The c.386A>C p.(Asn129Thr) variant in SMAD4 is likely to be pathogenic, causing Juvenile Polyposis Syndrome. A case report of a mosaic variant

Abstract

Background

Juvenile Polyposis Syndrome (JPS) is a rare autosomal dominant hereditary disorder characterized by the development of multiple hamartomatous gastrointestinal polyps. Here, we present a case of JPS with a mosaic variant in SMAD4.

Methods

Exome sequencing TRIO analysis, using germline DNA from the biological mother and father along with the index case (IC).

Results

A 46‐year‐old male with no family history of cancer presented with chronic iron deficiency anemia and was diagnosed with massive gastric polyposis (≥100 polyps). At the age of 59, he underwent a total gastrectomy, revealing numerous polyps occupying the entire gastric mucosa, including a 5 cm gastric hyperplastic polyp with high‐grade dysplasia and focal adenocarcinoma. TRIO analysis identified the c.386A>C p.(Asn129Thr) variant in the SMAD4 gene at an allele frequency (AF) of 22%, suggesting its mosaic origin. Subsequently, the variant was found in heterozygosity in the IC's son, who exhibited two subcentimeter polyps in the colon and seven inflammatory gastric polyps with gastric inflammatory areas and hyperplasia, suggesting that the c.386A>C p.(Asn129Thr) variant in SMAD4 segregated with the phenotype.

Conclusion

Our study provides evidence supporting the classification of the c.386A>C p.(Asn129Thr) variant in SMAD4 as a likely pathogenic variant. This finding contributes to improved accuracy in the diagnosis and genetic counseling of JPS.

Our study provides evidence supporting the classification of the c.386A>C p.(Asn129Thr) mutation in SMAD4 as a likely pathogenic variant. This finding contributes to improved accuracy in the diagnosis and genetic counseling of JPS.

Juvenile Polyposis Syndrome (JPS) (OMIM 174900) is a rare autosomal‐dominant hereditary disorder characterized by the development of multiple hamartomatous gastrointestinal polyps and a significant risk of gastrointestinal neoplasias (Chow & Macrae, 2005; Liu et al., 2021; Syngal et al., 2015). In 40%–60% of the patients diagnosed with JPS, a germline pathogenic loss of function mutation in either SMAD4 (#600993) or BMPR1A (#601299) can be identified (Calva‐Cerqueira et al., 2009). These genes encode proteins involved in the transforming growth factor‐beta (TGF‐β) signaling pathway (Zhao et al., 2018). SMAD4 mutations are associated with a higher incidence of gastric polyps and massive gastric polyposis (Blatter et al., 2020), and, unlike BMPR1A pathogenic carries, SMAD4 loss of function mutations have been associated with hereditary hemorrhagic telangiectasia (HHT) (OMIM 175050) (O'Malley et al., 2012). SMAD4 mutations have been described in JPS, with approximately 75% of cases inherited from a parent and 25% representing de novo pathogenic variants (Boland et al., 2022). Here, we describe a case of massive gastric polyposis in a patient harboring a mosaic variant identified by a Clinical Exome Sequencing TRIO analysis.

A 46‐year‐old male with no family history of cancer and chronic iron deficiency anemia was sent to the high‐risk clinic (Gastroenterology Department). The patient reported minor rectal bleeding. Due to the clinical symptoms, the patient underwent an upper oesophagogastroduodenoscopy and he was diagnosed with massive gastric polyposis (≥100 polyps). Biopsies revealed hyperplastic inflammatory polyps, some of them with low‐grade dysplasia. In addition, for the last 14 years, he has undergone surveillance colonoscopies with the removal of multiple polyps throughout the colon (16 adenomatous, 4 hamartomatous, and 19 inflammatory polyps), therefore, meeting diagnostic criteria for JPS (Boland et al., 2022). He had no hypoalbuminemia or edema, but hypogammaglobulinemia was noticed in a laboratory test. Helicobacter pylori tested negative.

