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Tuberous sclerosis complex (TSC) tumors are presently incurable despite a cytostatic response to mTOR pathway inhibition because recurrence of disease occurs after treatment is discontinued. Here, we explored the hypothesis that inhibiting tyrosine kinase activity in mesenchymal lineage-specific platelet-derived growth factor receptor β (PDGFRβ) signaling in TSC tumors is cytocidal and attenuates tumorigenesis at significantly higher levels than treatment with an mTOR inhibitor. Rapamycin-induced versus tyrosine kinase inhibitor (TKI)-induced renal angiomyolipoma (AML) and pulmonary lymphangioleiomyomatosis (LAM) tumor cells were comparatively analyzed using cell survival assays, RNA sequencing, and bioinformatics to distinguish tumoricidal mechanisms adopted by each drug type. The efficacy of imatinib therapy was validated against spontaneously developing renal cystadenomas in tuberous sclerosis Tsc2+/- mouse models (C57BL/6J mice; N = 6; 400 mg/kg/d; oral gavage) compared with Tsc2+/- mice treated with PBS (C57BL/6J mice; N = 6). Our study revealed that TKIs imatinib and nilotinib were cytocidal to both pulmonary LAM and renal AML cell cultures through the downregulation of the glycoprotein GPVI pathway and resultant disruption in mitochondrial permeability, increased cytosolic cytochrome C, and caspase 3 activation. Importantly, renal tumor growth was significantly attenuated in imatinib-treated Tsc2+/- mice compared with PBS treatment. The preclinical studies reported here provide evidence documenting the effectiveness of TKIs in limiting LAM and AML cell growth and viability with important clinical potential. Furthermore, these drugs elicit their effects by targeting a PDGF pathway-dependent apoptotic mechanism supporting the investigation of these drugs as a novel class of TSC therapeutics., (©2023 The Authors; Published by the American Association for Cancer Research.)

Tuberous sclerosis complex (TSC) is a rare autosomal dominant disorder involving multiple organ systems. TSC2 gene plays an important role in the development of TSC. The most common kidney manifestation of TSC is renal angiomyolipoma (RAML). TSC-RAML is more likely to be bilateral multiple tumors and tends to destroy the renal structure and damages renal function severely. As a result, patients with TSC-RAML often miss the opportunity for surgical treatment when TSC-RAML is diagnosed, causing difficulty in obtaining tumor specimens through surgery. Due to this difficulty, model cell lines must be constructed for scientific research. In this paper, TSC2 was knocked out in NIH-3T3 cell lines by CRISPR/Cas9 system. PCR, WB and mTOR inhibitor drug sensitivity test showed that the TSC2 knockout NIH-3T3 cells were successfully constructed. The ability of proliferation and invasion in TSC2 KO NIH-3T3 cells were higher than those in wild type group. The constructed KO cell line lay the foundation for further study of TSC., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Tuberous sclerosis complex (TSC) is an autosomal dominant disease characterized by multiorgan hamartomas, including renal angiomyolipomas and pulmonary lymphangioleiomyomatosis (LAM). TSC2 deficiency leads to hyperactivation of mTOR Complex 1 (mTORC1), a master regulator of cell growth and metabolism. Phospholipid metabolism is dysregulated upon TSC2 loss, causing enhanced production of lysophosphatidylcholine (LPC) species by TSC2-deficient tumor cells. LPC is the major substrate of the secreted lysophospholipase D autotaxin (ATX), which generates two bioactive lipids, lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P). We report here that ATX expression is upregulated in human renal angiomyolipoma-derived TSC2-deficient cells compared with TSC2 add-back cells. Inhibition of ATX via the clinically developed compound GLPG1690 suppressed TSC2-loss associated oncogenicity in vitro and in vivo and induced apoptosis in TSC2-deficient cells. GLPG1690 suppressed AKT and ERK1/2 signaling and profoundly impacted the transcriptome of these cells while inducing minor gene expression changes in TSC2 add-back cells. RNA-sequencing studies revealed transcriptomic signatures of LPA and S1P, suggesting an LPA/S1P-mediated reprogramming of the TSC lipidome. In addition, supplementation of LPA or S1P rescued proliferation and viability, neutral lipid content, and AKT or ERK1/2 signaling in human TSC2-deficient cells treated with GLPG1690. Importantly, TSC-associated renal angiomyolipomas have higher expression of LPA receptor 1 and S1P receptor 3 compared with normal kidney. These studies increase our understanding of TSC2-deficient cell metabolism, leading to novel potential therapeutic opportunities for TSC and LAM. SIGNIFICANCE: This study identifies activation of the ATX-LPA/S1P pathway as a novel mode of metabolic dysregulation upon TSC2 loss, highlighting critical roles for ATX in TSC2-deficient cell fitness and in TSC tumorigenesis., (©2020 American Association for Cancer Research.)

