A–H Effects on body weight (A), body weight change (B), food intake (C), body composition (fat and lean mass change) (D), glucose (upper panel) and insulin tolerance (E), fasting glucose (F), serum insulin levels (G), and HOMA-IR index (H) in PWA female mice daily administered with vehicle (saline) or GLP1/E (5, 10, and 25 nmol/kg) during 28 days. In contrast, the metabolic profile PNA mice was unaffected after intervention with lower doses of the dual- and triple-agonists, as well as with metformin (Fig. 4A–H), except for a significant decrease in basal glucose levels in GLP1/GIP-treated mice (Fig. 4F) and reduction in daily food intake in PNA mice treated with metformin (Fig. 4C). Finally, none of the multi-agonists at lower doses, nor metformin, had a beneficial effect on ovarian cyclicity or any other gonadal parameter, such as ovarian and uterus weights and circulating LH levels (Suppl. Fig. S5D–F). In contrast, the low dose of GLP1/GIP failed to persistently improve body weight, body composition, or food intake, but significantly ameliorated glucose handling and insulin sensitivity, and reduced basal glucose levels vs. the vehicle-treated PWA group (Fig. 3A–H). Of note, since we aimed to test the effects of compounds against the most unfavorable metabolic conditions, we assessed the potential therapeutic utility of GLP1 multi-agonists in androgenized animals fed on HFD (Suppl. Fig. S1), which phenocopy a predominant subset of women suffering PCOS, who display overweight or obesity and may be exposed to unhealthy Western diets39,40. Recent, as yet fragmentary evidence suggests that intervention with GLP1-RA, e.g., liraglutide, may reduce body weight and improve hyperandrogenism and menstrual irregularities in women with obesity and PCOS28,29,30,31,32; effects probably related to its anti-diabetic and weight-lowering properties. In women with PCOS and obesity, metformin has been shown to improve insulin sensitivity, reduce insulin and androgen levels, and ameliorate menstrual cyclicity and ovulation rates18,19,20. We assessed equimolar (10 nmol/kg) and effective doses of GLP1/E, GLP1/GIP, and GLP1/GIP/Glucagon, as documented in our previous pharmacological tests; body weight, food intake, and kaolin consumption were monitored along the first 72 h of treatments in PWA mice fed on HFD. To explore potential gastrointestinal distress linked to multi-agonist interventions, as a putative bias for interpretation of their effects, we conducted kaolin tests in PWA mice. As in previous experiments in PWA mice, no effects on the gonadal profiles were found after treatments with GLP1/E, independently of the dose (Suppl. Fig. S7D–F). (2) The intervention group received GLP-1RAs, while the control group received either metformin or placebo. The search strategy focused on randomized controlled trials (RCTs) comparing treatment with a combination of GLP-1RAs and metformin or placebo. Specifically, the analysis will evaluate changes in fasting glucose levels, body mass index (BMI), body weight, waist circumference (WC), insulin sensitivity as measured by the homeostatic model assessment of IR (HOMA-IR) and other relevant hormonal and metabolic markers. For PCOS patients with obesity and infertility, a combined treatment with metformin and liraglutide has been shown to effectively reduce body weight, improved glucose metabolism and relieved IR18. A systematic search of “PubMed”, “EMBASE”, “Cochrane Library”, “Web of Science” and “Google Scholar” was conducted up to October 2024 for randomized controlled trials involving adult women with PCOS treated with GLP-1RAs compared to metformin or placebo. The studies were selected based on inclusion and exclusion criteria, and those reporting relevant anthropometric and metabolic outcomes were included in the final analysis19. By using the data from multiple studies, in this analysis we attempted to clarify the role of IR in the pathogenesis of PCOS and assess the efficacy of GLP-1RAs in improving both the reproductive and metabolic outcomes for patients with PCOS. Further studies are needed to explore their long-term effects on glucose homeostasis and lipid profiles. Secondary outcomes included glucose homeostasis (fasting glucose, fasting insulin, OGTT results and HOMA-IR), hormone levels (DHEAS, SHBG, total and free testosterone and FAI), lipid profiles (total cholesterol, HDL, LDL and triglycerides) and safety. Notably, considerable metabolic improvement was achieved at relatively low doses of GLP1/E, thus increasing the safety margin of this compound, which did not cause uterotrophic effects per se. Balance between on- and off-target effects of the dual GLP1/E agonist, and the concurrence of desensitization at very high doses, might contribute to this phenomenon. Of note, most of the pharmacological testing of these GLP1-based compounds has been implemented so far in male models of obesity33,34,35,48, with limited insight into their effects in female-specific metabolic conditions. GLP-1-mediated delivery of tesaglitazar improves obesity and glucose metabolism in male mice Triagonist-treated mice showed a slight reduction of body weight along treatment (2%) and improved glucose tolerance, although no changes were detected in the other metabolic parameters analyzed (Fig. 3A–H and Suppl. Fig. S5A–C). For reference purposes, control non-androgenized mice are included (black bars and dashed lines). In order to assess integral glucose levels in the glucose tolerance test, the area under the curve (AUC) was calculated using the trapezoidal rule. However, the triagonist caused a substantial reduction in ovarian weight and a significant drop in serum LH levels (Suppl. Fig. S3D, F). Fig. 3. In addition, in order to compare the pharmacological efficacy of the multi-agonists with the standard treatment, subgroups of animals were orally treated with a daily dose of metformin (300 mg/kg); a dose adjusted to that employed in humans by means of the application of the body surface area normalization method71, frequently used in rodent models72,73. Prior to initiation of treatments, mice were randomized and distributed into groups allowing similar mean values of body weight and fat mass. This PWA model mimics a subtype of PCOS patients that exhibit chronic hyperandrogenism, with marked metabolic, hormonal, and reproductive/gonadal alterations, which represent the majority of cases. In PWA mice, metformin therapy caused very modest metabolic effects (Fig. 1A–G), denoted only by a moderate reduction in HOMA-IR (Fig. 1H), despite the dose employed was much higher (300 mg/kg) than the doses of multi-agonists. In addition, GLP1/E and GLP1/GIP/Glucagon significantly improved insulin sensitivity, as denoted by ITT and HOMA-IR index, and decreased insulin levels; effects that were not observed in GLP1/GIP-treated animals (Fig. 1E-lower panel, G, H). Yet, animals administered with GLP1/GIP or GLP1/E displayed reduced basal glucose levels at the end of treatment (Fig. 1F). In the PWA model, with overt metabolic and gonadal perturbations, chronic intervention with GLP1/E or the triagonist markedly reduced body weight, food intake, and circulating leptin levels (Fig. 1A–C and Suppl. Fig. S3A). A similar dose of E caused a slight reduction in body weight and fat mass, as well as a significant improvement in glucose tolerance and glucose levels (Fig. 7A–G), as well as a non-significant lowering of circulating leptin (Suppl. Fig. S9A). GLP1 alone significantly reduced body weight and fat mass, and improved glucose handling, as well as serum glucose and insulin concentrations, and insulin sensitivity in this PCOS model (Fig. 7A–H), together with a decrease in circulating leptin levels (Suppl. Fig. S9A). Since the 10 nmol/kg dose of GLP1/E was at least as effective as higher doses in improving the metabolic profile of PCOS mice, we compared its beneficial effects in terms of body weight, fat mass, glucose tolerance, insulin sensitivity, and other traits vs. each single constituent of the conjugate, namely GLP1 and estrogen (E). This suggests that while GLP-1RAs may not outperform metformin in managing hormone levels, they are still beneficial in populations not already treated with metformin23,24,25,26. When examining the effects on glucose homeostasis, the results