc N, Sethi JK, Gupta A, Antes TJ. Integrated systems for exosome investigation. Methods. 2015 Oct 01;87:31–45. doi: 10.1016/j.ymeth.2015.04.015.
    1. Wu Y, Deng W, Klinke DJ. Exosomes: improved methods to characterize their morphology, RNA content, and surface protein biomarkers. Analyst. 2015 Oct 07;140(19):6631–6642. doi: 10.1039/c5an00688k.
    1. McKiernan J, Donovan MJ, O'Neill V, Bentink S, Noerholm M, Belzer S, Skog J, Kattan MW, Partin A, Andriole G, Brown G, Wei JT, Thompson IM, Carroll P. A novel urine exosome gene expression assay to predict high-grade prostate cancer at initial biopsy. JAMA Oncol. 2016 Jul 01;2(7):882–889. doi: 10.1001/jamaoncol.2016.0097.
    1. McKiernan J, Donovan MJ, Margolis E, Partin A, Carter B, Brown G, Torkler P, Noerholm M, Skog J, Shore N, Andriole G, Thompson I, Carroll P. A prospective adaptive utility trial to validate performance of a novel urine exosome gene expression assay to predict high-grade prostate cancer in patients with prostate-specific antigen 2-10 ng/ml at initial biopsy. Eur Urol. 2018 Dec;74(6):731–738. doi: 10.1016/j.eururo.2018.08.019.

Source: PubMed

Patrocinadores y Colaboradores

Condiciones médicas

Intervenciones de drogas

3
Suscribir