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1 EARLY ONLINE RELEASE Note: This article was posted on the Archives Web site as an Early Online Release. Early Online Release articles have been peer reviewed, copyedited, and reviewed by the authors. Additional changes or corrections may appear in these articles when they appear in a future print issue of the Archives. Early Online Release articles are citable by using the Digital Object Identifier (DOI), a unique number given to every article. The DOI will typically appear at the end of the abstract. The DOI for this manuscript is doi: /arpa RS The final published version of this manuscript will replace the Early Online Release version at the above DOI once it is available College of American Pathologists

2 Resident Short Review Succinate Dehydrogenase Complex Succinate dehydrogenase (SDH) is uniquely tasked with a dual role in the essential energy-producing processes of a cell. Although SDH subunits and assembly factors form part of the same enzyme complex, mutations in their respective genes lead to significantly different clinical phenotypes. Remarkable discoveries in the last 17 years have led to the delineation of the SDH complex deficiency syndrome and its multiple pathogenic branches. Here we provide an updated overview of SDH deficiency in order to raise awareness of its multiple connotations including nonneoplastic associations and pertinent features of the continually growing list of SDHmutant tumors so as to better direct genetic counseling and predict prognosis. (Arch Pathol Lab Med. doi: /arpa RS) Succinate dehydrogenase (SDH) is the only enzyme complex that is involved in both the citric acid cycle and the electron transport chain. 1 Following the discovery of a germline SDHD mutation in 2000, there has been an explosion of data regarding its role in neoplasia, resulting in the eventual delineation of the SDH complex deficiency (SCD) syndrome. 2 SCD syndrome is an autosomal dominant disorder with incomplete penetrance characterized by the occurrence of multiple tumors including paragangliomas (PGLs), pheochromocytomas (PCCs), gastrointestinal stromal tumors (GISTs), renal cell carcinomas (RCCs), and others. It is caused by germline mutations in the gene family of mitochondrial complex II: succinate dehydrogenase subunits (SDH) or succinate dehydrogenase complex assembly factor 2 (SDHAF2). 3 In addition, aberrations in the SDH complex have also been linked to childhood neuromuscular diseases and male infertility. 4,5 Herein, we provide an overview of the physiologic role of SDH and the metabolic impact of its deficiency. Specifically, we focus on An Updated Review Mohamed Rizwan Haroon Al Rasheed, MBBS; Gabor Tarjan, MD the pertinent features of SDH-associated neoplasms and nontumoral implications of SDH aberrations. PHYSIOLOGY Succinate dehydrogenase, localized in the inner mitochondrial membrane, is uniquely tasked with a dual role in the essential energy-producing processes of a cell. 1 It is involved in the oxidation of succinate to fumarate in the citric acid cycle as well as in the reduction of ubiquinone (coenzyme Q) in the aerobic electron transfer chain, contributing to the generation of ATP by oxidative phosphorylation. 6 Structurally, it is a hetero-oligomer composed of 4 subunit proteins (SDHA, SDHB, SDHC, and SDHD). SDHA is a flavoprotein and SDHB is an iron-sulfur protein; together they form the main catalytic domain that oversees the oxidation of succinate to fumarate. SDHC and SDHD are the membrane-anchoring subunits of SDH and play a role in passing electrons through the electron transport chain. In addition, at least 2 other proteins, SDHAF1 and SDHAF2, support it by flavinating the SDHA subunit and promoting maturation of SDHB, thus enabling the assembly of the full SDH complex (Figure 1). 1,4,6 SDH DEFICIENCY AND NEOPLASIA Succinate dehydrogenase is the only respiratory chain complex whose subunits are entirely encoded by nuclear genes located in chromosome bands 5p15 (SDHA), 1p36 (SDHB), 1q21 (SDHC), 11q23 (SDHD), 19q13 (SDHAF1), and 11q12 (SDHAF2). 