Androgen Insensitivity Syndrome with Bilateral Gonadal Sertoli Cell Lesions, Sertoli–Leydig Cell Tumor, and Paratesticular Leiomyoma: A Case Report and First Systematic Literature Review

Androgen insensitivity syndrome (AIS) is a rare Mendelian disorder caused by mutations of the androgen receptor (AR) gene on the long arm of the X chromosome. As a result of the mutation, the receptor becomes resistant to androgens, and hence, karyotypically male patients (46,XY) carry a female phenotype. Their cryptorchid gonads are prone to the development of several types of tumors (germ cell, sex cord stromal, and others). Here, we report a 15-year-old female-looking patient with primary amenorrhea who underwent laparoscopic gonadectomy. Histologically, the patient’s gonads showed Sertoli cell hamartomas (SCHs) and adenomas (SCAs) with areas of Sertoli–Leydig cell tumors (SLCTs) and a left-sided paratesticular leiomyoma. Rudimentary Fallopian tubes were also present. The patient’s karyotype was 46,XY without any evidence of aberrations. Molecular genetic analysis of the left gonad revealed two likely germline mutations—a pathogenic frameshift deletion in the AR gene (c.77delT) and a likely pathogenic missense variant in the RAC1 gene (p.A94V). Strikingly, no somatic mutations, fusions, or copy number variations were found. We also performed the first systematic literature review (PRISMA guidelines; screened databases: PubMed, Scopus, Web of Science; ended on 7 December 2023) of the reported cases of patients with AIS showing benign or malignant Sertoli cell lesions/tumors in their gonads (n = 225; age: 4–84, mean 32 years), including Sertoli cell hyperplasia (1%), Sertoli cell nodules (6%), SCHs (31%), SCAs (36%), Sertoli cell tumors (SCTs) (16%), and SLCTs (4%). The few cases (n = 14, 6%; six SCAs, four SCTs, two SLCTs, and two SCHs) with available follow-up (2–49, mean 17 months) showed no evidence of disease (13/14, 93%) or died of other causes (1/14, 7%) despite the histological diagnosis. Smooth muscle lesions/proliferations were identified in 19 (8%) cases (including clearly reported rudimentary uterine remnants, 3 cases; leiomyomas, 4 cases). Rudimentary Fallopian tube(s) were described in nine (4%) cases. Conclusion: AIS may be associated with sex cord/stromal tumors and, rarely, mesenchymal tumors such as leiomyomas. True malignant sex cord tumors can arise in these patients. Larger series with longer follow-ups are needed to estimate the exact prognostic relevance of tumor histology in AIS.

Abstract: Androgen insensitivity syndrome (AIS) is a rare Mendelian disorder caused by mutations of the androgen receptor (AR) gene on the long arm of the X chromosome.As a result of the mutation, the receptor becomes resistant to androgens, and hence, karyotypically male patients (46,XY) carry a female phenotype.Their cryptorchid gonads are prone to the development of several types of tumors (germ cell, sex cord stromal, and others).Here, we report a 15-year-old female-looking patient with primary amenorrhea who underwent laparoscopic gonadectomy.Histologically, the patient's gonads showed Sertoli cell hamartomas (SCHs) and adenomas (SCAs) with areas of Sertoli-Leydig cell tumors (SLCTs) and a left-sided paratesticular leiomyoma.Rudimentary Fallopian tubes were also present.The patient's karyotype was 46,XY without any evidence of aberrations.Molecular genetic analysis of the left gonad revealed two likely germline mutations-a pathogenic frameshift deletion in the AR gene (c.77delT) and a likely pathogenic missense variant in the RAC1 gene (p.A94V).Strikingly, no somatic mutations, fusions, or copy number variations were found.We also performed the first systematic literature review (PRISMA guidelines; screened databases: PubMed, Scopus, Web of Science; ended on 7 December 2023) of the reported cases of patients with AIS showing benign or malignant Sertoli cell lesions/tumors in their gonads (n = 225; age: 4-84, mean 32 years), including Sertoli cell hyperplasia (1%), Sertoli cell nodules (6%), SCHs (31%), SCAs (36%), Sertoli cell tumors (SCTs) (16%), and SLCTs (4%).The few cases (n = 14, 6%; six SCAs, four SCTs, two SLCTs, and two SCHs) with available follow-up (2-49, mean 17 months) showed no evidence of disease (13/14, 93%) or died of other causes (1/14, 7%) despite the histological diagnosis.Smooth muscle lesions/proliferations were identified in 19 (8%) cases (including clearly reported rudimentary uterine remnants, 3 cases; leiomyomas, 4 cases).Rudimentary Fallopian tube(s) were described in nine (4%) cases.Conclusion: AIS may be associated with sex cord/stromal tumors and, rarely, mesenchymal tumors such as leiomyomas.True malignant sex cord tumors can arise in these patients.Larger series with longer follow-ups are needed to estimate the exact prognostic relevance of tumor histology in AIS.