were mixed. Future studies could help clarify which specific subgroups of patients benefited the most from GLP-1RAs, whether this would be based on the baseline BMI, metabolic health or duration of therapy. Polycystic ovary syndrome For glucose homeostasis, GLP-1RAs significantly reduced fasting insulin, glucose level at 2 h after OGTT, and HOMA-IR.Meta-analysis of the total cholesterol…In this case, 5-h-fasted mice were intraperitoneally administered with 0.75 U of insulin (Sigma-Aldrich, MO, USA) per kg body weight, and blood glucose concentrations were determined before (0) and at 15, 30, 60, and 120 min after insulin injection.From the Random Forest analysis, we selected the most significant proteins based on the Mean Decrease Accuracy, which were then used for subsequent pathway analysis.In terms of fatigue, 1 study (with 20 subjects from each of the groups) were included for analysis.In this single-center retrospective study, we examined weight loss effects of GLP-1 monotherapy versus metformin in patients with PCOS–O over a six-month interval.Four, 1 and 6 studies were categorized as having “high risk”, “medium risk” and “unclear risk” of detection bias, respectively.In terms of reduction in WC, 8 studies (with 232 and 189 participants from the experimental and control groups, respectively) were included for analysis. Various studies have shown that GLP-1 receptor agonists (GLP-1 RAs) increase post-prandial insulin release and inhibit glucagon secretion, all without the risk of hypoglycemia . Because GLP-1 secretion has an incretin mechanism that is independent of plasma glucose concentration, pharmacologic analogs have been developed to improve glycemic efficiency in individuals who have impaired glucose tolerance and type 2 diabetes (T1DM) . Glucagon-like-peptide-1 (GLP-1) is a regulatory GI incretin hormone that is secreted in response to oral glucose intake and stimulates the release of insulin from pancreatic beta cells 16-18. However, the majority of women suffering from PCOS and obesity fail to significantly reduce their body weight with lifestyle intervention. Weight loss and improvement of insulin resistance and circulating insulin levels are considered essential for the clinical management of hyperandrogenic women with PCOS and obesity, as this may contribute to attenuating androgen excess and improve ovarian function5,14,15,16. Insulin has stimulatory effects on ovarian androgen production5,11, and enhances the bioavailability of testosterone by decreasing the hepatic release of sex hormone-binding globulin (SHBG)12. We report herein a series of pharmacological and molecular studies in two validated mouse models of PCOS, generated by gestational (PNA) or postweaning (PWA) androgenization, addressing the effects of GLP1-based multi-agonist therapies on metabolic and reproductive traits of PCOS. These findings are further detailed in the boxplots, where the top significant proteins identified by Random Forest analysis demonstrated distinct expression patterns between treated and control groups. Random Forest analysis was performed to identify the most important proteins contributing to a distinction between treatment and control groups. E Boxplots depicting the expression levels of the most significant proteins identified by Random Forest analysis. GLP1/E improves metabolic and gonadal traits in a PCOS model of ovarian dysfunction In both tests, circulating glucose levels were measured by using a handheld digital glucometer (Roche Diagnostics; Barcelona, Spain) in blood samples taken from the tail vein. In this case, 5-h-fasted mice were intraperitoneally administered with 0.75 U of insulin (Sigma-Aldrich, MO, USA) per kg body weight, and blood glucose concentrations were determined before (0) and at 15, 30, 60, and 120 min after insulin injection. Glucose levels were determined in blood samples before (0) and at 15, 30, 60, and 120 min following glucose administration. Mice were fasted for 5 h at the beginning of the light cycle, and then intraperitoneally injected with 2 g of glucose per kg body weight. The random-effects model used in the analysis addresses this variation to some extent, but the underlying causes of the heterogeneity warrant further investigation.In terms of stomachache, 4 studies (with 229 subjects from each of the groups) were included for analysis.Given that these therapeutics have demonstrated efficacy in improving hyperlipidemia, promoting weight loss, and reducing glycated hemoglobin levels, the implementation of GLP-1 RA could lead to improved outcomes in the management of PCOS .6Meta-analysis of the total cholesterol (A), HDL (B), LDL (C) and TG (D) levels found in this study.Full size imageIn terms of reduction in total HDL levels, 9 studies (with 325 and 288 subjects from the experimental and control groups, respectively) were included for analysis.Therefore, in patients where the fasting glucose is a more dominant feature of their hyperglycemia, metformin may be more appropriate, while GLP-1RAs could be better suited for those with postprandial hyperglycemia.However, the majority of women suffering from PCOS and obesity fail to significantly reduce their body weight with lifestyle intervention.Clinically, these findings suggest that GLP-1RAs could be incorporated into PCOS management guidelines, particularly for patients who do not respond adequately to first-line therapies or present with postprandial hyperglycemia and central obesity.Additionally, reference lists of identified studies were manually checked for additional relevant articles. From the Random Forest analysis, we selected the most significant proteins based on the Mean Decrease Accuracy, which were then used for subsequent pathway analysis. This method strikes a balance by reducing the impact of large-fold changes while preserving the inherent variability of the data.In terms of statistical analysis, we followed two main approaches. Second, for data transformation, we applied a log transformation (base 10), which helps in stabilizing variance and normalizing data distribution, making the data more suitable for downstream analysis82. Data availability The random-effects model used in the analysis addresses this variation to some extent, but the underlying causes of the heterogeneity warrant further investigation. In terms of insomnia, 1 study (with 14 subjects from each of the groups) were included for analysis. In terms of fatigue, 1 study (with 20 subjects from each of the groups) were included for analysis. Meta-analysis of the total cholesterol (A), HDL (B), LDL (C) and TG (D) levels found in this study. Meta-analysis of the DHEAS (A), SHBG (B), total testosterone (C), free testosterone (D) and FAI (E) levels found in this study. Across the studies, hormonal profiles were examined in relation to GLP-1 receptor agonist interventions. Similarly, LIRA therapy resulted in improved mean blood glucose and insulin response following a 32-week intervention . Several studies evaluated the effect of GLP-1 RA on biochemical parameters that are indicative of metabolic health. GLP1 is a gut-derived polypeptide released from the intestinal L cells, capable of stimulating glucose-dependent insulin secretion and inhibiting food intake by acting on the pancreas and brain, respectively. Glucagon-like peptide-1 receptor agonists (GLP1-RA) are emerging as novel anti-diabetic and weight-loss medications for the treatment of T2D and obesity26. Yet, controversial outcomes on the efficacy of metformin have also been reported21,22,23,24, that might be attributed to heterogeneity of responses in some patient subgroups. However, the included studies generally lacked sufficient detail regarding these concurrent interventions, making it difficult to fully assess their potential impact on the outcomes. Notably, the high levels of heterogeneity in the studies analyzed as well as other measures such as variability in study populations, interventions and methodologies, could have affected the reliability of the results obtained. This meta-analysis study had some limitations regarding the efficacy of GLP-1RAs on the anthropometric measurements, glucose homeostasis and hormonal changes. In the meantime, the evidence supports GLP-1RAs as a strong alternative to metformin, especially