1. Cancer Biology
  2. Genetics and Genomics

Functional CDKN2A assay identifies frequent deleterious alleles misclassified as variants of uncertain significance

  1. Hirokazu Kimura
  2. Raymond M Paranal
  3. Neha Nanda
  4. Laura D Wood
  5. James R Eshleman
  6. Ralph H Hruban
  7. Michael G Goggins
  8. Alison P Klein
  9. Nicholas Jason Roberts  Is a corresponding author
  1. Johns Hopkins University, United States
  2. The Johns Hopkins University, United States
https://doi.org/10.7554/eLife.71137
Share this article
Cite this article
  1. Hirokazu Kimura
  2. Raymond M Paranal
  3. Neha Nanda
  4. Laura D Wood
  5. James R Eshleman
  6. Ralph H Hruban
  7. Michael G Goggins
  8. Alison P Klein
  9. Nicholas Jason Roberts
(2022)
Functional CDKN2A assay identifies frequent deleterious alleles misclassified as variants of uncertain significance
eLife 11:e71137.
https://doi.org/10.7554/eLife.71137
  2. Download BibTeX
  3. Download .RIS

Abstract

Pathogenic germline CDKN2A variants are associated with an increased risk of pancreatic ductal adenocarcinoma (PDAC). CDKN2A variants of uncertain significance (VUSs) are reported in up to 4.3% of patients with PDAC and result in significant uncertainty for patients and their family members as an unknown fraction are functionally deleterious, and therefore, likely pathogenic. Functional characterization of CDKN2A VUSs is needed to reclassify variants and inform clinical management. 29 germline CDKN2A VUSs previously reported in patients with PDAC or in ClinVar were evaluated using a validated in vitro cell proliferation assay. 12 of the 29 CDKN2A VUSs were functionally deleterious (11 VUSs) or potentially functionally deleterious (1 VUS) and were reclassified as likely pathogenic variants. Thus, over 40% of CDKN2A VUSs identified in patients with PDAC are functionally deleterious and likely pathogenic. When incorporating VUSs found to be functionally deleterious, and reclassified as likely pathogenic, the prevalence of pathogenic/likely pathogenic CDKN2A in patients with PDAC reported in the published literature is increased to up to 4.1% of patients, depending on family history. Therefore, CDKN2A VUSs may play a significant, unappreciated role in risk of pancreatic cancer. These findings have significant implications for the counselling and care of patients and their relatives.

Data availability

All data generated or analysed during this study are included in the manuscript, supporting files, and as Source data and Source code.

Article and author information

Author details

  1. Hirokazu Kimura

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  2. Raymond M Paranal

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  3. Neha Nanda

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  4. Laura D Wood

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  5. James R Eshleman

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  6. Ralph H Hruban

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    Ralph H Hruban, has the right to receive royalty payments from Thrive Earlier Diagnosis for the GNAS in pancreatic cysts invention in a relationship overseen by Johns Hopkins University..

  7. Michael G Goggins

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  8. Alison P Klein

    Department of Pathology, Johns Hopkins University, Baltimore, United States
    Competing interests
    No competing interests declared.

  9. Nicholas Jason Roberts

    The Johns Hopkins University, Baltimore, United States
    For correspondence
    nrobert8@jhmi.edu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8709-0664

Funding

The Sol Goldman Pancreatic Cancer Research Center (Pilot Grant)

  • Nicholas Jason Roberts

National Institutes of Health (S10 OD026859)

  • Nicholas Jason Roberts

The Rolfe Pancreatic Cancer Foundation

  • Nicholas Jason Roberts

National Institutes of Health (P50 CA622924)

  • Alison P Klein
  • Nicholas Jason Roberts

The Japanese Society of Gastroenterology

  • Hirokazu Kimura

The Japan Society for Promotion of Science

  • Hirokazu Kimura

The Joseph C Monastra Foundation

  • Nicholas Jason Roberts

The Geral O Mann Foundation

  • Nicholas Jason Roberts

Art Creates Cures Foundation

  • Nicholas Jason Roberts

Susan Wojcicki and Denis Troper

  • Nicholas Jason Roberts

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Emil Lou, University of Minnesota, United States