Introduction
Androgen insensitivity syndrome (AIS) is a rare disease caused by mutations of the androgen receptor (AR) gene located on the long arm of the X chromosome (Xq 11-12) [1][2][3][4].In a study, the incidence of AIS was 1:99,000 in genetically confirmed males [4].However, not all the reported studies included a molecular proof of the diagnosis, and there is a need for new clinic pathological data to elaborate proper conceptual approaches to this disease.
Clinically, these patients are genotypically males with a 46,XY karyotype, but they phenotypically present as females lacking the Müllerian derivates.Indeed, female-looking breasts and vulva are usually normal, while body hair could be less pronounced.The gonads are usually cryptorchid, identified in abdomino-pelvic or inguinal sites, while the uterus is absent and the vagina is typically blind and shortened; about 10% of patients may have one or both residual or well-formed Fallopian tubes [2].
Due to various types of mutations occurring in the locus of the AR gene (insertions and deletions), the phenotypical manifestations of the AIS may differ substantially; so, the disease itself is divided into broad entities, including complete (CAIS) and partial (PAIS) forms [3].Moreover, the clinical manifestations depend on the patient's age.In infants and pre-menarchal cases, the disease may be suspected during casual ultrasound investigations of the pelvis carried out for different reasons or manifested by inguinal hernias, including undescended testes.In menarchal patients, primary amenorrhea is the basic manifestation of the process.
Finally, single or multiple hamartomas or tumors may rarely develop in the cryptorchid gonads, including malignant neoplasms, which can present as pelvic masses or even cause distant metastases.In particular, various lesions containing Sertoli and/or Leydig cells have been described in patients with AIS, considered as hyperplastic, hamartomatous, or neoplastic, including Sertoli cell hyperplasia (SCHYP), Sertoli cell nodules (SCNs), Sertoli cell hamartomas (SCHs), Sertoli cell adenomas (SCAs), Sertoli cell tumors not otherwise specified (SCTs, NOS) or Sertoli-Leydig cell tumors (SLCTs); single or multiple, uni or bilateral, these lesions may appear as grossly detectable nodules or incidental histological findings.However, their diagnostic criteria may be subtle, challenging, and sometimes questionable in some cases, and their frequently small size and bland histology may result in a favorable prognosis; unfortunately, the rarity of these lesions and the lack of a systematic literature review on this topic have represented biases for further considerations [5][6][7].
We here present an unusual case of bilateral gonadal SCHs with associated SCAs, SLCT, and paragonadal leiomyoma in a patient with molecular confirmation of CAIS.Moreover, we performed the first systematic literature review of the features of Sertoli cell lesions in patients with AIS.

