Welcome everybody, I'm Andy Bauer from Children's Hospital of Philadelphia and I've been recruited to moderate this session which looks to be fabulous. So welcome, we have six excellent abstracts to go through, 10 minutes for presentations, five minutes for questions, and so this goes till 12.30. The first two, if we stick to the schedule, are on neuroregulation of puberty, two on CAH, and one on Turner's and one on treatment of genetic short stature. So I think this is really novel and up-to-date abstracts and congratulate those who are presenting today. So with that, we will get started and Stephanie Roberts is our first presenter. Thank you. Thank you for the invitation to speak today. The onset of puberty is marked by reactivation of gene rate secretion after a period of quiescence in childhood. When this occurs prematurely clinically, this manifests as central precocious puberty and conversely, if this is delayed or absent, this manifests as self-limited delayed puberty or IHH, respectively. Genetic variants, including McCORN Ring Finger Protein 3 or MCARIN3, which I've denoted here in the red boxes, have an important role in the timing of puberty on a population basis as well as in pubertal disorders. In rodents, the most potent activator of gene rate secretion is cispeptin. Specialized cispeptin neurons in the arcuate nucleus called CANDI neurons co-secrete neurokinin B as well as dynorphin. Loss of function mutations in MCARIN3 are the most common genetic cause of central precocious puberty with an estimated pooled prevalence of 9% and occurring in up to 46% of familial central precocious puberty cohorts. MCARIN3 acts as an inhibitor of gene rate secretion. As shown in the figure here, its action is mediated in part via actions on cispeptin and neurokinin B in the arcuate CANDI neurons. Additionally, MCARIN3's structure includes a ring finger domain which has E3 ubiquitin ligase activity which targets proteins for degradation. For this study, we aim to characterize the effects of MCARIN3 overexpression in the arcuate nucleus in post-pubertal mice. The rationale for this study is based on the age-related pattern of decline of MCARIN3 mRNA in the wild-type arcuate nucleus as well as the human and mouse genetic variants in the genes CIS1 and CIS1R, shown here in the red circle, that have been reported in both precocious puberty and hypogonadotropic hypogonadism based on if there are gain or loss-of-function mutations, respectively. Finally, overexpression of human MCARIN3 in the nematode C elegans has been previously demonstrated to result in delayed sexual differentiation. So we have previously generated a model of neonatal hypothalamic MCARIN3 overexpression. This was using recombinant viruses that are injected into wild-type mice into the lateral ventricle on postnatal day one, leading to widespread neuronal transduction. Shown here are the female mice, and you can see here the purple circles are MCARIN3 overexpressing mice, and the green are control-injected animals, and we demonstrated a significant delay in female mice in the mean age of vaginal opening and first estrus. This was found to be in association with a significant reduction in cispeptin protein, you can see here in the MCARIN3 overexpressing females at postnatal day 28, as well as neurokinin B protein expression in the arcuate nucleus in these animals. Not shown here is the reproductive phenotype of our male mice, which did not exhibit a delay in pubertal onset by puprucial separation. So for this study, we again used wild-type mice. We used mice that were postpubertal at three to four months of age that underwent bilateral stereotaxic injection targeting the arcuate nucleus of the hypothalamus. And again, we're using these overexpressing viruses. So you can see here our MCARIN3 overexpressing virus has the fluorescent tag, and then our control animals are also injected, but only with a virus that has EGFP. These animals subsequently underwent reproductive phenotyping and neuropeptide stimulation testing. So first I'm going to show the validation that we performed via immunohistochemistry, both for these viruses, as well as our injection coordinates. So on the left two panels here is a control-injected animal, and on the right panels is an MCARIN3 overexpressing animal. And you can see here that when we perform immunohistochemistry for EGFP protein, that both animals demonstrate EGFP protein expression, and this is an expected finding. In contrast, in the lower two panel, you can see absent MCARIN3 protein expression in our control-injected animals, and you can see easily appreciate the MCARIN3 protein in our MCARIN3 overexpressing animals. So this was the first step that we validated our model. So I'm going to start first with the reproductive phenotyping that we performed in females. So in the upper three panels, these are representative control-injected females that we followed the estrocyclicity for 55 days following injection. The bottom three panels are three representative MCARIN3 overexpressing females, and you can see here that the cycling is somewhat disrupted. When we look cumulatively at the mean percent time that our cohort spent, we find that in the MCARIN3 overexpressing females, they spend significantly less time in estrous and significantly more time in diastreous. We then performed general peptide stimulation testing, and so for this set of studies, these are gonad-intact females, and these studies were performed in the diastreous phase of the estrous cycle. So for these studies, we performed tail vein sampling to assess serum LH levels at time zero, 20, 40, and 60 minutes following administration of the neuropeptide, and I should note that this was peripheral administration, not ICV of these agents, and what you can see is that in response to the NK3R agonist synctide, this is a dose that's commonly cited in the literature. You can see that compared to the baseline, our control animals have a peak response at 20 minutes. This is a pattern that we typically see, but our MCARIN3 overexpressing females have a blended response. So there was significantly less LH at time 20 in our MCARIN3 overexpressing females compared to our controls. We then went on to challenge these animals with a higher dose of synctide just to see if we might be able to overcome this effect, and you can see here that actually now the control animals have an even higher response in their LH, but again, the MCARIN3 females at time 20 have a significant reduction in their LH level. In response to peripheral administration of cispeptin, we did not find any differences in response between the two groups. So because estradiol can impact the response to some of these agents, we've repeated these studies in animals that were over-ectomized and estradiol replaced, and again, we see the same study and the same response to synctide here in our MCARIN3 overexpressing females in a similar response in cispeptin to the gonad intact model. We also challenged these animals with GNOH, and you can see here that there was no difference in response between groups. So what about LH pulsatility? For these studies, these were in over-ectomized only females. You can see here three representative control-injected females with LH pulses denoted by the red star. And on the bottom are three representative MCARIN3 overexpressing females, and you can see these look fairly similar. So when we look at the whole cohort at the mean number of LH pulses, we see no difference between the two groups in the mean number of pulses or in the area under the curve. And for these LH pulsatility studies, we detail vein sampling every eight minutes over three hours. So putting these findings together, this really, again, localizes the mechanism of action in the candy neurons of the arcuate nucleus. So the next steps of the project is really to understand what's going on at the level of the protein. As I mentioned, MCARIN3 can target proteins for degradation. And so this is preliminary, but I'll just share with you today that the first protein we've looked at is neurokinin B. And so here you can see a control-injected animal. This is the arcuate nucleus, and you can hopefully appreciate here this bright red area is the neurokinin B protein by immunohistochemistry. In our MCARIN3 overexpressing female, you can see that there is a significant reduction in the protein that's visible. So what about the males? Does arcuate MCARIN3 overexpression also impact male mice via the same mechanism? So we did not observe a reproductive phenotype in our MCARIN3 overexpressing males. We looked at body weight over time, serum testosterone levels, and the response in LH to gonadectomy at three, seven, and 10 days. We also performed neuropeptides simulation testing, and I should note that this is in a gonad intact model. And interestingly, in the males, in response to synctide, you can see that we don't have that blunted response in the MCARIN3 overexpressing females that we noted in the females. And the response here to caspeptin and generH mimics what we saw in the females. So in summary, this is a novel post-pubertal mouse model to understand the mechanism of action of MCARIN3. And as I've shown in the schematic here on the right, the hypothalamic arcuate overexpression of MCARIN3 leads to a sex-specific response to the NK3R agonist synctide. We saw a reduced peak response to synctide in our MCARIN3 females, but not male mice, in a normal response to peripheral administration of caspeptin and generH. We also saw a reduction in neurokinin B protein expression in our