in cases where weight loss is a primary treatment goal. Overall, our meta-analysis underscores the value of GLP-1RAs as a multifaceted therapeutic option, particularly in PCOS patients with obesity and IR. On the other hand, while the triagonist, GLP1/GIP/Glucagon, was very effective in terms of body weight lowering, in keeping with previous references42, treatment of PWA mice led to a massive, likely detrimental weight loss, also at the expense of lean mass, that was not bound to clear amelioration of glucose intolerance in androgenized female mice. Intriguingly, GLP1/GIP had limited efficacy in our PCOS model with more severe metabolic complications, i.e., PWA mice fed HFD, suggesting that in the presence of persistently elevated androgen levels, its beneficial effects might be dampened. In contrast, PWA mice display overt metabolic perturbations, including overweight and impaired glucose homeostasis46, and may mimic the canonical human phenotype A of PCOS linked to hyperandrogenism, anovulatory cycles, and polycystic ovarian morphology45. Meta-analysis of the total cholesterol… Meta-analysis of the DHEAS ( A ), SHBG ( B ), total testosterone… Meta-analysis of the DHEAS ( … A meta-analysis of BMI changes found in this study ( A ), body… For glucose homeostasis, GLP-1RAs significantly reduced fasting insulin, glucose level at 2 h after OGTT, and HOMA-IR. In terms of the reduction in levels of free testosterone, 3 studies (with 39 and 41 subjects from the experimental and control groups, respectively) were included in the analysis.Adverse events information was available for 112 patients, where 49 had light side effects such as nausea and abdominal pain.However, the effects on waist circumference and blood pressure varied among studies 30,33,37.In PNA mice, treatment with the lower dose of GLP1/E had also superior metabolic efficacy vs. GLP1/GIP and the triple-agonist, promoting a sustained and significant decrease in body weight (8%), a marked reduction in fat tissue (≈40%) and food intake, as well as an improvement in glucose metabolism (Fig. 4A–H).For the subgroup analysis, the studies were classified based on whether the patients from the control group received either metformin or placebo.This effect was particularly pronounced when the control group received a placebo.To this end, we applied indirect calorimetry and infrared thermography systems in PWA mice, treated with equimolar doses (10 nmol/kg) of GLP1/E, GLP1/GIP, and GLP1/GIP/Glucagon, during the first 3 days of treatment; a period when body weight loss was particularly marked.Finally, due to the retrospective nature of our study, lifestyle modifications and dietary changes may not have been standardized across patients and could be considered a confounding variable in our analysis.4Meta-analysis of the fasting glucose (A), glucose at 2 h (B), fasting insulin (C), insulin at 2 h (D) and HOMA-IR (E) levels found in this study.Full size imageIn terms of reduction in glucose levels at 2 h after OGTT, 5 studies (with 205 and 213 subjects the experimental and control groups, respectively) were included for analysis. In contrast, no effects on ovarian function were observed after treatment with high doses of the different multi-agonists in this PCOS model (Suppl. Fig. S4D). In contrast, the triagonist induced a significant decrease in ovarian and uterus weight vs. vehicle-treated PNA mice (Suppl. Fig. S4A, B). As in PWA mice, such body weight loss was accompanied by a significant decrease in food intake and fat mass (Fig. 2C, D), while only mice treated with the triagonist displayed a significant reduction in lean mass (Fig. 2D). Our data also document changes in other specific hypothalamic signaling pathways that may be related to the metabolic effects of the multi-agonist, as is the case of calcium/calmodulin-dependent protein kinase 2 (CAMK2). In addition, HFD also promotes apoptotic processes at the hypothalamic level62, and weight-lowering therapies, as exercise, contribute to mitigate apoptosis in hypothalamic neurons60. It is well known that HFD-induced obesity promotes low-grade hypothalamic inflammation, causing deregulation of central mechanisms that control energy homeostasis59, and both physical exercise and some pharmacotherapies have been shown to improve systemic metabolism by attenuating hypothalamic inflammation60,61. The ultimate pathways whereby these molecular changes may translate into changes in body weight and metabolic (dys)function are yet to be fully disclosed, but alterations in the hypothalamic proteome involved in inflammatory and apoptotic processes, as well as in autophagy and vesicular trafficking, may play a role. In fact, only a modest improvement in insulin resistance was consistently detected in PWA mice following treatment with metformin, as reflected by a moderately reduced HOMA-IR index. Data from each cell of our framework underwent analysis to comprehensively describe the studies and findings regarding GLP-1 use among patients with PCOS. Moreover, it has been reported that the use of GLP-1 RAs resulted in weight loss via the hormone’s effects on appetite reduction, increased satiety, and significant control of plasma lipid profiles 20,21. It has been shown that 50% of patients with a PCOS diagnosis are overweight and obese 7,11 given that the insulin resistance implicated in the pathophysiology of this syndrome predisposes women to metabolic derangements such as hyperinsulinemia, hyperglycemia, impaired glucose tolerance, and dyslipidemia . In order to validate the PCOS models, non-androgenized controls treated with vehicle were also included in the different studies. To this end, we applied indirect calorimetry and infrared thermography systems in PWA mice, treated with equimolar doses (10 nmol/kg) of GLP1/E, GLP1/GIP, and GLP1/GIP/Glucagon, during the first 3 days of treatment; a period when body weight loss was particularly marked. Notably, treatments with the multi-agonists or GLP1 did not cause signs of nausea/malaise at any time-point tested (2–72 h), since pica behavior (i.e., kaolin intake) was similar, or even lower, in mice treated with GLP1/E, GLP1/GIP, GLP1/GIP/Glucagon, or GLP1 vs. the control PWA group (Suppl. Fig. S8C). In good agreement, intervention with the multi-agonists significantly reduced food intake from 2- to 72-h, while the anorectic effects of GLP1 manifested only from 2- to 24-h (Suppl. Fig. S8B). Chronic administration of GLP1/E significantly improved also glucose handling, as compared with the vehicle-treated PWA group, whereas no glucoregulatory actions were detected in mice treated with the dual or triple agonist (Fig. 1E-upper panel). In contrast, GLP1/GIP therapy was unable to consistently reduce body weight, food intake, or fat mass in PWA mice over the study period (Fig. 1A–D). In initial validation analyses, we documented that consumption of a high-fat diet (HFD) overtly exacerbated the metabolic profile of PWA animals (Suppl. Fig. S2A, C), while in PNA mice, the impact of HFD was of similar magnitude in control and prenatally-androgenized animals (Suppl. Fig. S2B). In this context, it has been demonstrated that treatments with either a GLP1/Estrogen (GLP1/E) conjugate, a unimolecular GLP1/GIP (gastric inhibitory peptide) dual agonist, or a GLP1/GIP/Glucagon triagonist, markedly reduced body weight and improved diabetic complications, including hyperinsulinemia, in preclinical models of obesity in males33,34,35. Of the GLP-1 patients included, 3 (15.7%) and 16 (84.2) were on semaglutide and liraglutide respectively. Baseline characteristics were compared between metformin monotherapy and GLP-1 monotherapy cohorts using contingency table chi-2 analysis for categorical variables and Fisher exact t-test for continuous variables. Patients treated with metformin for less than 180 days, semaglutide less than 30 days, or liraglutide less than 21 days were excluded from analysis. Patients treated with metformin included 500 mg and 1000 mg daily dosing. Patient charts were further reviewed to identify those treated with metformin or GLP-1 monotherapy (liraglutide or semaglutide) for weight loss. C Random Forest out-of-bag (OOB) error shows OOB error plots from Random Forest analysis for PWA (left) and PNA (right) conditions. The left heatmap compares PWA + Vehicle (control) against PWA + GLP1/E (treatment), and the right heatmap compares PNA + Vehicle (control) against PNA + GLP1/E (treatment). Heatmaps generated using Ward clustering and Euclidean distance measurement revealed distinct patterns of protein expression between treated and control groups in both PWA and PNA models. Treatment with a 10 nmol/kg dose of GLP1 also ameliorated estrous cyclicity, with 80% of PNA mice showing regular cycles (Fig. 9A–C), although no effects were seen in the rest of gonadal and hormonal parameters (Fig. 9D–G). Our sample size was limited given many patients with PCOS had been on multiple weight loss medications within a six-month period that would have confounded results. Our data suggests that GLP-1 agonists are likely better agents to facilitate meeting this weight loss threshold in patients with PCOS–O and can translate into long-term benefit. 