Carrier frequency and incidence of alpha-mannosidosis: population database-based study—focus on the East Asian and Korean population

Background: Alpha-mannosidosis caused by mutations in the MAN2B1 gene is a rare genetic disorder characterized by physical abnormalities and intellectual disabilities. The objective of this study was to analyze the carrier frequency and estimated incidence of alpha-mannosidosis in East Asian populations, as limited data exists on its incidence in this group. Methods: In this study, a total of 125,748 exomes from the gnomAD database was analyzed. Additionally, 5,305 data from the KOVA and 1,722 data from the KRGDB, both representing Korean populations, were included. Results: The global carrier frequency of alpha-mannosidosis in gnomAD was 0.23%; the highest carrier frequency was observed in the Finnish at 0.49%, and East Asians had the second highest carrier frequency at 0.30%. Globally, the approximate incidence of alpha-mannosidosis was calculated at 1 in 784,535, l in 166,801 Europeans (Finnish), and l in 431,689 East Asians. By integrating the data from the 8,936 Koreans in gnomAD Korean, KOVA and KRGDB, the carrier frequency of alpha-mannosidosis in the Korean population was 0.04% and estimated incidence was 1 in 19,963,024. Conclusion: This study is the first to investigate the carrier frequencies of alpha-mannosidosis in East Asians and Koreans, including specific subpopulations, utilizing gnomAD and the Korean genomic database. The variant spectrum of MAN2B1 genes in East Asians showed significant differences compared to other ethnic groups. Our data provide valuable reference information for future investigations into alpha-mannosidosis, aiding in understanding the genetic diversity and specific variants associated with the condition in East Asian populations.


Introduction
Alpha-mannosidosis is an autosomal recessive lysosomal storage disorder (OMIM #248500) caused by mutations in the MAN2B1 gene, resulting in reduced activity of the alphamannosidase enzyme (Malm and Nilssen, 1993).The deficient

Interpretation of MAN2B1 variants and comparative analysis with disease databases
Interpretation of all MAN2B1 (NM_000528.4)variants followed the 2015 ACMG/AMP guidelines, along with adherence to the Sequence Variant Interpretation general recommendations by ClinGen (https://clinicalgenome.org/ working-groups/sequence-variant-interpretation/, accessed on March 10, 2023).These guidelines recommend categorizing variants into five classes: pathogenic variant (PV), likely pathogenic variant (LPV), variants of uncertain significance, likely benign variant, and benign variant.Variant pathogenicity predictions were made using REVEL (Ioannidis et al., 2016) and SpliceAI (Jaganathan et al., 2019).We compared all MAN2B1 variants identified in population databases with previously classified disease-causing variants from ClinVar and the Human Gene Mutation Database (HGMD), both of which are prominent disease repositories.ClinVar (https://www.ncbi.nlm.nih.gov/clinvar/, accessed on March 2, 2023) offers a freely accessible archive containing variant classifications provided by clinical laboratories.The HGMD professional database (http://www.hgmd.org/, release 2022.04) is a comprehensive repository of germline variants, categorized into six groups, from which we focused solely on disease-causing mutations (DM).

Alpha-mannosidosis carrier frequency and incidence estimation
Carrier frequencies of alpha-mannosidosis were determined for the MAN2B1 gene using a population database.We included variants classified as PV and LPV according to the 2015 ACMG-AMP guideline interpretation, DM entries from HGMD, and PV and LPV annotations from ClinVar for the carrier frequency analysis.Subsequently, we estimated the incidence of alphamannosidosis based on the calculated frequency and the Hardy-Weinberg equilibrium principle (1 = p 2 + 2pq + q 2 ), where p represents the major allele frequency (non-disease) and q denotes the minor allele frequency (disease).The major allele frequency p was approximated to be close to 1, with 2pq indicating carrier frequency and q 2 signifying the disease state.We estimated carrier frequencies using the sum of alleles interpreted as PV/LPV based on the population for which exome sequencing was performed.This allowed us to calculate the carrier frequency and subsequently derive the q value.Ultimately, we used the q value to predict q 2 to estimate disease incidence.We conducted statistical analysis using MedCalc ver.11.5.1.0(MedCalc Software, Maiakerke, Belgium), and we computed 95% confidence intervals for carrier frequency and estimated disease incidence.