Case Description
A 15-year-old female-looking patient was referred to the gynecologist due to primary amenorrhea.Her maternal grandaunt had uterine agenesis.The patient was born after unremarkable gestation and delivery (unattended childbirth, timely delivered).At the age of 2 years, she underwent surgery for umbilical and left inguinal hernias; no nodules were found in the hernia sacs.At presentation, the patient did not have any evidence of chronic diseases and did not take any drugs on a regular basis; previous drug history was negative as well.
Upon clinical examination, the patient had a typical feminine habitus, showing welldeveloped breasts with pale nipples.Pubic hair was absent.A 6 cm long vaginal stump with a blind end was found during the gynecological exam.During the bimanual rectalabdominal investigation, the uterine body was not detected in the pelvis.Bilateral nodules (presumable gonads)-each of 4 cm in maximum size-were palpable at a high level of the pelvis, almost reaching the pelvic brim.Any other remarkable finding was identified on clinical examination, except for myopia of medium level.
Ultrasonographic investigation confirmed the uterine agenesis and identified the gonads at the entrance of the pelvis.The right gonad measured 31 × 12 × 11 mm, while the left one measured 34 × 18 × 21 mm, showing a nodule measuring 18 × 14 mm at one of the poles (Figure 1).Upon clinical examination, the patient had a typical feminine habitus, showing welldeveloped breasts with pale nipples.Pubic hair was absent.A 6 cm long vaginal stump with a blind end was found during the gynecological exam.During the bimanual rectalabdominal investigation, the uterine body was not detected in the pelvis.Bilateral nodules (presumable gonads)-each of 4 cm in maximum size-were palpable at a high level of the pelvis, almost reaching the pelvic brim.Any other remarkable finding was identified on clinical examination, except for myopia of medium level.
Ultrasonographic investigation confirmed the uterine agenesis and identified the gonads at the entrance of the pelvis.The right gonad measured 31 × 12 × 11 mm, while the left one measured 34 × 18 × 21 mm, showing a nodule measuring 18 × 14 mm at one of the poles (Figure 1).The serum testosterone level (15 nmol/L) exceeded the typical normal values for female patients (2.3 nmol/L), but it corresponded to the normal rates for boys of the same age.The serum levels of Luteinizing hormone (LH) (37.2 IU/L; normal values: 2.4-8.3IU/L) and anti-Müllerian hormone (AMH) (600 ng/mL; normal values: 10.6 ng/mL) were increased.
The patient underwent laparoscopic bilateral gonadectomy.During the procedure, the undescended gonads were found at the pelvic sidewalls.
Upon histological examination, both gonads revealed multiple nodules arising in a hamartomatous background (Figure 2).
Microscopically, the hamartomatous background was composed of an extensive proliferation of Sertoli, Leydig, and stromal cells, which were identified in variable proportions in different areas and surrounded by a loose or fibromatous stroma (Figure 3); in some areas, ovarian-type stroma was also identified.The serum testosterone level (15 nmol/L) exceeded the typical normal values for female patients (2.3 nmol/L), but it corresponded to the normal rates for boys of the same age.The serum levels of Luteinizing hormone (LH) (37.2 IU/L; normal values: 2.4-8.3IU/L) and anti-Müllerian hormone (AMH) (600 ng/mL; normal values: 10.6 ng/mL) were increased.
The patient underwent laparoscopic bilateral gonadectomy.During the procedure, the undescended gonads were found at the pelvic sidewalls.
Upon histological examination, both gonads revealed multiple nodules arising in a hamartomatous background (Figure 2).
Microscopically, the hamartomatous background was composed of an extensive proliferation of Sertoli, Leydig, and stromal cells, which were identified in variable proportions in different areas and surrounded by a loose or fibromatous stroma (Figure 3); in some areas, ovarian-type stroma was also identified.The Sertoli cells were arranged in compressed tubules separated by fibrous bands with foci of hyalinization and edema.Many tubules were wrapped by concentric layers of fibroblasts.The Sertoli cells were arranged in compressed tubules separated by fibrous ban with foci of hyalinization and edema.Many tubules were wrapped by concentric layer fibroblasts.The Sertoli cells were arranged in compressed tubules separated by fibrous bands with foci of hyalinization and edema.Many tubules were wrapped by concentric layers of fibroblasts.
Leydig cells were dispersed throughout the tubules in varying proportions.They were polygonal in shape, showing sharp cellular borders, broad granular light-brown cytoplasm, and centrally located round nuclei with inconspicuous nucleoli.We detected Reinke crystalloid in a small number of Leydig cells.
Spermatogonia were not found in any tubules of the gonads, in spite of careful search.The transition from the hamartomatous background to the tumoral nodules manifested with significant enlargement of the Sertoli cells, which often acquired a round acinar-like, complex/confluent, or solid pattern (Figures 4 and 5).
Leydig cells were dispersed throughout the tubules in varying proportions.They were polygonal in shape, showing sharp cellular borders, broad granular light-brown cytoplasm, and centrally located round nuclei with inconspicuous nucleoli.We detected Reinke crystalloid in a small number of Leydig cells.
Spermatogonia were not found in any tubules of the gonads, in spite of careful search.
