Background: About 10%-15% of GISTs do not harbour KIT/PDGFRA mutations (wild-type, WT). We analyzed the tumor transcriptome of two sporadic young adults WT GISTs patients using massively parallel sequencing approach in order to identify novel genetic mutations. Methods: Patients were a 28 years-old female (GIST_07) and a 30 years-old male (GIST_10). Primary tumors were in the stomach, multi-centric in GIST_07 and uni-centric in GIST_10, and presented metastases at diagnosis. RNA-Seq paired-end libraries were sequenced on the Illumina GAIIx system using a paired-end strategy at a read length of 75bp. Bioinformatic analysis highlighted 9 candidate genes common to both patients. Mutations in these genes were then filtered with SNP&GO, a tool suited to compute the likelihood of a mutation to be disease-related or not, depending on the protein sequence and its functional annotation. The identified candidate gene was analyzed by Sanger sequencing on tumor (T) and peripheral blood (PB) DNA from GIST_07 and GIST_10 patients, and on additional 8 tumor and 3 PB DNA from 8 sporadic WT GISTs patients. Immunohistochemistry (IHC) was also performed. Results: Only mutations in the sequence of SDHA were identified as disease-related with a high reliability index. GIST_07 tumor DNA was homozygous for an exon 9 p.S384X nonsense mutation; GIST_10 patient showed compound heterozygosity for a germinal exon 2 p.R31X nonsense mutation and a somatic exon 13 p.R589W missense mutation.Heterozygous nonsense mutations were already present in germline DNA in both patients. All mutations lead to functional inactivation of the protein. IHC revealed that both patients showed the down regulation of SDHA. The expanded study showed that 2/10 carried SDHA mutations in the tumor: compound heterozygosity for an exon 9 p.G419R and exon 13 p.E564K mutations in one patient, and a somatic heterozygous exon 5 p.R171C mutation in the other. Conclusions: Recently, SDHB and SDHC mutations were also reported in these patients. This the first study that identifies mutations on SDHA, in 4/10 WT GIST patients. This gene should be evaluated in larger seriesand be added in the sequencing screening of WT GISTs.

Identification of SDHA (subunit A of the succinate dehydrogenase) mutations in KIT/PDGFRA WT gastrointestinal stromal tumors (GISTs).

PANTALEO, MARIA ABBONDANZA;ASTOLFI, ANNALISA;INDIO, VALENTINA;PATERINI, PAOLA;FORMICA, SERENA;CASADIO, RITA;MARTELLI, PIER LUIGI;MALEDDU, ALESSANDRA;NANNINI, MARGHERITA;SANTINI, DONATELLA;CATENA, FAUSTO;CECCARELLI, CLAUDIO;FIORENTINO, MICHELANGELO;BIASCO, GUIDO
2011

Abstract

Background: About 10%-15% of GISTs do not harbour KIT/PDGFRA mutations (wild-type, WT). We analyzed the tumor transcriptome of two sporadic young adults WT GISTs patients using massively parallel sequencing approach in order to identify novel genetic mutations. Methods: Patients were a 28 years-old female (GIST_07) and a 30 years-old male (GIST_10). Primary tumors were in the stomach, multi-centric in GIST_07 and uni-centric in GIST_10, and presented metastases at diagnosis. RNA-Seq paired-end libraries were sequenced on the Illumina GAIIx system using a paired-end strategy at a read length of 75bp. Bioinformatic analysis highlighted 9 candidate genes common to both patients. Mutations in these genes were then filtered with SNP&GO, a tool suited to compute the likelihood of a mutation to be disease-related or not, depending on the protein sequence and its functional annotation. The identified candidate gene was analyzed by Sanger sequencing on tumor (T) and peripheral blood (PB) DNA from GIST_07 and GIST_10 patients, and on additional 8 tumor and 3 PB DNA from 8 sporadic WT GISTs patients. Immunohistochemistry (IHC) was also performed. Results: Only mutations in the sequence of SDHA were identified as disease-related with a high reliability index. GIST_07 tumor DNA was homozygous for an exon 9 p.S384X nonsense mutation; GIST_10 patient showed compound heterozygosity for a germinal exon 2 p.R31X nonsense mutation and a somatic exon 13 p.R589W missense mutation.Heterozygous nonsense mutations were already present in germline DNA in both patients. All mutations lead to functional inactivation of the protein. IHC revealed that both patients showed the down regulation of SDHA. The expanded study showed that 2/10 carried SDHA mutations in the tumor: compound heterozygosity for an exon 9 p.G419R and exon 13 p.E564K mutations in one patient, and a somatic heterozygous exon 5 p.R171C mutation in the other. Conclusions: Recently, SDHB and SDHC mutations were also reported in these patients. This the first study that identifies mutations on SDHA, in 4/10 WT GIST patients. This gene should be evaluated in larger seriesand be added in the sequencing screening of WT GISTs.
Proceedings of the 2011 ASCO annual meeting
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M. A. Pantaleo; A. Astolfi; V. Indio; P. Paterini; S. Formica; R. Casadio; P. Martelli; A. Maleddu; M. Nannini; A. P. Dei Tos; M. C. Heinrich; D. Santini; F. Catena; C. Ceccarelli; M. Fiorentino; M. di Battista; R. Moore; N. Thiessen; C. Gnocchi; G. Biasco
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11585/145750
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