Results
We conducted an analysis of 125,748 exomes, which included 9,197 East Asian exomes from the gnomAD database, focusing on variants within the MAN2B1 gene.Variant classification followed the 2015 ACMG/AMP guidelines, and we additionally compared these variants with data from two disease classification databases, HGMD and ClinVar (Table 1).Based on the 2015 ACMG/AMP guidelines, the global carrier frequency of alpha-mannosidosis was reported to be 0.23%.The carrier frequency and estimated incidence for each population are as follows (Figure 1): Among populations, the highest carrier frequency was observed in European (Finnish) at 0.49%.East Asians had the second highest carrier frequency at 0.30%.In contrast, the lowest carrier frequency was found among Ashkenazi Jewsh, 0.02%.The estimated incidence of alphamannosidosis was 1 in 784,535 worldwide, l in 166,801 European (Finnish) and l in 431,689 East Asians.According to ClinVar, the global carrier frequency for alpha-mannosidosis was reported as 0.16%, resulting in an estimated incidence of 1 in 1,505,514.HGMD data suggests a global carrier frequency of 0.67% for alphamannosidosis, resulting in an estimated incidence of 1 in 90,289.
According to 2015 ACMG/AMP guidelines, among East Asian populations in the gnomAD database, no PV/LPVs associated with alpha-mannosidosis were found in Koreans or Japanese (Supplementary Table S1).Therefore, the carrier frequency for alpha-mannosidosis in both populations was 0% in gnomAD.As a result, we could not evaluate the estimated incidence of alphamannosidosis in these specific East Asian populations based on the available data.The carrier frequency in other East Asian populations in gnomAD was 0.39%, and the estimated incidence was 1 in 265,372.
Based on the analysis conducted in accordance with the 2015 ACMG/AMP guidelines, the carrier frequency of alphamannosidosis in the Korean population database KOVA (n = 5,305) was determined to be 0.06%.Similarly, in the KRGDB database (n = 1,722), the carrier frequency was also 0.06%.The estimated incidence of alpha-mannosidosis in these populations was calculated to be 1 in 12,508,011 for the KOVA data and 1 in 11,861,136 for the KRGDB data (Supplementary Table S1).By integrating the data from the 8,936 individuals in gnomAD Korean, KOVA and KRGDB, the carrier frequency of alphamannosidosis in the Korean population was 0.04% and estimated incidence was 1 in 19,963,024.
Table 2 summarizes the PVs/LPVs in the MAN2B1 gene found in the gnomAD database.A total of 81 different variants were identified, comprising 284 alleles.Among them, 30 were missense variants (175 alleles), 22 were nonsense variants (48 alleles), 16 were frameshift variants (17 alleles), and 12 were splicing variants (43 alleles).Additionally, one synonymous variant was identified (1 allele).The most common variant worldwide was c.2248C>T, p.(Arg750Trp), with 59 alleles identified.This variant was frequently observed across multiple ethnicities but particularly in the European (Finnish) population.Following the most frequent variant, the next most common variants observed were c.565C>A, p.(Pro189Thr), and c.1830 + 1G>C, each found in 23 alleles.The c.565C>A variant was exclusively detected in Europeans (Finnish and non-Finnish), while the c.1830 + 1G>C variant was found only in East Asians.Additional analysis comparing PVs/LPVs in East Asians with those in other ethnic groups revealed significant distinctions.The PVs/ LPVs found in East Asian populations were absent in Latino, Ashkenazi Jewish, and South Asian populations.

