Polycystic Ovary Syndrome and Endometrial Cancer: A Scoping Review of the Literature on Gut Microbiota

Gut dysbiosis has been associated with polycystic ovary syndrome (PCOS) and endometrial cancer (EC) but no studies have investigated whether gut dysbiosis may explain the increased endometrial cancer risk in polycystic ovary syndrome. The aim of this scoping review is to evaluate the extent and nature of published studies on the gut microbiota in polycystic ovary syndrome and endometrial cancer and attempt to find any similarities between the composition of the microbiota. We searched for publications ranging from the years 2016 to 2022, due to the completion date of the ‘Human Microbiome Project’ in 2016. We obtained 200 articles by inputting keywords such as ‘gut microbiome’, ‘gut microbiota’, ‘gut dysbiosis’, ‘PCOS’, and ‘endometrial cancer’ into search engines such as PubMed and Scopus. Of the 200 identified in our initial search, we included 25 articles in our final review after applying the exclusion and inclusion criteria. Although the literature is growing in this field, we did not identify enough published studies to investigate whether gut dysbiosis may explain the increased EC risk in PCOS. Within the studies identified, we were unable to identify any consistent patterns of the microbiome similarly present in studies on women with PCOS compared with women with EC. Although we found that the phylum Firmicutes was similarly decreased in women with PCOS and studies on women with EC, there was however significant variability within the studies identified making it highly likely that this may have arisen by chance. Further research pertaining to molecular and microbiological mechanisms in relation to the gut microbiome is needed to elucidate a greater understanding of its contribution to the pathophysiology of endometrial cancer in patients with polycystic ovarian syndrome.


Introduction
The human body hosts trillions of microorganisms that play a pivotal role in modulating normal physiology and immune functions that are essential to our normal functioning; this effect is mediated through the production of bi-products and metabolites [1]. The term 'microbiome' refers to the microbes and their collective genomes within a community, while 'microbiota' refers to the "assemblage of microorganisms present in a defined environment" including bacteria, fungi, or archaea, but is more frequently used for bacteria composition and it refers to the microbes themselves in aggregate [2,3]. Up to 90% of the gut microbiota consists of two phyla, namely, Firmicutes and Bacteroidetes. The remaining 10% includes Actinobacteria, Proteobacteria, Fusobacteriota, and Verrucomicrobia [4].
Polycystic ovary syndrome (PCOS) is a disease of the hypothalamus-pituitary-ovarian (HPO) axis affecting about 20% of reproductive women worldwide [5,6]. PCOS is marked by anovulation, increased androgen secretion, and polycystic ovaries [7,8]. The presence of two out of the three characteristics of PCOS is needed to make a clinical diagnosis according to the Rotterdam criteria [9]. Other features include uneven gonadotropin secretion, i.e., increased luteinizing hormone (LH), increased LH:FSH (follicle-stimulating hormone) ratio, low sex hormone-binding globulin (SHBG), and chronic inflammation [10,11].
Endometrial Cancer (EC) is one of the most common malignancies occurring in women, accounting for about 142,000 cases and 42,000 deaths worldwide. Type 1 EC, which is the most common lesion is associated with an excellent prognosis, while Type 2 EC is often high grade and tends to recur [12]. Dumesic et al. suggested that women who were diagnosed with PCOS have a 2.7-fold increased risk of developing endometrial cancer (EC) [13]. Unfortunately, the exact mechanisms that increase the risk of EC in PCOS are unclear. The pathophysiology of the increased risk is thought to involve the exposure of the endometrium to abnormally high levels of estrogen because of anovulation unopposed to progesterone; however, this is uncertain [14].
The gut microbiome plays a pivotal role in modulating normal physiology and immune functions [1]. Dysbiosis of the gut microbiome with altered microbial composition and diversity has been associated with a myriad of disease conditions such as Type 2 Diabetes Mellitus, Inflammatory Bowel Disease, and severe conditions such as cancers [15].
Although PCOS and EC are different diseases, given the increased risk of EC in PCOS, a range of potential mechanisms has been explored as possible mechanisms underpinning the association, but not the possible role of gut microbiota.
Previous studies investigating the gut microbiome in PCOS [16][17][18][19] and EC patients [20] suggest a role for dysbiosis in the etiologies of both conditions. So far, no studies have sought to investigate whether commonalities in gut dysbiosis in PCOS and EC may explain the increased risk of EC in PCOS. However, alterations in alpha and beta diversity of the gut microbiome and the associated intestinal dysfunction have been postulated to play a role in the exacerbation of PCOS [21]. Gut dysbiosis results in abnormal activation of the immune system that interferes with the insulin receptors present in the body, causing hyperinsulinemia, which in turn elevates the secretion of androgens from the ovaries, preventing the formation of normal ovarian follicles [7]. With respect to EC, the most accredited theory with respect to the gut microbiota is that of the activity of the enzyme β-glucuronidase. Previous work by Baker et al. has shown a role for beta-glucuronidase produced by the gut microbiota in the regulation and deconjugation of estrogen into its active form [22]. Hence, with dysbiosis, the alteration in the modulation of this enzyme by the estrobolome could contribute to the development of endometrial hyperplasia and cancer [22].
As we did not identify any previous studies investigating whether commonalities in gut dysbiosis in PCOS and EC may explain the increased risk of EC in PCOS, we set out to perform a scoping review of the literature to determine the extent and nature of published studies on the gut microbiota in, PCOS and EC, and attempt to find any similarities between the composition of the microbiota to determine whether the gut microbiota may contribute to the pathogenesis of EC in those with PCOS.