1,5 Although SDH subunits and assembly factors form part of the same protein complex, mutations in their respective genes lead to remarkable differences in clinical phenotype. 1 SDH (which includes SDHA, SDHB, SDHC, and SDHD) and SDHAF2 are tumor suppressor genes. 1 As such, tumorigenesis occurs when a germline mutation in 1 of the alleles is followed by loss of heterozygosity of the other wild-type allele resulting in the loss of its respective protein. This, in turn, destabilizes the SDH complex and abolishes its Accepted for publication August 21, enzymatic activity. Germline mutations in SDH and From the Department of Pathology, University of Illinois at Chicago (Dr Haroon Al Rasheed); and the Department of Pathology, SDHAF2 have an autosomal dominant mode of inheritance John H. Stroger, Jr. Hospital of Cook County, Chicago, Illinois (Dr with incomplete penetrance. 7 In addition, inheritance of Tarjan). SDHD and SDHAF2 involves a parent-of-origin effect, such The authors have no relevant financial interest in the products or that although either parent can transmit it, an affected child companies described in this article. usually develops a neoplasm only if the mutated gene was Corresponding author: Mohamed Rizwan Haroon Al Rasheed, MBBS, Department of Pathology, University of Illinois at Chicago, received from the father, leading to possible generation 840 S Wood St, Suite 130 CSN, Chicago, IL ( skipping in affected families. However, unequal evidence of mharoon@uic.edu or mohamedrizwan@gmail.com). genomic imprinting as the underlying mechanism has not Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan 1

3 Figure 1. Succinate dehydrogenase (SDH) complex in the electron transport chain. Succinate dehydrogenase assembly factor 2 (SDHAF2) adds a flavin adenine dinucleotide (FAD) cofactor to SDHA to form an active SDHA flavoprotein. a-ketoglutarate is converted by a-ketoglutarate dehydrogenase (a-kgdh) to succinyl-coa, which in turn is converted by succinyl-coa synthetase (SCS) to succinate. SDHA oversees the oxidation of succinate to fumarate in the citric acid cycle, resulting in the reduction of FAD to FADH 2. 6 SDHAF1 is thought to interact with SDHB via insertion or retention of the iron-sulphur centers. 4 Electrons are transferred through a chain of cofactors including the triple iron-sulphur centers on SDHB to ubiquinone, bound to the membrane-anchoring subunits SDHC and SDHD, reducing it to ubiquinol. Ubiquinol transfers the electrons along the transport chain to complex III (not shown). 6 been identified and many other models have been hypothesized. 4,7 The list of SDH-mutant tumors has slowly expanded in the past 17 years and now includes SDH-deficient PGLs, PCCs, RCCs, GISTs, pituitary adenomas (PAs), and less commonly, thyroid tumors and neuroblastomas. Several models have been proposed to explain how the citric acid cycle dysfunction leads to tumor development. Of them, the 3 strongest mechanisms are decrease in apoptosis, increase in reactive oxygen species, and activation of a pseudohypoxia pathway. 6 Albayrak et al 8 reported that cells deficient in SDHC were defective in their apoptotic response to several stimuli, thus becoming the first group to indicate that SDH is a potential regulator of apoptosis. The same study also described that the reduction in SDH activity also resulted in an increased production of reactive oxygen species. Ishii et al 9 subsequently highlighted that the oxidative damage to nuclear DNA by the reactive oxygen species leads to increased mutations and tumorigenesis. An increasingly popular, but not mutually exclusive, third mechanism by which loss-of-function mutations of mitochondrial tumor suppressor genes contribute to cancer is known as pseudohypoxia. Pseudohypoxia occurs when hypoxia-inducible factor (HIF) pathways are constitutively activated, regardless of oxygen levels. In the citric acid cycle, a-ketoglutarate (a-kg; also known as 2-oxoglutarate) is converted by prolyl hydroxylases (PHDs) to succinate, which in turn undergoes oxidation by SDH to fumarate. a-ketoglutarate physiologically activates several a-kg dependent dioxygenases, including PHDs that promote HIF degradation, Jumonji-related histone demethylases (JMJDs), which demethylate histones, and the ten-eleven translocation family of DNA hydroxylases (TETs), which demethylate DNA. 10 Succinate, which accumulates as a result of the loss of SDH activity, leaves the mitochondria to enter the cytoplasm where it plays a key role in tumor pathogenesis via epigenetic modulation. Because of the structural similarity between succinate and a-kg, succinate competitively (1) inhibits PHDs, preventing HIF degradation; (2) inhibits JMJDs, preventing histone demethylation; and (3) inhibits TETs, preventing DNA demethylation. 