Genotype-first approach to identify associations between CDH1 germline variants and cancer phenotypes: a multicentre study by the European Reference Network on Genetic Tumour Risk Syndromes

Abstract

Background: Truncating pathogenic or likely pathogenic variants of CDH1 cause hereditary diffuse gastric cancer (HDGC), a tumour risk syndrome that predisposes carrier individuals to diffuse gastric and lobular breast cancer. Rare CDH1 missense variants are often classified as variants of unknown significance. We conducted a genotype-phenotype analysis in families carrying rare CDH1 variants, comparing cancer spectrum in carriers of pathogenic or likely pathogenic variants (PV/LPV; analysed jointly) or missense variants of unknown significance, assessing the frequency of families with lobular breast cancer among PV/LPV carrier families, and testing the performance of lobular breast cancer-expanded criteria for CDH1 testing.

Methods: This genotype-first study used retrospective diagnostic and clinical data from 854 carriers of 398 rare CDH1 variants and 1021 relatives, irrespective of HDGC clinical criteria, from 29 institutions in ten member-countries of the European Reference Network on Tumour Risk Syndromes (ERN GENTURIS). Data were collected from Oct 1, 2018, to Sept 20, 2022. Variants were classified by molecular type and clinical actionability with the American College of Medical Genetics and Association for Molecular Pathology CDH1 guidelines (version 2). Families were categorised by whether they fulfilled the 2015 and 2020 HDGC clinical criteria. Genotype-phenotype associations were analysed by Student's t test, Kruskal-Wallis, χ², and multivariable logistic regression models. Performance of HDGC clinical criteria sets were assessed with an equivalence test and Youden index, and the areas under the receiver operating characteristic curves were compared by Z test.

Findings: From 1971 phenotypes (contributed by 854 probands and 1021 relatives aged 1-93 years), 460 had gastric and breast cancer histology available. CDH1 truncating PV/LPVs occurred in 176 (21%) of 854 families and missense variants of unknown significance in 169 (20%) families. Multivariable logistic regression comparing phenotypes occurring in families carrying PV/LPVs or missense variants of unknown significance showed that lobular breast cancer had the greatest positive association with the presence of PV/LPVs (odds ratio 12·39 [95% CI 2·66-57·74], p=0·0014), followed by diffuse gastric cancer (8·00 [2·18-29·39], p=0·0017) and gastric cancer (7·81 [2·03-29·96], p=0·0027). 136 (77%) of 176 families carrying PV/LPVs fulfilled the 2015 HDGC criteria. Of the remaining 40 (23%) families, who did not fulfil the 2015 criteria, 11 fulfilled the 2020 HDGC criteria, and 18 had lobular breast cancer only or lobular breast cancer and gastric cancer, but did not meet the 2020 criteria. No specific CDH1 variant was found to predispose individuals specifically to lobular breast cancer, although 12 (7%) of 176 PV/LPV carrier families had lobular breast cancer only. Addition of three new lobular breast cancer-centred criteria improved testing sensitivity while retaining high specificity. The probability of finding CDH1 PV/LPVs in patients fulfilling the lobular breast cancer-expanded criteria, compared with the 2020 criteria, increased significantly (AUC 0·92 vs 0·88; Z score 3·54; p=0·0004).

Interpretation: CDH1 PV/LPVs were positively associated with HDGC-related phenotypes (lobular breast cancer, diffuse gastric cancer, and gastric cancer), and no evidence for a positive association with these phenotypes was found for CDH1 missense variants of unknown significance. CDH1 PV/LPVs occurred often in families with lobular breast cancer who did not fulfil the 2020 HDGC criteria, supporting the expansion of lobular breast cancer-centred criteria.

Funding: European Reference Network on Genetic Tumour Risk Syndromes, European Regional Development Fund, Fundação para a Ciência e a Tecnologia (Portugal), Cancer Research UK, and European Union's Horizon 2020 research and innovation programme.

