Introduction

Tuberous sclerosis complex (TSC) is a rare autosomal dominant disorder caused by pathogenic variations in the TSC1 or TSC2 gene. This results in hyperactivation of the mammalian/mechanistic target of rapamycin pathway, leading to hamartoma formation. The prevalence of TSC is estimated to be between 1/6800 and 1/15,000 and the incidence is estimated to be nearly 1:6000–10,000 live births [1,2,3]. It can affect all organ systems, leading to diverse clinical manifestations and has a broad variability, not only among individual patients but also within the affected families [1].

The 2012 International Tuberous Sclerosis Complex Consensus Conference provided recommendations to standardize the approach to manage this disorder. A system of major and minor criteria serves as a basis to establish the diagnosis clinically. Recommendations on surveillance and treatment mainly focus on these criteria [4]. However, the involvement of multiple organ systems at different stages of life presents a major challenge in the comprehensive clinical management of patients with TSC.

Over the last two decades, several clinical pathologies have been identified as associated with TSC, such as aortic and intracranial aneurysm [5, 6], arachnoid cysts [7, 8], lymphedema [9, 10], pancreatic endocrine tumor [11], pituitary adenoma [7], chordoma [12], and bone sclerotic foci [13]. Although these clinical pathologies are less frequent and not included as clinical diagnostic criteria, some of them can be life-threatening, while others may be challenging to integrate into a comprehensive clinical picture of the patients. Identification of these rare or less frequent manifestations and their clinical characteristics may help in their early diagnosis and contribute to ultimately preventing morbidity and mortality in patients with TSC.

A second important controversy in TSC is the fact that it remains a debate whether patients with TSC have an increased risk for malignant tumors. Until now, mostly renal cell carcinoma [14] has been identified in this context.

To the best of our knowledge, no systematic evaluation of these manifestations has been performed to date in a large patient cohort that might provide reliable results to aid clinicians in the management of TSC. Our aim is to present the rates and characteristics of various rare manifestations and malignancies observed in patients with TSC enrolled in the TuberOus SClerosis registry to increAse disease awareness (TOSCA) study and report differences in these rare manifestations and malignancies by sex, age, and TSC mutation.

Materials and methods

Study design, participants and data collection

A detailed methodology of the study has been published previously [15]. In brief, TOSCA is a multicenter, international disease registry structured to collect patient and disease information retrospectively and prospectively. It consists of a ‘core’ section and six ‘petals’ or ‘research projects’. In the core section of the study, information on the patient’s background, including demographics, familial and prenatal history, vital signs, and disease features, were collected at baseline and updated annually, wherever possible. Additional detailed information was collected in the six research projects that focused on subependymal giant cell astrocytoma, renal angiomyolipoma and lymphangiomyomatosis, genetics, TSC-associated neuropsychiatric disorders (TAND), epilepsy, and quality of life. Here, we present data on the rare manifestations, comorbidities and malignancies from the core section of the TOSCA registry.

Patients of any age with a documented visit for TSC in the preceding 12 months or newly diagnosed with TSC were enrolled into the TOSCA registry after obtaining written informed consent. Investigators collected data on rare manifestations by either selecting predefined items known to be associated with TSC or entering other items in a free text field.

For malignancies, data were collected for those patients who had either one or more of a number of predefined malignancies (renal, ovarian, testicular, and gastrointestinal malignancies). In addition, investigators could enter any other malignancy in a free text field.

Assessments

Demographic and baseline characteristics between the patients with and without rare manifestations, and between patients with and without malignancies, were compared. For the purpose of this study, we defined rare manifestations as all manifestations declared as rare and disease-related by the investigators that did not fit under major or minor TSC clinical criteria as outlined in the 2012 Tuberous Sclerosis Consensus Conference recommendations and that were not a clear sequelae of major or minor manifestations (such as epilepsy).

We have prospectively categorized the rare manifestations by sex and mutation type, and by several syndromal complexes, such as tumors, malformations, vascular malformations, cystic lesions, endocrine disorders, and others (Table 2). Rare manifestations were also categorized into the following organ classes: vasculature, ear, nose and throat, endocrine system, eye, heart, gastrointestinal, liver, lymphatic tissue, nervous system, skeletal, urogenital, and others (Additional File 1: Table S1).

Rare manifestations reported by investigators in the free text field of the case report form that did not have an unambiguous description, were clearly not rare manifestations, or were typical manifestations of TSC (major/minor diagnostic criteria or epilepsy) were excluded from the analysis (Additional File 1: Table S2).

Malignancies observed in the TOSCA cohort were reported by organ. Rare manifestations and malignancies (either predefined or open field) were grouped based on organ system, and further by age, sex, and genotype. Available information on the age at first TSC diagnosis, mutation type and patient who received treatment for rare manifestation and malignancies are reported.

