DnaJ Heat Shock Protein Family B Member 9 Is a Novel Biomarker for Fibrillary GN.

Fibrillary GN (FGN) is a rare primary glomerular disease. Histologic and histochemical features of FGN overlap with those of other glomerular diseases, and no unique histologic biomarkers for diagnosing FGN have been identified. We analyzed the proteomic content of glomeruli in patient biopsy specimens and detected DnaJ heat shock protein family (Hsp40) member B9 (DNAJB9) as the fourth most abundant protein in FGN glomeruli. Compared with amyloidosis glomeruli, FGN glomeruli exhibited a >6-fold overexpression of DNAJB9 protein. Sanger sequencing and protein sequence coverage maps showed that the DNAJB9 protein deposited in FGN glomeruli did not have any major sequence or structural alterations. Notably, we detected DNAJB9 in all patients with FGN but not in healthy glomeruli or in 19 types of non-FGN glomerular diseases. We also observed the codeposition of DNAJB9 and Ig-γ Overall, these findings indicate that DNAJB9 is an FGN marker with 100% sensitivity and 100% specificity. The magnitude and specificity of DNAJB9 overabundance in FGN also suggests that this protein has a role in FGN pathogenesis. With this evidence, we propose that DNAJB9 is a strong biomarker for rapid diagnosis of FGN in renal biopsy specimens.

Applying Standard Clinical Chemistry Assay Validation to Droplet Digital PCR Quantitative Liquid Biopsy Testing.

BACKGROUND: Droplet digital PCR (ddPCR) is an emerging technology for quantitative cell-free DNA oncology applications. However, assay performance criteria must be established in a standardized manner to harness this potential. We reasoned that standard protocols used in clinical chemistry assay validation should be able to fill this need. METHODS: We validated KRAS, EGFR, and BRAF quantitative ddPCR assays based on the Clinical Laboratory Improvement Act regulations for laboratory-developed tests in clinical chemistry and the matching Clinical and Laboratory Standards Institute guidelines. This included evaluation of limit of the blank (LOB), limit of detection (LOD), limit of quantification (LOQ), intraassay and interassay imprecision, analytical range, dilution linearity, accuracy (including comparison with orthogonal platforms), reference range study, interference, and stability studies. RESULTS: For the ddPCR assays, the LOB was 4 mutant copies, LODs were 12 to 22 copies, and LOQs were 35 to 64 copies. The upper limit of the dynamic range was 30000 copies, and dilutions were linear down to the LOQs with good accuracy of spike recovery of Horizon reference material. Method comparisons with next-generation sequencing and an alternative ddPCR platform showed complete qualitative agreement and quantitative concordance, with slopes of 0.73 to 0.97 and R 2s of 0.83 to 0.99. No substantial interferences were discovered. Wild-type copy numbers in plasma ranged from 462 to 6169/mL in healthy individuals. CONCLUSIONS: Standard clinical chemistry assay validation protocols can be applied to quantitative ddPCR assays. This should facilitate comparison of the performance of different assays and allow establishment of minimal significant change thresholds in monitoring applications.

Novel Type of Renal Amyloidosis Derived from Apolipoprotein-CII.

Amyloidosis is characterized by extracellular deposition of misfolded proteins as insoluble fibrils. Most renal amyloidosis cases are Ig light chain, AA, or leukocyte chemotactic factor 2 amyloidosis, but rare hereditary forms can also involve the kidneys. Here, we describe the case of a 61-year-old woman who presented with nephrotic syndrome and renal impairment. Examination of the renal biopsy specimen revealed amyloidosis with predominant involvement of glomeruli and medullary interstitium. Proteomic analysis of Congo red-positive deposits detected large amounts of the Apo-CII protein. DNA sequencing of the APOC2 gene in the patient and one of her children detected a heterozygous c.206A→T transition, causing an E69V missense mutation. We also detected the mutant peptide in the proband's renal amyloid deposits. Using proteomics, we identified seven additional elderly patients with Apo-CII-rich amyloid deposits, all of whom had kidney involvement and histologically exhibited nodular glomerular involvement. Although prior in vitro studies have shown that Apo-CII can form amyloid fibrils and that certain mutations in this protein promote amyloid fibrillogenesis, there are no reports of this type of amyloidosis in humans. We propose that this study reveals a new form of hereditary amyloidosis (AApoCII) that is derived from the Apo-CII protein and appears to manifest in the elderly and preferentially affect the kidneys.