Figure 1

Flowchart of the genotype–phenotype study and molecular and clinical features of the cohort of families with rare germline CDH1 variants (A) Flowchart of the genotype–phenotype study. (B) Relative frequency of variant groups and clinical classification in families fulfilling or not fulfilling the 2015 HDGC criteria. A χ² test was used to compare the frequency of variant clinical classifications between families fulfilling and not fulfilling the 2015 HDGC criteria (appendix p 53). 14 of 14 coding non-truncating variants of unknown significance were missense variants occurring in families fulfilling the 2015 HDGC criteria. 155 of 159 coding non-truncating variants of unknown significance were missense variants occurring in families not fulfilling the 2015 HDGC criteria. Details on variant type and correspondence with clinical criteria and clinical classification are depicted in appendix (pp 47, 54). ACMG–AMP=American College of Medical Genetics and Genomics and Association for Molecular Pathology. HDGC=hereditary diffuse gastric cancer. LBV/BV=likely benign or benign variant. PV/LPV=pathogenic or likely pathogenic variant. VUS=variant of unknown significance. *Probands could contribute more than one phenotype. †Breast cancer of unknown histotype. ‡Other phenotypes not including gastric cancer, breast cancer, colorectal cancer, or ovarian cancer (detailed in appendix p 3).

Figure 1

Flowchart of the genotype–phenotype study and molecular and clinical features of the cohort of families with rare germline CDH1 variants (A) Flowchart of the genotype–phenotype study. (B) Relative frequency of variant groups and clinical classification in families fulfilling or not fulfilling the 2015 HDGC criteria. A χ² test was used to compare the frequency of variant clinical classifications between families fulfilling and not fulfilling the 2015 HDGC criteria (appendix p 53). 14 of 14 coding non-truncating variants of unknown significance were missense variants occurring in families fulfilling the 2015 HDGC criteria. 155 of 159 coding non-truncating variants of unknown significance were missense variants occurring in families not fulfilling the 2015 HDGC criteria. Details on variant type and correspondence with clinical criteria and clinical classification are depicted in appendix (pp 47, 54). ACMG–AMP=American College of Medical Genetics and Genomics and Association for Molecular Pathology. HDGC=hereditary diffuse gastric cancer. LBV/BV=likely benign or benign variant. PV/LPV=pathogenic or likely pathogenic variant. VUS=variant of unknown significance. *Probands could contribute more than one phenotype. †Breast cancer of unknown histotype. ‡Other phenotypes not including gastric cancer, breast cancer, colorectal cancer, or ovarian cancer (detailed in appendix p 3).

Figure 2

Distribution of phenotypes and cancer average age of onset, in probands and relatives from germline CDH1 variant carrier families, and multivariable logistic regression models for the association of cancer phenotypes with PV/LPV and missense-variant of unknown significance Relative frequency of phenotypes, phenotype distribution, number of cases, and average age of onset in probands and relatives in families carrying PVs or LPVs molecularly classified as truncating (A) and in families carrying variants of unknown significance molecularly classified as missense (B) (data further detailed in appendix p 55). The left axis shows the relative frequency (%) of each phenotype out of the total number of phenotypes. Mean age of onset (SD) is shown for HDGC-associated phenotypes. Detailed data on the number of cases and average age of onset for all phenotypes are provided in the appendix (p 55). (C) Forest plot showing the results of a multivariable logistic regression model for the association of each phenotype with the presence of a PV/LPV vs a missense VUS in 856 patients with available age of disease onset and clinical phenotype. Probands could contribute more than one phenotype. HDGC=hereditary diffuse gastric cancer. PV/LPV=pathogenic or likely pathogenic variant. P=probands. R=relatives. VUS=variants of unknown significance. *Breast cancer of unknown histotype. †Other phenotypes not including gastric cancer, breast cancer, colorectal cancer, or ovarian cancer (detailed in appendix p 3).

