[en] Melanomas are highly immunogenic tumors that have been shown to activate the immune response. Nonetheless, a significant portion of melanoma cases are either unresponsive to immunotherapy or relapsed due to acquired resistance. During melanomagenesis, melanoma and immune cells undergo immunomodulatory mechanisms that aid in immune resistance and evasion. The crosstalk within melanoma microenvironment is facilitated through the secretion of soluble factors, growth factors, cytokines, and chemokines. In addition, the release and uptake of secretory vesicles known as extracellular vesicles (EVs) play a key role in shaping the tumor microenvironment (TME). Melanoma-derived EVs have been implicated in immune suppression and escape, promoting tumor progression. In the context of cancer patients, EVs are usually isolated from biofluids such as serum, urine, and saliva. Nonetheless, this approach neglects the fact that biofluid-derived EVs reflect not only the tumor, but also include contributions from different organs and cell types. For that, isolating EVs from tissue samples allows for studying different cell populations resident at the tumor site, such as tumor-infiltrating lymphocytes and their secreted EVs, which play a central anti-tumor role. Herein, we outline the first instance of a method for EV isolation from frozen tissue samples at high purity and sensitivity that can be easily reproduced without the need for complicated isolation methods. Our method of processing the tissue not only circumvents the need for hard-to-acquire freshly isolated tissue samples, but also preserves EV surface proteins which allows for multiplex surface markers profiling. Tissue-derived EVs provide insight into the physiological role of EVs enrichment at tumor sites, which can be overlooked when studying circulating EVs coming from different sources. Tissue-derived EVs could be further characterized in terms of their genomics and proteomics to identify possible mechanisms for regulating the TME. Additionally, identified markers could be correlated to overall patient survival and disease progression for prognostic purposes.
Disciplines :
Immunology & infectious disease Oncology Biotechnology Life sciences: Multidisciplinary, general & others
Author, co-author :
Al Hrout, Ala'a; Institute of Experimental Immunology, University of Zurich, Zurich, Switzerland.
Levesque, Mitchell P; Department of Dermatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.
CHAHWAN, Richard ; University of Luxembourg > Faculty of Science, Technology and Medicine (FSTM) > Department of Health, Medicine and Life Sciences (DHML) ; Institute of Experimental Immunology, University of Zurich, Zurich, Switzerland.
External co-authors :
yes
Language :
English
Title :
Investigating the tumor-immune microenvironment through extracellular vesicles from frozen patient biopsies and 3D cultures.
Yim KHW Hrout A Borgoni S Chahwan R. Extracellular vesicles orchestrate immune and tumor interaction networks. Cancers (Basel) (2020) 12:1–23. doi: 10.3390/cancers12123696
Bobrie A Colombo M Raposo G Théry C. Exosome secretion: molecular mechanisms and roles in immune responses. Traffic (2011) 12:1659–68. doi: 10.1111/j.1600-0854.2011.01225.x
Steinbichler TB Dudás J Skvortsov S Ganswindt U Riechelmann H Skvortsova II. Therapy resistance mediated by exosomes. Mol Cancer (2019) 18:58. doi: 10.1186/s12943-019-0970-x
Théry C Ostrowski M Segura E. Membrane vesicles as conveyors of immune responses. Nat Rev Immunol (2009) 9:581–93. doi: 10.1038/nri2567
Yim KHW Borgoni S Chahwan R. Serum extracellular vesicles profiling is associated with COVID-19 progression and immune responses. J Extracell Biol (2022) 1:e37. doi: 10.1002/jex2.37
Dellar ER Hill C Melling GE Carter DR Alberto Baena-Lopez L Elizabeth Dellar CR. Unpacking extracellular vesicles: RNA cargo loading and function. J Extracellular Biol (2022) 1:e40. doi: 10.1002/JEX2.40
Krzyzaniak O Yim KHW Hrout A Peacock B Chahwan R. Assessing extracellular vesicles in human biofluids using flow-based analyzers. bioRxiv (2022) 2022:7.20.500853. doi: 10.1101/2022.07.20.500853
Gide TN Wilmott JS Scolyer RA Long G v. Primary and acquired resistance to immune checkpoint inhibitors in metastatic melanoma. Clin Cancer Res (2018) 24:1260–70. doi: 10.1158/1078-0432.CCR-17-2267
Jenkins RW Barbie DA Flaherty KT. Mechanisms of resistance to immune checkpoint inhibitors. Br J Cancer (2018) 118:9–16. doi: 10.1038/bjc.2017.434
Pitt JM Vétizou M Daillère R Roberti MP Yamazaki T Routy B et al. Resistance mechanisms to immune-checkpoint blockade in cancer: tumor-intrinsic and -extrinsic factors. Immunity (2016) 44:1255–69. doi: 10.1016/j.immuni.2016.06.001
Mannavola F Tucci M Felici C Stucci S Silvestris F. MiRNAs in melanoma: a defined role in tumor progression and metastasis. Expert Rev Clin Immunol (2016) 12:79–89. doi: 10.1586/1744666X.2016.1100965