10 This collectively leads to increased HIF and global hypermethylation of target DNA and histones. Increased HIF in turn leads to activation of multiple target genes important in glycolysis, cell proliferation, angiogenesis, and apoptosis. 10 These molecular events are in turn reflected within the mitochondria of these unusual neoplasms, which display a hypoxic phenotype at the ultrastructural level. At least 1 study that compared SDH-mutant GISTs and PGLs has revealed that the cytoplasm of these SDH-deficient tumors contains increased numbers of structurally abnormal mitochondria in varying sizes with loss or complete absence of cristae. 11 In addition, many mitochondria also show an amorphous material forming membranous whorls or vacuoles. 11 Succinate dehydrogenase activity is typically assessed qualitatively via immunohistochemical stains for SDHB and SDHA proteins. These are sensitive and specific markers for their respective proteins, normally ubiquitously expressed in a granular cytoplasmic pattern localizing to the mitochondria. 12 Loss of SDH complex activity can be detected by corresponding loss of SDHB immunostaining. This is shown to significantly correlate with a germline mutation in SDH or SDHAF2, with rare instances of loss of staining in sporadic or non SDH-mutant cases, such as following SDHC promoter hypermethylation. 13 Nevertheless, this immunohistochemical test can be used as a screening method to guide genetic testing. 13 In contrast, loss of immunostaining for both SDHA and SDHB is only seen with an SDHA mutation, although rare cases of 2 Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan

4 sporadic and germline SDHA mutations do demonstrate retained SDHA immunostaining in the absence of SDHB staining. In other words, absence of SDHA staining is specifically associated with SDHA mutations. 12,14 Furthermore, elevated succinate to fumarate ratio is a consistent biomolecular phenotype of SDH-deficient tumors as demonstrated by Kim et al 15 using mass spectrometry to quantitatively analyze SDH dysfunction. Therefore, germline mutations in SDH as well as hypermethylation of the SDHC promoter can result in an elevated succinate to fumarate ratio, extending support to the above-described pseudohypoxia mechanism. 15 Syndromic Associations SCD syndrome, also known as hereditary paraganglioma/ pheochromocytoma syndrome or familial paraganglioma syndrome, is caused by germline loss-of-function mutations in SDH and SDHAF2. 3 Their syndromic associations with multiple tumors have only recently come to light, resulting in their segregation based on the underlying gene and the corresponding phenotypic correlation into 5 types that demonstrate unique clinical characteristics such as tumor site, multiplicity, function, risk of recurrence, and metastasis. 3,7,10 PGL type 1 (PGL1) is caused by an SDHD mutation, PGL2 by an SDHAF2 mutation, PGL3 by an SDHC mutation, PGL4 by an SDHB mutation, and PGL5 by an SDHA mutation. Phenotypically, all of them are associated with PGLs: 4 of them (except SDHAF2) are linked to PCCs, RCCs, and GISTs; and 3 (SDHA, SDHB, SDHD) are also associated with PAs, called the 3P association when there is a triad of PA, PGL, and PCC. 7,10,16 In terms of frequency, most SDH-mutated RCCs and thoracoabdominal extraadrenal PGLs, including urinary bladder PGLs, occur in PGL4 (SDHB mutated), solitary head and neck PGLs occur in PGL3, multiple head and neck PGLs occur in PGL1-2 (SDHD or SDHAF2 mutated), and GISTs occur in PGL5 (SDHA mutated). 