Linda G. Griffith was 11 years old when the pain started.

TE LG RAD T RELIE Downloaded via 148.251.232.83 on April 25, 2019 at 01:56:00 (UTC). ee https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles. CREDIT: CRI GA Painful and enigmatic, endometriosis presents major obstacles for women and drug developers Linda G. Griffith was 11 years old when the pain started. A searing sensation, as if a massive stone burned at the base of her spine, overtook her every time she had her period, along with fainting spells and what felt like the flu. As a teenager, she would cry so hard that her mother supplied her with gin; later, when her misery disturbed the halls of her dorm, Griffith resorted to monthly injections from her college infirmary of the opioid Demerol. By the time she entered graduate school in the 80s, when over-the-counter ibuprofen became available, she was devouring 30 to 50 pills each day of her period. he also took countless, often frustrating trips to various doctors. Among the more dubious theories put forward to explain her pain was that Griffith who rode a motorcycle, taught karate, and studied chemical engineering was rejecting her femininity. Everyone told her that her extreme symptoms were just a part of menstruation. o one wanted to believe there was something really wrong, says Griffith, who is now 58 and a professor of biological and mechanical engineering at Massachusetts Institute of Technology. inally, at age 28, after years of not being able to walk or even sit during her period, she had surgery in the hope of uncovering the cause of her anguish. At last, Griffith had a name for her disorder: endometriosis. My intestines were twisted together in scar tissue; the endometriosis was just everywhere, all over everything, she says. Through all those years of doctor s visits, she says, no one ever mentioned endometriosis to me. The chronic disease happens when endometrial tissue, the inner lining of the TIE GUYE, C&E WAIGT uterus that sheds each month if a woman does not become pregnant and turns into placenta if she does, grows where it shouldn t. Misplaced, the tissue gloms on to the outside of the uterus or other reproductive organs to form lesions. These lesions cause pain, heavy menstruation, nausea, bloating, and constipation symptoms commonly blamed on bad periods or conditions like irritable bowel syndrome. Endometriosis affects about one in 10 women of childbearing age, yet awareness of the disease is severely lacking. Even now, 30 years after Griffith was diagnosed, women still wait eight to 10 years on average for a diagnosis, which requires a surgery in which doctors look for telltale lesions by poking a camera into a patient s abdomen. But diagnosis doesn t guarantee relief. The limited treatment options cause awful side effects, and sometimes, these treatments are utterly ineffective. cientists, Griffith among them, are trying to improve this situation. Ramping up their search in recent years, researchers are In brief Although endometriosis affects about one in 10 women around the world, treatment options for the serious, often debilitating disorder are sorely lacking. The chronic condition happens when endometrial tissue escapes the uterus, wreaking havoc on reproductive organs. It can cause infertility and extreme pain in women, many of whom are disbelieved by doctors. n average, they end up waiting almost a decade for a diagnosis. And even when they are finally diagnosed, they are left with treatments that at best offer temporary relief; at worst, they trigger gut-wrenching side effects. But scientists are working hard to understand the enigmatic disease and develop new diagnostic tests and drugs, some of which are already making their way into the clinic. or millions of patients, relief from the frustrating disease can t come soon enough. CTBER 29, 2018 CE.AC.RG C&E 29

hunting for chemical clues called biomarkers that could be used to detect the disease; some of these biomarkers might double as targets at which drugmakers can aim future treatments. Meanwhile, drugmakers have already started pushing new compounds through the pipeline. In July, AbbVie won U.. ood & Drug Administration approval for rilissa, touted by the company as the first new endometriosis drug in more than 10 years, although it s not without side effects. And several other clinical candidates, including a handful with novel modes of attack, may follow suit. or women struggle to convince doctors of their condition, maintain a career, and start a family, a solution can t come soon enough. And as Deborah Bush, CE of the nonprofit Endometriosis ew Zealand, points out, the disease doesn t affect just women and girls: In the U.. alone, endometriosis causes an estimated $18 billion $22 billion in economic losses per year from women whose symptoms prevent them from working. It s a major global public health issue, Bush says. Tough choices In the past, women with endometriosis have been presented with two main treatment options: surgery or hormone-based treatments. Both are saddled with serious drawbacks. In the first, more invasive route, surgeons cut or burn away endometrial lesions, sometimes leading to scarring that can worsen patients pain. And for some people, the fix is just temporary