The Hallmarks of Endometriosis

In 2016, the kinase signaling pathways in endometriosis were investigated and it was concluded that the three main pathways to be targeted for treatment purposes are the IKKβ/NFκB, the MAPK, and the PI3K/AKT/mTOR pathway 20 . The literature on medications that specifically target the molecular and signaling pathways involved in the pathophysiology of endometriosis was thoroughly reviewed. The discussion included possible therapeutic targets, the molecules upstream and downstream that exhibit critical aberrant signaling, and the regulatory pathways that facilitate the expansion and maturation of endometriotic tissues and cells 21 . Recently, Shi also examined angiogenesis, lymphangiogenesis, neurogenesis, progesterone resistance, genetic alterations, estrogen-dependent induction of inflammation, imbalances in proliferation and apoptosis, and tissue remodeling in the pathogenesis of endometriosis. Additionally, the pharmacological mechanisms, constitutive relationships, and potential applications of each compound were studied as well 22 . Based on these works, a thorough search of both the ClinicalTrials.gov and the European Union Clinical Trials Register was conducted with a view to identifying completed and ongoing clinical studies investigating the role of the aforementioned pathways in patients with endometriosis. Table 1 and Table 2 briefly summarize the search results. Table 3 provides a brief overview of the relevant preclinical studies.

Over the past years, the scientific community has been able to investigate different molecular pathways and gain an insight into the (epi-)genetic and/or cellular mechanisms that seem to play a significant role in the genesis and progression of endometriosis. Of utmost significance, these pathomechanisms seem to pave new ways in the context of endometriosis diagnosis (as biomarkers) and therapy (as drug targets). The (epi-)genetic mechanisms are involved in the immunologic, immunohistochemical, histological, and biological aberrations of endometriosis 16 . Pelvic endometriosis has a complex pathogenesis and pathophysiological features. Two possible causes of the endometriotic lesions are in situ coelomic metaplasia of the peritoneal lining and transplantation of endometrial tissue through retrograde menstruation. In cases of extrapelvic lesions, vascular or lymphatic metastasis most likely happens infrequently. Through interacting molecular mechanisms that support cellular adhesion and proliferation, systemic and localized steroidogenesis, localized inflammatory response and immune dysregulation, as well as vascularization and innervation, superficial and deep endometriotic lesions seem to be established and maintained 17 . Endometriosis-related signaling pathways included estrogen-2, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), mitogen-activated protein kinase (MAPK), extracellular-signal regulated kinase (ERK), phosphatidylinositol 3-kinase (PI3K), protein kinase B (PKB/AKT) and mechanistic target of rapamycin (mTOR) (PAM), yes-associated protein (YAP), Wnt/β-catenin, Rho-associated protein kinase (ROCK), transforming growth factor β (TGF-β), vascular endothelial growth factor (VEGF), nitric oxide (NO), iron, cytokines and chemokines 18 . Despite being a benign condition, endometriosis exhibits malignant traits such as metastasis, hyperplasia, and cell invasion. This suggests a possible connection between endometriosis and particular signaling molecules and pathways that influence the invasion and metastasis of numerous common malignancies. The six biological abilities that are acquired throughout the multi-step development of human tumors are the hallmarks of cancer. The defining multiple characteristics provide a framework for understanding the complexity of neoplastic disease. The ability to maintain proliferative signaling, avoid growth suppressors, withstand cellular death, permit replicative immortality, trigger angiogenesis, and initiate invasion and metastasis are a few of them. These hallmarks are underpinned by inflammation, which supports several hallmark functions, and genome instability, which produces the genetic diversity that speeds up their acquisition. Two newer hallmarks of potential generality