7,17 Carney-Stratakis syndrome was initially described in 2002 to denote the familial occurrence of combined GIST and PGL. Since then, it has been demonstrated that most of these patients harbor mutations in 1 of the SDH genes. As such, Carney-Stratakis syndrome and PGL1-5 are now thought to represent parts of the same disease. 7 On the other hand, the similarly named Carney triad is a nonhereditary allelic condition that typically occurs in young females and is characterized by gastric GIST, PGL, and pulmonary chondroma. 7 In contrast to Carney-Stratakis syndrome, no inherited trait has been established in Carney triad although there is a possibility that deletions within the 1pcen13-q21 region, which harbors the SDHC gene, and epigenetic alteration of SDHC such as aberrant DNA hypermethylation, could be possible mechanisms for tumor development in Carney triad. 7,12 Cowden syndrome is an autosomal dominant disorder characterized by multiple hamartomas involving all 3 germ cell layers, caused mainly by germline mutations in PTEN. It confers a high risk of breast, thyroid, endometrial, renal, and colonic malignancies. 18 SDH genes, in particular SDHB and SDHD, have been identified as predisposing and modifier genes in a subset of patients with Cowden and Cowden-like syndrome, including in up to 10% of Cowden syndrome cases without PTEN mutations, ensuing an even higher risk of breast, kidney, and thyroid cancers over those with only germline PTEN mutations. 18 SDH-Deficient Paraganglioma and Pheochromocytoma The World Health Organization (WHO) defines PGL as a unique neuroendocrine neoplasm, usually encapsulated and benign, arising in specialized neural crest cells associated with segmental or collateral autonomic ganglia; consisting of uniform chief cells exhibiting neuronal differentiation forming compact nests (Zellballen) surrounded by sustentactular cells and a delicate capillary network. 18 They are classified by the site of origin, such as central nervous system PGL, head and neck PGL, (urinary) bladder PGL, mediastinal PGL, organ of Zuckerkandl PGL, extra-adrenal abdominal PGL, pelvic PGL, and pheochromocytoma (PGL of adrenal medulla). Most PGLs occur in adults and nearly all subtypes have a female predilection except Cauda equina PGL, which has a slight male predominance. 7,18 Paragangliomas stand on an elevated pedestal in the world of cancer biology; it is the first neoplasm in which a mitochondrial protein with a role in intermediary metabolism (SDHD) was shown to be a tumor suppressor. 1,2 Uniquely, they have the highest degree of heritability among human neoplasms, with as many as 40% associated with a germline mutation, including greater than 80% of these tumors in children with 7% to 13% of those presenting as an apparently sporadic tumor. 7,10 As a result, it is now recommended that all patients with PGL, including sporadic tumors, be referred for genetic testing. 7 Paragangliomas across the various sites share similar histologic and immunohistochemical features, being composed of 2 cell types: chief (type 1) cells and sustentacular (type 2) cells classically in a Zellballen (organoid or nesting) architectural pattern with a prominent vascular network separating the nests. 7,18 Immunohistochemically, the chief cells are positive for neuroendocrine markers, tyrosine hydroxylase, and GATA3, whilst the surrounding sustentacular cells express S100 and GFAP (glial fibrillary acidic protein). 7,19 Both cell types are negative for cytokeratins and p63, whilst reticulin would highlight the nesting pattern. As described above, when SDH mutated, the tumoral cells show loss of immunohistochemical staining for SDHB protein (Figure 2, A through C). 7,19 Comprehensive integrated analysis recently classified PCCs/PGLs into 4 molecularly defined groups: a kinase signaling subtype, a pseudohypoxia subtype, a Wnt-altered subtype, and a cortical admixture subtype. 20 The kinase signaling and Wnt-altered subtypes consist of PCCs, whilst the pseudohypoxia and cortical admixture subtypes include both PCCs and PGLs. 20 Germline mutations in SDHB and SDHD were found to be completely specific to the pseudohypoxia subtype, with most SDHB and SDHD germline mutations clustering within the hypermethylated subgroup. 