Statistical analysis

All patients, without any major protocol deviation, enrolled in the TOSCA clinical study were included for analysis. SAS® Version 9.2 or later was used to perform all statistical analyses. Continuous variables were summarized with descriptive statistics (n, mean, standard deviation, range [minimum and maximum] and median). Frequency counts and the percentage of patients within each category were used for categorical data. Demographic and baseline characteristics between patients with and without rare manifestations and between patients with and without malignancies were compared using chi-square test for association and Z-test for means, as appropriate. A p value < 0.05 was considered statistically significant.

Results

Patient demographics and clinical characteristics

A total of 2211 patients with TSC were enrolled in the TOSCA registry. Of those, 382 (17.3%) individuals had rare manifestations and 65 (2.9%) had malignancies reported by investigators. Demographic characteristics in patients with and without rare manifestations and malignancies are shown in Table 1.

Table 1 Demographic characteristics in patients with and without rare manifestations and in patients with and without malignancies
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In patients with rare manifestations, TSC was diagnosed later than in those without (median: 2 years vs. 1 year; p < 0.0001). The female to male ratio was higher in patients with rare manifestations than in those without (female vs. male; 58.4% vs. 41.6% compared to 50.8% vs. 49.2%, respectively; p = 0.007). There was no difference in the mutation types (TSC1, TSC2 or no mutation identified [NMI]) in participants with rare manifestations compared to individuals without (p = 0.687).

In patients with malignancies, TSC was diagnosed later compared to those without malignancies (median: 10.5 years vs. 1 year; p < 0.0001). We found a higher female to male ratio in patients with malignancies compared to individuals without malignancy (66.2%:33.8% vs. 51.7%:48.3%; p = 0.021). Considering differences in mutations, participants with malignancies had a significantly higher rate of TSC1 mutations than participants without malignancies (TSC1 vs. TSC2 vs. NMI; 32.3% vs. 48.4% vs. 19.4% and 18.5% vs. 64.7% vs.14.5%, respectively; p = 0.037).

Rare manifestations

In this study, 88 different manifestations were identified that were designated by the investigators as rare TSC-associated manifestations (Table 2). The five most frequent rare manifestations were bone sclerotic foci (39.5%), scoliosis (23%), thyroid adenoma (5.5%), adrenal angiomyolipoma (4.5%), hemihypertrophy and pNET (3.1% each) (Table 3).

Table 2 Rare manifestations observed in the study and grouped by tumors, malformations, endocrine disorders, cystic lesions, and others
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Table 3 Demographics of patients with the ten most common rare manifestations by median age, sex, mutational status, and treatment
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Of the 382 patients with rare manifestations, tumors and cystic lesions were reported in 268 (tumors in 248 patients and cystic lesions in 20 patients), with female patients being more commonly affected than males (60.5% vs. 39.5%). Malformations were reported in 124 patients, equally affecting both sexes (male vs. female; 51.3% vs. 48.7%) and a majority of patients having TSC2 mutation (85.2%). Malformations were observed at an earlier age (median age, 11 years), while tumors (median age, 28 years), cystic lesions (median age, 27 years), and endocrine disorders (median age, 31.5 years) were observed at a later age (Table 4).

Table 4 Malformations, tumors, and other manifestations in patients with rare manifestations
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As reported above, rare manifestations were predominant in female compared to male patients, except for scoliosis which was a little more frequent in male patients (51.1% vs. 48.9%; p = 0.733). Similar distribution patterns of TSC gene mutation were noted in patients with or without rare manifestation, i.e. rate of TSC2 mutation being more than TSC1, with the exception of patients with thyroid adenoma who had a higher rate of TSC1 mutations than TSC2 (57.1% vs. 42.9%; p = 0.512). Upon stratification by age group, rare manifestations were more common in adult patients (> 18 years) (Table 5).

Table  5 Rate of the ten most common rare manifestations by age group
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Malignancies

Malignancies were reported in 65 patients. Most frequent malignancies observed were renal cell carcinoma (47.7%), breast cancer (10.8%), and thyroid cancer (9.2%).

Altogether, the percentage of female patients was significantly higher in the group of participants with malignancies (66.2%) compared to participants without malignancies (51.7%; p = 0.021) (Table 1). The predominance of females was consistent in patients with renal cell carcinoma (64.5%), but didn’t reach a statistical significance. In addition, the TSC1:TSC2 ratio was significantly higher in patients with malignancies (TSC1 vs. TSC2 vs. NMI was 32.3% vs. 48.2% vs. 19.4%) compared to individuals without malignancies (18.5%:64.7%:14.5%; p = 0.037). Accordingly, the TSC1:TSC2 ratio was markedly higher in patients with renal malignancy (36.4% vs. 53.3%) than in patients without malignancy (18.5% vs. 64.5%). Thyroid carcinoma (n = 6) was exclusively reported in six female patients, of which half had TSC1 mutations (Table 6).

