Treatment of skeletal and non-skeletal alterations of Mucopolysaccharidosis type IVA by AAV-mediated gene therapy.

Mucopolysaccharidosis type IVA (MPSIVA) or Morquio A disease, a lysosomal storage disorder, is caused by N-acetylgalactosamine-6-sulfate sulfatase (GALNS) deficiency, resulting in keratan sulfate (KS) and chondroitin-6-sulfate accumulation. Patients develop severe skeletal dysplasia, early cartilage deterioration and life-threatening heart and tracheal complications. There is no cure and enzyme replacement therapy cannot correct skeletal abnormalities. Here, using CRISPR/Cas9 technology, we generate the first MPSIVA rat model recapitulating all skeletal and non-skeletal alterations experienced by patients. Treatment of MPSIVA rats with adeno-associated viral vector serotype 9 encoding Galns (AAV9-Galns) results in widespread transduction of bones, cartilage and peripheral tissues. This led to long-term (1 year) increase of GALNS activity and whole-body correction of KS levels, thus preventing body size reduction and severe alterations of bones, teeth, joints, trachea and heart. This study demonstrates the potential of AAV9-Galns gene therapy to correct the disabling MPSIVA pathology, providing strong rationale for future clinical translation to MPSIVA patients.

Plasma Proteomic Analysis in Morquio A Disease.

Mucopolysaccharidosis type IVA (MPS IVA) is a lysosomal disease caused by mutations in the gene encoding the enzymeN-acetylgalactosamine-6-sulfate sulfatase (GALNS), and is characterized by systemic skeletal dysplasia due to excessive storage of keratan sulfate (KS) and chondroitin-6-sulfate in chondrocytes. Although improvements in the activity of daily living and endurance tests have been achieved with enzyme replacement therapy (ERT) with recombinant human GALNS, recovery of bone lesions and bone growth in MPS IVA has not been demonstrated to date. Moreover, no correlation has been described between therapeutic efficacy and urine levels of KS, which accumulates in MPS IVA patients. The objective of this study was to assess the validity of potential biomarkers proposed by other authors and to identify new biomarkers. To identify candidate biomarkers of this disease, we analyzed plasma samples from healthy controls (n=6) and from untreated (n=8) and ERT-treated (n=5, sampled before and after treatment) MPS IVA patients using both qualitative and quantitative proteomics analyses. The qualitative proteomics approach analyzed the proteomic profile of the different study groups. In the quantitative analysis, we identified/quantified 215 proteins after comparing healthy control untreated, ERT-treated MPSIVA patients. We selected a group of proteins that were dysregulated in MPS IVA patients. We identified four potential protein biomarkers, all of which may influence bone and cartilage metabolism: fetuin-A, vitronectin, alpha-1antitrypsin, and clusterin. Further studies of cartilage and bone samples from MPS IVA patients will be required to verify the validity of these proteins as potential biomarkers of MPS IVA.

A Case Report of a Japanese Boy with Morquio A Syndrome: Effects of Enzyme Replacement Therapy Initiated at the Age of 24 Months.

BACKGROUND: Morquio A syndrome, mucopolysaccharidosis type IVA (MPS IVA), is a lysosomal storage disorder caused by the deficient activity of N-acetylgalactosamine-6-sulfatase (GalNac6S), due to alterations in the GALNS gene. This disorder results in marked abnormalities in bones and connective tissues, and affects multiple organs. Here, we describe the clinical course of a Japanese boy with MPS IVA who began enzyme replacement therapy (ERT) at the age of 24 months. PATIENT: the patient presented for kyphosis treatment at 22 months of age. An X-ray examination revealed dysostosis multiplex. Uronic acids were elevated in the urine and the keratan sulfate (KS) fraction was predominant. The leukocyte GalNac6S enzyme activity was extremely low. The patient exhibited the c.463G > A (p.Gly155Arg) mutation in GALNS. Based on these findings, his disease was diagnosed as classical (severe) Morquio A syndrome. An elosulfase alfa infusion was initiated at the age of 24 months. The patient's body height improved from -2.5 standard deviation (SD) to -2 SD and his physical activity increased during the first 9 months on ERT. However, he gradually developed paralysis in the lower legs with declining growth velocity, which required cervical decompression surgery in the second year of the ERT. The mild mitral regurgitation, serous otitis media, and mild hearing loss did not progress during treatment. CONCLUSION: early initiation of the elosulfase alfa to our patient showed good effects on the visceral system and muscle strength, while its effect on bones appeared limited. Careful observation is necessary to ensure timely surgical intervention for skeletal disorders associated with neurological symptoms. Centralized and multidisciplinary management is essential to improve the prognosis of pediatric patients with MPS IVA.