Materials and Methods
Institutional review board approval was not required for this study as it did not involve direct contact with patients, and it was a secondary review of primary studies in the literature.

Eligibility Criteria
The published articles reviewed in this study were limited to studies published between the years 2016 to 2022. This was because of the completion of the human microbiome project in 2016, which was a major milestone in enhancing our understanding of the human microbiome. We also limited the study to studies on humans and excluded all studies on animal models to maintain relevance to clinical practice as well as any review articles. The literature search was also limited to studies published in the English language. Articles had to be focused on the gut microbiota, PCOS, and EC as well as including specific keywords such as 'gut microbiome', 'gut microbiota', 'gut dysbiosis', 'PCOS', and 'endometrial cancer'.

Information Sources
To collect all the relevant published articles, we used two databases: PubMed and Scopus. PubMed is a free search engine to search medicine and biomedical journal literature. It searches several databases and interfaces Medline, directly. This search engine maps user's search terms to the Medical subject heading (Mesh) and text words in Medline records and then searches [23]. Scopus is an abstract and indexing database with full-text links [24]. The timeline of the articles was filtered in the search engines following our previously outlined eligibility criteria. Two reviewers individually navigated the databases and exported the relevant articles into a spreadsheet, and once data collection was completed, the articles were reviewed once more in a group setting, and duplicates were removed.

Search
The search strategy for PubMed involved using the following combination of keywords such as 'gut microbiome and pcos', 'gut microbiota and pcos', 'gut dysbiosis and pcos', 'gut microbiome and endometrial cancer', 'gut microbiota and endometrial cancer', 'gut dysbiosis and endometrial cancer'. The time range was limited to 2016 to 2022.
The search strategy for Scopus involved using the following combination of keywords such as 'gut microbiome and pcos', 'gut microbiota and pcos', 'gut dysbiosis and pcos', 'gut microbiome and endometrial cancer', 'gut microbiota and endometrial cancer', 'gut dysbiosis and endometrial cancer'. The time range was limited to 2016 to 2022. Publications were also limited to 'All open access articles' and by provided categories into 'intestinal flora', 'polycystic ovary syndrome', 'human studies', 'endometrial cancer'.

Selection of Sources of Evidence
Following the completion of the data search, articles presented by the databases were split into two halves and assigned to one of two reviewers to be screened individually. Reviewers first screened article titles and if keywords were present the reviewer would then screen the abstract followed by the discussion and conclusion of each study. Data were then extracted from chosen studies and then finally cross-reviewed by both reviewers. Duplicate publications were removed.

Data Charting
The selected articles were read by two reviewers to extract the required data. A table was constructed that included the required variables that were needed to be extracted from each article. Once data charting was completed individually, these data were further reviewed in a group setting.

Data Items
The data extracted included the authors' names and the study design. Other variables included the 'study sample size' (if applicable), which included the number and description of participants in the study; 'sequencing technique used' (if applicable), which was the method used for sequencing the bacterial composition obtained from the participants in the study; and 'results of the study' (here information regarding the microbial composition or microbial diversity changes in the gut was included as well as any relevant information regarding conclusions made with respect to the gut microbiota).

Synthesis of Results
Once data charting was complete, two tables were constructed. One for PCOS and the other for EC. Comparisons of the PCOS and EC data to identify any identify commonalities in the microbial composition and any associated changes were carried out. This comparison was then summarized narratively.  Figure 1 is an illustration of the PRISMA Chart describing the results from the literature search. Following our initial search, we identified 200 results that we then screened. After applying the exclusion criteria, we reduced the list to 153 articles. We then applied our inclusion criteria, which resulted in a list of 25 articles. Of these 25 articles, 23 pertained to gut microbial changes in PCOS and 2 to EC.

