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. 2022 Oct 31;17(10):e0276640. doi: 10.1371/journal.pone.0276640

Dp71 and intellectual disability in Indonesian patients with Duchenne muscular dystrophy

Kristy Iskandar 1,*, Agung Triono 1, Sunartini 1, Ery Kus Dwianingsih 2, Braghmandita Widya Indraswari 1, Ignatia Rosalia Kirana 1, Gabriele Ivana 1, Retno Sutomo 1, Suryono Yudha Patria 1, Elisabeth Siti Herini 1, Gunadi 3
Editor: Giulio Piluso4
PMCID: PMC9621454  PMID: 36315559

Abstract

Introductions

Duchenne muscular dystrophy (DMD) is an X-linked recessive progressive muscular disease marked by developmental delays due to mutations in the DMD gene, which encodes dystrophin. Brain comorbidity adds to the burden of limited mobility and significantly impacts patients’ quality of life and their family. The changes of expression of dystrophin isoforms in the brain due to DMD gene mutations are thought to be related to the cognitive and neurobehavior profiles of DMD.

Objectives

This cross-sectional study aimed to characterize cognitive and neurodevelopmental profiles of patients with DMD and to explore underlying genotype-phenotype associations.

Methods

Patients with DMD aged 5–18 years from Dr Sardjito Hospital and Universitas Gadjah Mada Academic Hospital from 2017–2022 were included. Multiplex ligation-dependent probe amplification and whole exome sequencing were used to determine mutations in the DMD genes. Cognitive function was measured by intelligence quotient testing using the Wechsler Intelligence Scale for Children and adaptive function tests with Vineland Adaptive Behavior Scales. The Autism Mental Status Exam and Abbreviated Conner’s Rating Scale were used to screen for autism spectrum disorder (ASD) and attention deficit and hyperactivity disorder (ADHD), respectively.

Results

The mean total IQ score of DMD patients was lower than that of the general population (80.6 ± 22.0 vs 100 ± 15), with intellectual disability observed in 15 boys (29.4%). Of the 51 patients with DMD, the Dp71 group had the lowest cognitive performance with a total IQ score (46 ± 24.8; p = 0.003), while the Dp427 group and Dp140 group’s total IQ scores were 83.0 ± 24.6 and 84.2 ± 17.5 respectively. There were no DMD patients with ASD, while 4 boys (7.8%) had comorbidity with ADHD.

Conclusion

Boys with DMD are at higher risk of intellectual disability. The risk appears to increase with mutations at the 3’ end of the gene (Dp71 disruption). Moreover, Dp71 disruption might not be associated with ADHD and ASD in patients with DMD.

Introduction

Duchenne muscular dystrophy (DMD) is a severe neuromuscular disorder that affects skeletal muscles in 1:5,000 births [1, 2]. DMD is an X-linked recessive progressive muscular disease due to mutations in the DMD gene, which causes the absence or damage of dystrophin [2, 3]. Dystrophin serves as a link between the internal cytoskeleton and extracellular matrix proteins, which stabilize muscle contractions. In patients with DMD, the absence of dystrophin results in muscle damage due to impaired membrane integration and cellular necrosis [3].

Dystrophin has an essential role in brain development and function. Furthermore, dystrophin-associated glycoprotein complexes are involved in ion channels and postsynaptic membrane receptors during synaptogenesis. Dystrophin protein (Dp) variants are named based on their weight in kilodaltons [4]. The intact dystrophin protein (Dp427) and other shorter isoforms (Dp260, Dp140, Dp116, Dp71) are expressed specifically in muscle, heart, and brain tissues [5].

Previous studies have shown that the distribution of dystrophin in the amygdala, hippocampus and cerebral cortex is associated with cognitive phenotypes [5]. The incidence of cognitive decline was higher in the deletions of exons 45–52, affecting the Dp140 and Dp71 isoforms, suggesting the importance of these isoforms in cognition and brain function [6, 7]. Mutations that cause changes in the dystrophin isoform Dp140 (exons 45–55) were found in 5 of 8 (62.5%) DMD patients with autism spectrum disorder (ASD), whereas mutations occurring between exons 31 and 62 had stereotypical symptoms of ASD [8].

The prevalence of ASD, attention deficit and hyperactivity disorder (ADHD), epilepsy and obsessive-compulsive disorder were higher in boys with dystrophinopathies, including DMD and Becker muscular dystrophy (BMD) compared to normal populations. Learning and behaviour problems in DMD can occur with or without cognitive impairment and can be detected early in development. Intellectual disability is common in children with DMD (30%), while in the general population, the prevalence is only 1% [6]. ADHD is the most common neurobehavioral comorbidity in DMD. As many as 30–50% of children with DMD also have ADHD [7], while the incidence of ASD in people with DMD is 4–37%. The average intelligence quotient (IQ) of people with DMD, which is 85, is lower than the population mean, while the IQ considered normal in the general population is 100±15 [8].