We discovered that 90.3% of patients with angiomyolipomas, lymphangioleiomyomatosis (LAM), and tuberous sclerosis complex (TSC) carry the arginine variant of codon 72 (R72) of TP53 and that R72 increases the risk for angiomyolipoma. R72 transactivates NOTCH1 and NODAL better than the proline variant of codon 72 (P72); therefore, the expression of NOTCH1 and NODAL is increased in angiomyolipoma cells that carry R72. The loss of Tp53 and Tsc1 within nestin-expressing cells in mice resulted in the development of renal cell carcinomas (RCC) with high Notch1 and Nodal expression, suggesting that similar downstream mechanisms contribute to tumorigenesis as a result of p53 loss in mice and p53 polymorphism in humans. The loss of murine Tp53 or expression of human R72 contributes to tumorigenesis via enhancing epithelial-to-mesenchymal transition and motility of tumor cells through the Notch and Nodal pathways. IMPLICATIONS: This work revealed unexpected contributions of the p53 polymorphism to the pathogenesis of TSC and established signaling alterations caused by this polymorphism as a target for therapy. We found that the codon 72 TP53 polymorphism contributes to TSC-associated tumorigenesis via Notch and Nodal signaling., (©2019 American Association for Cancer Research.)

Tuberous sclerosis complex (TSC) is a genetic disorder characterized by tumor formation in multiple organs, with over 80% of TSC patients developing angiomyolipomas (TSC-AMLs). However, the molecular events that contribute to TSC-AMLs are not well understood. Recent reports have demonstrated that microRNAs (miRNAs) are critical in TSC cortical tubers. However, little is known about the role of miRNAs in TSC-AMLs. In the current study, we analyzed changes in the miRNA and mRNA profiles in TSC-AMLs and matched normal adjacent tissues. A total of 15 differentially expressed miRNAs and 2664 mRNAs were identified. Using quantitative real-time PCR, we confirmed the results of the miRNA and mRNA profile experiments. Through bioinformatic analysis and luciferase reporter assays, we found that BCL2L11, an apoptotic activator, was the direct target of miR-9-5p, miR-124-3p, and miR-132-3p. Engineered expression of miR-9-5p, miR-124-3p, or miR-132-3p significantly regulated proliferation and apoptosis in Tsc2 -/- cells. Manipulated expression of BCL2L11 also led to proliferation and apoptosis alterations in Tsc2 -/- cells, in agreement with the effects of the above three miRNAs. In addition, BCL2L11 rescued the proliferation and apoptotic inhibition induced by miR-9-5p, miR-124-3p, and miR-132-3p in Tsc2 -/- cells. This study provides supportive evidence that miR-9-5p, miR-124-3p, and miR-132-3p play a role in TSC-AMLs through the regulation of BCL2L11.

The mechanism by which TSC2 inactivation or deficiency contributes to the pathology of tuberous sclerosis complex (TSC) is not fully clear. We show that renal angiomyolipomas from TSC patients and kidney cortex from Tsc2+/- mice exhibit elevated levels of reactive oxygen species (ROS). Downregulation of tuberin (protein encoded by TSC2 gene) in renal proximal tubular epithelial cells significantly increased ROS concomitant with enhanced Nox4. Similarly, we found elevated levels of Nox4 in the renal cortex of Tsc2+/- mice and in the renal angiomyolipomas from TSC patients. Tuberin deficiency is associated with activation of mTORC1. Rapamycin, shRNAs targeting raptor, or inhibition of S6 kinase significantly inhibited the expression of Nox4, resulting in attenuation of production of ROS in tuberin-downregulated proximal tubular epithelial cells. In contrast, activation of mTORC1 increased Nox4 and ROS. These results indicate that Nox4 may be a potential target for tuberin-deficiency-derived diseases. Using a xenograft model from tuberin-null tubular cells in nude mice, both anti-sense Nox4 and GKT137831, a specific inhibitor of Nox1/4, significantly inhibited the tumor growth. Thus, our results demonstrate the presence of an antagonistic relationship between tuberin and Nox4 to drive oncogenesis in the tuberin deficiency syndrome and identify Nox4 as a target to develop a therapy for TSC.