MCARIN3 overexpressing females. So this supports MCARIN3's inhibitory action in candine neurons in the arcuate nucleus. MCARIN3 overexpression may inhibit caspeptin synthesis or secretion, and or the action or expression of the NK3R, and these remain our future directions for this study. This also suggests that modulation of MCARIN3 expression may be a therapeutic target for reproductive disorders, such as an alternative to generH agonist. I'd like to thank Dr. Ursula Kaiser and the members of the laboratory who contributed to this work, as well as Dr. Sina Estevez, who helped with the production of our viruses for these studies and our sources of funding. Thank you. Nice work. We'll open for questions now. The center mic is the one that's active. Thanks for a great talk. Two quick questions. Firstly, how do you explain the sex differences, which is really interesting? And the second thing, I was just wondering, have you looked at the MC3 receptor, given its recent role in puberty, and is there any interaction between MKRN3 and the MC3 receptor? So I'll answer your second question first, if that's okay. So we have not, but that's a great suggestion. So thank you for that. And in response to your first question, of course, for the males, we did validate those animals to ensure that the injections were accurate and that they were, in fact, MKRN3 overexpressing. I think this question about potential sex differences with MKRN3 remains an area that we're very interested in understanding more. Certainly, the mouse models, at least by the measures that we perform, reproductive phenotyping in rodents, females seem to be more affected than the male mice. But whether that actually translates to differences in the mechanisms of action, that we still need to understand more. In humans, certainly, we know that loss of function mutations in this gene in association with precocious puberty, that actually, if you look at the meta-analysis that I showed from 2019, boys are actually more likely to harbor a mutation than girls. So both boys and girls are certainly affected by mutations in this gene. Great, thank you. Yonichi from NIH, thank you very much for your excellent talk. I'm sorry if you mentioned this, but did you mention what age you overexpressed MKRN3? Yeah, so for the studies that I showed for this presentation, we used wild-type mice at three to four months of age. Did you compare how it would be different between when you expressed earlier, like seven days old or even younger, and the later age? Did you see any differences? So the other model that we've generated is the neonatal model. It's not, of course, performing stereotoxic injection, because the animals are only one day of life. And that's because we want to maintain MKRN3 expression before it's actually declining. That graph that I showed where it's going down before the onset of puberty. So those methods are a bit crude in terms of the ways that we can overexpress MKRN3 at that age, but those are the two ages that we've looked at. Okay, I don't see any questions online either. I understand one of our speakers is gonna be a minute or two over, so if we can save a minute or two, I think we'll move forward and bring in our next speaker. Again, really elegant work. Thank you so much. Thank you. All right, our second speaker, Dr. Flavia Tonano. No. No? No. Okay. Okay. From the University of Sao Paulo. Yes. Okay. Same place. You start, or I start? They should start. Ah, okay. I'll restart the timer, though. Good morning, first I would like to thank the meeting organization for the opportunity to present this work. We have nothing to disclose. Familial central precocious puberty is defined by the early onset of secondary sexual characteristics in more than one family member. In this setting, relatives of first, second, or third degree of the province are usually diagnosed by clinically documented central precocious puberty or precocious menarche, which is the age at menarche less than or equal to nine years. Suggestive characteristics in males may be early few puberty, including full facial shaving and voice breaking, and early timing of growth spurt or growth complexion, and in both sexes and adult short stature. In the past, familial central precocious puberty was described in isolated case reports. Since DeVry and collaborators published the first study of prevalence estimates and modes of inheritance based on pedigree analysis, they suggested an autosomal dominant transmission with incomplete sex dependent penetrance and the predominance of female cases. Since that moment, the prevalence of familial central precocious puberty has been estimated in 25 to 27% of cases. In the last years, loss of function mutations in two imprinted genes have been described as the most prevalent monogenic causes of familial CPP. In MKRN3 and DLK8 dose genes, the inheritance is autosomal dominant and the transmission is exclusively paternal due to the maternal imprinting. In this situation, in both genes normally, the maternal allele is silenced and only the paternal allele is expressed. So if we have the mutation, if the patient had a mutation in the paternal allele, the CPP phenotype is expressed, but if the mutation is in the maternal allele, the CPP phenotype is not expressed because the maternal allele is already silenced by imprinting. Here our objectives were to estimate the prevalence of familial CPP in a large cohort, to characterize the modes of inheritance and transmission, to compare clinical and hormonal characteristics of the probands with familial CPP due to different modes of transmission, and to characterize the genetic basis of the involved families. To estimate the prevalence, we analyzed the two cohorts of CPP patients from Spain and Brazil, comprising 418 patients from 394 families. The Spanish patients were from the Spanish Puberi Registry, and the Brazilian patients were from the University of São Paulo Tertiary Hospital. All patients with criteria for familial CPP were included. They were all categorized in four distinct modes of transmission based on pedigree analysis, maternal, paternal, undetermined, and both maternal and paternal. In the last years, our group has been performing DNA sequencing analysis of MKRN3 and DLK1 genes by Sanger sequencing method and large-scale parallel sequencing methods. Here genetic findings identified in previous studies were pooled for a quantitative analysis. Our results identified a prevalence of familial CPP cases in 22% of cases when we considered precocious menarche as the age of menarche less than or equal to nine years. Of note, if we considered the age of menarche less than or equal to 10 years in those families, the prevalence of familial CPP was around 27%. The similar prevalence as Devry and collaborators found using the same criteria of 10 years. To characterize the families besides the Brazilian and Spanish families, we included 128 other families from distinctive countries who have been referenced to our laboratory for genetic analysis, compressing a total of 213 families. Maternal and paternal modes of transmission were the most prevalent, and we can see that they have similar rates. Looking to the number of affected generations in familial CPP, we may see that we found in paternal transmission, families had one or two generations affected, and that in maternal transmission, most families had at least two generations known to be affected. In both transmissions, maternal and paternal, the affected family members had a large female predominance. Maternal and laboratory features of problems with familial CPP were quite similar among maternal transmission families, paternal transmission families, and families with undetermined transmission. Genetic diagnoses of familial CPP have been identified in this multi-ethnic cohort of 213 families. They were previously published in studies from our group, and we pulled the results here together, and we found that loss-of-function mutations in MKRN3 were identified in 71 cases from 36 families, and loss-of-function mutations in DLK1 were identified in 12 cases from 6 families. So we could calculate the prevalence of MKRN3 and DLK1 mutations in familial CPP, and among all families, MKRN3 loss-of-function mutations are the cause of CPP in 21% of cases, and DLK1 loss-of-function mutations were the cause in 3%. If we will look just to the families with paternal transmission, MKRN3 mutations are the cause in 55% of families, and DLK1 mutations in 9% of families. In conclusion, the prevalence of familial CPP was 22%, and paternal and maternal transmissions had similar rates. A large female predominance was demonstrated among affected individuals of families with CPP. MKRN3 loss-of-function mutations represented the most common cause among familial CPP cases, 55% in families with paternal transmission. And the family history of pubertal development is essential, and it should be actively investigated for the precise etiological diagnosis of CPP. I'd like to thank to all patients, relatives, our colleagues from Sao Paulo University. A special thanks to our collaborators, especially to Professor Jesus Argenti from Argentina, who is a very nice collaborator. A very special thanks to my mentor, Professor Ana Cláudia and to our funding grants. Thank you. Open for questions. I'll start with one for Dr. Canton. So have you looked at treatment for these families that you see and response to treatment and any differences between the mutational status and or the transmission? Yes, they have the same response to treatment. They respond very well