The transition from the hamartomatous background to the tumoral nodules manifested with significant enlargement of the Sertoli cells, which often acquired a round acinar-like, complex/confluent, or solid pattern (Figures 4 and 5).All the nodules were unencapsulated.Most of them were well circumscribed, while one of the nodules in the left gonad seemed to show a more cellular and complex/diffuse pattern of growth, worth a diagnosis of a sex cord-stromal tumor; in our opinion, we favored a well-differentiated SLCT (Figures 6 and 7).
The Sertoli cells varied in size from cuboid to high prismatic, usually arranged in cords and solid tubules; depending on the areas, their cytoplasm was slightly eosinophilic, dense, or with signs of gradual vacuolization starting from apical to basal compartments.We also encountered some lipid-rich Sertoli cells.The nuclear/cytoplasmic ratio of tumor cells was very low.The nuclei were basally or centrally located, round in shape, with pale, finely dispersed chromatin and small, sometimes eosinophilic nucleoli.Some Leydig cells seemed to be intermixed.
The mitotic activity was still low in any areas of the sex cord-stromal tumor, as well as in the well-delimited nodules and in the hamartomatous background, revealing a maximum of 2 mitoses per 10 high-power field (HPF).Necrosis or other signs of discirculatory events were absent in any areas.All the nodules were unencapsulated.Most of them were well circumscribed, while one of the nodules in the left gonad seemed to show a more cellular and complex/diffuse pattern of growth, worth a diagnosis of a sex cord-stromal tumor; in our opinion, we favored a well-differentiated SLCT (Figures 6 and 7).
The Sertoli cells varied in size from cuboid to high prismatic, usually arranged in cords and solid tubules; depending on the areas, their cytoplasm was slightly eosinophilic, dense, or with signs of gradual vacuolization starting from apical to basal compartments.We also encountered some lipid-rich Sertoli cells.The nuclear/cytoplasmic ratio of tumor cells was very low.The nuclei were basally or centrally located, round in shape, with pale, finely dispersed chromatin and small, sometimes eosinophilic nucleoli.Some Leydig cells seemed to be intermixed.
The mitotic activity was still low in any areas of the sex cord-stromal tumor, as well as in the well-delimited nodules and in the hamartomatous background, revealing a maximum of 2 mitoses per 10 high-power field (HPF).Necrosis or other signs of discirculatory events were absent in any areas.Immunohistochemical stainings specific for germ cells (Oct-4, PLAP, c-kit) were negative in all the sex cord cells and did not reveal any in situ or invasive germ cell neoplasia.Conversely, marked positive reaction for inhibin-α, calretinin, and Melan-A was disclosed in Sertoli and Leydig cells; the proliferative index, according to the Ki67 level, was low, up to 4% mainly in basally located Sertoli cells (Figure 8).
The 1.5 cm paragonadal nodule (Figures 1, 2 and 9) was represented by bland, elongated cells arranged in fascicular structures and reminiscent of a smooth muscle neoplasm; mitotic activity and necrotic areas were not found.Immunohistochemical stainings specific for germ cells (Oct-4, PLAP, c-kit) were negative in all the sex cord cells and did not reveal any in situ or invasive germ cell neoplasia.Conversely, marked positive reaction for inhibin-α, calretinin, and Melan-A was disclosed in Sertoli and Leydig cells; the proliferative index, according to the Ki67 level, was low, up to 4% mainly in basally located Sertoli cells (Figure 8).
The 1.5 cm paragonadal nodule (Figures 1, 2 and 9) was represented by bland, elongated cells arranged in fascicular structures and reminiscent of a smooth muscle neoplasm; mitotic activity and necrotic areas were not found.
Bilateral rudimentary fallopian tubes were also identified in the paragonadal tissues (Figure 10).Cytogenetic analysis revealed a 46,XY karyotype.Next-generation sequencing (NGS) analysis of the tumor tissue (Table 1) searched for DNA mutations of 523 genes and 127 microsatellite loci by using the NextSeq 550 instrument (Illumina, San Diego, CA, USA) with the TruSight Oncology 500 DNA+RNA reagent kit (Illumina, San Diego, CA, USA) in accordance with the instructions of the manufacturer.NGS analysis identified a pathogenic variant in the AR gene (frameshift deletion generating a stop codon at position 34) and another likely pathogenic missense variant in the RAC1 gene, while no mutations were detected in any of the other 521 genes tested; both variants represented likely germline mutations.Strikingly, no somatic mutations, translocations, or copy number variations were identified in the tumor sample.Pathogenic germline AR variant confirms the diagnosis of classic AIS.Cytogenetic analysis revealed a 46,XY karyotype.Next-generation sequencing (NGS) analysis of the tumor tissue (Table 1) searched for DNA mutations of 523 genes and 127 microsatellite loci by using the NextSeq 550 instrument (Illumina, San Diego, CA, USA) with the TruSight Oncology 500 DNA+RNA reagent kit (Illumina, San Diego, CA, USA) in accordance with the instructions of the manufacturer.NGS analysis identified a pathogenic variant in the AR gene (frameshift deletion generating a stop codon at position 34) and another likely pathogenic missense variant in the RAC1 gene, while no mutations were detected in any of the other 521 genes tested; both variants represented likely germline mutations.Strikingly, no somatic mutations, translocations, or copy number variations were identified in the tumor sample.Pathogenic germline AR variant confirms the diagnosis of classic AIS.