Safety, immunogenicity, and clinical outcomes in patients with Morquio A syndrome participating in 2 sequential open-label studies of elosulfase alfa enzyme replacement therapy (MOR-002/MOR-100), representing 5 years of treatment.

Elosulfase alfa is an enzyme replacement therapy for Morquio A syndrome (mucopolysaccharidosis IVA), a multisystemic progressive lysosomal storage disorder. This report includes the primary treatment outcomes and immunogenicity profile of elosulfase alfa in patients with Morquio A syndrome from 2 sequential studies, MOR-002 (ClinicalTrials.govNCT00884949) and MOR-100 (NCT01242111), representing >5 years of clinical study data. MOR-002 was an open-label, single-arm phase 1/2 study that evaluated the pharmacokinetics, safety, immunogenicity, and preliminary efficacy of 3 sequential doses of elosulfase alfa (0.1, 1.0, and 2.0 mg/kg/week) in patients with Morquio A syndrome (n = 20) over 36 weeks, followed by an optional 36- to 48-week treatment period using elosulfase alfa 1.0 mg/kg once weekly (qw). During the 0.1 mg/kg dosing phase, 1 patient discontinued due to a type I hypersensitivity adverse event (AE), and that patient's sibling voluntarily discontinued in the absence of AEs. An additional patient discontinued due to recurrent infusion reactions during the 1.0 mg/kg continuation phase. The remaining 17 patients completed MOR-002 and enrolled in MOR-100, an open-label, long-term extension study that further evaluated safety and clinical outcomes with elosulfase alfa administered at 2.0 mg/kg qw. During the course of MOR-100, patients were given the option of receiving elosulfase alfa infusions at home with nursing assistance. Over the course of both studies, all patients experienced ≥1 AE and most patients experienced a drug-related AE, generally of mild or moderate severity. Hypersensitivity reactions reported as related to study drug occurred in 25% of patients. Thirteen patients who chose to receive infusions at home had the same tolerability and safety profile, as well as comparable compliance rates, as patients who chose to receive on-site infusions. All patients developed antibodies to elosulfase alfa. Positivity for neutralizing antibodies was associated with increased drug half-life and decreased drug clearance. Despite formation of antidrug-binding (total antidrug antibodies, TAb) and in vitro neutralizing antibodies (NAb) in all patients, these types of immunogenicity to elosulfase alfa were not correlated with safety or clinical outcomes. In contrast with the reported natural history of Morquio A, no trends toward decreasing endurance, respiratory function, or ability to perform activities of daily living were observed in this cohort over the 5-year period.

Voice alterations in patients with Morquio A syndrome.

