Paula Boaventura

Telomerase promoter mutations in cancer: an emerging molecular biomarker?

Cell immortalization has been considered for a long time as a classic hallmark of cancer cells. Besides telomerase reactivation, such immortalization could be due to telomere maintenance through the “alternative mechanism of telomere lengthening” (ALT) but the mechanisms underlying both forms of reactivation remained elusive. Mutations in the coding region of telomerase gene are very rare in the cancer setting, despite being associated with some degenerative diseases. Recently, mutations in telomerase (TERT) gene promoter were found in sporadic and familial melanoma and subsequently in several cancer models, notably in gliomas, thyroid cancer and bladder cancer. The importance of these findings has been reinforced by the association of TERT mutations in some cancer types with tumour aggressiveness and patient survival. In the first part of this review, we summarize the data on the biology of telomeres and telomerase, available methodological approaches and non-neoplastic diseases associated with telomere dysfunction. In the second part, we review the information on telomerase expression and genetic alterations in the most relevant types of cancer (skin, thyroid, bladder and central nervous system) on record, and discuss the value of telomerase as a new biomarker with impact on the prognosis and survival of the patients and as a putative therapeutic target.

TERT Promoter Mutations in Skin Cancer: The Effects of Sun Exposure and X-Irradiation

The reactivation or reexpression of telomerase (TERT) is a widespread feature of neoplasms. TERT promoter mutations were recently reported that were hypothesized to result from UV radiation. In this retrospective study, we assessed TERT promoter mutations in 196 cutaneous basal cell carcinomas (BCCs), including 102 tumors from X-irradiated patients, 94 tumors from patients never exposed to ionizing radiation treatment, and 116 melanomas. We sought to evaluate the effects of UV and X-ray irradiation on TERT mutation frequency. TERT mutations were detected in 27% of BCCs from X-irradiated patients, 51% of BCCs from nonirradiated patients, and 22% of melanoma patients. TERT mutations were significantly increased in non-X-irradiated BCC patients compared with X-irradiated BCC patients; the mutations also presented a different mutation signature. In nonirradiated patients, TERT mutations were more frequent in BCCs of sun-exposed skin, supporting a possible causative role of UV radiation. In melanoma, TERT promoter mutations were generally restricted to intermittent sun-exposed areas and were associated with nodular and superficial spreading subtypes, increased thickness, ulceration, increased mitotic rate, and BRAFV600E mutations. Our results suggest that various carcinogenic factors may cause distinct TERT promoter mutations in BCC and that TERT promoter mutations might be associated with a poorer prognosis in melanoma.

Head and neck basal cell carcinoma prevalence in individuals submitted to childhood X-ray epilation for tinea capitis treatment

BACKGROUND A higher prevalence for basal cell carcinoma (BCC) has been associated with radiation, namely with tinea capitis epilation treatment. OBJECTIVE To evaluate the prevalence of head and neck basal cell carcinoma (BCC) and to identify the major risk factors for BCC in individuals irradiated in childhood for tinea capitis treatment. METHODS We clinically observed 1,308 individuals from an original cohort of 5,356 irradiated between 1950 and 1963, registering previous skin lesions excisions and proposing for surgery all the suspicious lesions detected. In 585 participants, 47 with BCC, the skin pigmentation was measured. RESULTS The overall prevalence of BCC was 8.0% and of multiple BCC was 2.4%. Both total (14.7%) and multiple BCC (6.6%) were significantly more common in the individuals who had received a higher radiation dose. Multiple BCC was more prevalent (3.7%) in younger irradiated individuals and total BCC (9.4%) in women. Participants with BCC and without BCC presented similar skin pigmentation. CONCLUSION Younger age at irradiation, higher dose and female gender increased the risk of developing BCC in these irradiated individuals.

Head and neck lesions in a cohort irradiated in childhood for tinea capitis treatment