Hereditary Lysozyme Amyloidosis Variant p.Leu102Ser Associates with Unique Phenotype.

Lysozyme amyloidosis (ALys) is a rare form of hereditary amyloidosis that typically manifests with renal impairment, gastrointestinal (GI) symptoms, and sicca syndrome, whereas cardiac involvement is exceedingly rare and neuropathy has not been reported. Here, we describe a 40-year-old man with renal impairment, cardiac and GI symptoms, and peripheral neuropathy. Renal biopsy specimen analysis revealed amyloidosis with extensive involvement of glomeruli, vessels, and medulla. Amyloid was also detected in the GI tract. Echocardiographic and electrocardiographic findings were consistent with cardiac involvement. Proteomic analysis of Congo red-positive renal and GI amyloid deposits detected abundant lysozyme C protein. DNA sequencing of the lysozyme gene in the patient and his mother detected a heterozygous c.305T>C alteration in exon 3, which causes a leucine to serine substitution at codon 102 (Human Genome Variation Society nomenclature: p.Leu102Ser; legacy designation: L84S). We also detected the mutant peptide in the proband's renal and GI amyloid deposits. PolyPhen analysis predicted that the mutation damages the encoded protein. Molecular dynamics simulations suggested that the pathogenesis of ALys p.Leu102Ser is mediated by shifting the position of the central ß-hairpin coordinated with an antiparallel motion of the C-terminal helix, which may alter the native-state structural ensemble of the molecule, leading to aggregation-prone intermediates.

Clinical proteome informatics workbench detects pathogenic mutations in hereditary amyloidoses.

Shotgun proteomics of hereditary amyloid deposits generates all the information necessary to identify pathogenic mutant peptides and proteins. However, these mutant peptides are invisible to traditional database search strategies. We developed a two-pronged informatics workflow for detecting both known and novel amyloidogenic mutations from clinical proteomics data sets. We implemented the workflow in a CAP/CLIA certified clinical laboratory dedicated for proteomic subtyping of amyloid deposits extracted from formalin-fixed paraffin-embedded specimens. Performance of the workflow was characterized on a validation cohort of 49 hereditary amyloid samples, with confirmed mutations, and 85 controls. The sensitivity, specificity, positive predictive value, and negative predictive value of the known mutation detection workflow were determined to be 92%, 100%, 100%, and 96%, respectively. For novel mutation detection workflow, these performance parameters were 82%, 99%, 99%, and 90%, respectively. Validated workflow was applied to detect amyloidogenic mutations from a clinical cohort of 150 amyloid samples. The known mutation detection workflow detected rare frame shift mutations in apolipoprotein A1 and fibrinogen alpha amyloid deposits. The novel mutation detection workflow uncovered unanticipated mutations (W22G and C71Y) of the serum amyloid A4 protein present in patient amyloid deposits. In summary, clinical amyloid proteomics data sets contain mutant peptides of clinical significance that are recoverable with improved bioinformatics.

Allelic diversity in MCAD deficiency: the biochemical classification of 54 variants identified during 5 years of ACADM sequencing.