Table 6 Rate of malignancies in different organs in all patients, by sex and genotype
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Patients with malignancy appeared to be older (median 32.7 years) than those without (median 12 years; p < 0.0001). Overall, malignancies were more common in adult patients (≥ 18 years) compared with pediatric patients (< 18 years) (Table 7). However, it is important to state that nearly one-third of renal cell carcinoma cases, half of thyroid carcinomas, half of bone and soft tissue malignancies, one-fourth of ovarian malignancies, and all of the pancreatic malignancies occurred in children (≤ 14 years). Overall, 26% (n = 17) of malignancies were detected in patients < 18 years and 63.1% (n = 41) in those < 40 years (Table 7). Three patients (all female) had two different malignancies: one patient had ovarian and thyroid malignancies, the second patient had colon and breast malignancies, and the third patient had renal and breast malignancies. No patient died due to malignancies during the study.

Table 7 Malignancies by organ class and age groups
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Discussion

Rare manifestations

The consensus clinical TSC diagnostic criteria include commonly presented TSC manifestations and the surveillance recommendations mainly focus on these manifestations [4]. However, there are numerous additional manifestations of TSC that are reported. They may occur quite frequently (for example, bone sclerotic foci), but may be systematically missed as they are not usually of clinical relevance. This large cohort study provides a clear estimate of the frequency of rare manifestations. Bone sclerotic foci might be mainly detected as a by-product of thoracic computed tomography scans, which are recommended for adult women affected by TSC only in order to screen for lymphangioleiomyomatosis (LAM) [16]. This might explain the clear female predominance of this manifestation in our study. Although of minor clinical relevance, it is important to know about this rather frequent ‘rare manifestation’ as bone sclerotic foci could be misinterpreted as bone metastasis or bone secondaries, which might result in unnecessary and potentially invasive assessments [4, 17] in spite being a common benign manifestation of TSC. Expert advice should be sought in case of doubt in these patients.

In addition, bone sclerotic lesions are discussed to differentiate TSC-associated LAM from sporadic LAM [16]. Given its frequency, one could argue for introducing bone sclerotic foci to the clinical diagnostic criteria.

The consensus guidelines further recommend excluding TSC-associated manifestations, such as bone cysts, endocrinopathies, vascular aneurysms, and gastrointestinal polyps, from routine evaluation unless coupled with clinical symptoms or history due to the insufficient evidence of benefit [1].

TSC is a multisystem disorder based on defects in tumor suppressor genes. We therefore hypothesized that there might be additional rare manifestations and asked investigators to document clinical signs considered as possible TSC-associated rare manifestations.

In total, 88 different rare manifestations were recorded in 17.3% of patients in this study. This shows the complexity of the disease and highlights the limitations of systematic evaluation and treatment of rare manifestations. Most rare manifestations were more common in female patients and those with TSC2 mutations, which is in line with findings from previous literature on rare manifestations such as lymphedema and angiomyolipoma [10, 18]. Tumors and cystic lesions in a broad variety of organs seem to occur (or at least are detected) at higher ages. However, clinical significance seems limited in most cases, as treatment was reported for only 16% of tumors and 5% of cysts. It is worth noting that treatment rates differed markedly depending on organ system. Details are given in Additional File 1: Table S1.

A relevant number of rare manifestations were malformations (occurring at younger ages), of which scoliosis and hemihypertrophy were the most frequent. Diagnosis of musculoskeletal malformations can be performed easily via careful physical examination in most cases, which might contribute to detection at an earlier age. Whereas hemihypertrophy was clearly more frequent in our cohort compared to the overall population (1 in 86,000 live births), the rate of scoliosis did not differ from reports of adolescent idiopathic scoliosis (3.8% vs. 3.3%) [19, 20]. This might raise the question of whether scoliosis is a (rare) manifestation of TSC or a coincidental finding. Although causality cannot be proven by a registry study, it is worth mentioning that 22.7% of the scoliosis patients in our study required treatment (compared to 0.3% with adolescent idiopathic sclerosis). This possibly points toward a higher degree of severity or an underestimation of mild cases [21]. As previously reported, vascular malformations, including arterial aneurysms, can occur in patients but seem to be quite rare (n = 5 corresponding to 0.2% of the participants) (Table 5). Therefore, our data supports the 2012 consensus recommendations on not to perform the routine evaluation given the sparse numbers. If routine imaging of the brain is performed, it seems justifiable to screen for blood vessel abnormalities as 75% of arterial aneurysms in our cohort were reported to occur in the extra- or intracranial brain-supporting vessels [4].