Tuberous sclerosis syndrome (TSC) is an autosomal dominant tumor suppressor gene syndrome affecting multiple organs, including renal angiomyolipomas and pulmonary lymphangioleiomyomatosis (LAM). LAM is a female-predominant interstitial lung disease characterized by the progressive cyst formation and respiratory failure, which is also seen in sporadic patients without TSC. Mutations in TSC1 or TSC2 cause TSC, result in hyperactivation of mammalian target of rapamycin (mTOR), and are also seen in LAM cells in sporadic LAM. We recently reported that prostaglandin biosynthesis and cyclooxygenase-2 were deregulated in TSC and LAM. Phospholipase A2 (PLA2) is the rate-limiting enzyme that catalyzes the conversion of plasma membrane phospholipids into prostaglandins. In this study, we identified upregulation of adipocyte AdPLA2 (PLA2G16) in LAM nodule cells using publicly available expression data. We showed that the levels of AdPLA2 transcript and protein were higher in LAM lungs compared with control lungs. We then showed that TSC2 negatively regulates the expression of AdPLA2, and loss of TSC2 is associated with elevated production of prostaglandin E2 (PGE2) and prostacyclin (PGI2) in cell culture models. Mouse model studies also showed increased expression of AdPLA2 in xenograft tumors, estrogen-induced lung metastatic lesions of Tsc2 null leiomyoma-derived cells, and spontaneous renal cystadenomas from Tsc2+/- mice. Importantly, rapamycin treatment did not affect the expression of AdPLA2 and the production of PGE2 by TSC2-deficient mouse embryonic fibroblast (Tsc2-/-MEFs), rat uterine leiomyoma-derived ELT3 cells, and LAM patient-associated renal angiomyolipoma-derived "mesenchymal" cells. Furthermore, methyl arachidonyl fluorophosphate (MAFP), a potent irreversible PLA2 inhibitor, selectively suppressed the growth and induced apoptosis of TSC2-deficient LAM patient-derived cells relative to TSC2-addback cells. Our findings suggest that AdPLA2 plays an important role in promoting tumorigenesis and disease progression by modulating the production of prostaglandins and may serve as a potential therapeutic target in TSC and LAM.

Genetic studies have shown that the tuberous sclerosis complex (TSC) 1-TSC2-mammalian target of Rapamycin (mTOR) and the Hippo-Yes-associated protein 1 (YAP) pathways are master regulators of organ size, which are often involved in tumorigenesis. The crosstalk between these signal transduction pathways in coordinating environmental cues, such as nutritional status and mechanical constraints, is crucial for tissue growth. Whether and how mTOR regulates YAP remains elusive. Here we describe a novel mouse model of TSC which develops renal mesenchymal lesions recapitulating human perivascular epithelioid cell tumors (PEComas) from patients with TSC. We identify that YAP is up-regulated by mTOR in mouse and human PEComas. YAP inhibition blunts abnormal proliferation and induces apoptosis of TSC1-TSC2-deficient cells, both in culture and in mosaic Tsc1 mutant mice. We further delineate that YAP accumulation in TSC1/TSC2-deficient cells is due to impaired degradation of the protein by the autophagosome/lysosome system. Thus, the regulation of YAP by mTOR and autophagy is a novel mechanism of growth control, matching YAP activity with nutrient availability under growth-permissive conditions. YAP may serve as a potential therapeutic target for TSC and other diseases with dysregulated mTOR activity., (© 2014 Liang et al.)

Angiomyolipomas (AMLs) are associated with cell fibrosis in kidney of Tuberous Sclerosis Complex patients. The mechanism by which the fibrotic proteins accumulated in AMLs has not been explored. In the present study, we investigated the role of Akt/tuberin/mTOR pathway in the regulation cell fibrosis proteins. AML cells that expressed low levels of tuberin showed less expression of N-cadherin and higher of vimentin proteins compared to HEK293 cells. AML cells infected with Ad-tuberin showed a significant decrease in vimentin and an increase in N-cadherin protein expression. In addition, cells treated with rapamycin showed a significant increase in p-Akt and a decrease in p-p70S6K that was associated with a decrease expression of vimentin and a slight increase expression in N-cadherin. On the other hand, cells treated with Akt inhibitor revealed a significant decrease in p-Akt and p-p70S6K that was associated with a significant decrease in vimentin and an increase in N-cadherin expression. In addition, cells transfected with DN-Akt or DN-S6K show significant increase expression in N-cadherin and a decrease in vimentin. Moreover, cells transfected with siRNA against rictor or siRNA against raptor resulted in a decrease in vimentin and an increase N-cadherin expression. Kidney tumors from TSC patients showed significant decrease in N-cadherin and significant increased in vimentin protein expression compared to control kidney tissues. These data comprise the first report to provide the role of Akt/tuberin/mTORC1/2 in the regulation of N-cadherin and vimentin that are involved in the progression of fibrosis in kidney tumor of TSC patients.