84.2% and 57.8% achieved greater than 5% and 10% weight loss with GLP-1 monotherapy, respectively, versus metformin (25% and 16.6%). A significantly greater proportion of patients treated with GLP-1 monotherapy achieved a higher degree of weight loss in our study. To our knowledge, only one previous study by Jensterle et al. has sought to evaluate the effect of GLP-1, specifically liraglutide, versus metformin monotherapy on weight loss . Thus, integration of both hormones into a single molecule may not only maximize the metabolic efficacy of the individual signals, but allow also specific delivery of estrogen into GLP1 receptor-expressing cells, avoiding potential undesired effects of estrogen on reproductive tissues, as shown also in our studies. The logic behind this approach is that the concomitant activation of GLP1 and estrogen receptors may drive additional favorable effects, when compared with the individual activation of these pathways, which are known to conduct, on their own, beneficial metabolic actions on energy and glucose homeostasis52. Our data were congruent with those findings, suggesting that GLP1/E retains its powerful body weight-lowering effects in female models of persistent androgenization and PCOS. In addition, our data unambiguously documented that the effects of this and other multi-functional analogs on body weight and food intake are not merely due to food aversion or malaise linked to gastrointestinal distress, regarded as one of the main side-effects of GLP1-based therapy in humans49. In terms of reduction in total cholesterol, 10 studies (with 335 and 237 subjects from the experimental and control groups, respectively) were included for analysis. In terms of the reduction in FAI, 6 studies (with 195 and176 subjects from the experimental and control groups, respectively) were included in the analysis. In terms of the reduction in total testosterone, 9 studies (with 264 and 223 subjects from the experimental and control groups, respectively) were included in the analysis. Study characteristics Further prospective randomized controlled studies are needed to establish GLP-1 weight loss efficacy in patients with PCOS-O and clinically related outcomes. The included studies uniformly reported reductions in weight and body mass index (BMI) among patients who were prescribed GLP-1 RAs, specifically liraglutide or exenatide. While our methods focused on comparing GLP-1RAs to metformin or placebo, we acknowledge the possibility that additional background interventions may have been employed across some studies. The heterogeneity observed in the hormonal analysis may stem from differences in baseline hormonal levels and other concurrent therapies used by the subjects in the included studies. Moreover, the recent FDA approval of some GLP-1 RAs for weight management may have limited the availability of eligible studies within our time frame. The limited number of studies included in this scoping review may restrict the generalizability of our findings. Additionally, our findings, which show conflicting results regarding the effects of GLP-1 RAs on lipid profiles, underscore the need for more robust studies to clarify their impact on cardiovascular risk factors in PCOS 31,32,34,37. Future research endeavors should focus on conducting larger, placebo-controlled trials to establish the efficacy and safety of GLP-1 RAs in PCOS populations. GLP-1RAs effectively reduced body weight, BMI and insulin resistance in patients with PCOS, although they were accompanied by nausea, vomiting and dizziness. There was no significant difference in total cholesterol and HOMA-IR levels. We performed data extraction and quality assessment for studies that met the inclusion criteria. Our primary outcomes of interest included body mass index (BMI), triglycerides, waist circumference, total testosterone, total cholesterol, and HOMA-IR. This section collects any data citations, data availability statements, or supplementary materials included in this article. However, GLP1/E reduced testosterone levels in PNA mice, an effect mimicked by monotherapies and metformin (Fig. 8I); a similar profile was detected for serum levels of progesterone (Suppl. Table S1, lower panel). In contrast, metformin did not cause significant changes in any of the metabolic parameters under analysis (Fig. 8A–H). For reference purposes, values from control non-androgenized mice are also included (black bars and dashed lines). In order to calculate integral glucose levels in the glucose tolerance test, the area under the curve (AUC) was calculated using the trapezoidal rule. In this single-center retrospective study, we determined weight loss in patients with PCOS–O with GLP-1 monotherapy versus metformin. In this single-center retrospective study, we determined weight loss in patients with PCOS-O with GLP-1 monotherapy versus metformin. Notably, we found a significant increase in nausea events among participants receiving GLP-1RAs compared to those treated with metformin or placebo. This highlights the limited role that GLP-1RAs play in lipid management, suggesting that additional lipid-lowering therapies may be necessary for comprehensive metabolic control27,28,29. While total cholesterol and LDL levels were reduced, there was no significant differences in HDL and triglycerides. However, GLP-1RAs did show a favorable impact on these hormones when compared to placebo alone. Metformin primarily reduces hepatic glucose production, while GLP-1RAs enhance insulin secretion in response to meals20,21,22. GLP1/E significantly ameliorated also insulin sensitivity, as denoted by ITT and HOMA-IR, regardless of the dose, and reduced basal insulin levels (Fig. 5E, G, H); yet, the latter was significant only in mice treated with the highest dose. In good agreement, mice treated with GLP1/E displayed significant loss of fat mass, which was greater in animals exposed to the intermediate and high doses (Fig. 5D), together with a pronounced reduction in leptin levels (Suppl. Fig. S7A). The three doses of GLP1/E led to a very significant reduction of body weight along the treatment period (Fig. 5A), with a 10% drop in body weight for the highest dose, and an 8% reduction with the 10 and 5 nmol/kg doses relative to vehicle-treated PWA mice (Fig. 5B). No significant alterations were detected either in circulating FGF21 or adiponectin levels (Suppl. Fig. S5B, C). Statistically significant differences were assessed by one-way ANOVA followed by Tukey’s multiple comparison tests to analyze the effects of compound intervention vs. vehicle administration in PNA mice. The hormonal changes observed in our analysis further contribute to the therapeutic potential of GLP-1RAs, though the impact was less consistent compared to their effects on anthropometric measurements and glucose regulation. Therefore, in patients where the fasting glucose is a more dominant feature of their hyperglycemia, metformin may be more appropriate, while GLP-1RAs could be better suited for those with postprandial hyperglycemia. The consistent reductions in BMI and body weight across various studies highlight the potential of GLP-1RAs in managing PCOS-related conditions, even in populations where metformin has traditionally been the first-line therapy. A–H Effects on body weight (A), body weight change (B), food intake (C), body composition (fat