include reprogramming of energy metabolism and immune escape 19 . In this regard, Fig. 3 summarizes the corresponding hallmarks of endometriosis. Hallmarks of endometriosis and their potential targets for therapy. Captions in blue: completed clinical trials. Captions in green: ongoing clinical trials. Captions in black: experimental models in vitro/in vivo. Abbreviations: ADAM17 = A Disintegrin and Metalloprotease 17; COX-2 = Cyclooxygenase 2; E2 = Estrogen 2; EPAC1 = Exchange Protein Directly Activated by cAMP 1; ERK = Extracellular Signal-regulated Kinase; FAK = Focal Adhesion Kinase; FGFR2 = Fibroblast Growth Factor Receptor 2; GPR30 = G Protein-coupled Receptor 30; HIF = Hypoxia-inducible Factor; LXA4 = Lipoxin A4; MAPK = Mitogen-activated Protein Kinase; METTL3 = Methyltransferase-like 3; miR = micro RNA; MMP = Matrix Metallopeptidase; NF-κB = Nuclear Factor kappa B; NGF = Nerve growth factor; NLRP3 = NLR Family Pyrin Domain Containing 3; NO = Nitric Oxide; PGE2 = Prostaglandin E2; PKA = Protein Kinase A; PI3K/Akt/mTOR = Phosphatidylinositol 3-kinase/Protein Kinase B/mammalian Target of Rapamycin; PPAR = Peroxisome Proliferator-activated Receptor; PrPC = Cellular Prion Protein; Rap1 = Ras-associated Protein-1; ROCK = Rho-associated Protein Kinase; S1P = Sphingosine-1-Phosphate; SERCA = Sarcoplasmic/endoplasmic Reticulum Ca2+-ATPase; SF = Steroidogenic Factor; SHH = Sonic Hedgehog; SHP-1 = Src Homology Region 2 Domain-containing Phosphatase-1; sICAM-1 = soluble form of intercellular adhesion molecule-1; SMAD = Suppressor of Mothers against Decapentaplegic; SRC = Steroid Receptor Coactivator; TGF = Transforming Growth Factor; TIMP = Tissue Inhibitor of MMP; TNF = Tumor Necrosis Factor; VEGFR = Vascular Endothelial Growth Factor Receptor; ZEB1 = Zinc Finger E-Box Binding Homeobox 1

A great number of extensive review articles has so far been published on the role of signaling pathways/molecules in endometriosis 16 17 18 19 20 21 22 . Of note, given some shared molecular (genetic) mechanisms, endometriosis seems to be associated with various risk factors and other disease entities such as migraine, autoimmunity and chronic pelvic pain 23 24 25 26 ( Fig. 4 ). Chronic pain, for instance, seems to share similar pathomechanisms as endometriosis in terms of abundance of proinflammatory molecules, angiogenesis and estrogen-dependent pain meditation 23 . Even though endometriosis is not yet officially classified as an autoimmune disease, there are a number of similarities between the two conditions, including a predominance of females (and hormones), immunological abnormalities, genetic polymorphisms, as well as chronicity 24 . In the case of migraine, mechanisms associated with sex hormone activities, protein adhesion, phosphorylation, inflammation or immune dysregulation seem to play a similar role as in the pathogenesis of endometriosis 25 . Endometriosis is a disease condition encountering gynecologists every single day in both the outpatient and the clinic routine. Patients seek medical advice either because of the adverse pain symptoms and/or due to the unfulfilled desire to become pregnant. Unfortunately, most patients are very disappointed once they learn that surgery does not grant the end of the disease and that the only possible symptomatic treatment is hormone-based. In times of targeted treatment therapies and ample possibilities to investigate and discover novel therapeutic approaches (i.e. inflammation, apoptosis, angiogenesis, cellular adhesion, etc.), endometriosis represents a profound example of an understudied disease that to date may only be treated symptomatically. The present work aims at raising the awareness of both researchers and clinicians in this context and to highlight the need of further research in order to establish and launch targeted therapies for the successful treatment of endometriosis patients. All in all, we herein intended to summarize the current research status and to point out the field’s novel therapeutic approaches. However, the considerable side effects of these targeted therapies need to be further examined and taken into consideration in the context of risk–benefit calculation. Risk factors and associated conditions with endometriosis 14 17 23 24 25 26 .