Medium-chain acyl-coA dehydrogenase (MCAD) deficiency is a commonly detected fatty acid oxidation disorder and its diagnosis relies on both biochemical and molecular analyses. Over a 5-year period, sequencing all 12 exons of the MCAD gene (ACADM) in our laboratory revealed a total of 54 variants in 549 subjects analyzed. As most molecular ACADM testing is referred for the follow-up of an abnormal newborn screening result obtained from an asymptomatic newborn, the identification of a novel DNA variant, or "variant of unknown significance (VUS)," presents clinicians with a dilemma. Frequently, the results of molecular analyses are correlated to biochemical findings, such as the concentration of octanoylcarnitine (C8) in plasma and the excretion of hexanoylglycine (HG) in urine. Here, we describe the classification of genotypes harboring at least one VUS through the comparison of C8 and HG values measured in individuals who are carriers of, or affected with, MCAD deficiency on the basis of the following genotypes: c.985A>G/wildtype, c.199T>C/c.985A>G and c.985A>G/c.985A>G. Our findings emphasize the importance of obtaining both plasma and urine when following up positive newborn screening results and may influence the way physicians counsel their asymptomatic patients about MCAD deficiency after genetic analysis.

First Report of Bilateral External Auditory Canal Cochlin Aggregates ("Cochlinomas") with Multifocal Amyloid-Like Deposits, Associated with Sensorineural Hearing Loss and a Novel Genetic Variant in COCH Encoding Cochlin.

Pathogenic variants in COCH, encoding cochlin, cause DFNA9 deafness disorder with characteristic histopathologic findings of cochlin deposits in the inner and middle ears. Here, we present the first case of deafness associated with bilateral external auditory canal (EAC) cochlin deposits, previously unreported evidence suggestive of cochlin-derived amyloid formation, and a novel COCH variant. A 54-year-old woman presented with progressive sensorineural hearing loss and bilateral EAC narrowing by subcutaneous thickening. Excision and histologic evaluation of tissue from both EACs showed paucicellular eosinophilic aggregates containing multiple Congo red-positive foci with yellow and green birefringence under crossed polarization light microscopy. Mass spectrometry performed on both the Congo red-positive and Congo red-negative areas identified cochlin as the most abundant protein, as well as a low abundance of universal amyloid signature peptides only in the Congo red-positive areas. Peptides indicative of a canonical amyloid type were not detected. Electron microscopy showed haphazard, branched microfibrils (3-7 nm in diameter) consistent with cochlin, as well as swirling fibrils (10-24 nm in diameter) reminiscent of amyloid fibrils. Cochlin immunohistochemical staining showed positivity throughout the aggregates. Sequencing of the entire COCH gene coding region from the patient's blood revealed a novel variant resulting in a non-conservative amino acid substitution of isoleucine to phenylalanine (c.1621A>T, p.I541F) in the vWFA2 domain at the protein's C-terminus. Our findings reveal a new pathologic manifestation of cochlin, raise the possibility of previously undescribed cochlin-derived amyloid formation, and highlight the importance of thoroughly investigating all aggregative tissue findings in the practice of diagnostic pathology.

Amyloid Typing by Mass Spectrometry in Clinical Practice: a Comprehensive Review of 16,175 Samples.

OBJECTIVE: To map the occurrence of amyloid types in a large clinical cohort using mass spectrometry-based shotgun proteomics, an unbiased method that unambiguously identifies all amyloid types in a single assay. METHODS: A mass spectrometry-based shotgun proteomics assay was implemented in a central reference laboratory. We documented our experience of typing 16,175 amyloidosis specimens over an 11-year period from January 1, 2008, to December 31, 2018. RESULTS: We identified 21 established amyloid types, including AL (n=9542; 59.0%), ATTR (n=4600; 28.4%), ALECT2 (n=511; 3.2%), AA (n=463; 2.9%), AH (n=367; 2.3%), AIns (n=182; 1.2%), KRT5-14 (n=94; <1%), AFib (n=71; <1%), AApoAIV (n=57; <1%), AApoA1 (n=56; <1%), AANF (n=47; <1%), Aß2M (n=38; <1%), ASem1 (n=34; <1%), AGel (n=29; <1%), TGFB1 (n=29; <1%), ALys (n=15; <1%), AIAPP (n=13; <1%), AApoCII (n=11; <1%), APro (n=8; <1%), AEnf (n=6; <1%), and ACal (n=2; <1%). We developed the first comprehensive organ-by-type map showing the relative frequency of 21 amyloid types in 31 different organs, and the first type-by-organ map showing organ tropism of 18 rare types. Using a modified bioinformatics pipeline, we detected amino acid substitutions in cases of hereditary amyloidosis with 100% specificity. CONCLUSION: Amyloid typing by proteomics, which effectively recognizes all amyloid types in a single assay, optimally supports the diagnosis and treatment of amyloidosis patients in routine clinical practice.