and lean mass change) (D), glucose (upper panel) and insulin tolerance (E), fasting glucose (F), serum insulin levels (G), and HOMA-IR index (H) in PNA female mice daily administered with vehicle (saline), GLP1/E (50 nmol/kg), GLP1/GIP (3 nmol/kg), GLP1/GIP/Glucagon triagonist (3 nmol/kg) or metformin (300 mg/kg) during 28 days. A–H Effects on body weight (A), body weight change (B), food intake (C), body composition (fat and lean mass change) (D), glucose (upper panel) and insulin tolerance (E), fasting glucose (F), serum insulin levels (G), and HOMA-IR index (H) in PWA female mice daily administered with vehicle (saline), GLP1/E (50 nmol/kg), GLP1/GIP (3 nmol/kg), GLP1/GIP/Glucagon triagonist (3 nmol/kg) or metformin (300 mg/kg) during 28 days. Regarding gonadal effects, treatment with GLP1/E, GLP1/GIP, and metformin did not alter ovarian or uterus weights, nor did they change circulating LH levels in this hyperandrogenic model of PCOS (Suppl. Fig. S3D–F). A separate observational analysis by Rasmussen et al. demonstrated weight loss of 9 kg (9.1%) in patients with PCOS treated with combination therapy of liraglutide and metformin, with titration up to 1.8 mg of liraglutide . While side effects such as nausea and dizziness are more common, they are generally manageable and should be weighed against the potential metabolic improvements. More robust, long-term studies are needed to elucidate the long-term safety profiles of GLP-1RAs and their role in combination therapy for metabolic syndrome. Importantly, the funnel plot analysis demonstrated no evidence of publication bias, enhancing the credibility of our findings. Given that dyslipidemia is a key component of metabolic syndrome, and cardiovascular outcomes are of the utmost importance in managing such patients, a combination approach may yield better results than GLP-1RA monotherapy in this aspect30. E Boxplots depicting the expression levels of the most significant proteins identified by Random Forest analysis.Exclusion criteria were applied to studies involving children (≤ 18 years of age) or postmenopausal women over the age of 65.In this study of adult patients with PCOS–O, treatment with GLP-1 monotherapy resulted in a trend towards greater weight loss of 9.1 kg versus 4.9 kg in those treated with metformin.Notwithstanding, previous studies have documented beneficial effects of the activation of estrogen receptors in diet-induced obese females, by reducing body weight and fat accumulation and improving leptin resistance at central level53,54.The selected studies provided valuable insights into the use of long-acting GLP-1 RAs, including LIRA and EXE, across diverse geographic locations and study designs 30-37.GLP-1RAs are effective in reducing body weight, BMI, and insulin resistance in patients with PCOS, particularly those with obesity or metabolic complications.A visual representation of the eligibility of the quality of the clinical studies included is shown in Fig. All studies were published in English and exhibited wide geographic diversity. This study aimed to investigate the therapeutic efficacy of GLP-1 RAs use for weight loss in women diagnosed with PCOS. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. Notably, these effects were not mimicked by E alone, which failed to rescue cycle perturbations and had a discernible uterotrophic action, but were replicated, in terms of reversal of irregularities of the ovarian cycle, by GLP1 alone, which nonetheless did not elevate LH levels. Intriguingly, in our studies, hypothalamic CAMK2 expression was upregulated by GLP1/E; an effect that may be due to compensatory mechanisms in response to the marked reduction of body weight after chronic treatment with GLP1/E. Of note, different reports have also documented the limited efficacy of similar doses of metformin in terms of improvement of metabolic profiles in experimental models of diabetes in female rats56,57, as well as in PNA mice58, suggesting that the metabolic effects of metformin may be heterogeneous and inconsistent depending on the dose and preclinical model used. Univariate linear regression was used to identify correlations between treatment and weight loss.These results are clinically significant, as GLP-1RAs may offer an alternative approach for patients with PCOS.However, GLP-1RAs did show a favorable impact on these hormones when compared to placebo alone.In addition, we also evaluated the safety of the combined treatment of metformin and GLP-1RAs.Interestingly, reproductive responses were partially different in the PCOS model of preserved ovulation, since treatment of PNA mice with a low dose of GLP1/E was capable to elevate LH levels and reversed, to a large extent, the ovarian cycle irregularities observed in PNA mice treated with vehicle.Despite the growing popularity and promising outcomes of GLP-1 RAs as weight-loss adjuncts, there is a need for an examination of the available evidence to determine the safety and efficacy of these pharmacotherapies in patients with PCOS 23-26.Notably, treatments with the multi-agonists or GLP1 did not cause signs of nausea/malaise at any time-point tested (2–72 h), since pica behavior (i.e., kaolin intake) was similar, or even lower, in mice treated with GLP1/E, GLP1/GIP, GLP1/GIP/Glucagon, or GLP1 vs. the control PWA group (Suppl. Fig. S8C). In addition, changes in respiratory quotient pointed out greater fat oxidation after multi-agonist treatments, which may contribute to the beneficial effects of the compounds in the metabolic handling of PWA mice. Admittedly, these metabolic analyses were focused on the initial phase of treatments, as this period displayed the largest changes in body weight and is less likely affected by compensatory responses, which might confound the identification of primary mechanisms. Investigation of the mechanisms underlying the beneficial metabolic effects of multi-agonist therapies, conducted in PWA mice, as a genuine model for the metabolic alterations of PCOS36, revealed a predominant action in terms of suppression of food intake, with negligible effects in terms of energy expenditure, thermogenic function or locomotor activity. This is in contrast with the beneficial effects reported previously for the triagonist, in terms of reversion of adiposity excess and improved metabolic profile in diet-induced obese female mice42, suggesting that hyperandrogenism is bound to particular metabolic features that require specific multi-agonist treatments for maximum efficacy. Interestingly, clinical trials reported to date on the effects of tirzepatide did include obese men and women, but results were not segregated by gender47, thus making it difficult to anticipate whether a sex dimorphism exist in GLP1/GIP responses. For glucose homeostasis, we compared the results for the reduction in fasting glucose and insulin levels, their levels at 2 h after OGTT and HOMA-IR. Funnel plots of the reductions in BMI, body weight, WC, WHR and AG of the patients indicated no publication bias (Fig. S2). A meta-analysis of BMI changes found in this study (A), body weight changes (B), WC (C), WHR (D) and AG (E). A visual representation of the eligibility of the quality of the clinical studies included is shown in Fig. This is somewhat unsurprising, given the fact that persistently elevated androgen levels in PWA rodents have been shown to prevent ovulatory rescue after administration of more proxy activators of the reproductive axis, such as kisspeptin46. In parallel to their metabolic impact, the reproductive effects of the GLP1-based multi-agonists were also assessed in PNA and PWA models of PCOS, particularly for GLP1/E. All in all, while identification of the molecular mechanisms for its beneficial metabolic effects warrants further investigation, our proteomic data strongly suggest a distinctive role of central actions of GLP1/E for the amelioration of the metabolic profiles of our PCOS models. Pharmacological inhibition of hypothalamic CAMK2 has been shown to reduce food intake and body weight by decreasing NPY expression66. Reduced levels of CRP and improved endothelial function suggest potential cardio-protective effects of GLP-1 RAs in PCOS patients .In contrast, the metabolic profile PNA mice was unaffected after intervention with