20 At least 19 susceptibility genes have been identified to date with mutations, both hereditary and somatic, found in a mutually exclusive manner in these tumors, although the most commonly reported exception was a combination of SDHB germline and ATRX somatic mutation. 20 Analyzed gene sets also demonstrate that SDHB has the most common germline mutation (15 of 173 patients; 9%) and the highest number of copy number alterations (99 of 173 patients; 57%) in these tumors. 20 In contrast, SDH is generally rarely (SDHD and SDHB) or never (SDHA, SDHAF2, SDHC) mutated in the nonfamilial, sporadic tumors. 7,10 Furthermore, PGLs with germline Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan 3

5 mutations in SDHB and SDHD are also typically negative for epinephrine and metanephrine secretions. 20 Paragangliomas are generally slow growing with a low rate of recurrence following resection but it can be greater than 50% in patients with SDHB mutations. 7 Interestingly, SDHB-deficient urinary bladder PGLs, when compared with the SDHB-intact cases, are characterized by a large size, higher mitoses, frequent lymphovascular invasion, and metastases. 21 Given the frequent association with germline mutations, it is difficult to reliably distinguish PGL recurrence from a second primary, particularly if the involved gene is SDHB, SDHD, orsdhaf2. 7 All PGLs are now known to have some potential for metastasis, prompting the WHO to retire the term malignant paraganglioma in favor of metastasizing paraganglioma or paraganglioma with metastasis. 7 However, there are no agreed upon universal histologic criteria to predict metastasis and the overall likelihood of metastasis is still quite low with the highest risk in SDHB-mutant tumors. 7 As such, SDHB germline mutation is a marker of more aggressive disease. 20 SDH-Deficient Gastrointestinal Stromal Tumor SDH-mutated GISTs comprise about 7.5% of all gastric GISTs. 12,22 They typically occur in a younger and predominantly female population characteristically arising in the gastric antrum in a multinodular fashion with plexiform mural involvement. 12,22 They show an epithelioid morphology and are often associated with lymphovascular invasion and occasional lymph node metastasis, unlike conventional GISTs. 12,23 Immunohistochemically, most GISTs are positive for C-KIT and DOG1, variably positive for CD34, and rarely positive for smooth muscle actin or desmin. 12 Importantly, immunopositivity to C-KIT and DOG1 will not exclude an underlying SDH deficiency, as most SDH-deficient GISTs are also typically C-KIT and DOG1 positive. 23 SDHB immunoexpression is retained in all GISTs with KIT or PDGFRA mutations, whilst loss of SDHB expression in the neoplastic cells, with positive staining in fibrovascular septa and inflammatory cells, relates to an underlying SDH deficiency (Figure 3, A through C). 23,24 Conventional GISTs typically harbor KIT or PDGFRA mutations; 15% of adult GISTs and greater than 90% in children lack such mutations and are thus so-called wildtype GISTs. 23,24 Forty-two percent of these wild-type GISTs have an underlying SDH deficiency, most commonly as a result of a germline mutation in SDHA. 23,24 Unlike KITmutant GISTs, SDH-mutated GISTs are resistant to imatinib but respond better to second- or third-generation tyrosine kinase inhibitors. 23 National Comprehensive Cancer Network (NCCN) guidelines for risk stratification of conventional GISTs are based on tumor size, mitotic activity, and anatomic site with larger size, increased mitosis, and nongastric location all conferring a higher risk. 12 In contrast, SDH-deficient GISTs are quite unpredictable. Firstly, they are associated with a high rate (up to 82%) of distant metastasis, most often to the liver, regardless of conventional risk category. 25 Secondly, SDH-deficient GISTs may show an indolent course, metastasize even with low mitotic counts, have a very long latency before metastasizing, and survive for long periods after metastasis even without specific treatment. 12,23 Thirdly, SDHA mutations are associated with statistically significant better clinical outcome than KIT/PDGFRA mutations and KIT/PDGFRA wild-type without SDH deficiency. 