Tuberous sclerosis complex (TSC), an inherited tumor predisposition syndrome associated with mutations in TSC1 or TSC2, affects ∼1 in 6,000 individuals. Eighty percent of TSC patients develop renal angiomyolipomas, and renal involvement is a major contributor to patient morbidity and mortality. Recent work has shown that mammalian target of rapamycin complex 1 (mTORC1) inhibition caused angiomyolipoma shrinkage but that this treatment may cause cytostatic not a cytotoxic effect. Endoplasmic reticulum (ER) stress can develop in TSC-associated cells due to mTORC1-driven protein translation. We hypothesized that renal angiomyolipoma cells experience ER stress that can be leveraged to result in targeted cytotoxicity. We used immortalized human angiomyolipoma cells stably transfected with empty vector or TSC2 (encoding tuberin). Using cell number quantification and cell death assays, we found that mTORC1 inhibition with RAD001 suppressed angiomyolipoma cell proliferation in a cytostatic manner. Angiomyolipoma cells exhibited enhanced sensitivity to proteasome inhibitor-induced ER stress compared with TSC2-rescued cells. After proteasome inhibition with MG-132, Western blot analyses showed greater induction of C/EBP-homologous protein (CHOP) and more poly (ADP-ribose) polymerase (PARP) and caspase-3 cleavage, supporting ER stress-induced apoptosis. Live cell numbers also were decreased and cell death increased by MG-132 in angiomyolipoma cells compared with TSC2 rescued. Intriguingly, while pretreatment of angiomyolipoma cells with RAD001 attenuated CHOP and BiP induction, apoptotic markers cleaved PARP and caspase-3 and eukaryotic translation initiation factor 2α phosphorylation were increased, along with evidence of increased autophagy. These results suggest that human angiomyolipoma cells are uniquely susceptible to agents that exacerbate ER stress and that additional synergy may be achievable with targeted combination therapy.

Renal epithelioid angiomyolipoma (EAML) is a rare but distinct variant of angiomyolipoma, closely simulating renal cell carcinoma or sarcoma both clinically and histopathologically. This report presents an unusual case of unilateral simultaneous renal EAML and renal clear cell carcinoma. A 52-year-old man without any sign of tuberous sclerosis had a complaint of 6-month history of pain in left renal area and had macroscopic hematuria twice within the recent 1 month. Computed tomography showed the presence of 2 masses in the upper and lower portion of the left kidney. The patient underwent left radical nephrectomy. Histological examination revealed the upper mass was composed of medium to large epithelioid cells with clear or eosinophilic cytoplasm and numerous giant multinucleated cells. Adult-appearing adipose tissue and coagulative necrosis could also be observed focally in the mass. Immunohistochemically, the tumor cells in the upper mass showed positive reactions to actin, HMB-45, Melan-A, and CD68 but negative reactions to pan-cytokeratin (pan-CK), epithelial membrane antigen, and CD10. However, the lower mass was composed of diffusely monomorphic clear cells with strongly immunoreactive for pan-CK, vimentin, and CD10, whereas without expression for HMB-45 and actin. The patient showed no evidence of recurrence or metastasis during 1-year postoperative following-up period. To the authors' knowledge, this is the first report of coincidental renal EAML and clear cell carcinoma in the same kidney. Unlike classic triphasic angiomyolipoma, adjuvant therapy after resection should be considered for renal EAML because of its malignant potential, more aggressive behavior and poor prognosis.