Synthesis of Results
Once data charting was complete, two tables were constructed. One for PCOS and the other for EC. Comparisons of the PCOS and EC data to identify any identify commonalities in the microbial composition and any associated changes were carried out. This comparison was then summarized narratively. Figure 1 is an illustration of the PRISMA Chart describing the results from the literature search. Following our initial search, we identified 200 results that we then screened. After applying the exclusion criteria, we reduced the list to 153 articles. We then applied our inclusion criteria, which resulted in a list of 25 articles. Of these 25 articles, 23 pertained to gut microbial changes in PCOS and 2 to EC.

Characteristics of Sources of Evidence
The articles (n = 25) compiled and charted consist of several different study designs and varied samplings. The included articles were grouped into two main categories, i.e., those that discuss PCOS and others that discuss EC. Descriptive features such as the study authors, sample size, sequencing method, as well as study type pertaining to studies based on both PCOS as well as EC are summarized in Tables 1 and 2, respectively. The articles were original studies that had an objective relevant to the microbial composition change with regard to PCOS or EC. The charted findings for each of the compiled articles are

Characteristics of Sources of Evidence
The articles (n = 25) compiled and charted consist of several different study designs and varied samplings. The included articles were grouped into two main categories, i.e., those that discuss PCOS and others that discuss EC. Descriptive features such as the study authors, sample size, sequencing method, as well as study type pertaining to studies based on both PCOS as well as EC are summarized in Tables 1 and 2, respectively. The articles were original studies that had an objective relevant to the microbial composition change with regard to PCOS or EC. The charted findings for each of the compiled articles are summarized in Tables 1 and 2. The results outline the diversity and microbial changes in the gut microbiota pertaining to PCOS and EC.

Synthesis of Results
There were more (23 articles) relevant to PCOS and the microbiome compared to EC articles (2 articles). The frequency of mention of different microbial changes and the modal change that was discovered were noted in both tables. In PCOS, the most frequent mention of a decrease in microbial abundance was in Firmicutes [25][26][27][28] and Prevotellaceae [26,29,30], which was reported in 4 and 3 articles, respectively. Whilst opposingly, a frequency of mention of an increase in abundance in Bacteroides vulgatus [31,32], Escherichia [31,33], and Streptococcus [18,34,35] was present in 2 articles for each apart from Streptococcus (3 articles). The microbiome in women with EC, also, demonstrated a decrease in the Firmicutes to Bacteroidetes ratio [36].
Data from both tables regarding microbial changes only shared two common phyla, Bacteroidetes and Firmicutes. Results from the EC articles (1 article) [36] revealed an increase in the Bacteroidetes phylum while, opposingly, results from PCOS articles (2 articles) [26,34], revealed a decrease in the same phylum. This was one piece of evidence of opposing results when comparing PCOS to EC, with other instances of such results prevalent in the PCOS table of results. The phylum Firmicutes was also mentioned in PCOS and EC studies with the results being in agreement as 3 articles [25][26][27] from PCOS showed a decrease similar to the findings in one EC study [37].
There was variability in contradictions in findings pertaining to the similarities in microbial composition change between both diseases. Liang Z et al. [33] Case-Control study 20 women with PCOS (lean PCOS, PL, n = 10; overweight PCOS, PO, n = 10) and 20 healthy control women (lean control, CL, n = 10; overweight control, CO, n = 10)

S rDNA V3-V4 region
Increase in gamma-aminobutyric acid (GABA)-producing species in PCOS, including Parabacteroides distasonis, Bacteroides fragilis, and Escherichia coli. Decrease in alpha diversity of gut microbiota of the women with PCOS from controls.

PCOS patients and 26 control patients 16S rRNA
The abundance of Faecalibacterium, Bifidobacterium, and Blautia was shown to be higher in the control group, while that of Parabacteroides and Clostridium was in PCOS Zhou L et al. [46] Cross-sectional study 60 women with PCOS (30 obese and 30 non-obese) and 41 control women (30 healthy and 11 healthy but obese)

16S rRNA
Decreased abundance of phylum Synergistetes in women with PCOS, Lactococcus was the characteristic gut microbiota in NG (non-obese with PCOS), while Coprococcus_2 in OG (obese with PCOS) and decrease in Tenericutes in non-obese women with PCOS.
Zhou L et al. [25] Case-Control study 18 obese patients with PCOS and 15 obese control women without PCOS 16S rRNA Decreased ratio of Firmicutes/Bacteroides as well as increased abundance of Fusobacteria (p = 0.022), while a reduced abundance of Tenericutes (p = 0.018).