to GnRH analogues. Okay. Gordon Cutler, beautiful talk. Does the genetics depend at all on the age at which, in other words, there's a two or three year old or one year old with leprosy? Do you really have the same pattern of genetic prevalence as somebody who's 5 or 6? If I understood correctly what you were asking, we know that for MKR in 3, the median, the mean age of puberty onset for girls is 6.2 years and for boys 7.8. This is where our analysis. For DLK1, we have such a small number of cases, but it seems to be a little bit earlier, like 5.5 years in the cases we have. I only mentioned this that the first patient we treated began menstruating at, this is in 1970s with GnRH, at six months of age. And we saw a lot of the very young people originally, very different from what you see in practice. I'm just wondering if the two-year-olds, three-year-olds, and so on are genetically the same or whether they might have something different. Okay. One more question it looks like. Yeah, is this something that we should, I recently saw a patient that we suspected this was the case, started on GnRH agonist treatment. We presumed, you know, other than that perhaps idiopathic central precocious puberty, should we be referring these patients for genetic workup? And are there long-term sequelae we should be thinking about other than central precocious puberty that might change our management? Or would we treat with GnRH agonist otherwise? When should people be referred for genetic counseling and evaluation? And are there any long-term consequences? Okay, very nice question. So to refer, we believe that for when you have a familial case, it would be referred for genetic analysis. This is, and also when you have some syndromic features, also you should reference for analysis to study other kinds of syndromic central precocious puberty. But in the familial case, of course you should analysis for MKRN3 and DLK1, which were there. And this case are non-syndromic, are just like idiopathic case. Looking to the long-term outcomes, what we know for the DLK1 females is that they should, most of them have problems with metabolism later in life, such as overweight, obesity, a predominance of metabolic problems, health issues, dyslipidemia, and insulin resistance. So this should be monitored. For MKRN3, I would say that we didn't see any kind of different, distinct from other idiopathic CPP girls in long-term outcomes. Thank you for your presentation. Have you looked at family members who may not have been affected, who may also have these mutations to determine the penetrance? Do all the family members who have a mutation have? Not all, because as I showed you, if the mutation comes from the mother, it's in the maternal allele, the person, the individual, will not express. But is it guaranteed that if it comes from the father and it's in a girl, that it will be, that there will certainly have central precocious puberty? Certainly, certainly it will have. As far as we know, all cases... Have you looked at any sisters to see if that's... Yes, yes, we have some case, yes. Okay, thank you Dr. Cantan. We'll move on. Switching gears to CAH, and Dr. Jay Whitehead will present from Laurie Children's on 17-beta-hydroxysteroid dehydrogenase. All right, so my name is Jay Whitehead. I use they, them pronouns, and I'm coming from Laurie Children's Hospital. Thank you to the organizers for the opportunity to present our data today. So this work was done with the collaboration of the DSD Translational Research Network, a network of 15 academic hospitals, collaborating to improve the health, the health of people living with DSD. So we'll start with some background. So 17-beta-hydroxysteroid dehydrogenase type 3 deficiency, which we will, for obvious reasons, abbreviate as 17-beta, is a 46 XY DSD, resulting in testicular development and typical production of anti-malarian hormone. This leads to the regression of the uterus and cervix. There's no upper portion of the vagina present. On the other side of the slide, you'll see steroidogenesis proceeds as usual up until the action of the 17-beta enzyme, which typically converts androsinodione to testosterone. Due to the buildup of androsinodione above this blockade, the typical ratio of testosterone to androsinodione in this condition is low, less than 0.8. This leads to a phenotype of external genitalia at birth that can range from typical for a female infant to non-binary or atypical in appearance. At puberty, though, virilization occurs due to potentially multiple mechanisms, including other isoenzymes of 17-beta and the backdoor pathway of steroidogenesis. So for some additional background, 17-beta was first described in the late 1960s, but so far less than 20 cases have been published in the United States in particular. So it's thought to be relatively low prevalence in this country, but we hypothesize that that may not truly be the case. This condition can also present similarly to other 46-XY DSD, such as 5-alpha reductase deficiency or complete androgen insensitivity. So this chart review was done retrospectively, again with the collaboration of the DSD-TRN. We collected 16 patients. You'll see on the right-hand side of the slide the specific children's hospitals and the number of patients that each contributed. A few of the patients have been reported previously at endosociety meetings and abstracts, and this in total includes 10 patients who have not previously been reported. So this pie chart shows the age of initial workup for DSD, not specifically the DSD diagnosis, but the initial workup for any form of DSD and the chief complaint at that time. So roughly half the patients in this cohort presented or were worked up prenatally or shortly after birth, typically due to discordance between the karyotype, either from cell-free DNA testing and then the phenotype that was detected after birth or on prenatal ultrasound, or due to non-binary genitalia noted at birth. The three patients in yellow at the bottom were detected in early childhood due to testes that were identified at the time of inguinal hernia repair, and the remainder in green and purple were detected during puberty, either due to virilization and or primary amenorrhea. So this shows a general timeline of diagnosis and treatment. So each patient is represented by a bar here, purple if raised female and green if raised male. The timing of diagnosis of 17 beta is represented by the blue dot, and you can see the current age at the most recent follow-up on the x-axis in years. You'll notice that about half the patients on top are currently pre-pubertal, and the patients that are currently older at the time of the most recent follow-up, you'll notice that the diagnosis tended to be older, made at older ages, and typically around the time that management changed in terms of proceeding with gonadectomy or blockers and estrogen treatment, if desired by the patient. So physical exam findings in this condition are variable, as I alluded to earlier, and it also depends on the timing of the diagnosis in the exam. So in this sample, 13 patients were assigned female at birth, and in those, 11 of the 13 had cloromegaly at the time of the initial diagnosis by the DSD study team. The two that did not were examined in infancy, so something that can develop over time. And then at least 7 of the 10 had at least one palpable gonad at the time of the DSD team's exam in our records that are available, and the other six patients had had some surgical relocation of gonads, whether that's gonadectomy or inguinal hernia repair, and so we have some somewhat incomplete data on some of these points. This chart shows the hormonal testing that is available for these patients. In particular, in bold, the ratio of testosterone to androsinodione. So again, typically the ratio that's suspicious for the diagnosis is less than 0.8, and in these patients you'll see that most of them had ratios that were significantly less than 0.8. There are some caveats to hormonal testing, though. So for example, case number five would not have met criteria based on the hormonal data that's available, at least at this age. They were tested around one month of age in the mini-puberty, but they did have a genetic diagnosis confirmed. We know that patients commonly carry either incorrect diagnoses or less specific diagnoses. So three patients in this series, this slide in particular lists only the 12 that have genetically confirmed 17 beta, as opposed to hormonally diagnosed. Of these 12, three had a prior diagnosis of complete AIS. Some of them, such as case number two, carried this diagnosis for many years prior to additional testing. Three had a prior diagnosis of less specific 46 XY DSD of unknown etiology, again like case nine, from birth until age eight years when further workup was done. Of the four patients that are not listed on this slide that have not had genetically confirmed 17 beta, they have a hormonal diagnosis. One carried a diagnosis of partial androgen insensitivity from birth until age 16 when more hormonal testing was done. Two were in their teenage years when they first came to medical attention and they declined further genetic testing, but had hormone levels consistent with the diagnosis. And the fourth had a sibling that was genetically diagnosed and so did not have their own testing completed. There are some cases in which management may change significantly based on this updated diagnosis. Just as one example, case number 10 in this series was assigned female at birth and had a presumed diagnosis of complete androgen