Solid phase and solution synthesis of NvocLys(CO(CH2)5NH-NBD)OCH2CN, a trifunctional fluorescent lysine derivative.

Herein, we describe a general strategy for the facile synthesis of a multifunctional amino acid derivative bearing both fluorescent and photolabile groups such as the lysine derivative NvocLys(CO(CH2)5NH-NBD)OCH2CN (1) that can be used as a biophysical tool for studying protein structure. The synthetic strategy involves functionalization of the amine groups while the amino acid is attached to a solid support, followed by esterification of the carboxylic acid in solution. The solid support protects the caboxylic acid, preventing a side reaction associated with the synthesis in solution and obviating the need for chromatographic purification of several intermediates. This synthetic strategy can be used for the preparation of a variety of amino acid derivatives with unusual alpha-amine and side chain functionalities.

The Power of Proficiency Testing: Unraveling Single-Nucleotide Polymorphism Interference, With Potential Impact on Clinical Testing of Spinocerebellar Ataxia Type 3.

CONTEXT.­: The College of American Pathologists proficiency testing program has been instrumental in identifying problems in clinical testing. OBJECTIVE.­: To describe how this program was used to identify a single-nucleotide polymorphism that affects clinical testing for spinocerebellar ataxia type 3. DESIGN.­: A proficiency testing sample with discordant results for spinocerebellar ataxia type 3 analysis was further evaluated by targeted Sanger sequencing and genotype polymerase chain reaction using multiple DNA polymerases. RESULTS.­: Of 28 laboratories responding in the spinocerebellar ataxia type 3 Proficiency Survey, 18 reported an incorrect homozygous result and 10 reported the expected heterozygous result. A heterozygous single-nucleotide polymorphism complementary to the 3' end of a published forward primer was identified in the proficiency testing sample, which may have led to allele dropout. However, this primer was used by only 3 of 18 laboratories (16%) reporting a homozygous result. A new forward primer of identical sequence, except for the 3' end being complementary to the single-nucleotide polymorphism, showed the expected heterozygous pattern. The possibility of DNA polymerase 3'-5' exonuclease activity contributing to allele dropout was investigated by testing 9 additional polymerases with and without exonuclease activity. No clear pattern emerged, but enzymes with and without 3'-5' exonuclease activity yielded both homozygous and expected heterozygous results with the published forward primer. CONCLUSIONS.­: Proactive systematic primer sequence checking is recommended because single-nucleotide polymorphism interference may result in allele dropout and impact clinical testing. Allele dropout is also influenced by other factors, including DNA polymerase exonuclease activity.

Identification of aggressive Gardner syndrome phenotype associated with a de novo APC variant, c.4666dup.

Gardner syndrome describes a variant phenotype of familial adenomatous polyposis (FAP), primarily characterized by extracolonic lesions including osteomas, dental abnormalities, epidermal cysts, and soft tissue tumors. We describe a 2-yr-old boy presenting with a 2-cm soft tissue mass of the forehead. Pathologic evaluation revealed a nuchal-type/Gardner-associated fibroma. Sequencing of the APC gene revealed a pathologic variant c.4666dupA. Parental sequencing of both blood and buccal tissue supported the de novo occurrence of this pathologic variant. Further imaging revealed a number of additional lesions including a large lumbar paraspinal desmoid, a 1-cm palpable lesion posterior to the left knee, firm lesions on bilateral heels, and multiple subdermal lesions. Colonoscopy was negative. This case illustrates a genetic variant of Gardner syndrome resulting in an aggressive early childhood phenotype and highlights the need for an individualized approach to treatment.