Morquio A syndrome, or mucopolysaccharidosis (MPS IV A), is an inherited lysosomal storage disorder which belongs to the group of mucopolysaccharidoses (MPSs). It is caused by N-acetylgalactosamine-6-sulfatase (GALNS) activity deficiency, which results in impaired degradation of glycosaminoglycans (GAGs), including keratan sulfate (KS) and chondroitin-6-sulfate (CS). These compounds infiltrate and disrupt the architecture of the extracellular matrix, compromising the integrity of the connective tissue. Patients with Morquio A have also been noted for exhibiting abnormalities of the larynx and vocal tract. The aim of the study was to assess voice alterations using noninvasive acoustic and electroglottographic voice analysis. Electroglottographic signal and acoustic analyses revealed considerable changes in the voices of patients with Morquio A syndrome when compared to the voices of healthy controls. Affected patients tended toward tense voice, incomplete glottal closure, increased incidence of vocal fold nodules, dysphonia, and hoarse voice. Morquio A syndrome is characterized by connective tissue disease, which adversely affects voice quality. The use of objective voice analysis makes it possible to quantitatively monitor changes in the vocal apparatus over the course of disease progression, and also allows for assessment of the effects of the enzyme replacement therapy.

Elosulfase alfa.

Mucopolysaccharidosis type IVA (MPS IVA), also known as Morquio A syndrome, is an inherited, lysosomal storage disorder caused by genetic mutations in N-acetylgalactosamine-6-sulfatase (GALNS) enzyme gene. GALNS is essential for breakdown of glycosaminoglycans. The disease is characterized by the early onset of severe skeletal dysplasia resulting in significant disability by the second decade of life. Until recently there have been no available treatments other than surgery and palliative care. BioMarin Pharmaceutical developed elosulfase alfa, a recombinant human GALNS coproduced with sulfatase-modifying factor 1, as an enzyme replacement therapy for patients with MPS IVA. In clinical studies, enzyme replacement therapy with elosulfase alfa significantly improved physical endurance, respiratory function, growth and quality of life in patients with MPS IVA. Treatment increased clearance of glycosaminoglycans and induced gene expression consistent with improved chondrocyte function. Elosulfase alfa is approved for the treatment of MPS IVA in the U.S. and Europe.

Mucopolysaccharidosis IVA: correlation between genotype, phenotype and keratan sulfate levels.

Mucopolysaccharidosis IVA (MPS IVA) is caused by deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS), leading to systemic skeletal dysplasia because of excessive storage of keratan sulfate (KS) in chondrocytes. In an effort to determine a precise prognosis and personalized treatment, we aim to characterize clinical, biochemical, and molecular findings in MPS IVA patients, and to seek correlations between genotype, phenotype, and blood and urine KS levels. Mutation screening of GALNS gene was performed in 55 MPS IVA patients (severe: 36, attenuated: 13, undefined: 6) by genomic PCR followed by direct sequence analysis. Plasma and urine KS levels were measured by ELISA method. Genotype/phenotype/KS correlations were assessed when data were available. Fifty-three different mutations including 19 novel ones (41 missense, 2 nonsense, 4 small deletions, 1 insertion, and 5 splice-site) were identified in 55 patients and accounted for 93.6% of the analyzed mutant alleles. Thirty-nine mutations were associated with a severe phenotype and ten mutations with an attenuated one. Blood and urine KS concentrations in MPS IVA patients were age-dependent and markedly higher than those in age-matched normal controls. Plasma and urine KS levels in MPS IVA patients with the severe phenotype were higher than in those with an attenuated form. This study provides evidence for extensive allelic heterogeneity of MPS IVA. Accumulation of mutations as well as clinical descriptions and KS levels allows us to predict clinical severity more precisely and should be used for evaluation of responses to potential treatment options.

Deficiency in N-acetylgalactosamine-6-sulfate sulfatase results in collagen perturbations in cartilage of Morquio syndrome A patients.