insensitivity based on testes that were identified in inguinal hernia repair. However, around the time of puberty, at age 10 or 11, had unexpected masculinization that was undesired by the patient and so blocker treatment was initiated along with further diagnostics followed by estrogen replacement. So we know that the topic of gonadal management in DSD is complex. You're balancing risk of malignancy with considerations about future fertility, hormonal production, bodily autonomy. In this particular condition, germ cells can be found on pathology. Eight out of the 16 in this case series have so far had gonadectomy at the ages that are listed here in purple. None of them were found to have gonadal neoplasm and the best available guidelines at this point place it 17 beta in the intermediate risk category, but it's based on a relatively small number of cases of gonadal neoplasm. Five out of eight of the patients in this series did have immature germ cells that were found on pathology and seemingly more common in the younger age group at the time of gonadectomy. This is a representative from one of the patients that had gonadectomy at age 19 on pathology. They were found to have latex cell hyperplasia which tends to develop over time in patients with 17 beta and then on the right an atrophic seminiferous tubule with absence bromatogenesis, another relatively common finding. So in conclusion, diagnostic accuracy in DSD in general is key. It determines a lot of anticipatory guidance that we give to the families and patients, a lot of the prognosis in terms of risk of other organ system involvement, chance of fertility, gonadal malignancy risk, etc. We advocate for including 17 beta deficiency in your differential for 46 XY DSD. If hormonal testing, particularly HCG simulation test is pursued, to include androstenedione as one of the standard measurements there. Although we know that there are some caveats to hormonal diagnosis and so I think there's a very good argument to be made for proceeding straight to genetic testing, especially in an age when genetic testing is more available in many locations. And in the case in which HCG stimulation tests may produce or cause more virilization that may not be desired by the patient. Of course we need more data on long-term psychosocial outcomes, adult gender identity, and many other topics within this field. One of my colleagues, Dr. Jackie Papadakis, is leading a team that's working on analyzing more of the psychosocial data available for the patients in this cohort as well. So thank you so much for your attention. I'm happy to take any questions. Open to questions in the center. Center mic, if anyone has any. The one question I had to start with the answer in the last slide as far as biochem testing and genetics. But what about the topic of gonadectomy because the data that you showed was not entirely supportive. Is there a better way to think about screening rather than pursuing just based on risk? In terms of gonadal malignancy risk in particular? Correct. I think that in general in the in the world of DSD we have a lot to learn there in terms of if there's a better way that we can detect or predict the patients more at higher risk of gonadal malignancy. There may be newer diagnostics on the way I hope. In 17 beta in particular I think there's not there's not a lot of data on on true risk. I think there's also the consideration of the if the gonadal location helps influence the risk over time and so if the gonads are intra-abdominal versus if they're descended or palpable in terms of screening as well will influence risk over time and detection. A beautiful talk. I just this is a sort of a naive thing and maybe I didn't hear everything but would you assume then that given all the misdiagnoses that maybe this is underappreciated and because I always thought of it as extremely rare. I think we saw one or two cases maybe in the whole career. Do you think that a lot of these TFAMs and so forth might might be might actually just have very severe 17 blockage? Yeah I do wonder if if in general the diagnosis is underappreciated and I think it's interesting in this case series because I think we're in an age when the availability and cost availability is going up of genetic testing and cost is going down at the same time and so you can see the patients that were diagnosed at an older age more recently and those that were many more that were diagnosed very very young because of that availability of genetic testing. So otherwise they may not have been detected until virilization occurred at puberty or for other reasons and so I think we're in this we're in the sort of middle ground where we're starting to see more genetic diagnosis and probably a lot of patients that we may have presumed to have things like complete androgen insensitivity may in fact have actually had this diagnosis all along and we may never know because they may have had gonadectomy to not produce the the hormonal ratios that that would lead to suspicion for this diagnosis so. Okay just checking online. Okay so for that I think we'll finish and thank you again for a really beautiful talk. So our next speaker is Kiki Serafoglu from University of Minnesota and looking at 21-hydroxylase in a novel treatment. So we look forward to hearing this co-authored with 11 other institutes. Oh Ron is presenting. Ron Newfield from San Diego. So dear colleagues, ladies and gentlemen, good morning. Great to see so many of you in person. I'd like to present this pediatric trial and I'm thankful for the Endocrine Society for allowing us to present this data and I also like to thank all the dedicated investigators and research staff that made that happen. As you can see from my disclosures I've had a longtime interest in congenital adrenal hyperplasia and 21-hydroxylase congenital adrenal hyperplasia can result in impaired synthesis of cortisol and often aldosterone manifesting as salt wasting and also excess adrenal androgen production. For those of us who are treating CAH we know that treatment must balance the consequences of super physiologic corticodoses and the consequences of high ACTH and androgen excess as can be shown in that schema here. On the one hand chronic super physiologic glucocorticoid dosing can result in short stature in pediatrics, obesity, diabetes, increased cardiovascular risk, hypertension and osteoporosis and on the other hand under treatment can result in high ACTH and androgen excess that again pediatrics can lead to short stature, abnormal or early puberty as well as hirsutism in females, infertility, TART in male and also psychological effects. This leads us to Cronosophant which is an orally administered non-steroidal selective CRF1 receptor antagonist and CRF1 receptor antagonism in 21-hydroxylase CAH could inhibit ACTH release as shown in this schema and therefore reduce the androgen excess production from the adrenal glands. On the right hand side you can see that in a phase 2 study of Cronosophant adults with classic 21-hydroxylase efficiency experienced clinically meaningful reductions in ACTH, 17-hydroxyprogesterone, androstenedione and in females also of testosterone levels most of it in the 60 plus percent range and this was on a Cronosophant dose of 100 milligrams PID based on the average morning window values. So this leads us to a very similar study design in adolescents was Cronosophant in patients with classic 21-hydroxylase deficiency. As you can see the study design is very similar to the adults was a screening period followed by a baseline pharmacodynamic profile seven days before the first dose has been given followed by 14 days of Cronosophant and then a follow-up period. The key eligibility criteria were 14 to 17 years of age having classic 21-hydroxylase deficiency on a stable GC dose prior to the study entry and remaining on their home regimen including the fludrocortisone if they were on that. They all had to have a screening value of 17-hydroxyprogesterone greater than 800 nanograms per deciliter, cortisol less than 5 microgram per deciliter and ACTH of 20 micrograms per ml or more prior to the morning GC dose. The treatment was Cronosophant 50 milligram twice daily taken orally in the morning and in the evening was meals for 14 days. It's important to note that all of the participants remain on their home GC dose and through the corazon if they were on that. The hormone sample collection consisted of a 24-hour serial sampling at baseline, seven days before first dose of the drug, and then on day 14 for ACTH-17-hydroxyprogesterone, interesting down in testosterone, and the primary PD assessment was based on the morning window averaging the sample collected at 7 a.m. and 10 a.m. This slide illustrates the study population. We had eight participants, five of them were female, majority were Caucasian, the age was between 14 and 16. The height, weight, BMI, and BMI-Z score are as shown, and you can appreciate the weight median was 62 kilograms with quite a wide variability, and the same thing, the median BMI was quite above average at 1.2, was again, quite a big range. Only had one patient with a poorly defined adrenal crisis close to two years prior to study entry. Four out of the five females already reached menarche, and they all had regular cycle intervals, except for one patient that had up to 56 days in between cycles, but that was because she was less than a year since menarche. On the right-hand side of the table, you can see that six out of the eight were on a hydrocortisone-only regimen, two were on prednisone-only, and the