Novel V600E BRAF mutations in imatinib-naive and imatinib-resistant gastrointestinal stromal tumors

Abstract

BRAF and NRAS are commonly mutated in cancer and represent the most frequent genetic events in malignant melanoma. More recently, a subset of melanomas was shown to overexpress KIT and harbor KIT mutations. Although most gastrointestinal stromal tumors (GISTs) exhibit activating mutations in either KIT or PDGFRA, about 10% of the cases lack mutations in these genes. It is our hypothesis following the melanoma model that mutations in BRAF or NRAS may play a role in wild-type GIST pathogenesis. Alterations in RAS/MEK/ERK pathway may also be involved in development of imatinib resistance in GIST, particularly in tumors lacking secondary KIT or PDGFRA mutations. Imatinib-naive wild-type GISTs from 61 patients, including 15 children and 28 imatinib-resistant tumors without secondary KIT mutations were analyzed. Screening for hot spots mutations in BRAF (exons 11 and 15) and NRAS (exons 2 and 3) was performed. A BRAF exon 15 V600E was identified in 3 of 61 GIST patients, who shared similar clinical features, being 49- to 55-years-old females and having their tumors located in the small bowel. The tumors were strongly KIT immunoreactive and had a high risk of malignancy. An identical V600E BRAF mutation was also identified in one of 28 imatinib resistant GIST lacking a defined mechanism of drug resistance. In conclusion, we identified a primary BRAF V600E mutations in 7% of adult GIST patients, lacking KIT/PDGFRA mutations. The BRAF-mutated GISTs show predilection for small bowel location and high risk of malignancy. A secondary V600E BRAF mutation could represent an alternative mechanism of imatinib resistance. Kinase inhibitors targeting BRAF may be effective therapeutic options in this molecular GIST subset.

Figure 1

Pathologic findings in BRAF mutated GIST, showing a predominantly monomorphic spindle cell morphology (A: patient no.1, HE, ×200), with only focal epithelioid features and high mitotic activity (B: patient no. 1, HE ×400); one tumor showed focal areas of dystrophic calcification (C: patient 4, HE, ×100). Tumors were diffusely immunoreactive for CD117 (D: patient no. 2, ×200). Additional findings included: loss of P16 protein expression while PTEN expression was maintained (E, F: patient no. 1; ×200), as compared to patient no. 3 who had maintained P16 protein expression (G, ×200) and to patient no. 4 who lost PTEN (H, ×200).

Figure 2

Pre-imatinib pathologic findings revealed a high risk primary gastric GIST with epithelioid morphology (A, HE, ×200) and diffuse reactivity for CD117 (B, ×200) and PDGFRA (C, ×200). The post-imatinib resection for peritoneal metastasis showed foci of viable spindle cells with abundant eosinophilic cytoplasm and strap cells, resembling an embryonal rhabdomyosarcoma phenotype (D, HE, ×400), which was negative for CD117, PDGFRA, (E,F, ×200), but strongly positive for skeletal muscle markers (G, desmin, ×200; H, myogenin, ×200). The mutation analysis of this sample revealed in addition to a PDGFRA exon 18 deletion, a BRAF exon 15 V600E substitution (I, ABI sequence).