Age and Gender Associations of Virus Positivity in Merkel Cell Carcinoma Characterized Using a Novel RNA <i>In Situ</i> Hybridization Assay

Wang, Lisha; Harms, Paul W.; Palanisamy, Nallasivam; Carskadon, Shannon; Cao, Xuhong; Siddiqui, Javed; Patel, Rajiv M.; Zelenka-Wang, Sylvia; Durham, Alison B.; Fullen, Douglas R.; Harms, Kelly L.; Su, Fengyun; Shukla, Sudhanshu; Mehra, Rohit; Chinnaiyan, Arul M.

doi:10.1158/1078-0432.ccr-17-0299

Cited by 35 publications

(35 citation statements)

References 38 publications

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“…Currently, the methods used for the experimental detection or diagnosis of MCV in tumours include immunohistochemistry, PCR, RNA or DNA in situ hybridization and next-generation sequencing (NGS) 3,12,[33][34][35][36][37] . All of these assays vary considerably in terms of sensitivity and specificity for MCV detection in tumours.…”

Section: Discussionmentioning

confidence: 99%

“…Technical factors, including inefficient PCR amplification owing to mutations in the integrated MCPyV genome, primer incompatibilities, low purity of tumour samples or the detection of infectious wild-type MCPyV in the adjacent nonmalignant skin often confound the results for such tests 19,35 . Besides, qPCR does not allow for visual confirmation that positive results are associated with tumour cells; hence, background MCV presence cannot be excluded in MCC tumours with low signal 35 .…”

Section: Discussionmentioning

confidence: 99%

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DETECTing Merkel cell Polyomavirus in Merkel Tumours

Arora

Gupta

Vijaykumar

et al. 2019

Preprint

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Merkel cell carcinoma (MCC) is a rare, aggressive skin cancer caused either by Merkel cell polyomavirus (MCV) T antigen expression, post integration (~80% cases), or by UV mediated DNA damage. Interestingly, overall survival of patients suffering from MCV positive Merkel cell carcinoma is better, making this differential information of significant diagnostic and prognostic value. Also, MCV as a causative agent also provides a direct target for therapy in virus positive MCC patients. Currently, the methods used for diagnosis of MCV in tumours are often tedious, discordant and unreliable. In this study we used a guided molecular scissors based -DNA Endonuclease Targeted CRISPR Trans Reporter (DETECTR) technique to develop an in vitro molecular diagnostic tool for MCV positive MCC. DETECTR couples recombinase polymerase based amplification of target MCV DNA with Cas12a mediated detection. CRISPR diagnostics couple specific detection followed by cutting of the pathogenic DNA by the Cas enzyme -gRNA complex, with non-specific cutting of ssDNA that provides a measurable visual cue. To detect MCV DNA in MCC tumours, we designed Cas12a gRNAs targeting the MCV DNA and tested their targeting efficiency, and sensitivity using a fluorophore quencher labeled reporter assay. We show that this sophisticated MCV DETECTR system can detect MCV integrated in Merkel tumour rapidly, specifically and with femto-molar sensitivity. This new MCV DNA detecting system is promising and we hope it can be coupled with histopathological and immunohistochemical studies to diagnose the viral status of MCC in clinics in the near future.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

DETECTing Merkel cell Polyomavirus in Merkel Tumours

Arora

Gupta

Vijaykumar

et al. 2019

Preprint

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“…no. 322350; Advanced Cell Diagnostics, Newark, CA, USA) was performed using target probes to MCPyV on tissue sections according to the manufacturer's instructions and methods followed by Wang et al 24 Human peptidylprolyl isomerase B (Hs-PPIB) and bacterial dihydrodipicolinate reductase (DapB) were used as positive and negative controls, respectively. FFPE HeLa cells (Advanced Cell Diagnostics) were also used as negative and positive controls, as suggested by Advanced Cell Diagnostics.…”