Systematic Literature Review Method
We conducted a systematic literature review according to the PRISMA ("Preferred Reporting Items for Systematic Reviews and Meta-Analyses") guidelines (http://www.prisma-statement.org/;accessed on 7 December 2023) to identify the previously reported cases of Sertoli cell lesions in patients with AIS (Figure 11).
We conducted a systematic literature review according to the PRISMA ("Preferred Reporting Items for Systematic Reviews and Meta-Analyses") guidelines (http://www.prisma-statement.org/;accessed on 7 December 2023) to identify the previously reported cases of Sertoli cell lesions in patients with AIS (Figure 11).We searched for ("androgen insensitivity syndrome" OR "androgen resistance syndrome" OR "testicular feminization syndrome" OR "androgen receptor deficiency" OR "androgen insensitive syndrome" OR "Morris syndrome") AND (Sertoli OR Sertoli-Leyding OR "sex-cord" OR "sex cord") AND (tumor OR tumors OR tumour OR tumours OR nodule OR nodules OR adenoma OR adenomas OR hamartoma OR hamartomas) in the PubMed (all fields; 98 results; https://pubmed.ncbi.nlm.nih.gov,accessed on 7 December 2023), Scopus (Title/Abstract/Keywords; 119 results; https://www.scopus.com/home.uri,accessed on 7 December 2023) and Web of Science (Topic/Title; 74 results; https://login.webofknowledge.com,accessed on 7 December 2023) databases.No limitations or additional filters were set.The bibliographic research ended on 7 December 2023.We applied the following criteria: We searched for ("androgen insensitivity syndrome" OR "androgen resistance syndrome" OR "testicular feminization syndrome" OR "androgen receptor deficiency" OR "androgen insensitive syndrome" OR "Morris syndrome") AND (Sertoli OR Sertoli-Leyding OR "sex-cord" OR "sex cord") AND ( Two independent authors removed the duplicates and checked the titles and abstracts of all the retrieved results (n = 152).After applying the eligibility, inclusion, and exclusion criteria, they selected 82 relevant eligible papers; 74 articles were retrieved in full-text format, and their reference lists were manually examined to check for other potentially relevant studies [1,, while only abstracts or titles were available for the remaining 8 papers [79][80][81][82][83][84][85][86].Five of these eight articles reported relevant data in their abstract and were included in further analysis [79][80][81][82][83], while three references were excluded due to insufficient data, according to the inclusion/exclusion criteria [84][85][86].Finally, 79 articles were included in our study [1,, although 8 papers retrieved in full-text format were just included for the analysis of a few parameters, as they reported partially aggregated data [6,[72][73][74][75][76][77][78].The extracted results were checked and confirmed by two other authors.Data collection was study-and case-related.Categorical variables were analyzed as frequencies and percentages, whereas continuous variables were by ranges and mean values.Meta-analysis was not performed according to the few available data for comparisons, especially concerning follow-up.This study was not recorded in PROSPERO (https://www.crd.york.ac.uk/prospero/, accessed on 27 January 2024).
Only one patient showed multiple SCTs per gonad (three right and one left) [23], while the other cases with available information reported one SCT per gonad.
One was associated with intratubular germ cell neoplasia and Leydig cell hyperplasia [37], and one with left inguinal serous carcinoma of the tunica vaginalis (metastatic with carcinosis) [50].Follow-up was available in only four cases; three patients showed no evidence of disease 6 to 18 (mean 12) months after surgery [19,37,53], while the remaining patient recurred and died of another cause (inguinal serous carcinoma) after 24 months [50].
Finally, five sex cord-stromal tumors with Sertoli cells (four CAIS and one AIS, NOS) were reported, including two malignant cases [6,26]; follow-up was not available.
Another paper found expression of CYP11A1 and CYP17A1 in Leydig cells and HSD17B3 expression only in Sertoli cells of CAIS gonads [20].In the control normal testes, CYP11A1, CYP17A1, and HSD17B3 were detected only in Leydig cells; CYP19A1 was expressed by Leydig and Sertoli cells in the gonads of patients and controls.LHCGR was highly expressed by Leydig cells, while FSHR was not localized in CAIS gonads [20].

