Immune micro-environment analysis and drug screening for ovarian endometriosis

Background Patients of ovary endometriosis have an abnormal immune micro-environment, leading to endometrial tissue that from retrograde menstruation evade immune surveillance and subsequently develop into ectopic lesions.

This study aims to elucidate the crucial immune cells and molecular pathways that are associated with an aberrant immune micro-environment of endometriosis.

In this study, we identified differentially expressed genes between ovarian ectopic endometrial tissue (OVE) and eutopic endometrial tissue from patients with endometriosis (PE) and non-endometriosis patients (CON) by analyzing the mRNA sequencing data. Additionally, we used WGCNA(Weighted Gene Co-expression Network Analysis) to screen for key genes related to immune cell infiltration and compared the sub-types of infiltrating immune cells using CIBERSORT(cell-type identification by estimating relative subsets of RNA transcript). Subsequently, we conducted a single-cell analysis on the identified key genes. Furthermore, we analyzed potential drugs suitable for ovarian endometriosis treatment using pRRophertic.

Seven key genes associated with immune cell infiltration were screened out. The expression of these genes in OVE was significantly lower than that in PE and CON. These key genes were mainly enriched in the NK cell-mediated cytotoxicity pathway, especially for CD16 + CD56dim NK. Moreover, NK cells infiltration in ovarian endometriosis was significantly reduced compared with PE and CON, while M2 macrophage shown the opposite. Results of the single-cell analysis showed that the expression of the seven key genes in NK cells and monocyte-macrophages in OVE was significantly lower than that in PE or CON. Additionally, we identified potential drugs suitable for ovarian endometriosis treatment.

The decreased infiltration of NK cells and increased infiltration of M2 macrophages contribute to the evasion of immune surveillance against endometrial tissue, promoting the progression of OVE. Therefore, potential strategies for the treatment of OVE include increasing NK cell activation and decreasing M2 macrophage polarization. Similar content being viewed by others

