A Novel Method to Detect Copy Number Variation in Melanoma: Droplet Digital PCR for Quantitation of the CDKN2A Gene, a Proof-of-Concept Study

McFadden, Jason R.; Syku, Marie; Barney, Rachael E.; Stevanovic, Mirjana; Chaudhari, Advaita S.; O’Hern, Keegan; Chambers, Meagan; Baker, Catherine; LeBlanc, Robert E.; Doan, Linda; Tsongalis, Gregory J.; Hughes, Edward G.; Sriharan, Aravindhan

doi:10.1097/dad.0000000000002436

Cited by 2 publications

(4 citation statements)

References 23 publications

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“…successfully to detect CNVs in different malignancies, including melanocytic neoplasms. 5,6,[13][14][15] In our current study, we observed an acceptable concordance between ddPCR and CMA, with MYC and MYB exhibiting concordant results in 93.2% and 85.0% of cases, respectively. Similarly, studies have reported concordance rates ranging from approximately 70%-90% between FISH and CMA.…”

Section: Discussionsupporting

confidence: 68%

“…The most widely recognized CNVs in melanoma occur on chromosomal loci 6q23 (MYB), 6p24 (RREB1), 8q24 (MYC), 9q21 (CDKN2A), 11q13 (CCND1), and 12q13 (CDK4). 2,[4][5][6][7][8]11,12 The exact pathophysiologic mechanisms by which these genetic alterations contribute to melanomagenesis are not completely understood, but their implications in the evolution of melanoma are well-established. They constitute the basis of our current advances in the field of molecular genomics in pigmented skin lesions.…”

Section: Discussionmentioning

confidence: 99%

“…In our preliminary pilot studies, we established a successful correlation between digital PCR and CMA for the RREB1 and CDKN2A genes using a singlicate ddPCR assay. 5,6 Building upon this foundation, our current proof-of-concept investigation aimed to develop a multiplex ddPCR assay capable of simultaneous detecting chromosomal alterations on two genes within a single well/reaction. We specifically focused on targeting the MYC and MYB genes, two wellestablished contributors to the pathogenesis of melanoma.…”

Section: Introductionmentioning

confidence: 99%

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A novel method to assess copy number variations in melanocytic neoplasms: Droplet digital PCR for precise quantitation of MYC and MYB genes

Ramos‐Rodriguez,

McFadden,

Momtahen

et al. 2023

J Cutan Pathol

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IntroductionWhile most melanocytic neoplasms can be classified as either benign or malignant by histopathology alone, ancillary molecular diagnostic tests can be necessary to establish the correct diagnosis in challenging cases. Currently, the detection of copy number variations (CNVs) by fluorescence in situ hybridization and chromosomal microarray (CMA) are the most popular methods, but remain expensive and inaccessible. We aim to develop a relatively inexpensive, fast, and accessible molecular assay to detect CNVs relevant to melanoma using droplet digital polymerase chain reaction (ddPCR) technology.MethodsIn this proof‐of‐concept study, we evaluated CNVs in MYC and MYB genes from 73 cases of benign nevi, borderline melanocytic lesions, and primary and metastatic melanoma at our institution from 2015 to 2022. A multiplexed ddPCR assay and CMA were performed on each sample, and the results were compared.ResultsConcordance analysis of ddPCR with CMA for quantification of MYC and MYB CNVs revealed a sensitivity and specificity of 89% and 86% for MYC and 83% and 74% for MYB, respectively.ConclusionWe demonstrate the first use of a multiplexed ddPCR assay to identify CNVs in melanocytic neoplasms. With further improvement and validation, ddPCR may represent a low‐cost and rapid tool to aid in the diagnosis of histopathologically ambiguous melanocytic tumors.

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Section: Discussionsupporting

confidence: 68%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A novel method to assess copy number variations in melanocytic neoplasms: Droplet digital PCR for precise quantitation of MYC and MYB genes

Ramos‐Rodriguez,

McFadden,

Momtahen

et al. 2023

J Cutan Pathol

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“…Although several studies have identified copy number changes of CDK genes in tissue samples 39 – 41 , there is limited information on their capture from the plasma of MM patients. Thus, we decided to analyze CNVs in these genes in circulating ctDNA from MM patients and determine whether these aberrations may serve as a suitable biomarker for the noninvasive diagnosis of melanoma patients.…”

Section: Discussionmentioning

confidence: 99%

Novel liquid biopsy CNV biomarkers in malignant melanoma

Lukacova,

Hanzlikova,

Podlesnyi

et al. 2024

Sci Rep

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Malignant melanoma (MM) is known for its abundance of genetic alterations and a tendency for rapid metastasizing. Identification of novel plasma biomarkers may enhance non-invasive diagnostics and disease monitoring. Initially, we examined copy number variations (CNV) in CDK genes (CDKN2A, CDKN2B, CDK4) using MLPA (gDNA) and ddPCR (ctDNA) analysis. Subsequently, low-coverage whole genome sequencing (lcWGS) was used to identify the most common CNV in plasma samples, followed by ddPCR verification of chosen biomarkers. CNV alterations in CDK genes were identified in 33.3% of FFPE samples (Clark IV, V only). Detection of the same genes in MM plasma showed no significance, neither compared to healthy plasmas nor between pre- versus post-surgery plasma. Sequencing data showed the most common CNV occurring in 6q27, 4p16.1, 10p15.3, 10q22.3, 13q34, 18q23, 20q11.21-q13.12 and 22q13.33. CNV in four chosen genes (KIF25, E2F1, DIP2C and TFG) were verified by ddPCR using 2 models of interpretation. Model 1 was concordant with lcWGS results in 54% of samples, for model 2 it was 46%. Although CDK genes have not been proven to be suitable CNV liquid biopsy biomarkers, lcWGS defined the most frequently affected chromosomal regions by CNV. Among chosen genes, DIP2C demonstrated a potential for further analysis.

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A Novel Method to Detect Copy Number Variation in Melanoma: Droplet Digital PCR for Quantitation of the CDKN2A Gene, a Proof-of-Concept Study

Abstract: Supplemental Digital Content is Available in the Text.

Cited by 2 publications

References 23 publications

A novel method to assess copy number variations in melanocytic neoplasms: Droplet digital PCR for precise quantitation of MYC and MYB genes

A novel method to assess copy number variations in melanocytic neoplasms: Droplet digital PCR for precise quantitation of MYC and MYB genes

Novel liquid biopsy CNV biomarkers in malignant melanoma

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