Discussion
Molecular analysis and the identification of novel blood and tissue biomarkers are increasingly gaining a crucial role as diagnostic, prognostic, and/or predictive tools in managing tumors arising from various sites, including the genito-urinary and gynecological areas .
AIS is a rare disease associated with the derangement of the AR gene on the long arm of chromosome 17(q11-12).About 1000 mutations of this gene have been identified to date, and their different interconnections cause various clinical manifestations from partial to complete AIS [1,[110][111][112][113][114].In our series, data were available for just 11 patients; most of the cases were missense mutations or deletions [1,12,15,20,26,36,37,51,58].In our review, we reported a new AR gene mutation associated with the development of histologically confirmed Sertoli cell lesions, even if the molecular analysis was rarely performed in the cases included in our study.
Recently, an intermediate so-called mild type has been distinguished.Such patients produce the Müllerian-inhibiting factor (MIF), which hinders the development of Müllerian derivates.In rare cases, for some unknown reasons, the Fallopian tube may be formed, as in our case.To our review, rudimentary mono-or bilateral Fallopian tube(s) were described in 4% of the cases [10,28,34,36,59,71].One may hypothesize that in such patients, either the concentration of MIF is low or its receptors are absent in some embryonal structures.Our patient had a well-developed uterine tube with functionally full-fledged mucosa, represented by all types of tubal epithelium.Such patients have female phenotypes since the androgens produced by testicular tissue are completely converted into estrogens.This fact was proven by the corresponding test in our case.
The most important symptom of the disease is primary amenorrhea.For the proper diagnosis, we could stress the importance of the detection of gonadal nodules in inguinal hernial sacs, but cryptorchidism can also be asymptomatic [119].In our case, the clinicians overlooked this possibility, although the parents consulted the patient at the age of two years.
Tubular hamartomas (SCHs, Sertoli-Leydig hamartomas) are frequently bilateral and multiple, white, well-delimited, encapsulated testicular nodules composed of small and solid seminiferous tubules/cords with immature Sertoli cells and numerous interspersed Leydig cells; the tubular wall shows hyalinization, lacking elastic fibers and spermatogonia are absent or isolated in some lesions [5].
SCAs are usually solitary tumors, but they can be associated with SCHs.They consist of small, infantile seminiferous tubules but lacking germ cells and peritubular myofibroblasts.Tubules are arranged in a compact back-to-back pattern, and the basal lamina can be very prominent, sometimes forming a thick ring around small groups of Sertoli cells.Leydig cells or ovarian-like stroma are absent in the scant interstitium [5][6][7].
Like SCAs and SCHs, SCNs (Picks adenoma) are unencapsulated and composed of Sertoli cells arranged in well-formed tubules or cords that vaguely resemble immature Sertoli cells.The nuclei are bland hyperchromatic, oval to round in shape, frequently stratified with variable eosinophilic (hyaline) intraluminal material.The basal membrane of tubules can be thickened and invaginated within the lumen, mimicking Call-Exner bodies.Germ cells are absent or rarely admixed with immature Sertoli cells.SCHs, SCAs, and SCNs differ in size, as the first 2 entities are usually larger, while SCNs are frequently small, multifocal, and incidental.Unlike SCNs, SCHs and SCAs seem to show no pseudostratification nor intratubular nodular projections [5][6][7].Clusters of Leydig cells can be found between the tubules of SCNs (at least focally), but a non-neoplastic proliferation of Leydig cell hyperplasia is rarely identified [5][6][7].