AIM: To investigate extracellular matrix (ECM) characteristics of cortical bone and articular cartilage of patients with Morquio syndrome A, a lysosomal storage disease caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase. PATIENTS AND METHODS: Cartilage, bone, and fibroblasts from 2 unrelated patients with Morquio syndrome were used. Histological analysis on bone and cartilage was carried out by means of light and electron microscopy. Lysyl hydroxylation and cross-linking of collagen present in bone, cartilage, and fibroblast cultures was determined by reverse-phase high performance liquid chromatography. RESULTS: No histological or biochemical differences were seen in cortical bone; furthermore, no differences were seen in the amount and modification of collagen deposited by fibroblasts. Articular cartilage showed major differences: collagen fibrils show a wider range of fibril diameter, the fibrils are in mean thicker, the lysyl hydroxylation level of the triple helix is strongly decreased, the total amount of pyridinolines is in the lower ranges, and the ratio hydroxylysylpyridinoline to lysylpyridinoline is decreased. Changes were also observed with respect to the arrangement of proteoglycans in the extracellular matrix surrounding the chondrocytes. CONCLUSION: The collagen of bone and the collagen deposited by fibroblasts is normal, whereas the ECM of cartilage in Morquio syndrome A patients is affected. Thus, deficiency in N-acetylgalactosamine-6-sulfate sulfatase has an impact on the phenotypic properties of chondrocytes, resulting in the formation of cartilage that is more prone to degeneration, being an explanation for the occurrence of osteoarthritis in Morquio syndrome A patients at early age.

Mucopolysaccharidoses type I and IVA: clinical features and consanguinity in Tunisia.

UNLABELLED: Mucopolysaccharidoses (MPS) are a group of lysosomal storage disorders caused by the deficiency of specific enzymes which leads to the lysosomal accumulation of glycosaminoglycanes. Mucopolysaccharidosis type I or Hurler disease is characterized by the deficiency of alpha-l-iduronidase enzyme. Mucopolysaccharidosis type IVA or Morquio A disease is due to the lack of N-acetylgalactosamine-6-sulfate-sulfatase. Theses deficiencies result in a progressive accumulation of the substrates: dermatan and heparan sulfates for Mucopolysaccharidosis type I and keratan sulfate for MPS type IVA. This process leads to progressive and chronic course for visceral attacks of the affected organs such as lungs and heart. In the Hurler disease, the nervous system is particularly affected while in Morquio a disease, a skeletal dysplasia and a normal intelligence are characteristic. AIM OF THE STUDY: This study was carried out on MPS type I and MPS type IVA unrelated families recruited from many regions of Tunisia in order to determine the relation between consanguinity and these types of disorders. PATIENTS AND METHODS: Clinical and molecular analyses confirmed the diagnosis for four MPS type I and five MPS type IVA studied families. RESULTS: First cousins unions characterize all families except one Hurler family and one Morquio A family where the consanguinity is third cousin degree. CONCLUSION: MPS type I and type IVA seems to be associated with consanguinity in Tunisia.

Effect of 'attenuated' mutations in mucopolysaccharidosis IVA on molecular phenotypes of N-acetylgalactosamine-6-sulfate sulfatase.

Mucopolysaccharidosis IVA is an autosomal recessive disease caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS). Mutation screening of the GALNS gene was performed for seven MPS IVA patients with attenuated phenotypes from three unrelated families. Four of 5 missense mutations identified in this study (p.F167V, p.R253W, p.R380S, p.P484S) and two reported (p.F97V, p.N204K), associated with attenuated phenotypes, were characterized using in vitro stable expression experiments, enzyme kinetic study, protein processing and structural analysis. The stably expressed mutant enzymes defining the attenuated phenotype exhibited a considerable residual activity (1.2-36.7% of the wild-type GALNS activity) except for p.R380S. Enzyme kinetic studies showed that p.F97V, p.F167V and p.N204K have lower affinity to the substrate compared with other mutants. The p.F97V enzyme was the most thermolabile at 55 degrees C. Immunoblot analyses indicated a rapid degradation and/or an insufficiency in processing in the mutant proteins. Tertiary structure analysis revealed that although there was a tendency for 'attenuated' mutant residues to be located on the surface of GALNS, they have a different effect on the protein including modification of the hydrophobic core and salt-bridge formation and different potential energy. This study demonstrates that 'attenuated' mutant enzymes are heterogeneous in molecular phenotypes, including biochemical properties and tertiary structure.