Artemova D, Vishnyakova P, Khashchenko E et al (2021) Endometriosis and cancer: exploring the role of macrophages. Int J Mol Sci. https://doi.org/10.3390/ijms22105196 Borrelli GM, Kaufmann AM, Abrão MS et al (2015) Addition of MCP-1 and MIP-3β to the IL-8 appraisal in peritoneal fluid enhances the probability of identifying women with endometriosis. J Reprod Immunol 109:66–73 Celik O, Hascalik S, Elter K et al (2008) Combating endometriosis by blocking proteasome and nuclear factor-kappaB pathways. Hum Reprod 23:2458–2465 Chen B, Khodadoust MS, Liu CL, Newman AM et al (2018) Profifiling tumor infifiltrating immune cells with Cibersort. Methods Mol Biol 1711:243–259 Chen S, Liu Y, Zhong Z et al (2023) Peritoneal immune microenvironment of endometriosis: role and therapeutic perspectives. Front Immunol 14:1134663 Eskenazi B, Warner ML (1997) Epidemiology of endometriosis. Obstet Gynecol Clin N Am 24:235–258 Fernandez IZ, Baxter RM, Garcia-Perez JE et al (2019) A novel human IL2RB mutation results in T and NK cell-driven immune dysregulation. J Exp Med 216:1255–1267 Gabriel M, Fey V, Heinosalo T et al (2020) A relational database to identify differentially expressed genes in the endometrium and endometriosis lesions. Sci Data 7:284 Galkin AV, Melnick JS, Kim S et al (2007) Identification of NVP-TAE684, a potent, selective, and efficacious inhibitor of NPM-ALK. Ptoc Natl Acad Sci USA 104:270–275 Geeleher P, Cox N, Huang RS (2014) pRRophetic: an R package for prediction of clinical chemotherapeutic response from tumor gene expression levels. PLoS One. https://doi.org/10.1371/journal.pone.0107468 Hao Y, Hao S, Andersen-Nissen E et al (2021) Integrated analysis of multimodal single-cell data. Cell 184:3573-3587.e29 He S, Hu Q, Xu X et al (2020) Advanced glycation end products enhance M1 macrophage polarization by activating the MAPK pathway. Biochem Biophys Res Commun 525:334–340 Hirayama D, Iida T, Nakase H (2017) The phagocytic function of macrophage-enforcing innate immunity and tissue homeostasis. Int J Mol Sci 19:1–14 Ito K, Murphy D (2013) Application of ggplot2 to pharmacometric graphics. CPT Pharmacomet Syst Pharmacol 2:e79 Jeung IC, Chung YJ, Chae B et al (2015) Effect of helixor A on natural killer cell activity in endometriosis. Int J Med Sci 12:42–47 Jørgensen H, Hill AS, Beste MT et al (2017) Peritoneal fluid cytokines related to endometriosis in patients evaluated for infertility. Fertil Steril 107:1191–99.e2 Kaushal JB, Shukla V, Sankhwar P et al (2021) Targeted inhibition of TAK1 abrogates TGFβ1 non-canonical signaling axis, NFκB/Smad7 inhibiting human endometriotic cells proliferation and inducing cell death involving autophagy. Cytokine 148:155700 Kuncman Ł, Orzechowska M, Stawiski K et al (2022) The kinetics of fms-related tyrosine kinase 3 ligand (flt-3l) during chemoradiotherapy suggests a potential gain from the earlier initiation of immunotherapy. Cancers (basel). https://doi.org/10.3390/cancers14163844 Laganà AS, Salmeri FM, Ban Frangež H et al (2020) Evaluation of M1 and M2 macrophages in ovarian endometriomas from women affected by endometriosis at different stages of the disease. Gynecol Endocrinol 36:441–444 Langfelder P, Horvath S (2008) WGCNA: an R package for weighted correlation network analysis. BMC Bioinform 9:559 Lee S, Ku SK, Bae JS (2016) Anti-inflammatory effects of dabrafenib on polyphosphate-mediated vascular disruption. Chem-Biol Interact 256:266–273 Leenen S, Hermens M, de Vos van Steenwijk PJ et al (2021) Immunologic factors involved in the malignant transformation of endometriosis to endometriosis-associated ovarian carcinoma. Cancer Immunol Immun 70(7):1821–1829 Li B, Tan TB, Wang L et al (2019) p38MAPK/SGK1 signaling regulates macrophage polarization in experimental autoimmune encephalomyelitis. Aging (albany NY) 11:898–907 Li Q, Yuan M, Jiao X et al (2021) M1 macrophage-derived nanovesicles repolarize m2 macrophages for inhibiting the development of endometriosis. Front Immunol 12:707784 Lunemann S, Martrus G, Hölzemer A et al (2016) Sequence variations in HCV core-derived epitopes alter binding of KIR2DL3 to HLA-C∗03:04 and modulate NK cell function. J Hepatol 65:252–258 Lv X, Wang D, Ma Y et al (2018) Analysis of the oncogene BRAF mutation and the correlation of the expression of wild-type BRAF and CREB1 in endometriosis. Int J Mol Med 41:1349–1356 Ma J, Zhang L, Zhan H et al (2021) Single-cell transcriptomic analysis of endometriosis provides insights into fibroblast fates and immune cell heterogeneity. Cell Biosci 11:125 Michael F (2002) Corrgrams. Am Statistician 256:316–324. https://doi.org/10.1198/000313002533 Miller JE, Ahn SH, Marks RM et al (2020) IL-17A modulates peritoneal macrophage recruitment and m2 polarization in endometriosis. Front Immunol 11:108 Morikawa Y, Murakami M, Kondo H et al (2021) Natural killer cell group 7 sequence in cytotoxic cells optimizes