lesions can grow back. Griffith, for instance, has undergone nine surgeries in total, including two to remove lesions that persisted even after she had an emergency hysterectomy to remove her uterus. Christina ayse, who is 27 and was diagnosed with endometriosis six years ago, has already had two surgeries. After the second, in 2016, she was free from the pain that she describes as a sharp, shooting, stabbing, and burning feeling all at once. Though it was a welcome reprieve, she feared with each period that the pain would return. And exactly a year after the surgery, it did. ayse s doctor has recommended another surgery, this time one that cuts deeper into the tissue. But without insurance to fully cover the procedure, which she was told would run $26,000, she is left to live with the pain. The more common option for people with endometriosis is to take hormone-based drugs that suppress the estrogen and progesterone known to drive Linda Griffith compares a year of treatment for triple-negative breast cancer, during which she received a lot of sympathy and support, with 40 years of endometriosis, which she had to hide for many years. he says, TE CUMULATIVE UERIG AD METAL AGUI RM EDMETRII CMPLETELY, TTALLY DWAR MY EXPERIECE WIT TRIPLE- EGATIVE BREAT CACER. the endometrial lesions growth. f these, oral contraceptives are the first line of defense, which leaves some women to choose between starting a family and getting relief for their pain. Those who forgo treatment in order to conceive may still struggle: ne study found that 12% of women with endometriosis have issues with fertility, compared with 3% for those without endometriosis (Gynecol. bstet. Invest. 2017, DI: 10.1159/000452660). ayse and her husband have unsuccessfully tried to become pregnant for the past few years. Another option is to take noncontraceptive hormonal treatments called gonadotropin-releasing hormone (GnR) antagonists, which work by competing with natural GnR to block the release of hormones in the reproductive system. But they come with serious side effects. ayse tried one such GnR drug, AbbVie s Lupron Depot, and had terrible night sweats, mood swings, and hot flashes she was essentially plunged into menopause at age 21. Those six months were some of the hardest of her life, she says. Worst of all, the medication did nothing for her pain. AbbVie s new drug rilissa works in the same way and comes with similar symptoms, including bone loss, but it is packaged more conveniently as a pill instead of an injection like Lupron. While she waits for better treatments, ayse manages the pain with cannabis and Tylenol. he s hesitant to use opioids because of a family history of addiction, and the few times she has taken them, the pills made her severely nauseated. Like ayse, many women with endometriosis turn to alternative often unproven solutions when conventional treatments fail. Lindsey Peters, a 39-year-old communications professional, has tried all manner of treatments to control her symptoms, which include pain, cramps, bloating, and perpetual CREDIT: KATY WITTMA 30 C&E CE.AC.RG CTBER 29, 2018

Endometriosis by the numbers 10% in their reproductive years estimated to have endometriosis 30% experience pain during sex 53% experience menstrual pelvic pain 12% cannot become pregnant 8 10 years Average time it takes for a woman to be diagnosed with endometriosis $18 billion $22 billion Estimated economic loss per year in the U.. due to women with endometriosis who are unable to work ource: ertil. teril. 2017, DI: 10.1016/j.fertnstert.2017.01.009 CREDIT: CRITPER AYE bleeding, even while taking birth control pills. In addition to traditional options, Peters has tried probiotics, ginger, castor oil, magnesium, melatonin, flaxseed, milk thistle, and diindolylmethane. he doesn t eat dairy or red meat and has cut back on carbohydrates, sugar, and alcohol in the hope that this diet will help. Many of these treatment ideas are passed around online in endometriosis support groups, she says. We re so desperate for any type of help that we ll try it. Peters and ayse, like Griffith, also struggled to find sympathetic doctors. Peters, who has sought treatment from at least seven doctors, says one primary care physician refused to refer her to a specialist for months, saying that the symptoms were all in her head. It s affected everything, Peters says, who becomes emotional when describing socializing with friends and being physically active before the disease disrupted her life when she was in her early 30s. Dating is difficult, she says, because it takes energy that she simply doesn t have. It s made me into a recluse. Christina ayse started a business selling T- shirts and cushions for seat belts, which are often painful for people with endometriosis, and donates 10% of the proceeds to endometriosis research. he says, IT WAT KEEP ME GIG. Chasing causes With so many women s lives upended by their endometrial pain, new treatments are clearly needed, says Chandrakant Tayade, who has studied endometriosis for more than a decade at Queen s University in Canada. If we understand how and why they are getting the disease, he says, we can design better therapies, better diagnostics. Yet getting to the root of endometriosis has proved difficult. Women