26 NCCN risk stratification thus cannot accurately predict the behavior of SDH-deficient GISTs. 25 Given the multiple ramifications, it is recommended that SDHB immunohistochemistry be considered in all epithelioid gastric GISTs with a multinodular or plexiform growth pattern. 23 If SDHB shows negativity, the patient should be referred for genetic counseling, and subsequent loss of staining for SDHA could uncover an underlying SDHA mutation. 23 SDH-Deficient Renal Cell Carcinoma SDH-deficient RCC is defined per the WHO as a malignant epithelial tumor composed of vacuolated eosinophilic to clear cells defined by the loss of SDHB immunohistochemical staining. 19 It comprises up to 0.2% of all resected renal cell tumors with an overall risk of developing RCC in SDH deficiency being about 14% by age 70 years. Most patients are young adults with a slight male predominance and in nearly 30% of cases these tumors occur bilaterally. 3,17 Most SDH-deficient RCCs are unencapsulated and well circumscribed with a nested, solid, or tubular architecture and are composed of oncocytic cells with an eosinophilic cytoplasm and small round Fuhrman grade 2 nuclei without prominent nucleoli admixed with prominent intratumoral mast cells. 3 The most characteristic feature of this tumor is the presence of cytoplasmic inclusions, representing giant mitochondria, containing eosinophilic or flocculent to completely clear material. Immunohistochemically, in addition to loss of SDHB staining, SDH-deficient RCCs are also negative for CD117, cytokeratin 7, carbonic anhydrase 9 (CAIX), and S100. 3,27 Among the differential diagnoses of SDH-deficient RCCs is a novel form of fumarate hydratase deficient RCC that shows a similar low-grade oncocytic morphology but has a retained SDHB expression. 28 The overwhelming majority of these tumors have an underlying SDHB germline mutation and are thus associated most frequently with PGL4. Most cases are uniformly low grade and have an indolent clinical behavior but up to onethird, particularly those with dedifferentiation, high nuclear grade, and/or necrosis, can have an aggressive behavior with metastasis. 3,17,19 Thus, the WHO recommends that all patients with SDH-deficient RCCs be offered genetic testing, and even in the absence of an identified germline mutation be followed up long term for recurrence and surveillance of other SDH-deficient neoplasms. 19 SDH-Deficient Pituitary Adenoma In contrast to conventional PAs, SDH-deficient PAs may be larger, more likely to produce prolactin, display a more aggressive phenotype, be more resistant to somatostatin analogs, and more often require surgery. 16,29 However, the exact role of SDH deficiency in PAs remains controversial. On the one hand, Xekouki et al 16 note that while no SDH mutations are detected among sporadic PAs, up to 75% of familial cases are positive for a germline SDH mutation, particularly SDHB. Conversely, Gill et al 29 report that although SDH-deficient PAs can be identified by SDHB immunohistochemistry, they appear to be a rare event and can occur in the absence of germline mutation. Other SDH-Deficient Tumors Germline and somatic SDH variants occur in sporadic thyroid cancers and overall loss of SDH gene expression could represent a molecular signature of differentiated thyroid tumors. About 6% of thyroid cancers have a germline mutation of SDHB or SDHD and a further 5% 4 Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan

6 Figure 2. Succinate dehydrogenase A (SDHA) deficient paraganglioma. A, Centrally located chief cells with abundant eosinophilic cytoplasm surrounded by peripherally located slender, spindled sustentacular cells forming a nested (Zellballen) pattern separated by a prominent vascular network. B, Absence of immunostaining for SDHB protein in the neoplastic chief cells with retained positive granular cytoplasmic staining both in the sustentacular cells and vascular endothelial cells. C, Negative immunostaining for SDHA protein in the neoplastic chief cells appearing as a light, diffuse nongranular blush in contrast to the positive granular cytoplasmic staining both in the sustentacular cells and vascular endothelial cells, indicating an underlying