exocytosis of lytic granules essential for the perforin-dependent, but not fas ligand-dependent, cytolytic pathway. Immunohorizons 5:234–245 Pardo J, Balkow S, Anel A et al (2002) Granzymes are essential for natural killer cell-mediated and perf-facilitated tumor control. Eur J Immunol 32:2881–2887 Prescott J, Farland LV, Tobias DK et al (2016) A prospective cohort study of endometriosis and subsequent risk of infertility. Hum Reprod 31:1475–1482 Ramírez-Pavez TN, Martínez-Esparza M, Ruiz-Alcaraz AJ et al (2021) The role of peritoneal macrophages in endometriosis. Int J Mol Sci. https://doi.org/10.3390/ijms221910792 Robin X, Turck N, Hainard A et al (2011) pROC: an open-source package for R and S+ to analyze and compare ROC curves. BMC Bioinform 12:77 Shapouri-Moghaddam A, Mohammadian S, Vazini H et al (2018) Macrophage plasticity, polarization, and function in health and disease. J Cell Physiol 233:6425–6440 Shih AJ, Adelson RP, Vashistha H et al (2022) Single-cell analysis of menstrual endometrial tissues defines phenotypes associated with endometriosis. BMC Med 20:315 Sim MJ, Stowell J, Sergeant R et al (2016) KIR2DL3 and KIR2DL1 show similar impact on licensing of human NK cells. Eur J Immunol 46:185–191 Stoeckle C, Gouttefangeas C, Hammer M et al (2009) Cathepsin W expressed exclusively in CD8+ T cells and NK cells, is secreted during target cell killing but is not essential for cytotoxicity in human CTLs. Exp Hematol 37:266–275 Stuart T, Butler A, Hoffman P et al (2019) Comprehensive integration of single-cell data. Cell 177:1888-1902.e21 Syrop CH, Halme J (1987) Peritoneal fluid environment and infertility. Fertil Steril 48:1–9 Tan Y, Flynn WF, Sivajothi S et al (2022) Single-cell analysis of endometriosis reveals a coordinated transcriptional programme driving immunotolerance and angiogenesis across eutopic and ectopic tissues. Nat Cell Biol 24:1306–1318 Tao Y, Zhang Q, Huang W et al (2011) The peritoneal leptin, MCP-1 and TNF-α in the pathogenesis of endometriosis-associated infertility. Am J Reprod Immunol 65:403–406 Thiruchelvam U, Wingfield M, O’Farrelly C (2015) Natural killer cells: key players in endometriosis. Am J Reprod Immunol 74:291–301 Vallvé-Juanico J, Houshdaran S, Giudice LC (2019) The endometrial immune environment of women with endometriosis. Hum Reprod Update 25:564–591 Veljkovic DV, Dominovic M, Gulic T et al (2013) Granulysin expression and the interplay of granulysin and perforin at the maternal-fetal interface. J Reprod Immunol 97:186–196 Vera J, Paludo J, Kottschade L et al (2019) Case series of dabrafenib-trametinib-induced pyrexia successfully treated with colchicine. Support Care Cancer 27:3869–3875 Vora PA, Patel R, Dharamsi A (2020) Bortezomib - first therapeutic proteasome inhibitor for cancer therapy: a review of patent literature. Recent Pat Anticancer 15:113–131 Wang L, Li L, Li Y et al (2021) A history of endometriosis is associated with decreased peripheral nk cytotoxicity and increased infiltration of uterine cd68+ macrophages. Front Immunol 12:711231 Wex T, Bühling F, Wex H et al (2001) Human cathepsin W, a cysteine protease predominantly expressed in NK cells, is mainly localized in the endoplasmic reticulum. J Immunol 167:2172–2178 Wu T, Hu E, Xu S et al (2021) clusterProfiler 4.0: a universal enrichment tool for interpreting omics data. Innovation (n y) 2:100141 Yu G, Wang LG, Han Y et al (2012) clusterProfiler: an R package for comparing biological themes among gene clusters. OMICS 16:284–287 Yuan Z, Wang L, Wang Y et al (2014) Tubal origin of ovarian endometriosis. Mod Pathol 27:1154–1162 Zajec V, Mikuš M, Vitale SG et al (2022) Current status and challenges of drug development for hormonal treatment of endometriosis: a systematic review of randomized control trials. Gynecol Endocrinol 38:713–720 Zondervan KT, Becker CM, Missmer SA (2020) Endometriosis. N Engl JMed 382:1244–1256 Funding This study was supported by the grants from the National Natural Science Foundation of China (Grant no.: 82301856) and Guangdong Medical Science and Technology Research Fund (Grant no.: B2023145). Author information Authors and Affiliations Corresponding authors Ethics declarations Conflict of interest All the authors certify that there is no potential conflicts of interest with any financial or organization regarding manuscript. Additional information Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Supplementary Information Below is the link to the electronic supplementary material. Rights and permissions Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. About this article Cite this article Quan, Q., Gu, H., Wang, Y. et al. Immune micro-environment analysis and drug screening for ovarian endometriosis. Genes Genom 46, 803–815 (2024). https://doi.org/10.1007/s13258-024-01497-8 Received: Accepted: Published: Version of record: Issue date: DOI: https://doi.org/10.1007/s13258-024-01497-8