lower doses of the dual- and triple-agonists, as well as with metformin (Fig. 4A–H), except for a significant decrease in basal glucose levels in GLP1/GIP-treated mice (Fig. 4F) and reduction in daily food intake in PNA mice treated with metformin (Fig. 4C).Recent, as yet fragmentary evidence suggests that intervention with GLP1-RA, e.g., liraglutide, may reduce body weight and improve hyperandrogenism and menstrual irregularities in women with obesity and PCOS28,29,30,31,32; effects probably related to its anti-diabetic and weight-lowering properties.Its clinical manifestations are highly heterogeneous, primarily characterized by infrequent or absent ovulation, elevated androgen levels and polycystic ovarian changes1,2.In contrast, GLP1/GIP therapy was unable to consistently reduce body weight, food intake, or fat mass in PWA mice over the study period (Fig. 1A–D).The quality of included studies was assessed using the Cochrane risk-of-bias tool. Superior metabolic efficacy of GLP1/E vs. GLP1 or E monotherapies in an obese model of PCOS Weight loss facilitated by GLP-1 agonists in patients with PCOS-O is not well characterized.In sum, our data document the beneficial effects of GLP1-based multi-agonists, and particularly GLP1/E, in two models of PCOS, with the identification of optimal doses, main metabolic and reproductive effects, and tenable hypothalamic pathways targeted.The three doses of GLP1/E led to a very significant reduction of body weight along the treatment period (Fig. 5A), with a 10% drop in body weight for the highest dose, and an 8% reduction with the 10 and 5 nmol/kg doses relative to vehicle-treated PWA mice (Fig. 5B).Meta-analysis of the total cholesterol (A), HDL (B), LDL (C) and TG (D) levels found in this study.This method strikes a balance by reducing the impact of large-fold changes while preserving the inherent variability of the data.In terms of statistical analysis, we followed two main approaches.Regarding gonadal effects, treatment with GLP1/E, GLP1/GIP, and metformin did not alter ovarian or uterus weights, nor did they change circulating LH levels in this hyperandrogenic model of PCOS (Suppl. Fig. S3D–F).5Meta-analysis of the DHEAS (A), SHBG (B), total testosterone (C), free testosterone (D) and FAI (E) levels found in this study.Full size imageIn terms of the reduction in SHBG, 5 studies (with 151 and 153 subjects from the experimental and control groups, respectively) were included in the analysis. The most common abnormality in PCOS is excessive ovarian androgen secretion, present in up to 80% of patients9,10. We report here the hormonal, metabolic and gonadal responses to the glucagon-like peptide-1 (GLP1)-based multi-agonists, GLP1/Estrogen (GLP1/E), GLP1/gastric inhibitory peptide (GLP1/GIP) and GLP1/GIP/Glucagon, in two mouse PCOS models, with variable penetrance of metabolic and reproductive traits, and their comparison with metformin. Metabolic complications are common in patients suffering PCOS, including obesity, insulin resistance and type-2 diabetes, which severely compromise the clinical course of affected women. All data collection and analysis was performed following approval from the respective institutional review board. In terms of reduction in total HDL levels, 9 studies (with 325 and 288 subjects from the experimental and control groups, respectively) were included for analysis. In terms of the reduction in levels of free testosterone, 3 studies (with 39 and 41 subjects from the experimental and control groups, respectively) were included in the analysis. In terms of reduction in insulin levels at 2 h after OGTT, 5 studies (with 194 and 199 subjects from the experimental and control groups, respectively) were included for analysis. In terms of reduction in fasting insulin levels, 5 studies (with 208 and 212 subjects from the experimental and controls groups, respectively) were included for analysis. In this 12-week randomized study of 32 patients with PCOS–O, weight loss of 3 kg and 2.3 kg in those treated with liraglutide and metformin was seen. In this study of adult patients with PCOS–O, treatment with GLP-1 monotherapy resulted in a trend towards greater weight loss of 9.1 kg versus 4.9 kg in those treated with metformin. Five metformin and 17 GLP-1 monotherapy patients were excluded from analysis due to lack of six-month follow up data. The effects of GLP-1 agonists to facilitate weight loss in patients with PCOS and obesity (PCOS–O) is relatively unknown however. Two independent reviewers extracted the data from eligible studies using a standardized extraction protocol. Additionally, reference lists of identified studies were manually checked for additional relevant articles. In addition, we also evaluated the safety of the combined treatment of metformin and GLP-1RAs. This meta-analysis systematically assessed the impact of GLP-1RA therapy on anthropometric outcomes and key metabolic parameters in women with PCOS. However, while individual studies have shown promising results, the overall efficacy and safety of GLP-1RA therapy in women with PCOS require further comprehensive evaluation. Figures We next assessed the metabolic, gonadal, and hormonal effects of the intervention with equivalent doses (10 nmol/kg) of GLP1, E, or GLP1/E in PNA mice, which retain ovulatory capacity albeit with clear dysregulation of ovarian cyclicity. None of the compounds caused changes in testosterone levels, except for a modest but significant increase in GLP1-treated PWA mice (Fig. 7I). PWA mice were administered with equimolar doses of GLP1, E, or GLP1/E (10 nmol/kg), and results were compared with those of vehicle- or metformin-treated (300 mg/kg) PWA mice. A–G Effects on body weight change (A), food intake (B), body weight-correlated average energy expenditure (C), total locomotor activity (D), and respiratory quotient (E) in PWA female mice daily treated with vehicle, GLP1/E (10 nmol/kg), GLP1/GIP (10 nmol/kg), triagonist (10 nmol/kg), during 3 days. Treatment with the multi-agonists caused a very significant reduction in body weight during the 72-h treatment period, which was more pronounced in mice treated with GLP1/GIP/Glucagon (Fig. 6A). Thus, while up to 70% of PNA mice displayed irregular cyclicity, characterized by the appearance of only one generation of (mainly persistent regressing) corpora lutea, 100% of PNA mice treated with metformin showed regular ovarian cyclicity, denoted by the presence of two generations of corpora lutea, as a sign of preserved regular ovulations (Fig. 9A–C). Unlike the PWA model, circulating DHT levels were nearly undetectable and unaltered in PNA mice, regardless of the treatment (Suppl. Table S1, lower panel). As in the PWA model, the improvement caused by GLP1/E therapy in PNA mice in terms of body weight, fat mass, food intake, and insulin sensitivity largely exceeded that of GLP1 or E monotherapies (Fig. 8A–H). Chronic administration of GLP1 decreased body weight, fat mass, and food intake, whereas no glucoregulatory or insulin-sensitizing actions were detected (Fig. 8A–H), while treatment with a similar dose of E improved only insulin sensitivity (Fig. 8A–H). Dushyanth Srinivasan and Holly Lofton were involved in data collection, analysis, manuscript writing and editing. However, we note only three patients in the GLP-1 cohort were on semaglutide. Secondly, given our small sample size, the GLP-1 cohort was coupled to include both semaglutide and liraglutide usage and weight loss seen may not be accurately reflective of the efficacy of each medication. Baseline characteristics are described in Table 1 and were similar between metformin and GLP-1 cohorts with the exception of mean days on medication. Univariate linear regression was performed to determine associations between treatment cohort and weight loss at six months. Statistical analysis was performed using STATA software (StataCorp, College Station, TX). Treatments with any of the multi-agonists significantly reduced body weight from 24- to 72-h relative to control PWA animals, while GLP1 decreased body weight only from 24- to 48-h; the magnitude of such effect was much lower than of multi-agonists (Suppl. Fig. S8A). In addition, mice treated with any of the three low doses of GLP1/E displayed improved glucose handling