Development of MPS IVA mouse (Galnstm(hC79S.mC76S)slu) tolerant to human N-acetylgalactosamine-6-sulfate sulfatase.

Mucopolysaccharidosis IVA (MPS IVA) is an autosomal recessive disease caused by N-acetylgalactosamine-6-sulfate sulfatase (GALNS) deficiency. In recent studies of enzyme replacement therapy for animal models with lysosomal storage diseases, cellular and humoral immune responses to the injected enzymes have been recognized as major impediments to effective treatment. To study the long-term effectiveness and side effects of therapies in the absence of immune responses, we have developed an MPS IVA mouse model, which has many similarities to human MPS IVA and is tolerant to human GALNS protein. We used a construct containing both a transgene (cDNA) expressing inactive human GALNS in intron 1 and an active site mutation (C76S) in adjacent exon 2 and thereby introduced both the inactive cDNA and the C76S mutation into the murine Galns by targeted mutagenesis. Affected homozygous mice have no detectable GALNS enzyme activity and accumulate glycosaminoglycans in multiple tissues including visceral organs, brain, cornea, bone, ligament and bone marrow. At 3 months, lysosomal storage is marked within hepatocytes, reticuloendothelial Kupffer cells, and cells of the sinusoidal lining of the spleen, neurons and meningeal cells. The bone storage is also obvious, with lysosomal distention in osteoblasts and osteocytes lining the cortical bone, in chondrocytes and in the sinus lining cells in bone marrow. Ubiquitous expression of the inactive human GALNS was also confirmed by western blot using the anti-GALNS monoclonal antibodies newly produced, which resulted in tolerance to immune challenge with human enzyme. The newly generated MPS IVA mouse model should provide a good model to evaluate long-term administration of enzyme replacement.

Mouse model of N-acetylgalactosamine-6-sulfate sulfatase deficiency (Galns-/-) produced by targeted disruption of the gene defective in Morquio A disease.

Mucopolysaccharidosis IVA is an autosomal recessive disorder caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS), a lysosomal enzyme required for the stepwise degradation of keratan sulfate (KS) and chondroitin-6-sulfate (C6S). To generate a model for studies of the pathophysiology and of potential therapies, we disrupted exon 2 of Galns, the homologous murine gene. Homozygous Galns-/- mice have no detectable GALNS enzyme activity and show increased urinary glycosaminoglycan (GAGs) levels. These mice accumulate GAGs in multiple tissues including liver, kidney, spleen, heart, brain and bone marrow. At 2 months old, lysosomal storage is present primarily within reticuloendothelial cells such as Kupffer cells and cells of the sinusoidal lining of the spleen. Additionally, by 12 months old, vacuolar change is observed in the visceral epithelial cells of glomeruli and cells at the base of heart valves but it is not present in parenchymal cells such as hepatocytes and renal tubular epithelial cells. In the brain, hippocampal and neocortical neurons and meningeal cells had lysosomal storage. KS and C6S were more abundant in the cytoplasm of corneal epithelial cells of Galns-/- mice compared with wild-type mice by immunohistochemistry. Radiographs revealed no change in the skeletal bones of mice up to 12 months old. Thus, targeted disruption of the murine Galns gene has produced a murine model, which shows visceral storage of GAGs but lacks the skeletal features. The complete absence of GALNS in mutant mice makes them useful for studies of pharmacokinetics and tissue targeting of recombinant GALNS designed for enzyme replacement.

First-trimester diagnosis of Morquio disease type A.