Perivascular epithelioid cell (PEC) neoplasms are an unusual group of mesenchymal tumors thought to arise from the PEC, which characteristically expresses melanocytic markers with common coexpression of muscle markers. They show a wide morphologic spectrum and have been described in multiple anatomical locations, including the uterus. We report a 59-year-old woman with tuberous sclerosis, initially diagnosed with a uterine malignant mixed müllerian tumor, in whom the hysterectomy specimen also showed 3 other incidental tumors: a subserosal angiomyolipoma consisting of blood vessels of varying caliber, mature adipose tissue, and nests of spindle cells arranged in a perivascular location, a sclerosing PEComa consisting of small nests of clear epithelioid cells present in a background of marked sclerosis centered in the lower uterine segment/endocervix, and diffuse lymphangiomyomatosis involving the endocervix, lower uterine segment, and uterine corpus. This case provides further evidence of a common precursor cell of origin for these lesions.

Background: Tuberous sclerosis (TSC) related tumors are characterized by constitutively activated mTOR signaling due to mutations in TSC1 or TSC2., Methods: We completed a phase 2 multicenter trial to evaluate the efficacy and tolerability of the mTOR inhibitor, sirolimus, for the treatment of kidney angiomyolipomas., Results: 36 adults with TSC or TSC/LAM were enrolled and started on daily sirolimus. The overall response rate was 44.4% (95% confidence intervals [CI] 28 to 61); 16/36 had a partial response. The remainder had stable disease (47.2%, 17/36), or were unevaluable (8.3%, 3/36). The mean decrease in kidney tumor size (sum of the longest diameters [sum LD]) was 29.9% (95% CI, 22 to 37; n = 28 at week 52). Drug related grade 1-2 toxicities that occurred with a frequency of >20% included: stomatitis, hypertriglyceridemia, hypercholesterolemia, bone marrow suppression (anemia, mild neutropenia, leucopenia), proteinuria, and joint pain. There were three drug related grade 3 events: lymphopenia, headache, weight gain. Kidney angiomyolipomas regrew when sirolimus was discontinued but responses tended to persist if treatment was continued after week 52. We observed regression of brain tumors (SEGAs) in 7/11 cases (26% mean decrease in diameter), regression of liver angiomyolipomas in 4/5 cases (32.1% mean decrease in longest diameter), subjective improvement in facial angiofibromas in 57%, and stable lung function in women with TSC/LAM (n = 15). A correlative biomarker study showed that serum VEGF-D levels are elevated at baseline, decrease with sirolimus treatment, and correlate with kidney angiomyolipoma size (Spearman correlation coefficient 0.54, p = 0.001, at baseline)., Conclusions: Sirolimus treatment for 52 weeks induced regression of kidney angiomyolipomas, SEGAs, and liver angiomyolipomas. Serum VEGF-D may be a useful biomarker for monitoring kidney angiomyolipoma size. Future studies are needed to determine benefits and risks of longer duration treatment in adults and children with TSC., Trial Registration: Clinicaltrials.gov NCT00126672.

Background: The tuberous sclerosis complex (TSC) is caused by defects in one of two tumor suppressor genes, TSC-1 or TSC-2. TSC-2 gene encodes tuberin, a protein involved in the pathogenesis of kidney tumors, both angiomyolipomas and renal cell carcinomas. Loss of heterozygosity at the 8-oxoG-DNA glycosylase (OGG1) allele is found in human kidney clear cell carcinoma identifying loss of OGG1 function as a possible contributor to tumorigenesis in the kidney. Tuberin regulates OGG1 through the transcription factor NF-YA in cultured cells. The purpose of this study is to determine the effect of tuberin-deficiency on OGG1 protein and mRNA levels as well as on 8-oxodG levels in kidney tumors from patients with TSC. In addition we evaluated the phophorylation level of downstream targets of mTOR, phospho-S70K, in kidney tumor tissue from TSC patients., Results: Kidney angiomyolipoma tissue from TSC patients expresses significant levels of phopho-tuberin and low levels of tuberin compared to control kidney tissue. The increase in tuberin phosphorylation and the decrease tuberin expression are associated with decrease in OGG1 protein and mRNA levels in tumor samples compared to normal kidney samples. The decrease OGG1 expression is also associated with significant decrease in the transcription factor, NF-YA, expression in tumor samples compared to normal tissues. In addition, the levels of 8-oxodG are 4-fold higher in tumors compared to control samples. The significant increase of phospho-tuberin expression is associated with increase phosphorylation of S6K in tumor samples compared to controls. Cyclin D1 expression is also 3-fold higher in increase in the tumor tissues compared to normal kidney tissues., Conclusion: These data indicate that tuberin deficiency in angiomyolipoma enhances mTOR activation by phosphorylation of S6K and downregulation of protein and mRNA expression of OGG1 resulted in accumulation of oxidized DNA in patients with TSC. These data suggest that tuberin and OGG1 are important proteins in the pathogenesis of angiomyolipoma in TSC patients.

