1. Introduction

Deficient tissue-nonspecific alkaline phosphatase (TNSALP) enzyme activity is caused by a loss-of-function variant within the alkaline phosphatase (ALPL) gene and has been recognized as the underlying etiology of hypophosphatasia (HPP), a rare inborn-error-of-metabolism disease [1]. The resultant extracellular accumulation of TNSALP substrates, including inorganic pyrophosphate (a mineralization inhibitor) can impair the mineralization of teeth and bones. Clinical manifestations of HPP are extremely variable, ranging from perinatal death to adult dental problems and osteomalacia [2].

Six major classes of clinical manifestations of HPP have been reported, including lethal perinatal, prenatal (or perinatal) benign, infantile, childhood, adult, and odontohypophosphatasia (odonto-HPP) (Table 1). In lethal perinatal and infantile forms, variants usually affect both alleles of the ALPL gene. In milder subtypes (e.g., benign perinatal, childhood, adult, and odonto-HPP), the inheritance patterns may be autosomal dominant or autosomal recessive [3, 4]. Odonto-HPP is characterized by premature loss of primary teeth without skeletal abnormalities [5]. In most pediatric patients, at least one primary tooth (before the age of 5 years) is lost before resorption occurs because cementum hypoplasia impairs the connection between the tooth and the periodontal ligament. Typically, primary incisors are the first teeth affected, and the lower incisors are lost before the upper incisors [6]. To our knowledge, the genotype-phenotype relationship of odonto-HPP, a mild form of HPP, has not been described in the literature. Here, we describe a 4-year-old boy with odonto-HPP who carried two compound heterozygous missense variants in the ALPL gene.

2. Case report

A 4-year-old boy was referred to our hospital because he had experienced premature loss of primary teeth. He had no other clinical manifestations (e.g., involving other organs), and his body weight and height were normal. The proband had lost primary tooth 71 at the age of 18 months. Two years later, he lost teeth 72, 81, and 82. Subsequently, teeth 51 and 61 exhibited mobility, root exposure, and fan-shaped displacement. Three months later, these teeth were lost. Clinical examination of teeth 52 and 62 revealed grade II mobility, as well as gingival recession. Periodontal probing analysis showed that teeth 55 and 85 both had a probing depth of 5 mm; teeth 54, 65, 75, and 74 all had a probing depth of 4 mm. X-ray radiography revealed generalized horizontal alveolar bone loss comprising one-third of the bone in affected teeth (Fig. 1). The proband exhibited lower values of serum alkaline phosphatase (Table 2). Based on the clinical and biochemical findings, the proband was diagnosed with odonto-HPP.

Fig. 1.Clinical presentation of anterior dentition, gingival recession, and generalized horizontal bone loss.

The proband’s 8-year-old sister was asymptomatic. The proband’s mother exhibited periodontal disease and had lost four teeth between the ages of 36 and 38 years. As a child, the proband’s father had experienced intermittent spasms in the lower limbs while walking; these spasms had disappeared in adulthood. Moreover, the proband’s father had lost two teeth between the ages of 38 and 39 years. Biochemical tests showed that the proband’s mother had a serum alkaline phosphatase level within the normal range, whereas the proband’s father and sister both had low serum alkaline phosphatase levels. Furthermore, the proband’s mother had a serum phosphorus level within the normal range, but the serum phosphorus levels were elevated in the proband, proband’s father, and proband’s sister. The proband’s father and mother both had low levels of 25-hydroxyvitamin D, whereas the proband and his sister had normal levels of 25-hydroxyvitamin D. The calcium levels were normal in all family members. Pyridoxal 5’-phosphate/vitamin B6 levels were not analyzed. To explore the genetic etiology, peripheral blood samples from the family members were subjected to whole-exome sequencing and Sanger sequencing. Genetic analysis of the proband’s ALPL gene revealed two compound heterozygous missense variants: c.346G>A (p.Ala116Thr) and c.1563C>G (p.Ser521Arg). Each of these two variants was derived from one of his parents. The proband’s mother was a heterozygous carrier of c.1563C>G (p.Ser521Arg), whereas his father and sister were heterozygous carriers of c.346G>A (p.Ala116Thr) (Fig. 2).

Fig. 2.Sequencing analysis of the ALPL gene in the proband and his family members.