Consensus characterization of 16 FMR1 reference materials: a consortium study.

Fragile X syndrome, which is caused by expansion of a (CGG)(n) repeat in the FMR1 gene, occurs in approximately 1:3500 males and causes mental retardation/behavioral problems. Smaller (CGG)(n) repeat expansions in FMR1, premutations, are associated with premature ovarian failure and fragile X-associated tremor/ataxia syndrome. An FMR1-sizing assay is technically challenging because of high GC content of the (CGG)(n) repeat, the size limitations of conventional PCR, and a lack of reference materials available for test development/validation and routine quality control. The Centers for Disease Control and Prevention and the Association for Molecular Pathology, together with the genetic testing community, have addressed the need for characterized fragile X mutation reference materials by developing characterized DNA samples from 16 cell lines with repeat lengths representing important phenotypic classes and diagnostic cutoffs. The alleles in these materials were characterized by consensus analysis in nine clinical laboratories. The information generated from this study is available on the Centers for Disease Control and Prevention and Coriell Cell Repositories websites. DNA purified from these cell lines is available to the genetics community through the Coriell Cell Repositories. The public availability of these reference materials should help support accurate clinical fragile X syndrome testing.

A Patient With Hereditary ATTR and a Novel AGel p.Ala578Pro Amyloidosis.

Hereditary amyloidosis represents a group of diseases in which mutant proteins are deposited in various organs leading to their dysfunction. Correct identification of the amyloid-causing protein is critical because this will determine the optimal therapy for the patient. The most common type of hereditary amyloidosis is due to mutant transthyretin (ATTRm) deposition and often presents with heart failure or peripheral neuropathy. We report the first known case of a patient who had amyloidosis both due to a mutant transthyretin (p.Val122Ile) and due to a novel variant in the gelsolin gene (p.Ala578Pro). Both mutant proteins were identified by mass spectrometry analysis of amyloid deposits as well as sequencing of the genes. Molecular dynamic simulations suggest that the gelsolin p.Ala578Pro variant is likely amyloidogenic.

CDKN2A Germline Rare Coding Variants and Risk of Pancreatic Cancer in Minority Populations.

Background: Pathogenic germline mutations in the CDKN2A tumor suppressor gene are rare and associated with highly penetrant familial melanoma and pancreatic cancer in non-Hispanic whites (NHW). To date, the prevalence and impact of CDKN2A rare coding variants (RCV) in racial minority groups remain poorly characterized. We examined the role of CDKN2A RCVs on the risk of pancreatic cancer among minority subjects.Methods: We sequenced CDKN2A in 220 African American (AA) pancreatic cancer cases, 900 noncancer AA controls, and 183 Nigerian controls. RCV frequencies were determined for each group and compared with that of 1,537 NHW patients with pancreatic cancer. Odds ratios (OR) and 95% confidence intervals (CI) were calculated for both a case-case comparison of RCV frequencies in AAs versus NHWs, and case-control comparison between AA cases versus noncancer AA controls plus Nigerian controls. Smaller sets of Hispanic and Native American cases and controls also were sequenced.Results: One novel missense RCV and one novel frameshift RCV were found among AA patients: 400G>A and 258_278del. RCV carrier status was associated with increased risk of pancreatic cancer among AA cases (11/220; OR, 3.3; 95% CI, 1.5-7.1; P = 0.004) compared with AA and Nigerian controls (17/1,083). Further, AA cases had higher frequency of RCVs: 5.0% (OR, 13.4; 95% CI, 4.9-36.7; P < 0.001) compared with NHW cases (0.4%).Conclusions: CDKN2A RCVs are more common in AA than in NHW patients with pancreatic cancer and associated with moderately increased pancreatic cancer risk among AAs.Impact: RCVs in CDKN2A are frequent in AAs and are associated with risk for pancreatic cancer. Cancer Epidemiol Biomarkers Prev; 27(11); 1364-70. ©2018 AACR.