The perivascular epithelioid cell (PEC) is a cell type constantly present in a group of tumors called PEComas. PEC expresses myogenic and melanocytic markers, such as HMB45 and actin. Recently, recurrent chromosomal alterations have been demonstrated in PEC. At present, PEComa is a widely accepted entity. In the past 10 years, the use of this term has allowed to report and describe numerous cases permitting to start highlighting the biology of this group of lesions. PEComas are related to the genetic alterations of tuberous sclerosis complex (TSC), an autosomal dominant genetic disease due to losses of TSC1 (9q34) or TSC2 (16p13.3) genes which seem to have a role in the regulation of the Rheb/mTOR/p70S6K pathway. There are some open questions about PEComas regarding its histogenesis, the definition of epithelioid angiomyolipoma and the identification of the histological criteria of malignancy. An innovative therapeutic trial using rapamycin is under way for tumors occurring in TSC such as renal angiomyolipoma and lymphangioleiomyomatosis. Its success could provide the rationale for the use of the same drug in other lesions composed of PECs, especially in the malignant ones.

We report a case of uterine perivascular epithelioid cell tumor (PEComa) with malignant histological features in a 59-year-old woman with tuberous sclerosis (TBS). The patient also had extrapulmonary lymphangioleiomyomatosis involving pelvic lymph nodes, myometrium, cervix, and ovary ("pecomatosis"). The uterine tumor measured 2.6 cm and had marked nuclear pleomorphism, necrosis, and 2 mitoses per 50 high-powered field, with an occasional atypical mitosis and infiltrative borders. The nonneoplastic myometrium, the cervical wall, and the hilum of the ovary had multiple clusters of bland-looking epithelioid clear cells that ranged from 1 to 5 mm (pecomatosis). The uterine tumor cells were positive for HMB-45 (90%), Melan-A (70%), smooth muscle actin (50%), and estrogen receptor (30%). Of the 16 pelvic lymph nodes excised, 3 were involved with lymphangioleiomyomatosis that was positive for HMB-45 and estrogen receptor. This is only the second reported PEComa associated with pecomatosis and the fourth PEComa described in a patient with TBS. The clinical significance of pecomatosis is still uncertain but seems to be seen only in patients with TBS.

The pathology associated with tuberous sclerosis complex (TSC) shows diverse phenotypes that suggest abnormal signaling of multiple pathways. Besides the negative regulatory role of the TSC1/TSC2 proteins on mTOR, we have reported an effect on beta-catenin signaling at the level of the degradation complex in vitro. The TSC1/TSC2 complex associates with GSK3 and Axin and promotes beta-catenin degradation to inhibit Wnt-stimulated TCF/LEF-dependent transcription. Here, we show that beta-catenin and its effectors, cyclin D1 and connexin 43, were up-regulated in TSC-related angiomyolipomas and lymphangioleiomyomatosis. This was supported by the failure of three disease-causing TSC2 missense mutants to inhibit Wnt signaling. Further, the interaction between TSC1/TSC2 and components of the beta-catenin degradation complex was dependent on Wnt stimulation such that binding of tuberin to GSK3 and Axin was reduced in the presence of Wnt whereas the tuberin-Dishevelled interaction was increased. GSK3 activity played a role in regulating the assembly/stability of the degradation complex. Inhibition of GSK3 by lithium chloride reduced its association with TSC1 whereas disruption of GSK3-phosphorylation sites in TSC1 reduced interaction between TSC2 and TSC1. Collectively, our data provide further evidence that beta-catenin signaling plays a role in TSC pathogenesis in vivo and suggest a novel role of GSK3 in modulating the TSC1/TSC2 complex through TSC1 phosphorylation.

Angiomyolipoma of the liver or kidney is one of the clinical manifestations of tuberous sclerosis complex. However, concurrence of angiomyolipoma in both liver and kidney associated with tuberous sclerosis complex is a rare entity. Renal angiomyolipomas with large aneurysms confer a higher probability of rupture as compared to small aneurysms. Herein, we document a case of tuberous sclerosis coexisting with hepatic and renal angiomyolipoma in a 37 year-old woman who presented with an acute abdomen due to ruptured tumor. Computed tomography of the abdomen revealed multiple tumors over the bilateral kidneys and liver. A right nephrectomy was performed. During surgery, a liver biopsy was performed from which a preliminary diagnosis of necrosis was established. However, immunoreactivity staining using monoclonal antibody HMB-45 (Human Melanoma, Black) led to the final diagnosis of angiomyolipoma. We emphasized that pathologists and clinicians should be aware that cases of tuberous sclerosis complex may be associated with renal and hepatic angiomyolipoma. To avoid an inappropriate diagnosis, before diagnosing liver necrosis, immunohistochemical staining for HMB-45 is recommended.

