Carla G. Monico

Primary hyperoxaluria type 1: update and additional mutation analysis of the AGXT gene†

Primary hyperoxaluria type 1 (PH1) is an autosomal recessive, inherited disorder of glyoxylate metabolism arising from a deficiency of the alanine:glyoxylate aminotransferase (AGT) enzyme, encoded by the AGXT gene. The disease is manifested by excessive endogenous oxalate production, which leads to impaired renal function and associated morbidity. At least 146 mutations have now been described, 50 of which are newly reported here. The mutations, which occur along the length of the AGXT gene, are predominantly single‐nucleotide substitutions (75%), 73 are missense, 19 nonsense, and 18 splice mutations; but 36 major and minor deletions and insertions are also included. There is little association of mutation with ethnicity, the most obvious exception being the p.Ile244Thr mutation, which appears to have North African/Spanish origins. A common, polymorphic variant encoding leucine at codon 11, the so‐called minor allele, has significantly lower catalytic activity in vitro, and has a higher frequency in PH1 compared to the rest of the population. This polymorphism influences enzyme targeting in the presence of the most common Gly170Arg mutation and potentiates the effect of several other pathological sequence variants. This review discusses the spectrum of AGXT mutations and polymorphisms, their clinical significance, and their diagnostic relevance. Hum Mutat 30, 910–917, 2009.

Transplantation Outcomes in Primary Hyperoxaluria

Optimal transplantation strategies are uncertain in primary hyperoxaluria (PH) due to potential for recurrent oxalosis. Outcomes of different transplantation approaches were compared using life‐table methods to determine kidney graft survival among 203 patients in the International Primary Hyperoxaluria Registry. From 1976–2009, 84 kidney alone (K) and combined kidney and liver (K + L) transplants were performed in 58 patients. Among 58 first kidney transplants (32 K, 26 K + L), 1‐, 3‐ and 5‐year kidney graft survival was 82%, 68% and 49%. Renal graft loss occurred in 26 first transplants due to oxalosis in ten, chronic allograft nephropathy in six, rejection in five and other causes in five. Delay in PH diagnosis until after transplant favored early graft loss (p = 0.07). K + L had better kidney graft outcomes than K with death‐censored graft survival 95% versus 56% at 3 years (p = 0.011). Among 29 year 2000–09 first transplants (24 K + L), 84% were functioning at 3 years compared to 55% of earlier transplants (p = 0.05). At 6.8 years after transplantation, 46 of 58 patients are living (43 with functioning grafts). Outcomes of transplantation in PH have improved over time, with recent K + L transplantation highly successful. Recurrent oxalosis accounted for a minority of kidney graft losses.

International registry for primary hyperoxaluria

Background/Aims: Primary hyperoxaluria (PH) is an inherited disorder that causes calcium urolithiasis and renal failure. Due to its rarity, experience at most centers with this disease is limited. Methods: A secure, web-based, institutional review board/ethics committee and American Health Insurance Portability and Accountability Act (HIPAA)-compliant registry was developed to facilitate international contributions to a data base. To date 95 PH patients have been entered. Results: PH type was confirmed in 84/95 (PH1 79%, PH2 9%). Mean age ± SD at symptom onset was 9.5 ± 10.2 (median 5.5) years whereas age at diagnosis was 15.0 ± 15.2 (median 10.0) years. Urolithiasis was present at diagnosis in 90% (mean 7, median 1, stones prior to diagnosis) and nephrocalcinosis in 48%. Surprisingly 15% of the patients were asymptomatic at the time of diagnosis. Nineteen of the 95 patients were first recognized to have PH after they had reached end-stage renal disease, with the diagnosis made only after kidney transplantation in 7 patients. Patients were followed for 12.1 ± 10.6 (median 9.4) years. Thirty-four of 95 progressed to end-stage renal failure, before (19 patients) or after (15 patients) diagnosis. In the PH1 cohort actuarial renal survival was 64% at 30 years of age, 47% at 40 years, and 29% at 50 years. Conclusion: We have developed a PH registry, and demonstrated the feasibility of this secure, web-based approach for data entry. By facilitating accumulation of an increasing cohort of patients, this registry should allow more complete characterization of clinical expression of PH, an appreciation of geographic variability, and identification of treatment outcomes.