RET proto-oncogene genotyping using unlabeled probes, the masking technique, and amplicon high-resolution melting analysis.

Single bp mutations in the RET proto-oncogene can cause multiple endocrine neoplasia type 2 syndromes. The conventional approach for genotyping RET mutations is sequencing the exons. A closed-tube RET genotyping assay using a saturating DNA dye, unlabeled probes, and amplicon high-resolution melting analysis was developed. The method required two sequential polymerase chain reaction stages, a primary and secondary assay. The primary assay analyzed RET exons 10, 11, 13, 14, and 16 with a total of seven reactions using eight unlabeled probes. The primary assay genotyped wild-type exons, a common exon 13 polymorphism, and an exon 16 mutation, whereas other RET sequence variation was detected. The primary unlabeled probe data limited the possible genotypes for the detected RET sequence variation, which permitted genotyping in a secondary assay with only two to five reactions. Six probes were designed with the masking technique and masked selected sequence variations to allow unambiguous analysis of other mutations elsewhere under the probe. After this two-stage RET genotyping assay, less than 0.2% of exons tested would require sequencing for genotype. A blinded study generated from five wild type and 29 available RET sequence variation samples was 100% concordant with sequencing. Amplicon high-resolution melting analysis with unlabeled probes and the masking technique is a fast, accurate method for genotyping the >50 RET sequence variations.

Homozygous transthyretin mutation in an African American Male.

Cardiac amyloidosis of transthyretin type in the elderly may be senile or familial. The senile form is not typically associated with specific genetic changes. However, the familial form is and also occurs more frequently in African Americans than in the general population. One transthyretin mutation, V122I, is common in the African-American population, has a carrier frequency of 4%, and has marked cardiac specificity. Symptoms generally develop in the eighth and ninth decades. Here, we report the case of a 60-year-old African-American man who had a 2-year history of dyspnea and diffuse left ventricular wall thickening. Endomyocardial biopsy showed interstitial deposits of amorphous material confirmed as amyloid by Congo red staining and electron microscopy. Mass spectrometry showed a shift in protein mass of 14 d, indicative of transthyretin and confirming the production of abnormal protein. Bidirectional whole gene sequencing showed a homozygous mutation leading to a valine 122 isoleucine substitution (V122I). The 14-d mass shift observed using mass spectrometry is consistent with the V122I mutation. Homozygosity for the V122I mutation may be associated with earlier onset of cardiac disease. Transthyretin analysis should be considered for older African Americans with amyloid heart disease of transthyretin type.

Relationship between age-related macular degeneration-associated variants of complement factor H and LOC387715 with coronary artery disease.

OBJECTIVE: To determine whether either of the gene variants associated with age-related macular degeneration is associated with coronary artery disease (CAD). PATIENTS AND METHODS: This study consisted of 493 patients who underwent clinically indicated coronary angiography between June 1, 1998, and January 1, 1999. The Y402H variant of the complement factor H (CFH) gene and the A69S variant of the LOC387715 gene locus were examined by restriction fragment length polymorphism. Multiple logistic regression models were used to assess the association of CFH and LOC gene variants with CAD. Covariates with well-established associations with CAD were also evaluated. RESULTS: Seventy patients (14%) were homozygous for the histidine variant (HH) of CFH, 237 (48%) were heterozygous for the histidine variant (HY), and 186 (38%) were homozygous for the tyrosine variant (YY). Three hundred eight patients (62%) were homozygous for the alanine allele of LOC387715, 170 (34%) were heterozygous for Ala and Ser alleles, and 15 (3%) were homozygous for the serine variant. The overall association of the CFH genotype with CAD was not statistically significant (P=.08). However, some evidence (P=-.046) suggested that CAD was increased for the HH genotype compared to the homozygous wild-type YY genotype (odds ratio, 1.95; 95% confidence interval, 1.01-3.76). Male sex, hypertension, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, and age were other variables that demonstrated significant associations with CAD. The overall effect of the LOC genotype was not statistically significant (P=-.06). Heterozygosity for the serine variant was (P=-.02) associated with the absence of CAD vs the AS genotype (odds ratio, 0.59; confidence interval, 0.38-0.91; P=-.02). CONCLUSION: The CFH genotype may have an independent association with CAD, although our evidence did not show statistical significance. Controlling for known risk factors, the age-related macular degeneration-associated HH variant appears to be associated with CAD. The LOC387715 gene may also play a role in CAD.