GC dose and hydrocortisone equivalence represented as a medium was a range, min to max, was quite super physiological at 16.2. Following that, you can see that the androgens, ACTH, and the precursors at baseline, represented as a median and IQR, are quite elevated, but that was in part a study design that samples were drawn before their morning GC dose. This slide shows the two PD profile at baseline, represented in gray, and then on day 14, in what I'm told is a cyan color. The morning window is a peach colored, and you can see quite nicely meaningful reductions in ACTH, 17-hydroxyprogesterone, and androstenedione, especially during the morning window, and you can see that for both ACTH, 17-hydroxyprogesterone, and androstenedione, and you can appreciate also that the reduction was not only limited when you look at the androstenedione, not only to the morning window. This slide illustrates that the majority had a 50% or greater median reductions in dose hormones, including the androstenedione to testosterone ratio in males after the 14 days, and the darker color represents the average 24-hour change in PD, and the lighter color represents the change in it based on the morning window alone. This slide is a little bit busier, so I'd like to kind of guide you through that. On the bottom, you can see the numbers of the individual eight patients. Circled around are those that are showing response. Above each patient, the P represents those that were on prednisone, H represents those that were on hydrocortisone, and as you can see, the majority of them had a really good response, judged as 50% or greater reduction. Not everybody was a responder, so if you look, for example, on the left-hand side on patient number one, that did not seem to respond as well, that may have been due to the fact that that patient had the lowest 17-hydroxyprogesterone at baseline, well below 1,000, so did not have much room to start with to improve. Patient number six, for example, a male that did not seem to respond very well had a BMI of 38. This table illustrates the number of patients that actually achieved a 50% or greater reduction from baseline in those hormones. Five out of eight achieved that for ACTH, six out of eight for the 17-hydroxyprogesterone, four out of eight for the androstenedione, and three out of the five females achieved that for testosterone. In addition, two out of three of the male participants achieved a response for the androstenedione to testosterone ratio delta 4 to T, defined as 0.5 or greater at baseline and below 0.5 at day 14. Again, based on the average morning window values. Moving on to safety, chronesophon was generally well-tolerated with no treatment, emergent adverse events that were of any significance. They were mild and no safety concerns were based on the routine laboratory tests, vital signs, EKGs, or neuropsychiatric assessments. If you look at the specific AEs that were reported, one participant had a headache that was deemed possibly related to study medication simply because that patient had the headaches that were mild while on the study medication and they resolved once off the medication. One patient had both vomiting, fever, myalgias, and a headache during an acute viral illness that happened after 14 days of the drug, so it did not affect the hormonal profiles and that patient was stress-dosed. We had one patient with dizziness on day four that was very short-lived and resolved on its own without changing the GC dose or the study medication. So in conclusion, in this pediatric study, it was adolescents with classic 21-hydroxylase deficiency. We were able to show substantial and clinically meaningful median reduction between 57 and 76% in the morning levels of adrenal androgens and androgen precursors that were observed after 14 days of chronisophon treatment. These data were consistent with results from a prior study of chronisophons in adults with classic 21-hydroxylase. It is of note that the good results that we have observed in this study were achieved on 50 milligrams twice daily compared to the adult doses of 100 milligrams twice daily. And if you remember that initial slide about the average weight of these adolescents was very close to adult, so we achieved pretty good results on a lower dose. So all this leads us to conclude that further studies are really warranted to evaluate the potential of longer-term chronisophon therapy, and one such study is currently underway. This should hopefully allow us to see if being on chronisophon can afford sustained reduction in all adrenal-derived androgens, allow for lower, more physiological glucocorticoid dosing, something that we've all been waiting for, and above all, see if we can improve clinical outcomes in weight, stature, decreased metabolic risks, fertility, and so forth. Thank you for your attention. Thank you. Beautiful work. It looks like we have a question. Andrew? I'm Andrew Dauber from Children's National. So I know it's a little hard to predict the future, but if this medication proves to be efficacious and is approved, right now, you showed that it's efficacious in the patients who are not well-controlled or are requiring highest doses. Do you think this will change our approach in that for patients who are well-controlled, will we start with this and go to a physiologic dose of hydrocortisone with an extra medicine, or will we stay with our standard care and only use this for patients who are out of control? Where do you think we're gonna go in the field? Like you said, I think it's hard to predict the future, and that's why it's important to do those studies. But I believe that it will allow us to reduce glucocortisone doses. And when we say reduce, it doesn't necessarily mean to go from 10 to seven and a half. It may allow us to have more patients be able to do well on just BID dosing, for example, of hydrocortisone. So I think it's gonna be, it'll allow us much better quality of life for patients. And I do think it'll have a role also in those that are controlled on somewhat lower doses. From the adult data, are there any data looking at the stability of dose and also what occurs during stressful times, you know, stress dosing? So again, it's important to note that this is not a substitute. So you cannot have patients come off steroids, and then when they do have a stressful illness, they all still need to have that stress dosing. So that's very important to note. Gordon Cutler. I was curious about the lower dose in these adolescents. Are there sort of PK levels that sort of are the blood levels measurable for conussant font? Are there, is it possible that you could also explore the dose range more in the children? I assume this is done just for precautionary reasons in the children? Yeah, I mean, like I said, the PK study, I was not made, I was not privileged enough to see that. But I'm told that even the heavier subjects actually achieve pretty good drug levels. And so from what we're seeing from the PK studies, we're definitely getting two levels that should block the CRF receptor. Okay, so beautiful work, nice step forward. Thank you, Ron. Our next speaker is Dr. Andrew Dauber from Children's National, talking about treatment of non-achondroplasia genetic short stature. Awesome. All right, well, thank you very much. Thank you for the opportunity to present our work today. I'll be talking about fesorotide for selected genetic causes of short stature. So these are my disclosures. This study is funded by Biomarin, but it is an investigator-initiated study, and Biomarin has not played any role in the design, conduct, or analysis of the data, or preparing the presentation. So I'd like to start by showing you a picture of a histological section of the growth plate, pictured here on the left with its various zones, showing the chondrocytes lining up into their columns, hypertrophying, and ultimately leading to increased endochondrial ossification and bone growth. And if you zoom in on a single chondrocyte on the right, you see the FGFR3 receptor, which acts as a break to growth when activated, goes down if you follow that pathway over the right through that RAS, RAF, MEK, MAP kinase pathway, inhibiting chondrocyte hypertrophy. And on the upper right, you see CNP, or C-type natriuretic peptide, which binds to its receptor encoded by the NPR2 gene, and that opposes this pathway and kind of lifts up that break, thereby promoting growth. And visorotide is an analog of C-type natriuretic peptide, or CNP. And in a randomized, double-blind, placebo-controlled phase three study in children with achondroplasia, visorotide was shown to increase growth velocity over the first year by approximately 1.6 centimeters per year compared to placebo, and this subsequently recently led to the approval, both in the US and Europe, of visorotide for achondroplasia. And I and others have posited that this same medication and class of medications could be beneficial for individuals with other selected genetic causes of short stature, and our current study inclusion criteria include six different genetic causes of short stature. The first is hypochondroplasia, which are just milder activating mutations of the FJFR3 receptor, it's pretty obvious, the same hypothesis with achondroplasia. The next is individuals with CNP deficiency. If you're missing CNP, can we replace it? Now, there have only been around like five patients in the world described with that. We don't have any in the study, but if we find one, I would love to give them some and see what happens. But the next are heterozygous mutations