To characterize the relationship between genotype and phenotype, three-dimensional protein structure analysis was performed using UCSF Chimera software version 1.14 (Resource for Biocomputing, Visualization, and Informatics, San Francisco, CA, USA). c.346G>A (p.Ala116Thr) resulted in a change from alanine to threonine at amino acid 116 in the TNSALP protein. The hydrogen bond interactions between threonine-116 and threonine-113 were increased (purple dashed lines), compared with the hydrogen bond interactions between alanine-116 and threonine-113 in the wild-type protein. In the three-dimensional structure, amino acid 116 is located in the core region and is a buried residue. c.346G>A (p.Ala116Thr) causes side chain enlargement and hydrophilicity enhancement, leading to the disruption of structural stability (blue dashed lines). c.1563C>G (p.Ser521Arg) resulted in a change from serine to arginine at amino acid 521. The polarity (blue dashed lines) and hydrogen bond interactions (purple dashed lines) were not affected by the variant. The amino acid side chains were enlarged and the hydrophobicity was increased (dark blue dashed lines) in exposed residues at the C-terminus of the protein (Fig. 3). According to the American College of Medical Genetics guidelines [7], c.346G>A (p.Ala116Thr) was classified as “pathogenic”, whereas c.1563C>G (p.Ser521Arg) was classified as “unclear significance”.

Fig. 3.Three-dimensional modeling structures of ALPL variants. (A) Ribbon presentation of wild-type and mutant ALPL monomers. (B) Structural representation of amino acid 116 in wild-type and mutant proteins. (C) Structural representation of amino acid 521 in wild-type and mutant proteins.

3. Discussion

Variants in ALPL diminish expression of the gene, compromise its mRNA stability, and inactivate the enzyme by altering its various domains. Thus far, >400 (usually missense) ALPL-related genetic defects have been recorded. Our patient exhibited a novel combination of two compound heterozygous missense variants in the ALPL gene, c.346G>A (p.Ala116Thr) and c.1563C>G (p.Ser521Arg), which resulted in the odonto-HPP phenotype.

Genetic analyses by SIFT, Condel, and SpliceAI predicted that c.346G>A was deleterious; PhyloP Vertebrates, PhyloP Placental Mammals, and GERP++ predicted that the variant was evolutionary conserved. A previous study revealed heterozygous expression of c.346G>A in patients with odonto-HPP and childhood HPP [8], but the clinical significance of c.1563C>G was not previously described. The allele frequencies of c.1563C>G in humans were reported to be 0.000021, 0.000026, and 0.000297 in the Exome Aggregation Consortium database, Genome Aggregation Database, and GnomAD-EAS database, respectively. An in vitro study showed that the c.1563C>G variant impairs enzymatic alkaline phosphatase activity [9]. Our present findings indicate that c.346G>A is a pathogenic genetic alteration, whereas c.1563C>G might contribute to the dental phenotype when combined with c.346G>A.

Thus, genetic diagnosis might be considered when early tooth loss occurs in a patient with a positive family history. Pediatric dentists providing primary care should consider a diagnosis of odonto-HPP in children with premature loss of primary teeth; they should also ensure rapid referral for patients who require specialized medical management based on their inheritance patterns. Odonto-HPP is a progressive disease; thus, parents should be informed that the affected patient might manifest bone symptoms with maturity because of inorganic pyrophosphate accumulation. Alkaline phosphatase enzyme replacement therapy has been reported to improve bone hypomineralization in patients with HPP [10]. Although there are minimal published data concerning the management of oral health in patients with HPP, dental therapeutic options (e.g., promoting awareness of rigorous dental hygiene prevention, periodontal cleaning to control local determinants of alveolysis, early clinical management of caries caused by bacterial biofilm, orthodontic management, and functional rehabilitation) might delay disease progression in the oral cavity [11]. Therefore, pediatricians and pediatric dentists should collaborate to prepare appropriate therapeutic plans.

Abbreviations

HPP: hypophosphatasia; TNSALP: tissue-nonspecific alkaline phosphatase; ALPL: alkaline phosphatase; odonto-HPP: odontohypophosphatasia.

Availability of Data and Materials

Not applicable.

Author contributions

JX and JJ—conceived the ideas and wrote the manuscript. HWL, JJ and HYK—performed the clinical treatment. LSG and XHZ—conducted the data analysis and revised the manuscript.

Ethics approval and consent to participate

Research study protocols were approved by the Ethics Committee of Shenzhen Longgang E.N.T. Hospital. Reference number of approval of the Ethics Committee was ZSSOM2022-0001. The parents gave their written consent for their child’s personal or clinical details to be published in this study.

Acknowledgment

Not applicable.

Funding

This work was supported by Guangdong Basic and Applied Basic Research Foundation (2020A1515010237); Shenzhen Key Medical Discipline Construction Fund (No.SZXK039); Free Exploration Projects of Shenzhen Science and Technology Innovation Committee (JCYJ20180305163353862 and JCYJ20180305163259711); Special Fund for Science and Technology Development of Longgang District, Shenzhen (LGKCYLWS2021000031).

Conflict of interest

The authors declare no conflict of interest.