Mione M Bosserhoff A. MicroRNAs in melanocyte and melanoma biology. Pigment Cell Melanoma Res (2015) 28:340–54. doi: 10.1111/pcmr.12346
Mirzaei H Gholamin S Shahidsales S Sahebkar A Jafaari MR Mirzaei HR et al. MicroRNAs as potential diagnostic and prognostic biomarkers in melanoma. Eur J Cancer (2016) 53:25–32. doi: 10.1016/j.ejca.2015.10.009
Varamo C Occelli M Vivenza D Merlano M lo Nigro C. MicroRNAs role as potential biomarkers and key regulators in melanoma. Genes Chromosomes Cancer (2017) 56:3–10. doi: 10.1002/gcc.22402
Latchana N Abrams ZB Harrison Howard J Regan K Jacob N Fadda P et al. Plasma microrna levels following resection of metastatic melanoma. Bioinform Biol Insights (2017) 11. doi: 10.1177/1177932217694837
Mo MH Chen L Fu Y Wang W Fu SW. Cell-free circulating miRNA biomarkers in cancer. J Cancer (2012) 3:432. doi: 10.7150/jca.4919
Michniewicz AG Czyz M. Role of mirnas in melanoma metastasis. Cancers (Basel) (2019) 11. doi: 10.3390/cancers11030326
Dror S Sander L Schwartz H Sheinboim D Barzilai A Dishon Y et al. Melanoma miRNA trafficking controls tumour primary niche formation. Nat Cell Biol (2016) 18:1006–17. doi: 10.1038/ncb3399
la Shu S Yang Y Allen CL Maguire O Minderman H Sen A et al. Metabolic reprogramming of stromal fibroblasts by melanoma exosome microRNA favours a pre-metastatic microenvironment. Sci Rep (2018) 8:1–14. doi: 10.1038/s41598-018-31323-7
Chen G Huang AC Zhang W Zhang G Wu M Xu W et al. Exosomal PD-L1 contributes to immunosuppression and is associated with anti-PD-1 response. Nature (2018) 560:382–6. doi: 10.1038/s41586-018-0392-8
Poggio M Hu T Pai CC Chu B Belair CD Chang A et al. Suppression of exosomal PD-L1 induces systemic anti-tumor immunity and memory. Cell (2019) 177:414–27.e13. doi: 10.1016/j.cell.2019.02.016
Xie F Xu M Lu J Mao L Wang S. The role of exosomal PD-L1 in tumor progression and immunotherapy. Mol Cancer (2019) 18:1–10. doi: 10.1186/s12943-019-1074-3
Shultz LD Goodwin N Ishikawa F Hosur V Lyons BL Greiner DL. Human cancer growth and therapy in immunodeficient mouse models. Cold Spring Harb Protoc (2014) 2014:694–708. doi: 10.1101/pdb.top073585
Wang M Yao LC Cheng M Cai D Martinek J Pan CX et al. Humanized mice in studying efficacy and mechanisms of PD-1-targeted cancer immunotherapy. FASEB J (2018) 32:1537–49. doi: 10.1096/fj.201700740R
Olson B Li Y Lin Y Liu ET Patnaik A. Mouse models for cancer immunotherapy research. Cancer Discovery (2018) 8:1358–65. doi: 10.1158/2159-8290.CD-18-0044
Anton D Burckel H Josset E Noel G. Three-dimensional cell culture: a breakthrough. vivo Int J Mol Sci (2015) 16:5517–27. doi: 10.3390/ijms16035517
Unger C Kramer N Walzl A Scherzer M Hengstschläger M Dolznig H. Modeling human carcinomas: physiologically relevant 3D models to improve anti-cancer drug development. Adv Drug Delivery Rev (2014) 79:50–67. doi: 10.1016/j.addr.2014.10.015
Hrout A Cervantes-Gracia K Chahwan R Amin A. Modelling liver cancer microenvironment using a novel 3D culture system. Sci Rep 2022 12:1 (2022) 12:1–14. doi: 10.1038/s41598-022-11641-7
Ravi M Paramesh V Kaviya SR Anuradha E Paul Solomon FD. 3D cell culture systems: advantages and applications. J Cell Physiol (2015) 230:16–26. doi: 10.1002/jcp.24683
Ingram M Techy GB Saroufeem R Yazan O Narayan KS Goodwin TJ et al. Three-dimensional growth patterns of various human tumor cell lines in simulated microgravity of a NASA bioreactor. In Vitro Cell Dev Biol Anim (1997) 33:459–66. doi: 10.1007/s11626-997-0064-8
Mehta G Hsiao AY Ingram M Luker GD Takayama S. Opportunities and challenges for use of tumor spheroids as models to test drug delivery and efficacy. J Control Release (2012) 164:192–204. doi: 10.1016/j.jconrel.2012.04.045
Restivo G Tastanova A Balázs Z Panebianco F Diepenbruck M Ercan C et al. Live slow-frozen human tumor tissues viable for 2D, 3D, ex vivo cultures and single-cell RNAseq. Commun Biol (2022) 5. doi: 10.1038/s42003-022-04025-0
Neal JT Li X Zhu J Giangarra V Grzeskowiak CL Ju J et al. Organoid modeling of the tumor immune microenvironment. Cell (2018) 175:1972–1988.e16. doi: 10.1016/J.CELL.2018.11.021
Dong L Zieren RC Horie K Kim CJ Mallick E Jing Y et al. Comprehensive evaluation of methods for small extracellular vesicles separation from human plasma, urine and cell culture medium. J Extracell Vesicles (2020) 10:e12044. doi: 10.1002/JEV2.12044
Vogel R Savage J Muzard J della CG Vella G Law A et al. Measuring particle concentration of multimodal synthetic reference materials and extracellular vesicles with orthogonal techniques: who is up to the challenge? J Extracell Vesicles (2021) 10:e12052. doi: 10.1002/JEV2.12052
Crescitelli R Lässer C Lötvall J. Isolation and characterization of extracellular vesicle subpopulations from tissues. Nat Protoc 2021 16:3 (2021) 16:1548–80. doi: 10.1038/s41596-020-00466-1