We read with interest Shifra Shvarts and colleagues’ Historical Review on the tinea capitis treatment campaign in 1950s Serbia. Treatment of tinea capitis infection that included radiation was also used in Portugal in the same period, in accordance with the same Kienbock-Adamson technique. We had access to the registries of a cohort treated in the north of Portugal, which included patients’ details, treatment dates, tinea diagnoses (type of infection), and doses received (table). In March, 2006, we started to locate and contact the cohort members; this was a diffi cult task because 40–50 years have passed since their tinea capitis treatment. Nevertheless we have traced 3548 individuals, to whom we sent information letters with a free-phone contact number. This method allowed us to clinically examine 1287 individuals, all by the same clinician (TG), and report that 292 are dead and 85 are living abroad. We recommended neck ultrasounds, and 886 (70%) of the participants had the examination. A fi ne-needle aspiration biopsy was advised in 221 patients who had nodules with suspicious features. Surgery was proposed for 45 people whose biopsy samples showed malignant or follicular lesions. At clinical examination, 18 individuals had been previously diagnosed with thyroid carcinoma, and we diagnosed 15 more. In total, we recorded a 2·6% prevalence of thyroid carcinoma; similar to the 2·1% reported for survivors of the Hiroshima and Nagasaki atomic bombs in a survey study by Misa Imaizumi and colleagues that used a similar protocol (thyroid ultrasonography). If we exclude from our study the previous diagnoses, the prevalence decreases to 1·4%, which is similar to the 0·95% reported by Siegal Sadetzki and co-workers. Our data seem to agree with the high risk of thyroid tumours reported in Shvarts and colleagues’ study. We have observed in our cohort a high prevalence of meningiomas and basal-cell carcinoma (data not shown), not mentioned by Shvarts and colleagues. We have also shown in this cohort that the favus tinea infection gives an eight-times increase in risk of alopecia when compared with trichophytic tinea, even after adjustment for age and irradiation dose. Our data support and emphasise the arguments presented by Shvarts and colleagues, that physicians should be aware of particular subsets of population that might be at risk of late radiation-associated health eff ects. These data justify a close follow-up of the irradiated tinea capitis cohorts to identify those head and neck lesions that are undiagnosed. This work was supported by a grant from Fundação Calouste Gulbenkian (ref 76636) and Portuguese Foundation for Science and Technology (FCT) (project: PIC ⁄IC ⁄83154 ⁄2007), and further funding from the FCT by a grant to PB (SFRH ⁄BPD ⁄34276 ⁄2007). IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education, and is partly supported by the FCT. We thank all the individuals that agreed to participate in this study as well as all the physicians who provided us the clinical material and information.

Genetic alterations in thyroid tumors from patients irradiated in childhood for tinea capitis treatment

OBJECTIVE Exposure to ionizing radiation at young age is the strongest risk factor for the occurrence of papillary thyroid carcinoma (PTC). RET/PTC rearrangements are the most frequent genetic alterations associated with radiation-induced PTC, whereas BRAF and RAS mutations and PAX8-PPARG rearrangement have been associated with sporadic PTC. We decided to search for such genetic alterations in PTCs of patients subjected in childhood to scalp irradiation. DESIGN We studied 67 thyroid tumors from 49 individuals irradiated in childhood for tinea capitis scalp epilation: 36 malignant (12 cases of conventional PTC (cPTC), two cPTC metastases, 20 cases of follicular variant PTC (FVPTC), one oncocytic variant of PTC and one follicular carcinoma) and 31 follicular thyroid adenomas. METHODS The lesions were screened for the BRAF(V600E) and NRAS mutations and for RET/PTC and PAX8-PPARG rearrangements. RESULTS BRAF(V600E) mutation was detected in seven of 14 (50%) cPTC and two of 20 FVPTC (10%) (P=0.019). NRAS mutation was present in one case of FVPTC (5%). RET/PTC1 rearrangement was found, by RT-PCR, in one of 17 cases (5.9%) and by fluorescence in situ hybridization in two of six cases (33%). PAX8-PPARG rearrangement was not detected in any carcinoma. None of the follicular adenomas presented any of the aforementioned genetic alterations. CONCLUSIONS The prevalence of BRAF(V600E) mutation in our series is the highest reported in series of PTCs arising in radiation-exposed individuals. The prevalence of RET/PTC1 rearrangement fits with the values recently described in a similar setting.

TERT promoter mutations: A genetic signature of benign and malignant thyroid tumours occurring in the context of tinea capitis irradiation

