Italiano et al. BMC Cancer 2012, 12:408 http://www.biomedcentral.com/1471-2407/12/408 Page 5 of 7 Figure 4 Immunohistochemistry showed concomitant loss of SDHA and SDHB expression in SDHA-mutant GIST. Complete loss of expression for SDHA (A) and SDHB was noted in the germline SDHA-mutant GIST (B), while normal liver showed preserved reactivity. In the somatic SDHA-mutant GIST there was partial loss of SDHA expression (C), while SDHB staining was absent (D). SDHA expression was retained in a WT pediatric GIST (E), while SDHB reactivity was lost (F, with internal positive control). In the control KIT exon 11-mutant GIST from a young adult both SDHA (G) and SDHB protein expressions were preserved (H). enzyme complex linked to the respiratory chain and a member of the Krebs cycle. It consists of 4 subunits: the flavoprotein subunit (SDHA), the iron-sulfur protein subunit (SDHB), and the integral membrane protein subunits (SDHC and SDHD). Mutations of one of the gene encoding these subunits impair the activity of this complex and lead to the stabilization and activation of HIF-1a, which in turn activates cell proliferation and angiogenesis [12-15]. In addition to paragangliomas and pheochromocyto- mas, a number of other solid tumors have been asso- ciated with mutations in genes encoding the succinate dehydrogenase complex (SDH) complex II. These in- clude gastrointestinal stromal tumors (GIST) [11,16], renal tumors [26], thyroid tumors [26-28], testicular seminoma [29]. The best known association between SDH complex II germline mutations and other tumors is represented by the Carney–Stratakis syndrome (or dyad) which is characterized by the occurrence of KIT and PDGFRA WT GIST and paraganglioma. This syndrome is associated with germline point mutations or large deletions of the genes encoding the SDHB, SDHC or SDHD subunits [30]. Strikingly, inactivating germline mutations in SDHB or SDHC genes have been also iden- tified in sporadic WT GISTs occurring in patients with- out a personal or family history of paraganglioma [11]. The SDHA gene encodes the major catalytic subunit of the succinate dehydrogenase complex II. Germline mutations in SDHA are associated with neurodegenera- tive diseases such as an early-onset encephalopathy, known as Leigh syndrome [31-34] and a late-onset optic atrophy, ataxia and myopathy [35]. Until recently, no genetic link between SDHA and cancer could be estab- lished. However, two recent studies allowed the identifi- cation of SDHA germline mutations in at least 3% patients with apparently sporadic cases of paraganglioma or pheochromocytoma [36]. Interestingly, four cases of sporadic KIT and PDGFRA WT GIST occurring in one pediatric and three young adult patients have also been associated with germline mutation of SDHA [16,17]. In the present study, we investigated a series of 17 appar- ently sporadic and Carney’s triad-related KIT and PDGFRA WT GIST for SDHA mutations and found an additional two cases with mutations in this gene. These were exclusively present in apparently sporadic cases oc- curring in young adults. The p.Arg31X SDHA germline mutation identified in our study leads to a truncated protein [16]. An identical mutation has been previously reported in four Dutch patients with paraganglioma and in one young adult patient with sporadic WT GIST [16,36]. The second SDHA mutation identified in our study (p.D38V) has been reported as a single nucleotide polymorphism. However, none of the other 16 GIST cases tested showed this change and this mutation was found only in the tumor DNA, but not in corresponding normal DNA of the patient. Furthermore, this tumor showed significant loss of SDHA protein expression by both western blot and IHC, suggesting a functional im- pact of this genetic alteration. But since only one source of normal DNA was analyzed in this patient, we cannot formally exclude the possibility of germline mosaicism. Moreover, since we have sequenced only SDHA exons 2, 9 and 13, we cannot exclude also the presence of a germline mutation in one of the other exons.