in the receptor, in the NPR2 gene, and these are actually quite common. It's been estimated that approximately one in 50 individuals with idiopathic short stature have heterozygous mutations in the NPR2 gene, that's pretty impressive. And the thought here is that as long as you have one wild type receptor, that if there's a decrement in signaling here, if we can augment that back closer to normal with a CNP analog, it could rescue the short stature. The next is the class of Rhazopathies. So the one we're most familiar with is Noonan syndrome, but the Rhazopathies are also due to have their, at least major growth effect due to over activation of this RAS, RAF, MEK, MAP kinase pathway. And can we now directly inhibit that with a CNP analog? Shox gene mutations, as Shox is involved in affecting FGFR3 expression. And finally, agrican deficiency, one of my personal favorites. And agrican encodes a proteoglycan in the extracellular matrix of the chondrocytes, sorry, of the growth plate around the chondrocytes. And there's some animal data that shows that there's increased signaling phospho-ERK levels, which is also influenced by this pathway. So the inclusion criteria for our study are children between the ages of three and 11 for males, 10 for females who are prepubertal and have heights below minus 2.25 standard deviations. They must have a mutation documented in one of those six categories. They cannot be concurrently treated with growth hormone, but briar treatment is okay. There is, the design of the study has a six month run in observation period so that we can get a baseline growth velocity as this is quite a heterogeneous cohort. And then we start everyone on vasorotide at a dose of 15 micrograms per kilogram per day as a daily injection subcutaneous therapy, which is the same dose used in the phase three study for achondroplasia. We then follow patients for one year. The primary study endpoints are first safety, looking at the incidence of adverse events, and then secondary looking at efficacy with changes in growth velocity at 12 months and changes in height standard deviation score. We're also looking at body proportions, bone age ratios, and exploratory endpoints include pharmacokinetics, pharmacodynamic markers, bone density, and quality of life surveys. So here are our baseline demographics. The target enrollment for the study is 35 subjects. We currently have 34. Number 35 is scheduled later this week. So hopefully we will be close to enrollment soon. And we have 21 males and 13 females in the study. The majority, as you can see, have hypochondroplasia, 26 of them, three with NUNIN, three with NPR2 mutations, and currently two with AGRICAN, the last hopefully scheduled patient has an AGRICAN gene mutation as well. This is the age distribution. So as you can see, we go all across the age distribution with the majority of the patients being six or younger, but patients all the way up to somebody the week before their 11th birthday. And then on the right is the height standard deviation score. So these bars represent a height of that standard deviation or lower. So we have one patient below minus five standard deviations, four between minus four and minus five, 16 between minus three and minus four, and then the remainder between minus 2.25 and minus three. So here is the six-month growth velocity data for the initial 12 subjects in the study. The study's obviously progressing along. So on the left are the individuals just with hypochondroplasia. The eight patients pictured here, and you can see there is some variability in the response, but there's a mean increase in growth velocity of 1.8 centimeters per year, but with some wide variability, but similar to what was seen, perhaps slightly better than achondroplasia. But really remarkable on the right, and I just combined NUNIN and NPR2 just for the sake of the slide, but they have nothing to do with each other. The two individuals in blue are the subjects with NUNIN syndrome who are growing initially at growth velocities of around nine and close to 12 centimeters per year at six months, and then two patients with NPR2 mutations who also have had really, I would say, remarkable increases in growth velocity. One of the subjects was close to 11 when he started, really was not growing at all for the first six months, and went up to a growth velocity of just under 10 centimeters per year. So the mean increase for that group, for whatever it's worth, is 6.1 centimeters per year, but I wouldn't make anything of those stats. So hot off the presses, these are our 12-month data of our first nine subjects. So in that group, you see the one patient with the largest blue line from the bottom to the top is that same NPR2 patient who's still at the end of 12 months is growing around 10 centimeters per year. The other two patients with the largest increase are those with, one of those is an NPR2 patient and one with Nuenen, and then clustered in the middle are the hypochondroplasia patients. So for the six hypochondroplasia patients at 12 months, we're seeing a mean increase now of 1.9 centimeters per year with less variability as well. So encouraging data. This is the height standard deviation data. So I've plotted here, as you see in the baseline observation period, the height standard deviation does not improve, which is not surprising. It doesn't get that much worse either. But then after treatment, all of the subjects have an increase in their height standard deviation score with, again, some variability. In the hypochondroplasia, very small number, still six subjects. There's a mean increase of 0.3 standard deviations. This is just data from 12 subjects, so very preliminary, two with Nuenen, two with NPR2, and eight with hypochondroplasia, looking at the generation of urine CGMP, which is a pharmacodynamic biomarker of CMP action. And what I think is very interesting is the difference in the curves. So the top curve are the two patients with Nuenen syndrome who had very large variability. One of the patients had extremely high CGMP generation. The hypochondroplasia is in the blue in the middle, showing pretty tight results in what was similar to what was seen previously. But the two patients with NPR2 mutations had actually very low levels of CGMP generation, despite actually their very good growth velocities. But interesting to think about that a little bit more mechanistically. So the pharmacokinetic parameters were very similar to what's been previously published for visorotide in children with achondroplasia, and we're continuing to gather that data and we'll present more details when we have a larger cohort. The CMAX and AUC did correlate strongly with the peak increase in the CGMP, but these PK parameters were not significantly correlated with the change in growth velocity. We are going to be analyzing another pharmacodynamic marker called collagen 10 marker, or CXM, and we have not yet, we're batching the samples, so have not yet received any results on that. In terms of safety, it's overall very well tolerated. We had injection site reactions in around 44% of the subjects, very common. They were all mild and self-resolved without intervention. No episodes of symptomatic hypotension. We did have one subject who had an episode of syncope and two episodes of presyncope, one of which was during the observation period after a blood draw. There was one serious adverse event of viral-induced ITP, which we've determined was unrelated to the study drug, and the patient was subsequently restarted and has been doing great with no subsequent platelet issues. And one subject who had baseline scoliosis, which did worsen with significant growth with an NPR2 mutation. So in conclusion, the Soratide appears to be well tolerated with similar safety profiles to previous reports. Our preliminary results suggest a positive response in all subgroups with inter-individual variability and a robust initial response in patients with NPR2 mutations in Noonan syndrome, and of course, we are awaiting data on additional subjects. For the sake of time, I will skip that last slide, and I am happy to answer any questions. Thank you for your questions. This is great work, and congratulations. My question is regarding the Noonan syndrome patients, the two of them. Could you dissect a little bit in the type of mutation that they had, and none of these patients entered puberty while they were treated? Great questions. Okay, so all three Noonan's patients have BTPN11 mutations, so that's all I can comment on in, of course, very small numbers. One of the oldest subjects, one of the ones who did have very big growth velocity, they had to be still at the initiation of day one, pre-pubertal for the study. His testes were 4 cc's at 6 months or 12 months, so at his most recent visits. So he did enter puberty, but at 4 cc testes, I do not think that can explain his tremendous increase in growth velocity. Jeff Barron, NIH, so yeah, beautiful study, very exciting, Andrew. It looked like the urinary cyclic GMP peaked at about 2 hours, which you said was similar to the data in achondroplasia. Does that suggest to you that the duration of action really is not ideal, and that to get a greater benefit in terms of growth velocity, that we need a longer acting form, CMP form? I think that's a great question. So, Vasorotide has a longer half-life than native CMP, which is like minutes, right? But it still only lasts around for a few hours, right? So I do think that it is theoretically possible and worth exploring whether a longer acting form of CMP, which are in