The most common renal lesions of tuberous sclerosis complex, an autosomal-dominant syndrome resulting from losses of TSC1 (9q34) or TSC2 (16p13.3), are renal cysts and angiomyolipomas. Epithelial neoplasms are less common. The TSC2 gene lies adjacent to PKD1, the major gene responsible for autosomal-dominant polycystic kidney disease. Recently, a deletion mutation disrupting both TSC2 and PKD1 has been described in young children with tuberous sclerosis complex with severe renal cystic disease. This disease has been termed the TSC2/PKD1 contiguous gene syndrome. We describe the lesions in the resected kidneys of two adults with TSC2/PDK1 contiguous gene syndrome, at the time of the nephrectomies: a 31-year-old man and his 44-year-old mother. The four kidneys were enlarged reniform masses composed of cysts lined by flattened, cuboidal, or, infrequently, large deeply eosinophilic epithelial cells. The kidneys also contained numerous classic angiomyolipomas and rare intraglomerular microlesions. In the son the largest tumor was a monotypic epithelioid angiomyolipoma. In the wall of his left renal pelvis there was a plaque-shaped, HMB-45-positive localized lesion of lymphangioleiomyomatosis. This is the first description of the renal lesions in adults with genetically confirmed TSC2/PDK1 contiguous gene syndrome. The pathologic findings highlight the importance of thorough sampling for histology in polycystic kidney diseases and indicate that the observation of an angiomyolipoma in biopsy material from patients with enlarged cystic kidneys should suggest the diagnosis of TSC2/PKD1 contiguous gene syndrome, even in cases without ultrasonographic and macroscopic evidence of angiomyolipoma.

We report a case of secondary parathyroid hyperplasia in a 49-year-old man with tuberous sclerosis. Two parathyroid glands had collections of large, eosinophilic ganglion-like endocrine cells that to our knowledge have not been previously described at this site. These cells are morphologically similar to those of subependymal giant cell astrocytoma, tubers, and atypical angiomyolipoma, all of which may arise in the setting of tuberous sclerosis. These large, eosinophilic ganglion-like cells found in different affected organs appear to be distinctive of tuberous sclerosis. We suggest these large eosinophilic cells arise from a common stem cell precursor that acquires variable phenotypes according to alterations in the cellular microenvironment.

A recent case of angiomyolipoma (AML) with a prominent component of polygonal epithelioid cells is described. A 27-year-old Japanese male with tuberous sclerosis presented with massive abdominal tumors increasing progressively in size. The patient died of respiratory disturbance and the autopsy revealed massive tumors in the bilateral kidneys, liver and lymph nodes, subependymal giant cell glioma of the brain and lymphangiomyomatosis of the lungs. The giant tumors were an unusual type of AML with a component of polygonal epithelioid cells, which showed a hepatocellular carcinoma-like pattern in some areas. Smooth muscle components comprising spindle cells, short or plump spindle cells and polygonal epithelioid cells frequently exhibited positive staining for HMB-45 but negative staining for epithelial cell markers. The unusual AML presented in this case was thought to be of low-grade malignancy and slow growing. It has been suggested that angiomyolipomas with diffuse areas of epithelioid cell component are potentially malignant. Immunostainings positive for HMB-45 but negative for epithelial cell markers are considered to be useful in differentiating AML with polygonal epithelioid cell component from other tumors, especially from renal cell carcinoma and hepatocellular carcinoma.

Angiomyolipomas can occur sporadically or in association with tuberous sclerosis complex (TSC). TSC is an autosomal dominant disorder characterized by seizures, mental retardation, and benign tumors of the brain, heart, kidney, and skin. Angiomyolipomas are more common in women than in men, suggesting a possible hormonal influence on tumor growth. In this study, 35 angiomyolipomas from 23 patients were immunostained with antibodies to estrogen receptor (ER) and progesterone receptor (PR). Eleven angiomyolipomas (31%) contained clusters of PR-immunoreactive smooth muscle cells. None contained ER-immunoreactive cells. Of the 21 tumors from patients with TSC, 11 (48%) were PR immunoreactive. All of the PR-immunoreactive angiomyolipomas were from women younger than 50 years of age, and all except one of these women had TSC. This study suggests that hormonal factors play a role in the pathogenesis of TSC-associated angiomyolipomas.