Primary hyperoxaluria type III gene HOGA1 (Formerly DHDPSL) as a possible risk factor for idiopathic calcium oxalate urolithiasis

BACKGROUND AND OBJECTIVES Primary hyperoxaluria types I and II (PHI and PHII) are rare monogenic causes of hyperoxaluria and calcium oxalate urolithiasis. Recently, we described type III, due to mutations in HOGA1 (formerly DHDPSL), hypothesized to cause a gain of mitochondrial 4-hydroxy-2-oxoglutarate aldolase activity, resulting in excess oxalate. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS To further explore the pathophysiology of HOGA1, we screened additional non-PHI-PHII patients and performed reverse transcription PCR analysis. Postulating that HOGA1 may influence urine oxalate, we also screened 100 idiopathic calcium oxalate stone formers. RESULTS Of 28 unrelated hyperoxaluric patients with marked hyperoxaluria not due to PHI, PHII, or any identifiable secondary cause, we identified 10 (36%) with two HOGA1 mutations (four novel, including a nonsense variant). Reverse transcription PCR of the stop codon and two common mutations showed stable expression. From the new and our previously described PHIII cohort, 25 patients were identified for study. Urine oxalate was lower and urine calcium and uric acid were higher when compared with PHI and PHII. After 7.2 years median follow-up, mean eGFR was 116 ml/min per 1.73 m(2). HOGA1 heterozygosity was found in two patients with mild hyperoxaluria and in three of 100 idiopathic calcium oxalate stone formers. No HOGA1 variants were detected in 166 controls. CONCLUSIONS These findings, in the context of autosomal recessive inheritance for PHIII, support a loss-of-function mechanism for HOGA1, with potential for a dominant-negative effect. Detection of HOGA1 variants in idiopathic calcium oxalate urolithiasis also suggests HOGA1 may be a predisposing factor for this condition.

Implications of Genotype and Enzyme Phenotype in Pyridoxine Response of Patients with Type I Primary Hyperoxaluria

Background: Marked hyperoxaluria due to liver-specific deficiency of alanine:glyoxylate aminotransferase activity (AGT) characterizes type I primary hyperoxaluria (PHI). Approximately half of PHI patients experience improvement in the degree of hyperoxaluria following pyridoxine (VB6) treatment. Recently, we showed an association between VB6 response and the commonest PHI mutation G170R, with patients possessing one or two copies showing 50% reduction or complete to near complete normalization of oxaluria, respectively. Two patients showed responses varying from this pattern. To further clarify the molecular basis of VB6 response in PHI, we performed additional genotyping. Methods: 23 PHI patients diagnosed via hepatic enzyme analysis, hyperoxaluria and hyperglycolic aciduria or homozygosity for a known mutation, availability of pre- and post-VB6 24-hour urine oxalate and GFR >40 ml/min/1.73 m2 were included. Data was retrieved retrospectively, oxalate measured by oxalate oxidase, and genotyping performed by PCR-based methods. Results: VB6 response was associated with the G170R and F152I mutations. Eight new sequence changes were detected. Conclusions: In PHI, two mutations resulting in AGT mistargeting are associated with VB6 response. Whether this favorable effect is specific to the peroxisomal-to-mitochondrial mistargeting caused by these changes or due to another mechanism remains to be determined.

Phenotypic and Functional Analysis of Human SLC26A6 Variants in Patients With Familial Hyperoxaluria and Calcium Oxalate Nephrolithiasis

BACKGROUND Urinary oxalate is a major risk factor for calcium oxalate stones. Marked hyperoxaluria arises from mutations in 2 separate loci, AGXT and GRHPR, the causes of primary hyperoxaluria (PH) types 1 (PH1) and 2 (PH2), respectively. Studies of null Slc26a6(-/-) mice have shown a phenotype of hyperoxaluria, hyperoxalemia, and calcium oxalate urolithiasis, leading to the hypothesis that SLC26A6 mutations may cause or modify hyperoxaluria in humans. STUDY DESIGN Cross-sectional case-control. SETTING & PARTICIPANTS Cases were recruited from the International Primary Hyperoxaluria Registry. Control DNA samples were from a pool of adult subjects who identified themselves as being in good health. PREDICTOR PH1, PH2, and non-PH1/PH2 genotypes in cases. OUTCOMES & MEASURES Homozygosity or compound heterozygosity for SLC26A6 variants. Functional expression of oxalate transport in Xenopus laevis oocytes. RESULTS 80 PH1, 6 PH2, 8 non-PH1/PH2, and 96 control samples were available for SLC26A6 screening. A rare variant, c.487C-->T (p.Pro163Ser), was detected solely in 1 non-PH1/PH2 pedigree, but this variant failed to segregate with hyperoxaluria, and functional studies of oxalate transport in Xenopus oocytes showed no transport defect. No other rare variant was identified specifically in non-PH1/PH2. Six additional missense variants were detected in controls and cases. Of these, c.616G-->A (p.Val206Met) was most common (11%) and showed a 30% reduction in oxalate transport. To test p.Val206Met as a potential modifier of hyperoxaluria, we extended screening to PH1 and PH2. Heterozygosity for this variant did not affect plasma or urine oxalate levels in this population. LIMITATIONS We did not have a sufficient number of cases to determine whether homozygosity for p.Val206Met might significantly affect urine oxalate. CONCLUSIONS SLC26A6 was effectively ruled out as the disease gene in this non-PH1/PH2 cohort. Taken together, our studies are the first to identify and characterize SLC26A6 variants in patients with hyperoxaluria. Phenotypic and functional analysis excluded a significant effect of identified variants on oxalate excretion.