Malignancies

The TSC consensus guidelines do not specifically comment on the surveillance of malignancies in patients with TSC [4]. It is still a matter of debate whether TSC patients have an increased risk of malignant tumors [14].

This is the first study to report malignancies in a large and multinational cohort of patients with TSC. No deaths were reported due to malignancy during the study. Although, the overall rate of malignancies in our study was higher than reported by Jozwiak et al. [22] (2.9% vs. 1.1%), but taken as a whole, still low. Notably, TOSCA participants, including the cohort of patients with malignancy, were markedly younger (median age 31 years) compared to the overall population. This might explain why several other tumor types were found to be predominant in this cohort compared to the overall population. The most frequent cancer entities in the overall population, such as lung, colorectal, and prostate cancer (in men), occur at older median ages. Only breast cancer, which is the most frequent cancer in the overall female population, was also a frequent cancer type seen in our cohort [23].

The most frequent type of malignancy in our cohort was renal cell carcinoma, affecting 1.4% (31 patients) of participants, majorly observed in pediatric and young adults (10 patients, ≤ 18 years). The rate of renal cell carcinoma considerably exceeds the prevalence in the overall population. Similar findings have been reported previously; Yang et al. (2014) referred to an overall incidence of 2% to 4% for renal malignancies [23] and Peron et al. (2016) reported an overall renal cell carcinoma rate of 2.1% [14]. TSC-associated renal malignancies have been reported to be more frequently observed in females, which is in contrast with the overall population where the rate of renal malignancy is higher in males (6.6 vs. 3.9 per 100,000) (2). We, too, observed a higher rate of renal malignancy in female patients (64.5%).

All six thyroid cancer cases in our study were observed in females only, which seems to be more frequent than in overall population (compare Table 6). Another frequent tumor type observed in our cohort was breast cancer (females only), which is, however, the most frequent cancer type in the overall population. Interestingly, patients with PTEN hamartoma tumor syndromes are at higher risk for both aforementioned tumor entities (cumulative cancer risk at age 70: 77% for female breast cancer and 38% for thyroid cancer) [25]. Analogical to the TSC1 and TSC2 complex, PTEN is another negative regulator of the mTOR pathway.

It is important to be aware of malignancies in patients with TSC, even at young ages. The occurrence of renal cell carcinoma in children has been reported previously [18] and was evident in our cohort as well. Malignancies were not limited to adult individuals with TSC; approximately 25% of malignancies in our study were observed in individuals younger than 18 years and 63.1% in participants younger than 40 years, similar to Peron et al. [14]. Given the overrepresentation of young patients in our study (63.3% were younger than 19 years and 89.5% were younger than 41 years at inclusion), overall malignancy rates in TSC patients might be underestimated as most malignant tumors occur at higher ages.

In this study, the genotype–phenotype correlations were difficult to establish due to the low number of patients with malignancies. We report 32.3% of patients with malignancy harboring TSC1 mutations compared to only 18.5% of patients who did not have malignancy. A previous study similarly reported a predominance of TSC1 mutations in patients with malignancies [14]. TSC1 contributes to both tumor-suppressive and pro-metastatic action of the TGF-β-Smad pathway and works independently of TSC2, which is essential for cellular growth arrest and epithelial to mesenchymal transition [26]. This could be one possible reason for TSC1 predominance in patients with malignancies in our study. However, further research will be necessary to elucidate the molecular mechanisms.

The evaluation of rare manifestations in this study had several limitations. First, a sign or symptom was recorded as a rare manifestation of TSC per the investigator’s definition. Second, no systematic screening for specific manifestations was performed in the study, except investigations performed in routine clinical practice. The first bears the risk of falsely collating a coincident symptom (especially if recorded infrequently) as a rare manifestation, and the second might result in underestimation of clinically asymptomatic manifestations. With respect to malignancies, investigators were not asked to provide histology findings and no central review of histology could be performed. The study was not designed to monitor malignancy treatments and their outcomes; hence we cannot provide further details. The required frequency of radiological tests to monitor cerebral and renal manifestations in TSC patients might introduce a lead time bias that may affect the conclusions of the study with respect to age at diagnosis. Also, the chance of long-term genetic damage caused by the radiological dose at relatively young ages might be a factor to consider. Although potentially affected by selection bias, it is of note that no study participant died from cancer during the study. Moreover, the TOSCA registry was not specifically designed for rare manifestations, hence the incidence of rare manifestations may be underreported.

Conclusion

Rare manifestations occur in a relevant percentage of TSC patients. However, variability is high and further systematic evaluations are required to shape diagnostic and surveillance strategies. Malignancies affected about 2% of the participants in our study. Compared to the overall population, malignant tumors occurred earlier in age and were more common in females and participants with TSC1 mutation.