and basal glucose levels, with higher responses in mice treated with 10 and 25 nmol/kg doses (Fig. 5E, F). In PNA mice, treatment with the lower dose of GLP1/E had also superior metabolic efficacy vs. GLP1/GIP and the triple-agonist, promoting a sustained and significant decrease in body weight (8%), a marked reduction in fat tissue (≈40%) and food intake, as well as an improvement in glucose metabolism (Fig. 4A–H). As in previous experiments, intervention with an effective dose of metformin had minimal effects on the metabolic profile (Fig. 3A–G), except for marginal improvement of insulin resistance, denoted by a modest reduction of HOMA-IR (Fig. 3H). Yet, the efficiency of such interventions is, in many cases, limited, and additional, second-line treatments are frequently implemented, that mainly involve the use of metformin, as insulin-sensitizer, with variable efficacy in terms of improvement of reproductive and metabolic alterations. Considering that obesity and/or insulin resistance are predominant among patients with PCOS, lifestyle interventions aimed at lessening the metabolic burden of the syndrome are often at the first-line approach45. Our data delineate the therapeutic superiority of the di-agonist, GLP1/E, in the management of the metabolic alterations of PCOS, with beneficial effects also on ovarian function in our PCOS model of preserved ovulatory function. To our knowledge, there is only one study that has investigated the effects of GLP-1 monotherapy, specifically liraglutide, versus metformin . Both liraglutide and semaglutide have demonstrated meaningful weight loss in patients with obesity and without diabetes, ranging up to 8% and 15% of baseline body weight 13,14. With the additional benefits of weight loss, metformin has long been a part of the PCOS treatment paradigm. We identified 12 and 19 patients that were treated with metformin and GLP-1 monotherapy respectively. Our investigation delved into the clinical effects experienced by women of diverse racial and ethnic backgrounds with PCOS who were prescribed GLP-1 RAs for weight loss. In terms of reduction in fasting glucose levels, 6 studies (with 252 and 235 subjects from the experimental and control groups, respectively) were included for analysis. This meta-analysis aimed to evaluate the efficacy and safety of glucagon-like peptide-1 receptor agonists (GLP-1RAs) when compared to metformin and placebo in the management of body weight, glucose homeostasis and hormone levels in women polycystic ovary syndrome (PCOS). In contrast, high doses of GLP1/E had negligible effects on glucose control, insulin sensitivity, or circulating glucose and insulin levels (Fig. 2E–G), whereas metformin had no effects on any of the metabolic endpoints (Fig. 2A–H). Conducted the proteomic analysis by LC-MS/MS and participated in data analysis, with in-depth proteomic data analysis and interpretation being conducted by M.M.-O. The source data supporting the findings of this article are included in this article and its supplementary information files. These analysis were carried out employing R (version 4.0.5) and subsidiary packages85.Graphs and statistical analyses were performed using GraphPad Prism software V8.2.1 (Boston; MA, USA). Overall, the combination of these approaches (i.e., univariate analysis and Random Forest) leverages the strengths of both univariate and multivariate analyses, leading to a more robust and comprehensive understanding of the data. Other side effects, including fatigue, headache and insomnia, did not show significant differences when compared to the control group. Additionally, vomiting occurrences were also elevated, although no significant differences were observed in subgroup analyses when comparing to those given either placebo or metformin. This effect was particularly pronounced when the control group received a placebo. Future studies evaluating the combination of GLP-1RAs with statins or other lipid-lowering agents could provide further insights into the optimal treatment strategies for such patients. To assess the weight-lowering mechanisms linked to multi-agonists therapy, an indirect calorimetry system Oxylet ProTM (PANLAB; Barcelona, Spain) was used. Body weight changes were monitored daily along the treatment, and food and kaolin intakes were measured at 2-, 4-, 8-, 24-, 48-, and 72-h. PWA and control non-androgenized mice were exposed to kaolin pellets (Research Diets, K50001) 10 days prior to the start of the treatments to allow for adaptation to kaolin clay. Similarly, AMH and FGF21 serum levels were quantified by using commercial ELISA kits provided by Ansh Labs (TX, USA) and EMD Millipore Corporation (MO, USA), respectively. As internal control, in PWA mice, generated by chronic exposure to exogenous DHT, circulating levels of this androgen were consistently elevated, without differences among the different experimental groups (Suppl. Table S1, upper panel). As in previous experiments in PWA mice, no gonadal effects were detected following treatment with the different drugs, except for a significant reduction in ovarian weight and an increase in uterus weight in E-treated mice (Suppl. Fig. S9D–G). Conversely, intervention with the multi-agonists did not alter mean energy expenditure or locomotor activity in this PCOS model (Fig. 6C, D), suggesting that, at least in the initial phase of treatment, these compounds drive their weight-lowering effects mostly by reducing food consumption, without inducing thermogenic responses. In terms of the reduction in SHBG, 5 studies (with 151 and 153 subjects from the experimental and control groups, respectively) were included in the analysis. In terms of the reduction in DHEAS, 6 studies (with 128 and 110 subjects from the experimental and control groups, respectively) were included in the analysis. In terms of reduction in HOMA-IR, 10 studies (with 338 and 296 subjects from the experimental and control groups, respectively) were included for analysis. In terms of reduction in AG, 3 studies (with 117 and 121 subjects from the experimental and control groups, respectively) were included for analysis. With respect to the reduction in WHR, 4 studies (with 225 and 210 participants from the experimental and control groups, respectively) were included for analysis. In terms of diarrhea, 9 studies (with 369 and 319 subjects from the experimental and control groups, respectively) were included for analysis. In terms of vomiting, 7 studies (with 381 and 329 subjects from the experimental and control groups, respectively) were included for analysis. In terms of nausea, 11 studies (with 429 and 378 subjects from the experimental and control groups, respectively) were included for analysis. Its clinical manifestations are highly heterogeneous, primarily characterized by infrequent or absent ovulation, elevated androgen levels and polycystic ovarian changes1,2. Meta-analysis of nausea ( A ), vomiting ( B ), diarrhea ( C … Meta-analysis of nausea ( A … Meta-analysis of the total cholesterol ( A ), HDL ( B ), LDL… However, it is important to note the heterogeneity observed in some of the outcomes, particularly for BMI and body weight reductions. These results are clinically significant, as GLP-1RAs may offer an alternative approach for patients with PCOS. Meta-analysis of nausea (A), vomiting (B), diarrhea (C), constipation (D), stomachache (E), bloating (F), dizziness (G), fatigue (H), headache (I) and insomnia (J) found in this study. Funnels plot of the reduction in DHEAS, SHBG, total testosterone, free testosterone and FAI levels indicated no publication bias (Fig. S6). This analytical procedure allowed accurate determination of testosterone, dihydrotestosterone, androstenedione, progesterone, 17-alpha hydroxyprogesterone, and estradiol levels in serum samples from PNA and PWA mice. In order to accurately perform a comprehensive analysis of sex steroid profile in both PCOS models, specific protocols of LC-MS/MS were applied as described elsewhere78. To determine the effects of multi-agonists treatment on thermogenesis, infrared pictures were taken with a handheld infrared thermal camera (FLIR Systems E54; Oregon, USA). Home-cage locomotor activity was measured also along the treatments by means of an extensiometric weight transducer