development by multiple companies, could have even better growth effects. So I would love to see that happen. Okay, your next study, great. I don't know. Great talk. I'm just going to return your question about predicting the future. So in Noonan syndrome, we all know there is increased risk of malignancies, especially in the mutation type that you have for your patients. And so when we're giving him growth, when we're addressing the height from the very one focus strategy versus in this strategy, which actually blocks the pathway. So would you speculate in terms of future safety and cancer risks if there is? Yeah, so I mean, I would just echo what you said. I don't want to be alarmist about growth hormone. I think in general, the data about growth hormone are very reassuring. But there are at least theoretical risks. And in certain populations that are predisposed to cancer, I am hesitant to use growth hormone for those risks, although I do prescribe it in Noonan syndrome. I do think that a CMP-based therapy should theoretically be safer. But we have a total of zero long-term data on CMP therapy. So I think we just have to acknowledge that. Now, that being said, there are patients, human patients, who have activating mutations in the NPR2 gene. There are human patients who have overexpression of CMP lifelong. And to my knowledge, in none of those papers have there been any reports of malignancy. So that at least is somewhat reassuring to me. But again, the total number of patients we're talking about in the world with those types of mutations, like 10. So yes, I think this could be a theoretical benefit. And it's definitely one that has come up in discussions with families and with support groups, with the Rhizopathies Network and stuff, about why this could potentially be beneficial. But until right now, we didn't know if it was going to work. And we still have very preliminary data. So I think it's something that needs to be investigated more in the future. Thank you. Hi. Camelia Kamoon, a fellow at the Children's Hospital of Philadelphia. Super interesting work. I was curious if you could comment on the distribution between boys and girls in the study. It looked like, if I'm not mistaken, there were two-thirds of the individuals in the study were boys. Is that related to who was interested in participating or just kind of chance? And as a follow-up to that question, do you have any plans moving forward to try to make sure that there is maybe a more even split so that any sex differences in response to treatment or pharmacokinetics could be best understood? Yeah. That's a good question. And first of all, let me congratulate you on your work on looking at these disparities in growth hormone treatment. So this is the numbers, 21 and 13. I actually think for a growth study of phase two to have gotten 13 girls into the study is really awesome. And these are rare diseases, right? They had to have documents and mutations. Anybody who approached who was eligible, we took into the studies. We have patients flying in from all over the country and, frankly, patients who have relocated from other countries in the world to be part of the study. So I think it was just what interest and what people pursued. For this study, we're going to be not close to closing enrollment. We'll see if we open other cohorts in the future. But I am hopeful that for rare diseases, there will be less of a sex bias. But I don't know. Thank you. And it looks like we have one last question. Yeah, one last question. Hi. Great talk. I'm very excited to see possible other indications for CMP. I had a quick question about some of the adverse events. I noticed that there was about two or three cases of presyncope. And given that CMP has that possible for hypertension, but there were no cases of hypertension, any idea about what may have caused those presyncope? Sure. So basically what happens is there's one patient who had a presyncopal event, actually the oldest patient in the study, when he had a blood draw during the observation period. At his first blood draw, he just was about to pass out. So that, I think, had absolutely nothing to do with medication. The other two events, the syncope and presyncope, were in the same patient. And that same patient, this little girl with hypochondriplasia, both of them occurred in the clinic visits after we had put in her IV. But she had gotten the medication, and I think could be medication-related. And she was running around like a little, you know, all over the place. She couldn't sit still. And in the six months in between, when she was on the medication at home, she never had any symptoms or events. So I honestly don't know. We checked her blood pressure. She never had hypotension during those events. She wasn't tachycardic. I'm not really sure what happened. But they were both pretty shortly after she had gotten her injection. She had also been getting PK samples and was running around. I don't know. You know, I don't know. But I can't definitively link it to any hypotension, because we never documented hypotension. So that's why there's that discrepancy. Okay. Thanks a lot. That's what happened. Appreciate it. All right. So that's it. Thank you. So our last talk, Alex Heron from University of Cincinnati. We'll talk about Madelung deformity and Turner syndrome. Thank you very much. I'd like to extend a thank you to ENDO 2022 for allowing us to present our work here today. And also would like to extend a thank you to my collaborators, Dr. Kevin Little, Dr. Felipe Black-Lajal, and Dr. Ira Scott-Mark Little, who could not be here with us today, but without whom this work would not have been possible. I have nothing to disclose. So our work was centered around a subset of patients with Turner syndrome, and as I'm sure many are aware, the etiology of this syndrome involves the partial or complete loss of the second X chromosome in biological females via a variety of methods, meiotic non-disjunction, or the formation of an isochromosome, to name a few. This loss results in many different medical and developmental disorders that present in several key organ systems, including cardiac malformations, such as coarctation of the aorta or a bicuspid aortic valve, and skeletal deformities such as short stature or a broad shield chest. A specific skeletal dysplasia that appears in these patients is made lung deformity, and that's the focus of our work here today. This deformity is characterized by a growth deficiency in the distal radial physis that causes excessive radial inclination and volar tilt, with result in ulnar carpal impaction. And so if we look at the bone age here on the left, we note that the radius, due to a lack of growth, slopes, and that sloping causes the ulna to compact into the carpal bones, which is characteristic of the disorder. This disorder is, or deformity rather, is perhaps most well-known for its association with Larry Weal dyschondriosis, which involves mutation of the Sjock's gene, as the previous speaker had detailed. This is located on the tip of the isochromosome and contributes to the growth and maturation of the skeleton, in particular, the long bones of the arm and the leg. So it's thought that the loss, the absence of the second copy of the Sjock's gene contributes to the formation of Madelung in Turner syndrome patients. Importantly, in patients with Madelung deformity, surgical intervention can halt and reverse it, but it's most effective if it's implemented in the early stages of the deformity. And so despite a considerable body of literature that highlights the presence of Madelung deformity in Turner syndrome patients, despite its relatively low prevalence, it's often underdiagnosed. And we theorize this was in part due to a lack of data documenting the epidemiology and the natural history of its progression. And so really our study sought to look at the frequency of clinical diagnosis in patients meeting radiographic measurement thresholds for Madelung deformity, and also to plot the time course of these radiographic measurements to document its progression and its development. So in terms of our methods, we began by assembling a database of 264 patients with Turner syndrome, and we excluded 77 of these patients due to a lack of relevant bone imaging. Our next step was to calculate values using the bone age for two generally accepted parameters used in the diagnosis of Madelung deformity, ulnar tilt and lunate subsidence. Using these measurements and widely accepted threshold values, seven patients out of the 187 were determined to have the deformity. And then for those patients, we conducted chart review to determine if a diagnosis of Madelung deformity was made by the radiologist, and then plotted the measurements over time to document the time course. So just to dive a little bit deeper into the measurements themselves, again the two that we used were ulnar tilt and lunate subsidence. And ulnar tilt is defined as the complement of the acute angle between the longitudinal axis of the ulna and a line tangent to the proximal surfaces of the scaphoid and lunate bones. And so if we look here on the diagram, attempt to use the laser pointer here, this is the longitudinal axis, so that is that line, and then we drew a tangent line to the scaphoid and lunate bone. That defined the angle A, and the angle that defines ulnar tilt is the one highlighted by the arrow, which is the complement of angle A. We also used lunate subsidence, and this is defined as the distance in millimeters between the most proximal part of the lunate and a line