s symptoms vary wildly, and researchers have struggled to categorize their seemingly random presentation in patients. The frustrating fact is that scientists still don t know exactly what causes this enigmatic disease. The most accepted theory and Tayade emphasizes that it s still a theory about the cause of endometriosis was suggested by John A. ampson in 1927. e proposed that the condition occurs because of something called retrograde menstruation, a phenomenon in which endometrial tissue flows backward into the body instead of out, spreading debris into the abdominal cavity where it can collect on nearby organs. But retrograde menstruation happens naturally in up to 90% of women, and only about 10% of women have endometriosis. o why don t all women with retrograde menstruation also have endometriosis? cientists say the disease must have some other driver, though they can t quite agree on what that is. Tayade is among those that believe the difference lies in the patient s immune system. ormally, immune cells search and destroy any misplaced molecules. But in endometriosis, researchers think some malfunction could be keeping them from efficiently sweeping away debris. Another idea put forward by researchers is that the endometrial tissue in some women might be stickier, and thus primed for lesion building, than the tissue in oth- CTBER 29, 2018 CE.AC.RG C&E 31

ers. ther groups point to diet or genetic or environmental factors. ne study of about 60,000 women observed an increased risk of endometriosis for women reporting abuse earlier in life (um. Reprod. 2018, DI: 10.1093/humrep/dey248). till others theorize that the disease arises when cells on the surface of other organs or residual stem cells turn into endometrial tissue, which they say is supported by the rare (fewer than 10) cases of endometriosis in men (Int. J. Urol. 2012, DI: 10.1111/j.1442-2042.2012.03064.x). This diversity of opinion on endometriosis s underlying cause is partly why doctors still rely on laparoscopic surgery to diagnose the disease. Although researchers have come up with a laundry list of possible biomarkers cancer antigens, glycoproteins, microras, cytokines associated with immune and inflammation pathways, hormones, proteins that stimulate formation of blood vessels, and more none of these has gathered enough interest or data to be tested in the clinic. As the authors of a 2015 review article conclude: Despite the plethora of studies on endometriosis biomarkers, neither a single biomarker nor a panel of biomarkers has been validated for a noninvasive diagnostic test with sufficient sensitivity and specificity (BioMed Res. Int. 2015, DI: 10.1155/2015/130854). Whether a universal set of biomarkers even exists is up for debate, says Griffith, the MIT professor. he points out that the disease is broken down into four stages according to the number and location of lesions. But these stages don t neatly correlate with the severity of one s symptoms. ow could it possibly be one disease? Griffith asks. Instead of classifying the disease only by lesions size and location, which she compares to classifying cancers according to tumors physical attributes, she believes endometriosis should also be classified by its molecular mechanisms. ince entering the endometriosis research field in 2009, Griffith s research group has taken a systems biology approach to map the networks of cytokines and proteases in inflammatory pathways of the disease to then sort these pathways into a classification system. To study these pathways, the researchers are developing synthetic tissue cultures that can mimic endometrial cells. owever, Griffith acknowledges this type of organ-on-a-chip research for endometriosis still has a long way to go. Tayade, whose lab has identified several cytokines as potential biomarkers, remains optimistic. Advances in sequencing technology, which allows researchers to quickly look for genetic variations in patient samples, have accelerated discovery. Whereas Lindsey Peters started experiencing severe pelvic pain, cramps, and continuous bleeding. After months of pushing back against disbelief from her doctor, she was finally diagnosed with endometriosis. he says, IT TK A LT CVICIG. researchers used to be able to look at only a few genes at a time, his lab now uses a platform called anotring, which can evaluate hundreds of genes involved in inflammation in a sample. Tayade says despite the complicated nature of the disease, systematic studies and new technology will reveal targets for new drug classes within the next five years. Regardless, he says, I m not going to quit until I find something. The way forward Even if researchers figure out a good drug target say, an immune pathway gone awry the field faces other obstacles in translating that into a treatment. Translatability is often defined by the quality of animal models, explains Werner Lanthaler, CE of Evotec, which since 2012 has been working with Bayer to bring new endometriosis drugs to market. Replicating endometriosis in animals has been challenging, mainly because mice, drug developers favorite test subjects, don t menstruate. cientists have devised makeshift mouse models by injecting or suturing endometrial tissue onto mouse organs, but these models suffer