Figure 3

CDH1 PV/LPV carrier families and lobular breast cancer-related criteria (A) CDH1 single-nucleotide and copy-number variants occurring in PV/LPV carrier families not fulfilling the 2020 HDGC criteria. PV/LPVs found in the cohort are shown by CDH1 exon location. The size of each circle is proportional to the number of families found to carry a particular variant. (B) Proposed lobular breast cancer-expanded criteria and proportion of the 176 PV/LPV carrier families explained by these criteria. Lobular breast cancer-centred criteria 3, 4, and 5 are proposed as an addition to the 2020 HDGC criteria. dup=duplication. EX=exon. del=deletion. HDGC=hereditary diffuse gastric cancer. PV/LPV=pathogenic or likely pathogenic variant. *Families resolved by lobular breast cancer-centred criteria. †One of two families is resolved by the lobular breast cancer-centred criteria.

Figure 4

Distribution of phenotypes in probands and relatives from families carrying germline CDH1 PV/LPVs and having gastric cancer only, gastric cancer and breast cancer, and breast cancer only, with reference to HDGC clinical criteria Relative frequency of phenotypes, phenotype distribution, number of cases, and average age of onset of probands and relatives in families carrying PV/LPVs and having gastric cancer only, gastric cancer and breast cancer, or breast cancer only (data further detailed in appendix pp 62–63). The left axis shows the relative frequency (%) of each phenotype out of the total number of phenotypes. Mean age of onset (SD) is shown for HDGC-associated phenotypes. Detailed data on number of cases and average age of onset for other phenotypes are provided in appendix (pp 62–63). Probands could contribute more than one phenotype. HDGC=hereditary diffuse gastric cancer. PV/LPV=pathogenic or likely pathogenic variant. P=probands. R=relatives. VUS=variants of unknown significance. *Breast cancer of unknown histotype. †Other phenotypes not including gastric cancer, breast cancer, colorectal cancer, or ovarian cancer (detailed in appendix p 3).

Figure 4

Distribution of phenotypes in probands and relatives from families carrying germline CDH1 PV/LPVs and having gastric cancer only, gastric cancer and breast cancer, and breast cancer only, with reference to HDGC clinical criteria Relative frequency of phenotypes, phenotype distribution, number of cases, and average age of onset of probands and relatives in families carrying PV/LPVs and having gastric cancer only, gastric cancer and breast cancer, or breast cancer only (data further detailed in appendix pp 62–63). The left axis shows the relative frequency (%) of each phenotype out of the total number of phenotypes. Mean age of onset (SD) is shown for HDGC-associated phenotypes. Detailed data on number of cases and average age of onset for other phenotypes are provided in appendix (pp 62–63). Probands could contribute more than one phenotype. HDGC=hereditary diffuse gastric cancer. PV/LPV=pathogenic or likely pathogenic variant. P=probands. R=relatives. VUS=variants of unknown significance. *Breast cancer of unknown histotype. †Other phenotypes not including gastric cancer, breast cancer, colorectal cancer, or ovarian cancer (detailed in appendix p 3).

Figure 5

CDH1 variant classification and distribution according to compliance with 2020 HDGC clinical criteria, and performance evaluation of HDGC clinical criteria sets (A) Alluvial plot (created with the visualisation platform developed by Mauri and colleagues) displaying the number of families carrying CDH1 variants and fulfilling 2020 HDGC criteria or lobular breast cancer-expanded criteria. Variants are categorised according to the American College of Medical Genetics and Genomics and Association for Molecular Pathology (version 2). (B) Receiver operating characteristic analysis comparing the AUCs for four different clinical criteria sets (lobular breast cancer-expanded vs 2020 HDGC criteria Z=3·54, AUC difference 0·042 [95% CI 0·02–0·06], p=0·0004). (C) Forest plot representing the lobular breast cancer-expanded clinical criteria equivalence test. The equivalence limit (δ) was set to 0·12. HDGC=hereditary diffuse gastric cancer. LBV/BV=likely benign or benign variant. PV/LPV=pathogenic or likely pathogenic variant. VUS=variants of unknown significance.