Discussion
Within this study, the carrier frequency and estimated incidence of alpha-mannosidosis were analyzed using gnomAD and the Korean database.According to our evaluation of gnomAD data, the global carrier frequency of alpha-mannosidosis was established at 0.23%, whereas in East Asians, it was slightly higher at 0.30%.Ashkenazi Jewish individuals exhibited the lowest carrier frequency, which was recorded at 0.02%.Carrier frequency studies of alpha-  mannosidosis are rare, which poses challenges in directly comparing our findings with existing literature.However, considering the reported incidence rates of alpha-mannosidosis from previous studies, which range from approximately 1 in 300,000 to 1 in 1,000,000 (Malm et al., 1995;Meikle et al., 1999;Meikle et al., 2004;Poupetová et al., 2010), our estimated incidence of 1 in 784,535 is consistent.
The carrier frequency and incidence rates of alpha-mannosidosis in East Asian populations were examined for the first time in this study.Our findings reveal high carrier frequency of alpha-mannosidosis in East Asians (0.30%).However, our study also reveals that alphamannosidosis is significantly rare among Koreans, one of the major East Asian groups.By incorporating data from the KOVA and KRGDB databases, our analysis of 8,936 Koreans showed a carrier frequency of 0.04%.This carrier frequency is as low as the lowest frequency observed in Ashkenazi Jewish (0.02%) in the gnomAD.This highlights a distinct pattern in the carrier frequency and estimated incidence of alphamannosidosis among ethnic groups and emphasizes the need for population-specific studies to better understand the disease's distribution and genetic factors contributing to its occurrence.
Based on data obtained from the Korean Statistical Information Service (http://kosis.kr/; accessed on June 20, 2023), as of 2021, Korea had a total population of 51.7 million and recorded 260,562 births.Considering the carrier frequency determined in this study, it is estimated that there are approximately 21,000 carriers in the entire population and 104 carriers among newborns each year.The estimated incidence of alpha-mannosidosis in Korea, calculated using the Hardy-Weinberg equilibrium, is approximately 0.01 cases per year.Thus, the expected number of eligible patients for enzyme replacement therapy (ERT) in Korea is likely to be quite small.
The ACMG practice resource categorizes conditions into tiers based on carrier frequency for carrier screening during pregnancy and preconception (Gregg et al., 2021).Alpha-mannosidosis is not included in the Tier 3 genes recommended for screening.However, the BabySeq project, a randomized controlled trial for genetic newborn screening, classifies the MAN2B1 gene as Category A, emphasizing its significance in newborn screening for highly penetrant childhood-onset disorders (Ceyhan-Birsoy et al., 2017).The rarity and variable severity of alpha-mannosidosis can lead to a diagnostic delay of over a decade (Guffon et al., 2019).Early diagnosis is crucial for optimal treatment outcomes, including interventions such as allogeneic hematopoietic stem cell transplantation or ERT (Mynarek et al., 2012;Borgwardt et al., 2018).Recognizing the importance of early detection and intervention is essential for the wellbeing of at-risk infants.Notably, our study revealed that, among East Asians, the carrier frequency is the secondhighest following the European (Finnish) population.This highlights the importance of active genetic screening in the East Asian region as it is likely to contribute to the identification and treatment of affected individuals.Among the PVs/LPVs in the MAN2B1 gene identified in the gnomAD database, the most prevalent variant worldwide was c. 2248C>T, p.(Arg750Trp).This finding aligns with previous studies indicating this as the most frequently observed variant among alphamannosidosis patients (Riise Stensland et al., 2012).The c.2248C>T, p.(Arg750Trp) variant is common in European (Finnish) populations but has been observed in other ethnic groups, including East Asian and African populations.This indicates that the variant is not limited to a specific racial or ethnic background.The next most frequently observed variants were c.565C>A, p.(Pro189Thr) and c.1830 + 1G>C.Among these, the c.1830 + 1G>C variant was the second most common variant found in alpha-mannosidosis patients (Riise Stensland et al., 2012).The c.565C>A, p.(Pro189Thr) variant is specifically observed in East Asian populations and has not been reported in other ethnic groups.Since previous studies mainly included Western alpha-mannosidosis patients, the results indicate a significant concordance between the variants identified in population databases and those found in actual cases of alpha-mannosidosis.This highlights the utility of population databases as valuable resources for studying the prevalence and distribution of specific mutations in different populations.
This study has several limitations.Firstly, this study did not assess structural variations, such as large deletions or insertions in the MAN2B1 gene, which may have led to an underestimation of the carrier frequency of alpha-mannosidosis.While reported cases of alphamannosidosis resulting from structural mutations in the MAN2B1 gene are limited (Riise Stensland et al., 2012;Wiesinger et al., 2020), this is likely to have had a minimal impact on the underestimation of carrier frequency.Second, since the estimated incidence of alpha-mannosidosis in this study was derived entirely from genetic information, the accuracy of the estimated incidence may be impaired.Lastly, in gnomAD, for subpopulations with smaller sample sizes such as Koreans and Japanese, there were instances where PV/LPV were not identified.As a result, this has led to limitations in accurate assessment and estimation of carrier frequency.
Nevertheless, this study effectively verified the carrier frequency and estimated incidence of alpha-mannosidosis across ethnicities.Moreover, this is the largest study in East Asians, including Koreans, to analyze the MAN2B1 gene.To our knowledge, there is a lack of largescale population studies on the carrier frequency and estimated incidence of alpha-mannosidosis, specifically in Koreans.Therefore, we believe that this study provides a more accurate prediction of the carrier frequency in both East Asians and Koreans.Given the recent advancements in treatments for alpha-mannosidosis, identifying the carrier frequency and incidence for effective management and intervention is increasingly important.

Conclusion
This study is the first to investigate the carrier frequencies of alphamannosidosis in East Asians, including Koreans, utilizing gnomAD and the Korean genomic database.Our findings confirmed that, among various ethnic groups, East Asians are second in terms of carrier frequency of alpha-mannosidosis.However, among East Asians, Koreans exhibited significantly lower carrier frequencies.This implies that Koreans have a notably lower prevalence of alpha-mannosidosis than other populations, including other East Asian populations.The variant spectrum of MAN2B1 genes in East Asians showed significant differences compared to other ethnic groups.Our data provide valuable reference information for future investigations into alpha-mannosidosis, aiding in understanding the genetic diversity and specific variants associated with the condition in East Asian populations.

FIGURE 1
FIGURE 1Carrier frequency and estimated incidence of alpha-mannosidosis in gnomAD according to population.Circle size: carrier frequency of alphamannosidosis.

TABLE 1
Carrier frequency and estimated incidence of alpha-mannosidosis in gnomAD.

TABLE 2
Pathogenic variants and likely pathogenic variants in gnomAD.

TABLE 2 (
Continued) Pathogenic variants and likely pathogenic variants in gnomAD.

TABLE 2 (
Continued) Pathogenic variants and likely pathogenic variants in gnomAD.