from lack of reproducibility. Endometriosis does occur naturally in baboons, however. A model developed by Asgi T. azleabas at Michigan tate University establishes endometriosis in baboons using a technique that mimics retrograde menstruation (he supports ampson s theory of the cause of endometriosis). But baboon studies are expensive and, for ethical reasons, can be run on only a small scale. azleabas estimates that he has conducted studies on about 200 baboons over the course of his 20-year career. A major limitation of baboon and other animal models for endometriosis is that it s hard to quantify the test subjects pain relief. But to gain approval, endometriosis treatments need to show reductions in pain or infertility, tephen Palmer says. Palmer, like azleabas, has studied endometriosis for about two CREDIT: MEREDIT LEE 32 C&E CE.AC.RG CTBER 29, 2018

Clinical candidates to treat endometriosis Approved earlier this year, AbbVie s rilissa is the first new endometriosis drug in more than 10 years. Drugmakers hope these candidates will follow suit. 3 C 3 C 3 C C 3 ource: clinicaltrials.gov AME Dienogest Vilaprisan decades; he started in industry and is now at Baylor College of Medicine s Center for Drug Discovery. But the model for measuring infertility, essentially evaluating embryo implantation, he says, is very complicated and hardly used. o instead, researchers look for measurable outcomes that can act as a surrogate for pain, he says. Most drug candidates are evaluated by their ability to shrink lesions in animals, Palmer says, but that connection to pain isn t necessarily straightforward. Lesions located on ovaries might cause infertility but no pain, while lesions close to a nerve might be extremely painful but not affect fertility. ome mouse models attempt to BAY1817080 Linzagolix Relugolix C 3 C 3 BAY1902607 BAY2328065 C 3 Bayer MAKER Bayer/Evotec bseva Myovant ciences Bayer TATU I MECAIM ACTI Progestin Purino P2X3 antagonist Gonadotropinreleasing hormone antagonist Gonadotropinreleasing hormone antagonist teroidal selective progesterone modulator Bayer/Evotec Phase I ot disclosed Bayer/Evotec Phase I ot disclosed precisely install lesions, but they re imperfect, he says. Evotec and Bayer have also put better, more affordable animal models at the top of their research agenda. ne thing very clear to Bayer and to us, Lanthaler says, was that we cannot rely on any of the published animals from academic groups in endometriosis. Because the published protocols lacked consistency, the team decided to develop its own mouse model, the details of which are proprietary. Evotec s ability to develop animal models combined with Bayer s expertise in women s reproductive health was crucial for their unique alliance, he says. In 2012, the team of nearly 80 discovery scientists from both companies set a goal: It would put three novel non-hormonal-based candidates for endometriosis into clinical trials within five years. Last year the scientists hit their goal. Two of the candidates in Phase I and II clinical trials target the P2X3, a known pain target, and another candidate aims to inhibit the enzyme AKR1C3, which has also been implicated in cancer. Palmer and his team have also made some progress. The Baylor researchers are working on compounds that block a signaling kinase involved in inflammation called c-jun -terminal kinase. ne such compound showed promise in animal and early-stage human trials but ultimately didn t reduce pain sufficiently, Palmer says. The researchers are now screening more-potent Jun kinase inhibitors. e says the team is also in talks with extgen Jane, a start-up developing a smart tampon that could test for various reproductive diseases, to identify a biomarker in the inflammatory pathway for endometriosis. till others continue to focus on hormonal-based therapies. bseva and Myovant ciences both have such compounds moving through the clinic, and another company, called orendo Pharma and based in inland, is investigating compounds that interrupt local estrogen production at the lesions themselves. Meanwhile, Evotec and Bayer hope to eventually shift their focus from compounds that target pain associated with endometriosis to drugs that may shrink endometrial lesions, Lanthaler says. owever, he s cautious about spreading the news in endometriosis forums before they have a product with a clear proof of concept and profile in the clinic. It s just too severe a disease to raise patients hopes too early, he says. The mysteries behind endometriosis, the lack of endometriosis research funding, and patients ongoing suffering from endometriosis are all incredibly frustrating, Endometriosis ew Zealand s Bush says. Rather than despair about the challenges of the disease, she says, advocates should focus on helping patients, especially young ones, recognize the symptoms early and get appropriate treatment in a timely manner. If we educate well, if we manage well when these young people present, if we treat them with the expertise that all women deserve, then we re on the right path to doing as much as we can while the researchers figure out what s going on, she says. It s not K to say to a patient, Wait 10 years for an answer. We need to get in there now. CTBER 29, 2018 CE.AC.RG C&E 33