integrated below each cage. During this 72-hour period, body weight and food intake were daily monitored, and oxygen consumption (VO2) and carbon dioxide production (VCO2) were measured every 9 min. 5Meta-analysis of the DHEAS (A), SHBG (B), total testosterone (C), free testosterone (D) and FAI (E) levels found in this study.Full size imageIn terms of the reduction in SHBG, 5 studies (with 151 and 153 subjects from the experimental and control groups, respectively) were included in the analysis. Some studies demonstrated improved glucose tolerance and insulin homeostasis with EXE compared to metformin interventions, along with reductions in fasting glucose and HbA1c levels . Similarly, all included studies that utilized LIRA as an intervention reported weight loss among PCOS patients 31,32,34-36. In line with our short-term metabolic analyses, previous studies in New Zealand obese and diet-induced obese male mice suggested that GLP1/E acts primarily at central levels to reverse metabolic alterations by decreasing food intake and body weight, while the potential beneficial effects on pancreatic beta-cells were possibly secondary and indirectly-mediated33,48. The dose-findings studies reported here not only documented the superior efficacy of GLP1/E in the management of metabolic alterations in our obese PCOS model, including lowering body weight and adiposity, as well as improvement of glucose handling and insulin resistance, but also highlighted a non-monotonic effect of this dual agonist in PNA mice, with optimal effects being achieved with submaximal doses. Interestingly, reproductive responses were partially different in the PCOS model of preserved ovulation, since treatment of PNA mice with a low dose of GLP1/E was capable to elevate LH levels and reversed, to a large extent, the ovarian cycle irregularities observed in PNA mice treated with vehicle. In addition, despite previous reports on the capacity of GLP1 on its own to activate the female reproductive axis67, this effect was not detected in our study, with no long-term impact on LH or AMH levels, which might suggest a state of refractoriness due to persistent androgenization. Importantly, the vast majority of the available PCOS models are generated by exogenous administration of androgens during specific developmental windows, which hampers evaluation of the effects of treatments on endogenous androgen secretion and ovarian function. The inability of the multi-agonists to restore gonadal function in PWA mice is likely due to chronic exposure to fixed concentrations of the exogenous androgen, DHT, which may limit the eventual beneficial effects of the compounds at the gonadal level, even when substantial metabolic improvements are achieved. However, the effects on waist circumference and blood pressure varied among studies 30,33,37. Across the studies, the use of EXE in women with PCOS consistently resulted in weight and BMI reduction 30,33, with some studies indicating significant improvements compared to alternative treatments . Relevant data from the studies were entered into a Microsoft Excel (Microsoft Corporation, Redmond, Washington, United States) extraction form for further analysis. Authors five through nine were responsible for completing the data extraction of selected studies, which included details such as study design, recruitment/data collection period, country, setting, sample size and characteristics, study objectives, and outcomes. We conducted a scoping review to provide a summary of the current available evidence regarding the relevance of GLP-1 agonist use for weight loss in patients diagnosed with PCOS. While GLP-1RAs showed significant reductions in BMI, body weight and WC compared to either metformin or placebo, the lack of consistency in fasting glucose and hormone levels suggested that the impact of the interventions might have varied significantly across the different patient groups. The findings from our meta-analysis suggested that GLP-1RAs were effective in improving the anthropometric parameters such as BMI, body weight, WC, WHR and AG, particularly when compared to the patients given either metformin or placebo. In terms of reduction in glucose levels at 2 h after OGTT, 5 studies (with 205 and 213 subjects the experimental and control groups, respectively) were included for analysis. For subgroup analysis, the studies were classified based on whether the control group received metformin or placebo. 4Meta-analysis of the fasting glucose (A), glucose at 2 h (B), fasting insulin (C), insulin at 2 h (D) and HOMA-IR (E) levels found in this study.Full size imageIn terms of reduction in glucose levels at 2 h after OGTT, 5 studies (with 205 and 213 subjects the experimental and control groups, respectively) were included for analysis. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. Clinically, these findings suggest that GLP-1RAs could be incorporated into PCOS management guidelines, particularly for patients who do not respond adequately to first-line therapies or present with postprandial hyperglycemia and central obesity. The observed heterogeneity across studies highlights the importance of individualized treatment strategies based on patient profiles and therapeutic goals. Yet, while GLP1/E and GLP1/GIP reduced body weight by 10% at the end of the treatment, the triagonist caused a 22% reduction (Fig. 2B). In PNA mice, that display modest metabolic dysregulation and overt gonadal perturbations, all multi-agonists decreased body weight (Fig. 2A, B). Statistically significant differences were assessed by one-way ANOVA followed by Tukey’s multiple comparison tests to analyze the effects of compound intervention vs. vehicle administration in PWA mice. Moreover, no effects were observed in ovarian cyclicity and histology following pharmacological treatment in any of the experimental groups (Suppl. Fig. S3G). Moreover, none of the treatments altered circulating adiponectin levels, and only mice treated with triagonist showed a significant elevation in serum levels of FGF21 (Suppl. Fig. S3B, C). GLP-1 is an incretin that delays gastric emptying, improves glucose-stimulated insulin secretion, and reduces appetite. More recently, glucagon-like peptide-1 (GLP-1) agonists such as liraglutide and semaglutide have been approved for chronic weight management. Weight loss of 5–10% is thought to reduce androgen levels, and improve metabolic markers and fertility 4,7,8. The quality of included studies was assessed using the Cochrane risk-of-bias tool. The primary outcomes included reductions in BMI, body weight, WC, WHR and AG. Reduced levels of GLP-1 in PCOS patients suggest that it may be involved in the pathogenesis and progression of the condition, and that it is potentially linked to obesity and IR. Meta-analysis of the fasting glucose ( A ), glucose at 2 h ( … Meta-analysis of the fasting glucose… Despite the growing popularity and promising outcomes of GLP-1 RAs as weight-loss adjuncts, there is a need for an examination of the available evidence to determine the safety and efficacy of these pharmacotherapies in patients with PCOS 23-26. Given that these therapeutics have demonstrated efficacy in improving hyperlipidemia, promoting weight loss, and reducing glycated hemoglobin levels, the implementation of GLP-1 RA could lead to improved outcomes in the management of PCOS . The serendipitous weight loss effects of semiglutide have prompted a robust investigation to conclude its significance as a therapeutic for weight management. For comparative purposes, a group treated with a similar dose of a GLP1 analog and a non-androgenized control group treated with a vehicle were also included. For calculation, 5-h fasting blood samples were collected from the tail, and glucose and insulin levels were determined using a digital glucometer (Roche) and a commercial ultrasensitive insulin ELISA kit (ALPCO; NH, USA), respectively. Weight loss facilitated by GLP-1 agonists in patients with PCOS-O is not well characterized. While the current evidence is encouraging, further research is warranted to elucidate both short- and long-term adverse effects of GLP-1 RA therapy for PCOS. Although some short-term adverse effects were noted, long-term effects of GLP-1 RAs use remain undetermined.