Endometriosis describes a disease characterized by the colonization of endometrium-like lesions outside the uterine cavity. Ectopic lesions were thought to represent solely lesions on the peritoneum of the internal genital organs (endometriosis genitalis externa), but in the meantime a migration of endometrial-like cells into the myometrium has been also described, hence rendering adenomyosis uteri (= endometriosis genitalis interna) a distinct disease entity. However, since endometriotic lesions may also infiltrate deeply into organs (mostly bowel, bladder or ureter) (deep infiltrating endometriosis) or even spread to the diaphragm or the umbilicus (extragenital endometriosis), symptoms are often extremely complicated 1 . A clinical/intraoperative distinction is made between the following four major entities of endometriosis depending on localization and extent: superficial, ovarian, uterine and deep infiltrating endometriosis. Deep infiltrating endometriotic lesions exceed the surface (usually the peritoneum) and invade into neighboring tissue or organs with an infiltration depth of at least 0.5 cm ( Fig. 1 ) 2 3 . The most widely used clinical/intraoperative classification is the rASRM score, the revised classification of the American Society for Reproductive Medicine (formerly the American Fertility Society) 2 4 . The rASRM score describes peritoneal and ovarian endometriosis. Deep endometriosis is included in the calculation of the numerical value, but no mapping or classification can be derived from it. To remedy this deficiency, a German-speaking working group has developed the Enzian classification. This classifies deep lesions in 3 anatomical levels or compartments (A: rectovaginal septum/vagina, B: sacrouterine ligament/pelvic wall, C: rectum). #Enzian represents since 2021 a novel comprehensive classification that included the superficial endometriosis, ovarian and with the Enzian classification, hence constituting a more rounded classification system, which, nevertheless, does not incorporate the two major symptoms of endometriosis: pain and infertility 5 . Deep infiltrating endometriosis-Intraoperative laparoscopic views (Patient collective-Department of Gynecology, University Hospital of Erlangen). Diagnosis of endometriosis is based on a detailed medical history, a thorough gynecological clinical examination including vaginal and rectovaginal or rectal palpation, a transvaginal and/or even transrectal sonographic evaluation, a renal ultrasonography with a view to ruling out asymptomatic urinary retention caused by deep infiltrating endometriosis of the ureter, magnetic resonance imaging, as well as a histological examination 6 7 8 . The diagnosis of a deep infiltrating endometriosis is mainly clinical – by describing the clinical symptoms (although not specific), inspection with two-leaf specula and vaginal and rectal palpation. Vaginal sonography should be performed first as an imaging measure, not least because of the simultaneous possibility of identifying ovarian endometriomas. Furthermore, deep rectal infiltration can be easily diagnosed by an experienced physician. If rectal endometriosis is suspected, an endosonography and/or colorectoscopy is often automatically arranged. However, endometrial infiltration of the mucosa is rather rare. A colorectoscopy should be performed in the presence of intestinal bleeding and whenever a bowel resection is intended in the case of suspected bowel infestation in order to rule out primary bowel pathologies such as polyps, tumors or inflammatory bowel diseases 9 . Fig. 2 summarizes the possible diagnostic approaches based on the four major endometriosis entities. Taken altogether, endometriosis genitalis (including vaginal endometriosis) is mainly associated with dysmenorrhea and dyspareunia, deep infiltrating endometriosis correlates with dysuria and dyschezia, while extragenital endometriosis (in organs other than the bladder or the bowel) requires a symptom-oriented examination 6 7 8 9 10 . Importantly, superficial peritoneal endometriosis might remain obscure until the performance of a diagnostic laparoscopy. Diagnostic approaches based on the four major endometriotic entities. Conservative options include medical and complementary procedures, reproductive medicine measures as well as multimodal pain management models (i.e. heat application, physical exercises, etc.) and psychotherapy in the wider context of the bio-psycho-social model. Surgical options include organ-preserving or radical and, if necessary, interdisciplinary ablation or excision of endometriosis lesions, preferably in certified endometriosis facilities 11 . Established pharmacologic approaches include either analgesics from the group of non-steroidal anti-inflammatory drugs (NSAIDs) or hormone therapy. NSAIDs pursue a symptomatic therapeutic approach 12 . As non-hormonal options treat purely symptomatically, hormone therapy is generally used. Established hormonal options include progesterons in the first-line therapy, as well as oral contraceptives and Gonadotropin-Releasing Hormone (GnRH) (ant-)agonists in the second-line therapy. There are no objectifiable differences with regard to the reduction of typical pain symptoms. There are differences in terms of undesirable side effects, the duration of possible use and the costs. Drug therapy is only effective while it is being taken, after which symptoms may recur immediately 13 . Progestogens (especially dienogest 2 mg) are the options for first-line therapy. They produce hypoestrogenism through anovulation. In oral contraceptives, they are part of a fixed combination of ethinylestradiol or estradiol valerate. When selecting progestogen, secondary treatment goals such as the treatment of skin blemishes can also be taken into account. Long-cycle use is more effective than cyclical use in reducing symptoms typical of endometriosis and should be favored 11 13 14 15 .