Targeted Long-Read Bisulfite Sequencing Identifies Differences in the TERT Promoter Methylation Profiles between TERT Wild-Type and TERT Mutant Cancer Cells

Background: TERT promoter methylation, located several hundred base pairs upstream of the transcriptional start site, is cancer specific and correlates with increased TERT mRNA expression and poorer patient outcome. Promoter methylation, however, is not mutually exclusive to TERT activating genetic alterations, as predicted for functionally redundant mechanisms. To annotate the altered patterns of TERT promoter methylation and their relationship with gene expression, we applied a Pacific Biosciences-based, long-read, bisulfite-sequencing technology and compared the differences in the methylation marks between wild-type and mutant cancers in an allele-specific manner. Results: We cataloged TERT genetic alterations (i.e., promoter point mutations or structural variations), allele-specific promoter methylation patterns, and allele-specific expression levels in a cohort of 54 cancer cell lines. In heterozygous mutant cell lines, the mutant alleles were significantly less methylated than their silent, mutation-free alleles (p < 0.05). In wild-type cell lines, by contrast, both epialleles were equally methylated to high levels at the TERT distal promoter, but differentially methylated in the proximal regions. ChIP analysis showed that epialleles with the hypomethylated proximal and core promoter were enriched in the active histone mark H3K4me2/3, whereas epialleles that were methylated in those regions were enriched in the repressive histone mark H3K27me3. Decitabine therapy induced biallelic expression in the wild-type cancer cells, whereas the mutant cell lines were unaffected. Conclusions: Long-read bisulfite sequencing analysis revealed differences in the methylation profiles and responses to demethylating agents between TERT wild-type and genetically altered cancer cell lines. The causal relation between TERT promoter methylation and gene expression remains to be established

The influence of tumor regression, solar elastosis, and patient age on pathologists\u27 interpretation of melanocytic skin lesions.

It is not known whether patient age or tumor characteristics such as tumor regression or solar elastosis influence pathologists\u27 interpretation of melanocytic skin lesions (MSLs). We undertook a study to determine the influence of these factors, and to explore pathologist\u27s characteristics associated with the direction of diagnosis. To meet our objective, we designed a cross-sectional survey study of pathologists\u27 clinical practices and perceptions. Pathologists were recruited from diverse practices in 10 states in the United States. We enrolled 207 pathologist participants whose practice included the interpretation of MSLs. Our findings indicated that the majority of pathologists (54.6%) were influenced toward a less severe diagnosis when patients were70 years of age, or by the presence of tumor regression or solar elastosis (58.5%, 71.0%, and 57.0%, respectively). Generally, pathologists with dermatopathology board certification and/or a high caseload of MSLs were more likely to be influenced, whereas those with more years\u27 experience interpreting MSL were less likely to be influenced. Our findings indicate that the interpretation of MSLs is influenced by patient age, tumor regression, and solar elastosis; such influence is associated with dermatopathology training and higher caseload, consistent with expertise and an appreciation of lesion complexity

Revision of the Melanocytic Pathology Assessment Tool and Hierarchy for Diagnosis Classification Schema for Melanocytic Lesions: A Consensus Statement

IMPORTANCE A standardized pathology classification system for melanocytic lesions is needed to aid both pathologists and clinicians in cataloging currently existing diverse terminologies and in the diagnosis and treatment of patients. The Melanocytic Pathology Assessment Tool and Hierarchy for Diagnosis (MPATH-Dx) has been developed for this purpose. OBJECTIVE To revise the MPATH-Dx version 1.0 classification tool, using feedback from dermatopathologists participating in the National Institutes of Health-funded Reducing Errors in Melanocytic Interpretations (REMI) Study and from members of the International Melanoma Pathology Study Group (IMPSG). EVIDENCE REVIEW Practicing dermatopathologists recruited from 40 US states participated in the 2-year REMI study and provided feedback on the MPATH-Dx version 1.0 tool. Independently, member dermatopathologists participating in an IMPSG workshop dedicated to the MPATH-Dx schema provided additional input for refining the MPATH-Dx tool. A reference panel of 3 dermatopathologists, the original authors of the MPATH-Dx version 1.0 tool, integrated all feedback into an updated and refined MPATH-Dx version 2.0. FINDINGS The new MPATH-Dx version 2.0 schema simplifies the original 5-class hierarchy into 4 classes to improve diagnostic concordance and to provide more explicit guidance in the treatment of patients. This new version also has clearly defined histopathological criteria for classification of classes I and II lesions; has specific provisions for the most frequently encountered low-cumulative sun damage pathway of melanoma progression, as well as other, less common World Health Organization pathways to melanoma; provides guidance for classifying intermediate class II tumors vs melanoma; and recognizes a subset of pT1a melanomas with very low risk and possible eventual reclassification as neoplasms lacking criteria for melanoma. CONCLUSIONS AND RELEVANCE The implementation of the newly revised MPATH-Dx version 2.0 schema into clinical practice is anticipated to provide a robust tool and adjunct for standardized diagnostic reporting of melanocytic lesions and management of patients to the benefit of both health care practitioners and patients