OBJECTIVE The aim of this study is to evaluate the frequency and molecular characteristics of TERTp mutations in thyroid adenomas and carcinomas occurring in the low-dose radiation exposure tinea capitis setting. DESIGN AND METHODS Twenty-seven patients with 34 well-differentiated thyroid carcinomas and 28 patients with 29 follicular adenomas diagnosed in a Portuguese tinea capitis cohort were studied. Blood samples were obtained from all the patients. Screening for TERTp mutations was performed by PCR amplification followed by Sanger sequencing. A series of 33 sporadic thyroid adenomas was used as control. RESULTS TERTp mutations were detected in six of the 28 patients with adenoma (21.4%) and in four of the 27 patients with carcinoma (14.8%). Three tumours (two carcinomas and one adenoma) had the tandem mutation -124/-125 GG>AA (30.0%), whereas the remaining seven had the -124G > A. The 20.7% frequency of TERTp mutations in adenomas contrasts with the absence of mutations in the adenomas from the control group and from most series on record, whereas the one found in carcinomas (11.8%) is similar to those reported in the literature for sporadic carcinomas. CONCLUSION TERTp mutations, including the tandem mutation -124/-125 GG>AA not described previously in thyroid tumours, appear to represent a genetic signature for thyroid tumours in patients submitted to low-dose X-ray irradiation. The high frequency of TERTp mutations in the adenomas of our cohort contrasts with their absence in sporadically occurring, as well as in adenomas of the Chernobyl series.

Alopecia in women submitted to childhood X-ray epilation for tinea capitis treatment

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IL6-174 G>C Polymorphism (rs1800795) Association with Late Effects of Low Dose Radiation Exposure in the Portuguese Tinea Capitis Cohort.

Head and neck cancers, and cardiovascular disease have been described as late effects of low dose radiation (LDR) exposure, namely in tinea capitis cohorts. In addition to radiation dose, gender and younger age at exposure, the genetic background might be involved in the susceptibility to LDR late effects. The -174 G>C (rs1800795) SNP in IL6 has been associated with cancer and cardiovascular disease, nevertheless this association is still controversial. We assessed the association of the IL6-174 G>C SNP with LDR effects such as thyroid carcinoma, basal cell carcinoma and carotid atherosclerosis in the Portuguese tinea capitis cohort. The IL6-174 G>C SNP was genotyped in 1269 individuals formerly irradiated for tinea capitis. This sampling group included thyroid cancer (n = 36), basal cell carcinoma (n = 113) and cases without thyroid or basal cell carcinoma (1120). A subgroup was assessed for atherosclerosis by ultrasonography (n = 379) and included matched controls (n = 222). Genotypes were discriminated by real-time PCR using a TaqMan SNP genotyping assay. In the irradiated group, we observed that the CC genotype was significantly associated with carotid plaque risk, both in the genotypic (OR = 3.57, CI = 1.60–7.95, p-value = 0.002) and in the recessive (OR = 3.02, CI = 1.42–6.42, p-value = 0.004) models. Irradiation alone was not a risk factor for carotid atherosclerosis. We did not find a significant association of the IL6-174 C allele with thyroid carcinoma or basal cell carcinoma risk. The IL6-174 CC genotype confers a three-fold risk for carotid atherosclerotic disease suggesting it may represent a genetic susceptibility factor in the LDR context.

Is Low-Dose Radiation Exposure a Risk Factor for Atherosclerotic Disease?

Nontargeted late effects of radiation include an increased risk of cardiovascular disease, although this is still debatable in the context of low-dose radiation. Tinea capitis patients treated in childhood with X rays to induce scalp epilation received a low dose of radiation to their carotids. To better clarify this issue, we evaluated carotid atherosclerosis in a cohort of such patients treated in 1950–1963 in Portugal. A group of 454 individuals randomly chosen from previously observed Portuguese tinea capitis patients and a control group mainly composed of their spouses (n = 280) were enrolled. Cardiovascular risk factors such as waist circumference, body mass index, blood pressure and tobacco consumption, as well as biochemical measurements were obtained. Ultrasound imaging of carotid arteries for intima media thickness and stenosis evaluation were performed according to a standardized protocol. In comparison to the control group, the irradiated cohort members were significantly older, more frequently never smokers, hypertensive, and presented higher glycated hemoglobin and alkaline phosphatase levels. In addition, the irradiated cohort showed a higher frequency of carotid stenosis ≥30% than the nonirradiated group (13.9% vs. 10.7%), although this was not significant (P = 0.20). Stenosis was ≥50% in 2.9% of the irradiated group and 0.4% of the nonirradiated group (P = 0.02). Likewise, the frequency of intima media thickness ≥1 mm was significantly higher in the irradiated group (16.8% vs. 10.7%; P = 0.02). Multivariate analysis, including other cardiovascular risk factors, showed that exposure to low-dose radiation increased the risk of carotid stenosis by ≥50% [odds ratio (OR) = 8.85; P = 0.04] and intima media thickness by ≥1 mm (OR = 1.82; P = 0.02). These findings confirm that low-dose exposure is a risk factor of carotid atherosclerotic disease.