Version history

  1. Received: June 9, 2021
  2. Accepted: January 6, 2022
  3. Accepted Manuscript published: January 10, 2022 (version 1)
  4. Version of Record published: February 8, 2022 (version 2)

Copyright

© 2022, Kimura et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,310
    views
  • 314
    downloads
  • 7
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Hirokazu Kimura
  2. Raymond M Paranal
  3. Neha Nanda
  4. Laura D Wood
  5. James R Eshleman
  6. Ralph H Hruban
  7. Michael G Goggins
  8. Alison P Klein
  9. Nicholas Jason Roberts
(2022)
Functional CDKN2A assay identifies frequent deleterious alleles misclassified as variants of uncertain significance
eLife 11:e71137.
https://doi.org/10.7554/eLife.71137

Share this article

https://doi.org/10.7554/eLife.71137

Categories and tags

Research organism

Further reading

    1. Cancer Biology
    2. Cell Biology

    A syngeneic spontaneous zebrafish model of tp53-deficient, EGFRvIII, and PI3KCAH1047R-driven glioblastoma reveals inhibitory roles for inflammation during tumor initiation and relapse in vivo

    Alex Weiss, Cassandra D'Amata ... Madeline N Hayes
    Research Article

    High-throughput vertebrate animal model systems for the study of patient-specific biology and new therapeutic approaches for aggressive brain tumors are currently lacking, and new approaches are urgently needed. Therefore, to build a patient-relevant in vivo model of human glioblastoma, we expressed common oncogenic variants including activated human EGFRvIII and PI3KCAH1047R under the control of the radial glial-specific promoter her4.1 in syngeneic tp53 loss-of-function mutant zebrafish. Robust tumor formation was observed prior to 45 days of life, and tumors had a gene expression signature similar to human glioblastoma of the mesenchymal subtype, with a strong inflammatory component. Within early stage tumor lesions, and in an in vivo and endogenous tumor microenvironment, we visualized infiltration of phagocytic cells, as well as internalization of tumor cells by mpeg1.1:EGFP+ microglia/macrophages, suggesting negative regulatory pressure by pro-inflammatory cell types on tumor growth at early stages of glioblastoma initiation. Furthermore, CRISPR/Cas9-mediated gene targeting of master inflammatory transcription factors irf7 or irf8 led to increased tumor formation in the primary context, while suppression of phagocyte activity led to enhanced tumor cell engraftment following transplantation into otherwise immune-competent zebrafish hosts. Altogether, we developed a genetically relevant model of aggressive human glioblastoma and harnessed the unique advantages of zebrafish including live imaging, high-throughput genetic and chemical manipulations to highlight important tumor-suppressive roles for the innate immune system on glioblastoma initiation, with important future opportunities for therapeutic discovery and optimizations.

    1. Cancer Biology
    2. Cell Biology

    Cancer: Modeling glioblastoma

    Ian Lorimer
    Insight

    Establishing a zebrafish model of a deadly type of brain tumor highlights the role of the immune system in the early stages of the disease.

    1. Cancer Biology

    YAP/TAZ enhances P-body formation to promote tumorigenesis

    Xia Shen, Xiang Peng ... Chen-Ying Liu
    Research Article

    The role of processing bodies (P-bodies) in tumorigenesis and tumor progression is not well understood. Here, we showed that the oncogenes YAP/TAZ promote P-body formation in a series of cancer cell lines. Mechanistically, both transcriptional activation of the P-body-related genes SAMD4A, AJUBA, and WTIP and transcriptional suppression of the tumor suppressor gene PNRC1 are involved in enhancing the effects of YAP/TAZ on P-body formation in colorectal cancer (CRC) cells. By reexpression of PNRC1 or knockdown of P-body core genes (DDX6, DCP1A, and LSM14A), we determined that disruption of P-bodies attenuates cell proliferation, cell migration, and tumor growth induced by overexpression of YAP5SA in CRC. Analysis of a pancancer CRISPR screen database (DepMap) revealed co-dependencies between YAP/TEAD and the P-body core genes and correlations between the mRNA levels of SAMD4A, AJUBA, WTIP, PNRC1, and YAP target genes. Our study suggests that the P-body is a new downstream effector of YAP/TAZ, which implies that reexpression of PNRC1 or disruption of P-bodies is a potential therapeutic strategy for tumors with active YAP.