SCTs, NOS usually occurs in clusters usually lacking intratubular arrangement without prominent internalized basement membrane component and commonly lack fetal phenotype.The tumor nuclei are usually bland (round to ovoid, small hyperchromatic), but sometimes worrisome prominent nucleoli may appear; the cytoplasm is frequently clear and abundant, but it may be foamy, lipidized, eosinophilic or scant; hyaline globules are common.Tumor cells are arranged in a variable combination of tubular, cord, tubule-papillary, macro-or micro-cystic, nested whorled, trabecular, retiform, solid, and pseudopapillary patterns.The tumor stroma may show basement membrane-like material around tubules, or it could be sclerotic (if >50% of the tumor, a sclerotic variant could be diagnosed), angiomatous, myxoid, or edematous; inflammatory cells may be present.Malignant SCTs are frequently > 5 cm in size and poorly circumscribed, with extratesticular extension and necrotic areas [5][6][7].
In our case, nine SLCTs were reported (4%), occurring only in patients with CAIS (age range: 15-80 years, mean 31 years) [1,12,30,33,34,41,45,64].SLCTs could be well, moderately, or poorly differentiated; none of the reported cases seemed to be intermediate or poorly differentiated.Well-differentiated SLCTs usually show open or compressed Sertoli cell tubules, admixed with clusters of Leydig cells in the intervening stroma, without significant atypia or mitotic activity.We have to, however, disclose that the differential diagnosis between the abovementioned sex cord entities in patients with AIS could be not so easy, and some reported diagnoses may indeed represent the same kind of lesions; a spectrum of entities may also be possible.
Interestingly, all the cases of our analysis (six SCA [11,24,63,65,66,68], four SCTs [19,37,50,53], two SLCTs [33,34], and two SCH [52,59]) who had been followed-up for 2 to 49 (mean 17) months showed no evidence of disease (13/14, 93%) or died of another cause (1/14, 7%) [50], despite the histological diagnosis.Even if this result could question the neoplastic nature of some lesions, longer follow-up studies of larger series should be performed, as only 6% of the cases analyzed in our series had available follow-up data.We feel that it could be imprudent to consider all the sex cord lesions arising from patients with AIS without a malignant potential; true malignant tumors may occur, indeed.
Including our paratesticular case, four clearly reported leiomyomas had been described in patients with AIS with gonadal Sertoli cell lesions [28,38,46].The cells were positive for markers of smooth muscle differentiation and showed low signs of proliferative activity without any other malignant features.
Nuclear atypia, necrosis, or significant mitotic activity were not reported in the literature cases, as well as unusual areas of adipose, chondroid, or osseous metaplasia that may rarely be identified in smooth muscle neoplasms arising in other sites [148][149][150][151].
Rudimentary uterine remnants or areas of smooth muscle hyperplasia were described as well; however, only 8% of cases analyzed in our systematic literature review were associated with smooth muscle proliferations/tumors [6,10,21,28,30,38,39,43,46,56,59,69,73,78,83].It is possible that this variety of terminology may reflect the same type of lesions as it is not completely clear if they are hamartomatous/embryological rests or true benign neoplasms; inter-observer variability may represent a diagnostic bias.
The post-surgery rehabilitation of such patients is very important and must be preceded by careful psychological preparation for further surgical intervention, including plastic operation of the vagina and general personological orientation of the patient.
Limitations of our analysis include the inability to compare the data of several cases, especially regarding follow-up and prognostic information; indeed, Sertoli cell lesions and AIS are both rare pathological findings, and they are more infrequently associated.Multicenter large series are lacking.Finally, the diagnostic criteria of some Sertoli cell lesions may be subtle and variably interpreted with a potential diagnostic overlapping among these different entities, thus maybe representing a diagnostic bias.Conversely, we feel that a point of strength of our study is represented by the detailed morphological, immunohistochemical, and genetic analysis of this very rare disease.We also reported the results of the first systematic literature review on this topic.