In September 2021, at the age of 59, the patient presented with impaired anemia (hemoglobin 6.6 g/dL, hematocrit 24.3%), and upper endoscopy showed a nonresectable giant polyp (≈5 cm) in the proximal gastric body (Figure 1a).

FIGURE 1.

(a) Gastroscopy showing multiple gastric polyps. The nonresectable giant polyp (≈5 cm) in the proximal gastric body is denoted by an asterisk. (b) Electropherogram of the SMAD4 sequence in the adenocarcinoma sample, patient blood (c), and the hyperplasic polyp (d).

The patient underwent a total gastrectomy. Macroscopic examination of the gastrectomy specimen showed uncountable polyps that practically occupied the entire gastric mucosa. The histologic exam demonstrated a 5 cm gastric hyperplasic polyp with high‐grade dysplasia and focal adenocarcinoma. Noteworthy, 12 months after surgery, the patient has completely recovered from the anemia (hemoglobin 15.6 g/dL).

The patient was referred for genetic testing. Pretest genetic counseling was carried out and clinical consent for genetic testing was obtained. Written informed consent was also obtained from the patient for publication of this case report as well as the accompanying images. Germline DNA was extracted from peripheral blood using the Maxwell® RSC whole blood DNA kit. Multigene panel testing using the Hereditary Cancer Solution (HCS) panel (Sophia Genetics) was performed. This panel is designed to detect single‐nucleotide variants (SNVs), small insertion or deletion events (Indels), and copy number variations (CNVs). As well as Alu insertions, confident discrimination between variants in PMS2CL from the causative ones in PMS2 (#600259) and reliable assessment of the Boland inversion in the MSH2 (#609309) gene. Information regarding genes included in this panel is available in the Supporting Information. The study revealed no pathogenic mutations in PTEN (#608309), APC (#611731), MUTYH (#604933), or CDH1 (#192090), excluding the diagnosis of Cowden Syndrome, Familial Adenomatous Polyposis or Hereditary Diffuse Gastric Cancer. Since SMAD4 and BMPR1A were not included in the panel and since the patient had no family history of gastric polyposis, a clinical exome sequencing TRIO analysis was conducted, using germline DNA from the biological mother and father along with the index case (IC) using the Clinical Exome Solution v.3 (Sophia Genetics), which targets over 4000 genes, and a NextSeq 550Dx® (Illumina) sequencer. Data were analyzed using the Sophia DDM‐V4 (Sophia Genetics) data analysis platform. Detailed information regarding TRIO analysis is available in the Supporting Information. Written informed consent was obtained from progenitors. Variant analysis comparing the findings in the IC against the progenitors identified the c.386A>C p.(Asn129Thr) variant in SMAD4 (NM_005359.6) in the IC at an allele frequency (AF) of 22%. Sanger sequencing further confirmed these results. Therefore, the c.386A>C p.(Asn129Thr) variant was cataloged as mosaicism, indicating that this variant occurred during embryonic development, leading to a mixture of cells with different genetic information. No pathogenic mutations were identified in BMPR1A.

The genetic study of the piece of the gastrectomy was also conducted. Samples from the adenocarcinoma and a hyperplasic polyp were analyzed by Sanger Sequencing. The variant was present in both samples. In addition, visual inspection of the electropherogram revealed that the peak height corresponding to the cytosine was higher in the adenocarcinoma sample (representing approximately 2/3 of the adenine peak height) compared with the hyperplasic polyp (Figure 1b).