Cellular Neurothekeoma in a Female with Guillain-Barré Syndrome: A Case Report and Review of the Literature

Cellular neurothekeoma is a rare cutaneous tumor that occurs more frequently in women. A 68-year-old female with a history of left nasal alar basal cell carcinoma and Guillain-Barré syndrome presented to the clinic with a 3-mm firm skin-colored papule with scattered telangiectasias. Histopathologic examination with immunochemistry of the lesion was consistent with cellular neurothekeoma. It stained positive for microphthalmia transcription factor and NKI-C3 and negative for HMB-45 and S-100. The lesion was excised with 3-mm margins, and no recurrence was noted within 1 year of follow-up. We present a case of cellular neurothekeoma in a patient with a history of Guillain-Barré syndrome as well as a review of the literature. Our case report is unique in that no prior association has been found in the literature between cellular neurothekeoma and Guillain-Barré syndrome. © 2014 S. Karger AG, Base

Vessel co-option and angiotropic extravascular migratory metastasis: a continuum of tumour growth and spread?

Two fields of cancer research have emerged dealing with the biology of tumour cells localised to the abluminal vascular surface: vessel co-option (VCo), a non-angiogenic mode of tumour growth and angiotropic extravascular migratory metastasis (EVMM), a non-hematogenous mode of tumour migration and metastasis. VCo is a mechanism by which tumour cells gain access to a blood supply by spreading along existing blood vessels in order to grow locally. Angiotropic EVMM involves “pericytic mimicry” (PM), which is characterised by tumour cells continuously migrating in the place of pericytes distantly along abluminal vascular surfaces. When cancer cells are engaged in PM and EVMM, they migrate along blood vessels beyond the advancing front of the tumour to secondary sites with the formation of regional and distant metastases. In the present perspective, the authors review the current scientific literature, emphasising the analogies between embryogenesis and cancer progression, the re-activation of embryonic signals by “cancer stem cells”, and the important role of laminins and epithelial-mesenchymal-transition. This perspective maintains that VCo and angiotropic EVMM constitute complementary processes and represent a continuum of cancer progression from the primary tumour to metastases and of tumour growth to EVMM, analogous to the embryonic development program

Cutaneous melanoma with brain metastasis: Report of 193 patients with new observations

BACKGROUND: Brain metastasis is a common endpoint in patients suffering from malignant melanoma. However, little is known about factors that predispose to brain metastases. OBJECTIVE: We performed a retrospective clinical and pathological investigation of melanoma patients with brain metastases in order to better characterise this patient population. METHODS: 193 melanoma patients with brain metastasis histologically diagnosed between 1990 and 2015 at the University Hospital Zurich were retrospectively identified and further specified for sex, age at diagnosis and detection of brain metastasis, and localisation. In addition, data were extracted regarding the subtype of primary melanoma, Breslow tumour thickness, Clark Level, mutation status, extent of metastatic spread and history of a second melanoma. RESULTS: We found a significant male predominance (n = 126/193; 65%; p < 0.001). Breslow tumour thickness showed a wide range from 0.2 to 12.0 mm (n = 99; median 2.3 mm). 14 of 101 melanomas (14%) were classified as T1, thereof 11 (79%) were found in men. In 32 of 193 patients (17%), the primary melanoma was unknown. CONCLUSIONS: Of special interest in our series is the high incidence of male predominance (79%) in cases of thin metastasing melanoma (14%), implicating genetic or epigenetic (hormonal) gender differences underlying tumour progression. Additionally, the high percentage of unknown primary melanoma (17%), at least partly representing completely regressed melanomas, indicates the importance of immune surveillance in melanoma progression

TERT promoter mutations are predictive of aggressive clinical behavior in patients with spitzoid melanocytic neoplasms

Spitzoid neoplasms constitute a morphologically distinct category of melanocytic tumors, encompassing Spitz nevus (benign), atypical Spitz tumor (intermediate malignant potential), and spitzoid melanoma (fully malignant). Currently, no reliable histopathological criteria or molecular marker is known to distinguish borderline from overtly malignant neoplasms. Because TERT promoter (TERT-p) mutations are common in inherently aggressive cutaneous conventional melanoma, we sought to evaluate their prognostic significance in spitzoid neoplasms. We analyzed tumors labeled as atypical Spitz tumor or spitzoid melanoma from 56 patients with available follow-up data for the association of TERT-p mutations, biallelic CDKN2A deletion, biallelic PTEN deletion, kinase fusions, BRAF/NRAS mutations, nodal status, and histopathological parameters with risk of hematogenous metastasis. Four patients died of disseminated disease and 52 patients were alive and disease free without extranodal metastasis (median follow-up, 32.5 months). We found TERT-p mutations in samples from the 4 patients who developed hematogenous metastasis but in none of tumors from patients who had favorable outcomes. Presence of TERT-p mutations was the most significant predictor of haematogenous dissemination (P < 0.0001) among variables analyzed. We conclude that TERT-p mutations identify a clinically high-risk subset of patients with spitzoid tumors. Application of TERT-p mutational assays for risk stratification in the clinic requires large-scale validation