perpendicular to the longitudinal axis of the ulna and through its distal articular surface. And importantly, this value is negative, or negative in that it confirms a diagnosis of Madelung if the ulna extends distal to the proximal surface of the lunate. And I should have noticed on the previous slide, I apologize, the threshold that we had used for ulnar tilt was 33 degrees, and the threshold that we used for lunate to confirm the diagnosis of Madelung was less than zero millimeters. We used these measurements, or selected them from a series of several available parameters as they were proven to be reproducible and reliable in a 2005 study conducted by McCarroll et al that used four independent raters which performed these measurements on a series of bone radiographs, independent of one another, and then compared the values using a correlation coefficient which was determined to be quite high. So there are several other measurements that can be used to make the diagnosis of Madelung, but these two are the most reproducible. And then additionally, the thresholds that we used were chosen based on the results of another study that his team performed, which showed that above these values there was no difference in the frequency of diagnosis between evaluators asked to make a diagnosis of Madelung based on the radiograph alone, with evaluators using both clinical and history and physical information and the radiograph. So just to get some audience participation, I have two radiographs from patients in our study, and I was just curious, via a show of hands, based on those two measurements that we just discussed, who here believes that this patient does have Madelung deformity? So who says yes? Okay, who says no? Looks like the yeses have it, and I would agree. As you can see, the ulnar tilt value here is quite pronounced, it's well over 33 degrees, and we'd note that the ulna itself is, the distal surface of the ulna extends past the proximal surface of the lunate, meaning that the lunate subsidence value is less than zero. So both values here meet the threshold values for inclusion, or to make the diagnosis of Madelung. Second example, who here would say that this patient does have Madelung? How about does not have Madelung? Yeah, we agreed. So here you'll note that the value for ulnar tilt is well below 33 degrees, and the distal surface of the ulna does not extend beyond the proximal surface of the lunate, so the lunate subsidence value is above zero, and it was not included as a diagnosis here. So just to detail our results, again focusing on the two aims of our study, the first being diagnostic frequency. We found that only one out of the seven patients who met the radiographic criteria for diagnosis of MD were actually given a clinical diagnosis. And perhaps most interestingly, these six undiagnosed patients had an average ulnar tilt and lunate subsidence value of 40.5 degrees and negative 4.4 millimeters as of the date of their most recent bone age. So despite the fact that they had values well beyond the threshold criteria, they were not given a clinical diagnosis, which suggests that it's definitely underdiagnosed in this patient population. We also then plotted time course values of the measurements for both ulnar tilt and lunate subsidence, and for patients with ulnar tilt, we were able to include four. We had to exclude three due to a lack of historical data and an inability to trend that data in those patients. We noted that they crossed the average, excuse me, the threshold for ulnar tilt at an average age of 15.3 years old, and that the average maximum read-to-read growth rate of 5.3 degrees occurred at an average age of 15.3 years old. In lunate subsidence, we were able to plot the data for three of our patients. These patients crossed the threshold of zero, I should say millimeters, at an average age of 13.6 years old, and the average read-to-read growth rate of 3.5 millimeters occurred at an average age of 14.2 years old. So in terms of our overall conclusions, again, the first name in the data there suggests that this deformity is significantly under-diagnosed in patients with Turner, highlighting the need for better screening tools to help make the diagnosis. We would suggest including physical examination, looking for the characteristic wrist deformity, and a careful analysis of bone age radiographs at an earlier age with a specific focus on trends in order to better implement corrective surgery when it's more likely to succeed. We'd also recommend careful scrutiny of yearly bone age radiographs from 10 to 15 years of age to ensure that the diagnosis is not missed, and obtaining bilateral wrist radiographs, both posterior, anterior, and lateral views for verification. And the lateral views are important here. As I mentioned before, there are a number of other measurements that can be used to help confirm the diagnosis of a lung, and by getting a lateral view, that enables us to conduct those measurements as well. And then finally, future research should focus on documenting disease progression using radiographic trends, as well as assessing how onset and progression are affected by therapeutic interventions such as growth hormone, or perhaps, as our last speaker outlined, that medicine. And with that, that's all I have. So thank you for your time. Thank you. Very nice work. We have some questions in the middle. Hi, I'm Sarah. I'm a Pediatric Endocrine Fellow at Walter Reed, and I have two questions for you. One is why you guys didn't include those shocks deficiency, because you can see that in boys and girls, and with them having more endogenous puberty, you might catch more matalung. And then also, what does it matter? If you correct surgically a matalung deformity, is it just cosmetic, or are they really having range of motion issues? Yeah, those are both good questions, and I'd say to answer your first one, we just focused on Turner patients, because that was the patient population we were interested in. Dr. Gutmark-Little focuses most of her work on that patient population, so that was our focus. I think our next steps for this research are to extend it, as you suggested, to patients with a broader shocks deficiency, and then also to focus on how growth hormone influences those developmental processes. And in terms of your second question, certainly it can be a combination of both. Patient outcomes are important from both a cosmetic and functional perspective, so we'd want to treat both of those things. I don't know that it heavily weights one way or another, but certainly both of them can be important at that stage of a patient's life. Thank you. Yeah, you're welcome. Eunice from NIH, thank you very much for your very interesting talk. I think it's really important to notice these x-ray findings, given the prevalence of Turner syndrome very often, but we don't necessarily look at these findings. So my question is that, did you look at the timing of epiphyseal capping, and there was a growth plate closure on the medial side and lateral sides, because the growth plate on the lateral side is normal, but the medial side is defect, so there may be some different timing of capping and disclosure. I mean, the epiphyseal closure on both sides, yeah. I'm sorry, I wasn't able to make out the second part of your question. Could you repeat it for me? Sure, so the growth plate defect is present in the medial side of it. So the epiphyseal capping, so the epiphyseal capping occurs before the growth plate closure. Yep. So did you look at the timing, but it has different timing? Yeah, that's gonna be something that we're focused on in the next round of studies. I think for us, really, because the patient population was so small, the seven, we were kind of limited in the conclusions we were able to draw. So in working with Dr. Little and Dr. Gutmark-Little, what we would like to do next is extend it to multiple sites and bring in more of that information. So that'll be the next round for us. Gordon Cutler, beautiful talk, and I'm embarrassed to say that, of course, I've heard of an adalong deformity, but I don't think we've ever made the diagnosis that I can recall, and we followed over 100 to adult height in certain trials, so I'm pretty sure our radiologists have missed it. But just by way of education, could you say more about the functional, I'm just surprised, that's why we come to this thing, to learn things, but nobody's ever mentioned, to my knowledge, any of our patients complaining about risk problems, some of them must have had this. What exactly goes wrong, can they not type, or does it hurt, or what happens? Yeah, that's a great question, and so as part of Dr. Little's work, he's an orthopedist, and his wife is an endocrinologist, Dr. Gutmark-Little is an endocrinologist, and they do work with Turner syndrome patients, and he had explained that, he'd be better suited to talk about this, but he had explained that in the surgical corrections that he does, it was a range of motion defect. So he released the Vickers ligament, and shrunk the radius bone to give the patient more range of motion, and more functionality, yeah. All right, so one last question, you're a medical student, ortho or endo? We won't hold you to it. Endo. All right, good, whew, all right. All right, thank you again to everyone, fabulous session, and enjoy the rest of the meeting.

Chronosophant

ACTH

21-hydroxylase deficiency

adolescents

hormone levels

hydrocortisone

Madelung deformity

Turner syndrome

underdiagnosis

screening tools

early detection

surgical intervention