SDHA mutation (hematoxylin-eosin, original magnification 3400 [A]; brown chromogen, original magnification 3400 [B and C]). Figure 3. Succinate dehydrogenase A (SDHA) deficient gastrointestinal stromal tumor (GIST). A, Epithelioid GIST with uniform nuclei and abundant eosinophilic cytoplasm located in the gastric wall. B, Absence of immunostaining for SDHB protein in the neoplastic cells with positive granular cytoplasmic staining in background plasma cells and fibrovascular septa. C, Similar absence of immunostaining for SDHA protein in the neoplastic cells with positive granular cytoplasmic staining in background plasma cells and fibrovascular septa, indicating an underlying SDHA mutation (hematoxylin-eosin, original magnification 3200 [A]; brown chromogen, original magnification 3400 [B and C]). Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan 5

7 have somatic SDHC mutations. 30 Consistent loss of either SDHC or SDHD gene expression is associated with earlier disease onset and higher pathologic TNM stage in both papillary and follicular thyroid tumors. 30 Although further studies focusing on the SDH transcript levels and immunohistochemical SDHB protein expression are to be fully explored, early studies point toward papillary histology as the major subtype. 31 Amongst patients with Cowden syndrome/cowden-like syndrome with germline SDHB or SDHD mutations, papillary thyroid carcinoma has an elevated standardized incidence ratio across all individuals, in contrast to the increased likelihood of follicular histology in PTEN mutation carriers. 30,31 Neuroblastoma and PCC have the same embryonal origin. In addition, neuroblastomas frequently exhibit deletions of chromosome band 1p36, which is also the region for both the SDHB gene and KIFBb gene. However, studies exploring the association between SDH mutations and neuroblastomas are relatively limited with most articles focusing solely on SDHB. Although a handful of reports do report an association between germline SDHB mutation and neuroblastoma, larger studies have so far provided no definitive evidence for SDHB mutations in sporadic or germline neuroblastoma. 32,33 Amongst other carcinomas, loss of SDHB has been reported in 1 prostatic adenocarcinoma (1 of 57; 1.8%), a gastric EBER-positive lymphoepithelial carcinoma, and a testicular seminoma (1 of 40; 2.5%). 34 Given the nature of the reported study, these findings are likely coincidental and thus require further research to elucidate the clinical significance and molecular genetics behind these SDHBnegative tumors. 34 NONNEOPLASTIC IMPLICATIONS OF SDH ABERRATIONS A homozygous SDHA mutation in 2 patients with Leigh syndrome was the first nuclear gene mutation causing a mitochondrial respiratory chain deficiency to be reported in humans. 35 Since then, a rare familial neonatal isolated cardiomyopathy and a late-onset neurodegenerative disease characterized by progressive optic atrophy, ataxia, and myopathy have also been linked to missense mutations of SDHA with subsequent decrease in SDH activity. In addition, mutations of SDHAF1 and SDHB have also been associated with specific types of infantile leukoencephalopathy. 4 As mentioned earlier, SDH is the only mitochondrial respiratory complex whose subunits are entirely encoded by nuclear genes; the efficiency of the spermatogenic process, as measured by the sperm concentration in ejaculate, has been correlated with the nuclear-encoded mitochondrial enzyme activities. 5 Therefore, Bonache et al 5 performed an exhaustive analysis to further explore the association, shedding light on 50 sequence variations involving SDHA through SDHD of which one, SDHA c.456þ32g.a, showed significant genotype association with nonobstructive male infertility. CONCLUSIONS The multitude of SDH alterations described here have opened up potential targets for future therapies: decitabine, a demethylase inhibitor, has shown a role in reversing SDHmutant methylation profiles in mouse models 10 ; a glutaminase inhibitor is currently undergoing trial as an adjunct to chemotherapy in SDH-mutated tumors; and a third trial using a DNA methyltransferase inhibitor, guadecitabine, to target SDH-deficient tumors is also in the preliminary stages. 