Comparison of three methods for genotyping the UGT1A1 (TA)n repeat polymorphism.

OBJECTIVES: The UGT1A1 promoter contains a (TA)n repeat polymorphism. The 7 repeat allele is associated with decreased enzyme activity and patients homozygous for this allele treated with irinotecan may experience life-threatening toxicity. Here, we have compared three methods [DNA sequencing, fragment analysis, and the Invader assay (Third Wave Technologies)] for genotyping this polymorphism. RESULTS: All of the DNA samples (n=119) had concordant genotype calls between the sequencing and size-based methods. The Invader method was also concordant if the genotypes were 6/6, 6/7, or 7/7. Both the size-based method and the Invader method had straightforward data analysis, while interpretation of the sequencing results was occasionally more challenging. The Invader method required more concentrated DNA for analysis, was more expensive, and had a limited genotyping spectrum. CONCLUSION: All three methods were valuable for genotyping the UGT1A1 (TA)n repeat, with the sequencing and size-based assays having the fewest drawbacks.

LOC387715/HTRA1 and complement factor H variants in patients with age-related macular degeneration seen at the mayo clinic.

PURPOSE: To confirm association of the complement factor H allelic variant (CFH Y402H) and the LOC387715/HTRA1 (LOC387715 A69S) risk alleles with age-related macular degeneration (AMD). STUDY POPULATION: Study of 89 Caucasian patients with neovascular (exudative) AMD and 232 Caucasian controls. METHODS: The Y402H variant of CFH gene and A69S variant of LOC387715/HTRA1 gene locus were examined. RESULTS: For CFH, the odds ratio for the homozygous variant was 4.97 (CI 2.52 to 9.79). For LOC387715/HTRA1 the odds ratio for the homozygous risk variant was 7.75 (CI 3.46 to 17.35). The odds ratio for heterozygous carriers was 3.35 (CI 1.91 to 5.90).

Mitochondrial DNA sequence data reveals association of haplogroup U with psychosis in bipolar disorder.

Converging genetic, postmortem gene-expression, cellular, and neuroimaging data implicate mitochondrial dysfunction in bipolar disorder. This study was conducted to investigate whether mitochondrial DNA (mtDNA) haplogroups and single nucleotide variants (SNVs) are associated with sub-phenotypes of bipolar disorder. MtDNA from 224 patients with Bipolar I disorder (BPI) was sequenced, and association of sequence variations with 3 sub-phenotypes (psychosis, rapid cycling, and adolescent illness onset) was evaluated. Gene-level tests were performed to evaluate overall burden of minor alleles for each phenotype. The haplogroup U was associated with a higher risk of psychosis. Secondary analyses of SNVs provided nominal evidence for association of psychosis with variants in the tRNA, ND4 and ND5 genes. The association of psychosis with ND4 (gene that encodes NADH dehydrogenase 4) was further supported by gene-level analysis. Preliminary analysis of mtDNA sequence data suggests a higher risk of psychosis with the U haplogroup and variation in the ND4 gene implicated in electron transport chain energy regulation. Further investigation of the functional consequences of this mtDNA variation is encouraged.