Section: Rna-in Situ Hybridizationmentioning

confidence: 99%

Low prevalence of Merkel cell polyomavirus in human epithelial thymic tumors

et al. 2019

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Background The etiology of thymic epithelial tumors is unknown. Murine polyomavirus strain PTA has been shown to induce thymomas in mice. Recently, using diverse molecular techniques, we reported the presence of human polyomavirus 7 (HPyV7) in thymic epithelial tumors. In the present study, we investigated the prevalence of Merkel cell polyomavirus (MCPyV) in thymic epithelial tumors. Methods Thirty‐six thymomas were screened for MCPyV by PCR and subsequently tested by DNA and RNA in situ hybridization and immunohistochemistry. Twenty‐six thymomas were diagnosed with myasthenia gravis (MG). Results MCPyV DNA was detected by PCR in 7 (19.4%) of the 36 thymic epithelial tumors and in six of these, the presence of MCPyV was confirmed by fluorescence situ hybridization. Of these, 3 (28.6%) revealed weak MCPyV LT‐antigen protein expression. In addition, one of the MCPyV positive thymomas tested positive for MCPyV LT RNA with RNAscope. Of interest, two out of the three thymomas that previously tested positive for MCPyV by immunohistochemistry also tested positive for HPyV7. One of the 11 MG‐negative and 2 of the 25 MG‐positive were positive for MCPyV. Conclusions MCPyV DNA and MCPyV protein expression can be detected in human epithelial thymoma; however, to a far lesser extent than HPyV7. Our data strongly indicate that because of its infrequent detection and weak expression, MCPyV is unlikely to play an important role in the etiopathogenesis of human thymomas.

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“…1 MCPyV, present in 60% to 80% of MCC tumors, may be detected by immunohistochemistry, RNA in situ hybridization, or polymerase chain reaction (PCR). 1,2,23 MCCIS displays similar patterns of diagnostic marker expression, although staining may be more focal, and cytoplasmic cytokeratin staining may be more frequent. 22 MCPyV has been reported in association with one case of MCCIS; other cases have been negative for MCPyV, or were not tested.…”

Section: Introductionmentioning

confidence: 99%

“…Of note, MCC may express certain lymphoid markers such as TdT or PAX5; however, CD45, CD3, and CD20 are reliably negative . MCPyV, present in 60% to 80% of MCC tumors, may be detected by immunohistochemistry, RNA in situ hybridization, or polymerase chain reaction (PCR) . MCCIS displays similar patterns of diagnostic marker expression, although staining may be more focal, and cytoplasmic cytokeratin staining may be more frequent .…”

Section: Introductionmentioning

confidence: 99%

Merkel cell carcinoma arising in association with cutaneous T‐cell lymphoma: A potential diagnostic pitfall

et al. 2019

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Merkel cell carcinoma (MCC) is a rare, aggressive cutaneous neuroendocrine carcinoma with increased prevalence in patients with immunosuppression or B‐cell neoplasms. To the best of our knowledge, an association with cutaneous T‐cell lymphoma (CTCL) has not been previously described. In this report, we present two cases of MCC arising in the setting of CTCL. The first case was a female during her 70s with previously diagnosed stage IVA1 Sezary syndrome. Biopsy of a scaly patch showed two distinct abnormal cell populations. The first population consisted of hyperchromatic dermal and epidermotropic lymphocytes, expressing CD3 and CD4 with diminished CD7. The second population consisted of intraepidermal clusters of larger atypical cells that expressed synaptophysin, neurofilament, CK20, and Merkel cell polyomavirus transcript. The combination of findings was consistent with intraepidermal MCC in a background of CTCL. Excision showed residual intraepidermal MCC without dermal involvement. The second case was a male during his 50s with a longstanding history of mycosis fungoides, who presented with a new lesion on his right thigh. Biopsy and excision showed dermal MCC without secondary involvement by CTCL. Our cases show that MCC may rarely occur in the setting of T‐cell lymphoma, and that intraepidermal MCC may mimic epidermotropic T‐cells.

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Age and Gender Associations of Virus Positivity in Merkel Cell Carcinoma Characterized Using a Novel RNA In Situ Hybridization Assay

Cited by 35 publications

References 38 publications

DETECTing Merkel cell Polyomavirus in Merkel Tumours

DETECTing Merkel cell Polyomavirus in Merkel Tumours

Low prevalence of Merkel cell polyomavirus in human epithelial thymic tumors

Merkel cell carcinoma arising in association with cutaneous T‐cell lymphoma: A potential diagnostic pitfall

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