Genetic determinants of urolithiasis

Urolithiasis affects approximately 10% of individuals in Western societies by the seventh decade of life. The most common form, idiopathic calcium oxalate urolithiasis, results from the interaction of multiple genes and their interplay with dietary and environmental factors. To date, considerable progress has been made in identifying the metabolic risk factors that predispose to this complex trait, among which hypercalciuria predominates. The specific genetic and epigenetic factors involved in urolithiasis have remained less clear, partly owing to the candidate gene and linkage methods that have been available until now, being inherently low in their power of resolution and in assessing modest effects in complex traits. However, together with investigations of rare, Mendelian forms of urolithiasis associated with various metabolic risk factors, these methods have afforded insights into biological pathways that seem to underlie the development of stones in the urinary tract. Monogenic diseases account for a greater proportion of stone formers in children and adolescents than in adults. Early diagnosis of monogenic forms of urolithiasis is of importance owing to associated renal injury and other potentially treatable disease manifestations, but diagnosis is often delayed because of a lack of familiarity with these rare disorders. In this Review, we will discuss advances in the understanding of the genetics underlying polygenic and monogenic forms of urolithiasis.

Primary hyperoxaluria type 1 and brachydactyly mental retardation syndrome caused by a novel mutation in AGXT and a terminal deletion of chromosome 2

Primary hyperoxaluria type 1 (PH1) is an autosomal recessive disorder caused by mutations in the alanine:glyoxylate aminotransferase (AGXT) gene, located on chromosome 2q37. Mutant AGXT leads to excess production and excretion of oxalate, resulting in accumulation of calcium oxalate in the kidney, and progressive loss of renal function. Brachydactyly mental retardation syndrome (BDMR) is an autosomal dominant disorder, caused by haploinsufficiency of histone deacetylase 4 (HDAC4), also on chromosome 2q37. It is characterized by skeletal abnormalities and developmental delay. Here, we report on a girl who had phenotypes of both PH1 and BDMR. PCR‐sequencing of the coding regions of AGXT showed a novel missense mutation, c.32C>G (p.Pro11Arg) inherited from her mother. Functional analyses demonstrated that it reduced the enzymatic activity to 31% of the wild‐type and redirected some percentage of the enzyme away from the peroxisome. Microsatellite and array‐CGH analyses indicated that the proband had a paternal de novo telomeric deletion of chromosome 2q, which included HDAC4. To our knowledge, this is the first report of PH1 and BDMR, with a novel AGXT mutation and a de novo telomeric deletion of chromosome 2q.

Primary hyperoxaluria: new directions for diagnosis and treatment

Abstract Primary hyperoxaluria type 1 (PH1) is a rare genetic autosomal recessive disorder caused by deficient function of the liver-specific metabolic enzyme alanine:glyoxalate aminotransferase (AGT). AGT normally catalyzes the breakdown of glyoxalate to glycine; in PH 1 , AGT function is abnormal and oxalate accumulates within the body. Oxalate can only be removed from the body by renal excretion, leading to hyperoxaluria with deposition of oxalate in the kidneys and progressive nephrocalcinosis, urolithiasis and renal failure. As renal function declines, oxalate deposition rapidly occurs in tissues throughout the body and results in systemic multiorgan dysfunction and early mortality. Diagnosis of PH1Sn the past was based on clinical presentation in conjunction with high plasma and urinary oxalate measurements. Confirmation of the diagnosis was made by examining liver biopsies for AGT activity. Treatment was primarily based on measures to increase urine flow and decrease oxalate crystallization within the kidney tubules. Pyridoxine therapy was empirically found to be helpful in a number of cases. Renal replacement therapy (dialysis and renal transplantation) met with limited success due to the low rate of oxalate clearance with both haemo- and peritoneal dialysis and the rapid deposition of oxalate within the transplanted kidney. Over the last 5 years, new understanding of the structure and function of AGT and the molecular basis of AGT dysfunction has resulted in the development of new diagnostic techniques which can in some cases eliminate the need for invasive liver biopsies. In addition, characterization of a known genetic polymorphism in combination with a specific amino acid mutation has been found to correlate with response to pyridoxine therapy in a third of patients. Improved long term results in patients who have progressed to renal failure have been reported with combined liver–kidney transplantation in conjunction with aggressive perioperative measures to reduce serum oxalate levels and systemic oxalate stores. Some experience with preemptive liver transplant prior to the development of renal failure has been reported. The role of pre-emptive liver transplantation remains uncertain given the unpredictability of the timing of renal failure and risks of liver transplantation and life-long immune suppression. Continuing efforts at developing specific gene therapy are ongoing.