It should be noted that mosaicisms are sometimes missed on sequencing analyses, potentially resulting in the underdiagnosis of genetic conditions. Moreover, pathogenic mosaic variants are associated with highly variable clinical expressivity depending on the tissue affected and the proportion of cells with the variant, which may be inferred by the AF. Mosaic mutations in SMAD4 can lead to a wide range of clinical presentations, including different types and numbers of polyps, as well as other gastrointestinal and non‐gastrointestinal manifestations. In this way, the study with capsule endoscopy and thyroid ultrasound showed no significant findings. Chest and abdominal CT discarded pulmonary arteriovenous malformation. Magnetic resonance imaging ruled out brain arteriovenous malformations and further dermatology physical examination did not reveal any other mucocutaneous lesions, such as telangiectasias. Since pathogenic mutations in SMAD4 have also been associated with aortopathy and mitral valve dysfunction (Heald et al., 2015), a transthoracic echocardiogram was also performed that did not reveal any pathological findings.

Finally, the offspring of the CI were offered genetic testing. Written informed consent was obtained. One son of the CI aged 30 years was diagnosed as a SMAD4 c.386A>C p.(Asn129Thr) carrier. Gastroenterological examination revealed two subcentimeter polyps in the colon and seven inflammatory gastric polyps with gastric inflammatory areas and hyperplasia. The patient did not present with anemia.

The c.386A>C p.(Asn129Thr) variant is absent in population databases (PM2) (gnomAD v2.1.1 and UK Biobank last accessed June 2023). This amino acid position is highly conserved. Seventeen out of 20 in silico tools (PolyPhen‐2, DEOGEN2, EIGEN, EIGEN PC, Mutation assessor, MutPred, PrimateAI, EVE, FATHMM‐MKL, FATHMM‐XF, LIST‐S2, LRT, M‐CAP, PROVEAN, SIFT, SIFT4G, and BLOSUM) predict a damaging effect of the variant on protein function (PP3). Metascores such as REVEAL also classified the variant as damaging. Taking into account that this is a mosaic variant, that the patient's phenotype is highly concordant with JPS caused by a pathogenic mutation in the SMAD4 gene (PS2), and that it segregates with the phenotype (PP1), the variant was cataloged as a likely pathogenic (class 4) following the American College of Medical Genetics and Genomics (ACMG) recommendations (Richards et al., 2015).

In conclusion, here we report a mosaic variant in SMAD4 in a patient diagnosed with JPS. Clinical diagnosis of JPS remains challenging since polyps show a wide histological variation. Moreover, diagnosis of JPS due to mosaic mutations in SMAD4 can be challenging, as the mutation may not be present in all cells of the body and may not be detectable by standard genetic testing methods. However, TRIO clinical exome sequencing in highly selected patients may result in high molecular diagnostic yield. Finally, we classify the c.386A>C p.(Asn129Thr) variant in SMAD4 as a likely pathogenic, allowing for more accurate diagnosis and genetic counseling.

AUTHOR CONTRIBUTIONS

Conceptualization, FV, AHT, and AR; NGS analysis, EGB, NGS, MP, JLC, and AR; Clinical data acquisition FV, AHT, and AS; writing—original draft, FV, AHT, and AR. All authors have read and agreed to the published version of the manuscript.

CONFLICT OF INTEREST STATEMENT

The authors have declared no conflict of interest.

ETHICAL COMPLIANCE

The study was approved by the ethics committee of Hospital Puerta de Hierro (internal code: PI_169‐18). Pretest genetic counseling was carried out and clinical consent for genetic testing was obtained. Written informed consent was also obtained from the patient for publication of this case report as well as the accompanying images.

Supporting information

Data S1: Supporting Information.

Valentín, F. , de Tejada, A. H. , Gonzaléz‐Vioque, E. , García‐Simón, N. , Sánchez, A. , & Romero, A. (2024). The c.386A>C p.(Asn129Thr) variant in SMAD4 is likely to be pathogenic, causing Juvenile Polyposis Syndrome. A case report of a mosaic variant. Molecular Genetics & Genomic Medicine, 12, e2348. 10.1002/mgg3.2348

DATA AVAILABILITY STATEMENT

The data supporting the study's findings are available from the corresponding author, AR, upon reasonable request.