Since the introduction in 1990 of a novel fluorogenic substrate for galactose-6-sulphate sulphatase we have used this substrate for prospective prenatal diagnosis in 10 pregnancies at risk for Morquio disease type A. Chorionic villi were analysed in five cases. The results indicated an affected fetus in one pregnancy which represents the first case of first-trimester diagnosis of this disorder; heterozygosity was demonstrated in two cases. Following amniocentesis, two affected fetuses and one heterozygote were diagnosed. The results of the present prospective prenatal analyses confirm our previous retrospective studies and demonstrate the reliability and convenience of the 4-methylumbelliferyl substrate.

The mouse N-acetylgalactosamine-6-sulfate sulfatase (Galns) gene: cDNA isolation, genomic characterization, chromosomal assignment and analysis of the 5'-flanking region.

Deficiency of lysosomal enzyme N-acetylgalactosamine-6-sulfate sulfatase (GALNS) leads to mucopolysaccharidosis IV A (MPS IV A), for which there is no definitive treatment so far. Although a number of mutations of the GALNS gene of MPS IV A patients have been described, pathogenesis of the disorder still remains elusive. In order to facilitate in vivo studies using model animals for MPS IV A, we isolated and performed molecular characterization of the mouse homolog of human GALNS. The 2.3-kb cDNA contains a 1560-bp open reading frame encoding 520 amino acid residues. The coding region has 84% similarity to the human GALNS cDNA at amino acid level. The mouse Galns gene was mapped by interspecific backcross analysis to the distal region of chromosome 8 where it co-segregates with Aprt. Northern blot analysis showed a wide expression of a single-copy gene, being higher especially in liver and kidney. The Galns gene was isolated from S129vJ genomic library and its genomic organization was characterized. The mouse Galns gene was about 50-kb long and organized into 14 exons and 13 introns. All intron-exon splice junctions conformed to the GT/AG consensus sequence except exon 8/intron 8 junction. Primer extension shows multiple transcription initiation sites between -44 and -75 although major transcription initiation site was observed at -90 bp from the ATG codon. The 5'-flanking region lacks canonical TATA and CAAT box sequences, but is G+C rich with 10 GC boxes (potential Sp1 binding sites), characteristic of a housekeeping gene promoter.

Combined adenine phosphoribosyltransferase and N-acetylgalactosamine-6-sulfate sulfatase deficiency.

We describe a Czech patient with combined adenine phosphoribosyltransferase (APRT) deficiency (2,8-dihydroxyadenine urolithiasis) and N-acetylgalactosamine-6-sulfate sulfatase (GALNS) deficiency (mucopolysaccharidosis Type IVA, Morquio disease A). Adenine and its extremely insoluble derivative, 2,8-dihydroxyadenine, were identified in the urine, and APRT deficiency was confirmed in erythrocytes. There was excessive excretion of keratan sulfate in the urine, and GALNS deficiency was confirmed in leukocytes. GALNS and APRT are both located on chromosome 16q24.3, suggesting that the patient had a deletion involving both genes. PCR amplification of genomic DNA indicated that a novel junction was created by the fusion of sequences distal to GALNS exon 2 and proximal to APRT exon 3, and that the size of the deleted region was approximately 100 kb. The deletion breakpoints were localized within GALNS intron 2 and APRT intron 2. Several other genes, including the alpha subunit of cytochrome B (CYBA), which is deleted or mutated in the autosomal form of chronic granulomatous disease, are located in the 16q24.3 region, but PCR amplification showed that this gene was present in the proband. A patient with hemizygosity for GALNS deficiency and APRT deficiency has been reported from Japan recently. These findings indicate that: (i) APRT is located telomeric to GALNS; (ii) GALNS and APRT are transcribed in the same orientation (centromeric to telomeric); and (iii) combined APRT/GALNS deficiency may be more common than hitherto realized.

Dental findings in three siblings with Morquio's syndrome.