We report the clinicopathologic, immunohistochemical, ultrastructural, and genetic features of an unusual renal tumor composed of large, atypical, densely packed, clear/eosinophilic epithelioid cells. Three patients, two men and one woman (ages 31, 36, and 60 years of age, respectively), had abdominal pain. Morphologically, all cases showed aggressive features (largeness, atypical cells, sarcomatoid features, necrosis, and, in one case, invasion of the renal vein). Despite the marked morphologic resemblance of these tumors to high-grade sarcomatoid renal cell carcinoma, their phenotype (HMB45+, CD68+/-, actin+/-, and vimentin and keratin negative) is in contrast to that observed in epithelial tumors and parallels the phenotypic profile of angiomyolipoma. Ultrastructural analysis showed the presence of glycogen, mitochondria, and prominent electron-dense, membrane-bound granules in the neoplastic cells, and the absence of melanosomes or premelanosomes. Genetic study, performed using polymerase chain reaction from paraffin sections, showed a loss of heterozygosity at the TSC2-containing region on 16p in one case, and on 3p in two cases, showing that multiple genetic alterations are taking place in these tumors. Follow-up has shown local recurrence in one case after 6 years, and the patient died 1 year later of cardiorespiratory failure. The other two patients are well after 26 and 10 months. All three patients were evaluated for signs of tuberous sclerosis, and findings were negative. We suggest that these tumors should be considered close relatives of the angiomyolipoma variants, composed purely of perivascular epithelioid cells. More cases and longer follow-up durations are needed to fully evaluate its prognostic implication.

Angiomyolipoma of the kidney is a clonal neoplasm, apparently part of a family of neoplasms derived from perivascular epithelioid cells. Early angiomyolipomas are small nodules composed of HMB-45-reactive spindle cells in the renal capsule, cortex, or medulla. Most angiomyolipomas are asymptomatic, and they are more common than previously appreciated, approaching 13 per 10,000 adults. They are much more prevalent in patients with tuberous sclerosis, where they often are accompanied by cysts and occasionally by renal cell carcinoma. That they are rarely diagnosed before puberty in patients without tuberous sclerosis, that large ones are more common in women than men, and that they occasionally grow rapidly during pregnancy suggest that hormones may play a role in stimulating the growth of angiomyolipoma. Pathologists should recognize that the presence of multiple angiomyolipomas is, at the least, presumptive evidence for the diagnosis of tuberous sclerosis and include this in their reports. Typical angiomyolipomas are benign but may have alarming properties: nuclear pleomorphism and mitotic activity, extension into the vena cava (21 cases), and spread to regional lymph nodes (39 cases), without malignant progression. The most common serious complication of renal angiomyolipoma is hemorrhage. Epithelioid angiomyolipoma is a recently recognized variant with malignant potential. Angiomyolipoma and lymphangiomyoma are closely related, and tumors with features of both have occurred. Angiomyolipomas have occurred in other organs in association with renal angiomyolipoma.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by seizures, mental retardation, and tumors of skin, brain, heart, and kidney. In this study, we focused on two of the most frequent tumors in TSC patients, renal angiomyolipomas and subependymal giant cell astrocytomas (SEGAs). Two questions were addressed. First, is loss of tuberin, the product of the TSC2 gene, seen in both renal and central nervous system tumors from TSC patients? Second, when loss of tuberin occurs, does it affect each of the cell types seen in these tumors? We used a loss of heterozygosity approach to identify tumors from TSC2 patients. We found loss of tuberin immunostaining in the spindle and epithelioid cells but not in the giant cells of six TSC2 SEGAs. We also found loss of tuberin immunostaining in all three cell types (smooth muscle, fat, and vessels) of six TSC2 angiomyolipomas. Chromosome 16p13 loss of heterozygosity occurred in both spindle and epithelioid cells of a SEGA and in smooth muscle and fat but not the vessels of two angiomyolipomas. These results support a two-hit tumor suppressor model for the pathogenesis of SEGAs and angiomyolipomas. The vascular elements of angiomyolipomas and the giant cells of SEGAs may be reactive rather than neoplastic.