Discussion
Our scoping review identified 25 articles on the gut microbiome in women with PCOS and women with EC, with most (23 articles) of the studies published on PCOS. Although the literature is growing in this field, we did not identify enough published studies, in particular, original research papers, to enable us to investigate whether gut dysbiosis may explain the increased EC risk in PCOS.
Within the studies identified, we were unable to identify any consistent patterns of the microbiome similarly present in studies on women with PCOS compared with women with EC. Although we found that the phylum Firmicutes was similarly decreased in women with PCOS and studies on women with EC, there was, however, significant variability within the studies identified making it highly likely that this may have arisen by chance.
Although we did not identify enough published studies to enable us to investigate whether gut dysbiosis may explain the increased EC risk in PCOS, previous studies on the microbiome in PCOS and EC suggest that it is not unreasonable to investigate the gut microbiome as a possible link between PCOS and EC. The major microbial changes observed in women with PCOS and its consequences are as follows: an increase in Escherichia and Shigella, which causes an alteration in the short-chain fatty acids, impacting metabolism, immune response, and gut barrier permeability [21,48]; an increase in Prevotellaceae, resulting in a profound and unfavorable inflammatory response to the patient [48][49][50]; and an increase in Bacteroides vulgates causing a subsequent reduction in the levels of glycodeoxycholic and tauroursodeoxycholic acid [48].
Boutriq S et al. showed that, normally, estrogen is first conjugated (inactivated) by the liver and this conjugated estrogen is transported to the intestine for its excretion. However, due to dysbiosis of the estrobolome, this conjugated/inactivated estrogen can be converted back to its active form by the process of deconjugation under the influence of certain enzymes produced by the gut microbiota (beta-glucuronidase), leading to high levels of activated estrogen in the blood [20]. Bacterial species responsible for the deconjugation of the conjugated-estrogen complex are Clostridia and Ruminococcacaeae. They are known to influence tumorigenesis by the process of deconjugation [51].
The findings from our scoping review, that the phylum Firmicutes was similarly decreased in women with PCOS and in studies on women with EC, are, however, inconsistent with the gut microbiome underpinning the association between PCOS and EC. This is because obesity, a known risk factor for EC and PCOS, is associated with an increase in the levels of Firmicutes with an increased ratio of Firmicutes to Bacteroidetes [52].
Whilst the strength of this scoping review was in its originality, the low number of publications identified was a limitation. Studies pertaining to the microbiome, specifically, the gut microbiota and its implications in gynecological cancers, are still developing, and this will explain the low number of articles identified, especially those related to the gut microbiome of EC patients. In addition, differences in the patients' characteristics across studies as well as a low sample size for most of the studies leading to low statistical power may create a bias. These may also explain the differences in findings across studies, which made comparative analysis challenging The role of an altered gut microbial diversity and composition as one of the mechanisms in the development of EC in those diagnosed with PCOS has hitherto remained unexplored. In this scoping review, we have assessed the existing literature in an attempt to address this pertinent research question. While the literature is growing in this field, we did not identify enough published studies to enable a conclusive answer regarding the potential association to be provided. Although our findings showed that the phylum Firmicutes was similarly decreased in women with PCOS and studies on women with EC, there was, however, significant variability within the studies identified, making it highly likely that this may have arisen by chance. This highlights the need for robust studies aimed at investigating the role of dysbiosis of the gut microbiome in the development of EC and PCOS. Importantly, such studies should investigate the potential role of dysbiosis in stimulating the mechanisms at play in the pathophysiology of EC in patients with PCOS. Furthermore, we speculate that the vaginal microbiota of these patients could also yield important information; although, none of the studies we identified investigated this. We, therefore, highlight this as a gap in the literature and recommend that future studies should incorporate the investigation of both gut and vaginal microbiomes to provide a holistic picture of the extent of dysbiosis in these patients. From a clinical perspective, these studies are important as the potential intervention of reversing dysbiosis via microbiome replacement approaches, whereby the use of probiotics and fecal transplantations could be of value in reducing the development of EC in patients with PCOS. Although data from this scoping review do not address such clinical applicability, the findings enabled us to identify gaps in the literature and areas for future research.

Conclusions
In conclusion, this scoping review has identified important gaps in the existing literature regarding the gut microbiome in EC and PCOS patients. Our findings indicate a need for more robust studies to address this important research question of the role of dysbiosis in PCOS patients and the link with EC with particular emphasis on elucidating the possible mechanisms involved.  Institutional Review Board Statement: Not applicable because this study did not involve humans or animals.