Conclusions
In conclusion, the androgen insensitivity syndrome in its complete form (CAIS) may be associated not only with sex cord-stromal tumors (such as SCTs or SLCTs) but also with rare mesenchymal tumors.Our case represents the fourth description in the literature of leiomyomas.The patient also belongs to the 10% of cases with preserved Fallopian tubes, the resistance of which to the Müllerian-inhibiting factor is still unclear.
A larger centralized series with longer follow-up should study the prognostic relevance of Sertoli cell lesions in patients with AIS.

Figure 5 .
Figure 5. Histological examination.Details of other nodules showing a diffuse or more confluent growth of Sertoli cells with less evident Leydig cells.Some lumens were identified within a more complex growth (B) (hematoxylin and eosin; (A): 10×; (B): 10×) (previously unpublished original photos).

Figure 5 .
Figure 5. Histological examination.Details of other nodules showing a diffuse or more confluent growth of Sertoli cells with less evident Leydig cells.Some lumens were identified within a more complex growth (B) (hematoxylin and eosin; (A): 10×; (B): 10×) (previously unpublished original photos).

J 33 Figure 5 .
Figure 5. Histological examination.Details of other nodules showing a diffuse or more confluent growth of Sertoli cells with less evident Leydig cells.Some lumens were identified within a more complex growth (B) (hematoxylin and eosin; (A): 10×; (B): 10×) (previously unpublished original photos).

Figure 10 .
Figure 10.The rudimentary Fallopian tubes revealed a well-shaped wall of the uterine tube containing all three types of epithelial cells in the mucosa (Hematoxylin and eosin, 4×; previously unpublished, original photo).

Figure 10 .
Figure 10.The rudimentary Fallopian tubes revealed a well-shaped wall of the uterine tube containing all three types of epithelial cells in the mucosa (Hematoxylin and eosin, 4×; previously unpublished, original photo).

Figure 11 .
Figure 11.Systematic literature review: PRISMA flow chart.Our retrospective observational study was conducted via the PICO process: • Populations: human patients with AIS with a diagnosis of a gonadal Sertoli cell lesion; • Intervention: any; • Comparison: none; • Outcomes: clinical outcomes (status at last follow-up, and survival and recurrence rates).

Figure 11 .
Figure 11.Systematic literature review: PRISMA flow chart.Our retrospective observational study was conducted via the PICO process: • Populations: human patients with AIS with a diagnosis of a gonadal Sertoli cell lesion; • Intervention: any; • Comparison: none; • Outcomes: clinical outcomes (status at last follow-up, and survival and recurrence rates).
ACMG: American College of Medical Genetics and Genomics.
ACMG: American College of Medical Genetics and Genomics.

Table 2 .
Results of series with partially aggregated data.

Table 3 .
Clinical features of patients with non-aggregated data.

Table 4 .
Clinic-pathologic features of patients with matched data.

Table 5 .
Cases with details of mutations in AR gene.