Three siblings with Morquo's syndrome are described. Cultured fibroblasts from the youngest sibling demonstrated a total absence of N-acethylgalactosamine-6-sulphate-sulphatase whereas beta-galactocidase activity was normal, thus verifying the diagnosis of MPS-IV A. Dental features such as pointed cusps, spade-shaped incisors, thin enamel and pitted buccal surfaces were observed in all three children. Furthermore, in all three siblings the TMJ was affected with severe resorption of the head of the condyle. Histological examination of exfoliated primary molars showed a band of increased porosity following the striae of Retzius in the outer part of the enamel. These developmental disturbances were occasionally associated with minor localized defects in the enamel surface. The importance of close monitoring of dental development and regular dental care in order to prevent attrition of the teeth, loss of vertical face height and subsequent risk of TMJ dysfunction is emphasized.

Mucopolysaccharidosis IVA: four new exonic mutations in patients with N-acetylgalactosamine-6-sulfate sulfatase deficiency.

We report four new mutations in Japanese patients with mucopolysaccharidosis IVA (MPSIVA) who were heterozygous for a common double gene deletion. A nonsense mutation of CAG to TAG at codon 148 in exon 4 was identified, resulting in a change of Q to a stop codon and three missense mutations. V (GTC) to A (GCC) at codon 138 in exon 4, P (CCC) to S (TCC) at codon 151 in exon 5, and P (CCC) to L (CTC) at codon 151 in exon 5. Introduction of these mutations into the normal GALNS cDNA and transient expression in cultured fibroblasts resulted in a significant decrease in the enzyme activity. V138A and Q148X mutations result in changes of restriction site, which were analyzed by restriction-enzyme assay. P151S and P151L mutations that did not alter the restriction site were detected by direct sequencing or allele specific oligohybridization. Detection of the double gene deletion was initially done using Southern blots and was confirmed by PCR. Haplotypes were determined using seven polymorphisms to the GALNS locus in families with the double gene deletion. Haplotype analysis showed that the common double gene deletion occurred on a single haplotype, except for some variation in a VNTR-like polymorphism. This finding is consistent with a common founder for all individuals with this mutation.

Mucopolysaccharidosis IVA: submicroscopic deletion of 16q24.3 and a novel R386C mutation of N-acetylgalactosamine-6-sulfate sulfatase gene in a classical Morquio disease.

The N-acetylgalactosamine-6-sulfate sulfatase (GALNS) gene, which is responsible for autosomal recessive mucopolysaccharidosis IVA (MPSIVA), has been assigned to the long arm of chromosome 16, subregion 24.3, an area where the adenine phosophoribosyltransferase (APRT) gene and renal dipeptidase (DPEP I) gene are also localized. Molecular genetic studies on a severely affected patient with MPSIVA (Morquio disease), without karyotypic abnormality, revealed a partial submicroscopic deletion of 16q24.3 and a single point mutation on the other allele, with no functional GALNS activity. The patient, his mother, and siblings were hemizygous for GALNS and APRT loci, evidenced by informative RFLP and gene dosage analyses combined with a fluorescence in situ hybridization, utilizing a partial genomic clone of GALNS, but heterozygosity was retained at the DPEP I locus and proximal D16S7. Haplotyping of the family members revealed recombinational events between DPEP I locus and three other polymorphic loci on the paternal chromosome, localizing GALNS gene on the proximal side to DPEP I gene. As estimated from the genetic distance between two flanking markers of proximal D16S7 and distal DPEP I locus, size of the deletion was less than 3Mb. Mother of the boy and two older siblings were asymptomatic, despite this interstitial deletion of the Giemsa-light G band. The remaining paternal allele had no gene rearrangement but GALNS activity was not encoded as Arginine at 386 was replaced with Cysteine (R386C), suggesting this alteration accounts for the severe phenotype. Allelic loss of APRT is frequently observed in cancer tissues, thereby suggesting that the tumor suppressor gene locates near the APRT locus. No family member has evidence of any malignant disease. This study is apparently the first documentation of interstitial deletion of 16q24.3, involving GALNS and APRT genes.