20,36 Given the broad range of tumors with an underlying SDH mutation that are now becoming apparent, as well as the reduced penetrance of certain mutations, a trial to evaluate screening methods has already been completed. 37 However, a clear consensus on screening and follow-up is yet to emerge. Increasing awareness about the occurrence of newly associated tumors is imperative and identification of SDH deficiency is critical to determine prognosis and direct genetic counseling. 25,33 We would like to thank John Hart, MD, at University of Chicago, Chicago, Illinois, and Jason L. Hornick, MD, PhD, at Brigham & Women s Hospital, Boston, Massachusetts, for the images in Figures 2 and 3. References 1. Hensen EF, Bayley J-P. Recent advances in the genetics of SDH-related paraganglioma and pheochromocytoma. Fam Cancer. 2011;10(2): doi: /s Baysal BE, Ferrell RE, Willett-Brozick JE, et al. Mutations in SDHD, a mitochondrial complex II gene, in hereditary paraganglioma. Science. 2000; 287(5454): Amin MB, Tickoo SK, eds. Succinate dehydrogenase complex deficiency syndrome. In: Diagnostic Pathology Genitourinary. 2nd ed. Salt Lake City: Elsevier; 2016: Hoekstra AS, Bayley J-P. The role of complex II in disease. Biochim Biophys Acta. 2013;1827(5): doi: /j.bbabio Bonache S, Martínez J, Fernández M, Bassas L, Larriba S. Single nucleotide polymorphisms in succinate dehydrogenase subunits and citrate synthase genes: association results for impaired spermatogenesis. Int J Androl. 2007;30(3): doi: /j x. 6. Gottlieb E, Tomlinson IPM. Mitochondrial tumour suppressors: a genetic and biochemical update. Nat Rev Cancer. 2005;5(11): doi: / nrc Kimura N, Capella C, Gill A, et al. Paraganglion tumors. In: El-Naggar AK, Chan JKC, Grandis JR, Takata T, Slootweg PJ, eds. WHO Classification of Head and Neck Tumours. 4th ed. Lyon, France: IARC Press; 2017: World Health Organization Classification of Tumours; vol Albayrak T, Scherhammer V, Schoenfeld N, et al. The tumor suppressor cybl, a component of the respiratory chain, mediates apoptosis induction. Mol Biol Cell. 2003;14(8): doi: /mbc.e Ishii T, Yasuda K, Akatsuka A, Hino O, Hartman PS, Ishii N. A mutation in the SDHC gene of complex II increases oxidative stress, resulting in apoptosis and tumorigenesis. Cancer Res. 2005;65(1): Dahia PLM. Pheochromocytoma and paraganglioma pathogenesis: learning from genetic heterogeneity. Nat Rev Cancer. 2014;14(2): doi: /nrc Szarek E, Ball ER, Imperiale A, et al. Carney triad, SDH-deficient tumors, and Sdhbþ/- mice share abnormal mitochondria. Endocr Relat Cancer. 2015; 22(3): doi: /erc Miettinen M, Corless CL, Debiec-Rychter M, et al. Gastrointestinal stromal tumors. In: Fletcher CDM, Bridge JA, Hogendoorn PCW, Mertens F, eds. WHO Classification of Tumours of Soft Tissue and Bone. 4th ed. Lyon, France: IARC Press; 2013: World Health Organization Classification of Tumours; vol van Nederveen FH, Gaal J, Favier J, et al. An immunohistochemical procedure to detect patients with paraganglioma and phaeochromocytoma with germline SDHB, SDHC, or SDHD gene mutations: a retrospective and prospective analysis. Lancet Oncol. 2009;10(8): doi: /s (09) Miettinen M, Killian JK, Wang Z-F, et al. Immunohistochemical loss of succinate dehydrogenase subunit A (SDHA) in gastrointestinal stromal tumors (GISTs) signals SDHA germline mutation. Am J Surg Pathol. 2013;37(2): doi: /pas.0b013e Kim E, Wright MJ, Sioson L, et al. Utility of the succinate:fumarate ratio for assessing SDH dysfunction in different tumor types. Mol Genet Metab Rep. 2017; 10: doi: /j.ymgmr Xekouki P, Szarek E, Bullova P, et al. Pituitary adenoma with paraganglioma/pheochromocytoma (3PAs) and succinate dehydrogenase defects in humans and mice. J Clin Endocrinol Metab. 2015;100(5):E710 E719. doi: /jc Gill AJ, Hes O, Papathomas T, et al. Succinate dehydrogenase (SDH)- deficient renal carcinoma: a morphologically distinct entity: a clinicopathologic series of 36 tumors from 27 patients. Am J Surg Pathol. 2014;38(12): doi: /pas Arch Pathol Lab Med Succinate Dehydrogenase Complex: An Updated Review Rasheed & Tarjan

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