Materials and methods

Participants

This cross-sectional study was conducted on patients aged 5–18 years old who were diagnosed with DMD. Diagnoses were confirmed through a DMD gene mutation test from January, 2017 to June, 2022 at Dr. Sardjito General Hospital and Universitas Gadjah Mada Academic Hospital, Yogyakarta, Indonesia [9]. Multiplex ligation-dependent probe amplification (MLPA) was used to determine exon deletions or duplications in the DMD gene. If MLPA showed no deletions or duplications, point mutations were discovered using whole-exome sequencing (WES) at 3billion, Inc, Seoul, South Korea. Patients who are not able to perform cognitive and neurobehavior assessments were excluded.

Patients older than 12 years old and/or their parents or guardians (for patients <12 years old) signed a written informed consent form to be included in this study. The Institutional Review Board of the Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada/Dr. Sardjito General Hospital approved this study (KE/FK/0894/EC/2020 and KE/FK/0180/EC/2021).

Data collection and variables

Baseline characteristics of the patient were collected using questionnaires. Cognitive function was measured by IQ score using the Wechsler Intelligence Scale for Children (WISC). The Vineland Adaptive Behavior Scales (VABS) were used to evaluate standardized adaptive behaviour measurements. Further screening on ASD and ADHD was conducted using Autism Mental Status Exam (AMSE) and Abbreviated Conner’s Rating Scale (ACRS), respectively. Cognitive function, ASD and ADHD were assessed based on the Diagnostic and Statistical Manual of Mental Disorders-5th edition (DSM-5) criteria. We divided patients into three groups based on the mutations in the DMD gene: (1) Dp427 group (mutation before the 31st exon), (2) Dp140 group (mutation between the 31st to 62nd exons), and (3) Dp71 group (mutation after 63rd exon).

Statistical analysis

Frequency was presented as percentages. Normality of the data was checked using the Shapiro-Wilk test, with p-values >0.05 considered as normally distributed data. Data were presented as mean ± standard deviation (SD) for normally distributed data and median (minimum-maximum) for nonnormally distributed data. The differences between mutation groups were analyzed using Kruskal-Wallis tests and Fisher Exact tests for categorical data, while Independent T-test, one-way ANOVA and Kruskal-Wallis tests were used to analyze numeric data, with p-values <0.05 considered as statistically significant. Data entry and analysis were performed using SPSS v.25 (IBM Corp., Armonk, NY).

Results

Data were obtained from 62 patients diagnosed with DMD based on genetic mutation analysis at Dr. Sardjito General Hospital and Universitas Gadjah Mada Academic Hospital, Yogyakarta, Indonesia from January 2017 to June 2022. Eight patients were excluded due to no cognitive or neurobehavior data available, while 3 patients were excluded due to BMD phenotype. Out of 51 patients with DMD who underwent genetic testing, exon deletions were found in 36 (70.6%) patients, exon duplications were found in 11 (21.6%) patients, a point mutation was found in 4 (7.8%) patients, and intron mutation was found in 1 (1.9%) patient (Fig 1).

Fig 1. Distribution of types of mutation in patients with DMD.

Fig 1

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The patients were divided into three groups based on dystrophin isoform disruption: 10 patients were in the Dp427 group, 37 patients were in the Dp140 group, and only four patients were in the Dp71 group. The detail of the mutation identified in all subjects was presented in Table 1.

Table 1. Mutation identified.

Name Age Mutation
Dp427 (N = 10)
MFR 13 Deletion of exons 8–30
AAZ 15 Deletion of exons 3–17
KN 8 Deletion of exon 1
LR 13 Deletion of exons 5–7
MT 12 Deletion of exons 1–4
D 13 Deletion of exons 15
APS 8 Deletion of exons 2–15
KM 10 Deletion of exons 8–29
RP 8 Duplication of exons 10–17
Dp140 (N = 37)
NRR 9 Deletion of exon 45
ARP 8 Deletion of exons 46–50
PBT 13 Deletion of exons 7–43
RWN 9 Deletion of exons 46–50
AP 11 Deletion of exon 51
BWC 13 Deletion of exons 49–50
MHH 16 Deletion of exons 53–54
NPP 11 Deletion of exons 46–51
SY 8 Deletion of exons 46–50
AM 13 Deletion of exons 48–50
FS 17 Deletion of exons 60
MDZ 10 Deletion of exon 52
YSF 15 Deletion of exons 46–48
GAP 12 Deletion of exon 51
RGM 12 Deletion of exons 42–52
NSA 17 Deletion of exons 17–43
AR 13 Deletion of exons 3–44
DNM 12 Deletion of exons 51–54
MKH 7 Deletion of exons 53–55
ZRA 10 Deletion of exons 45–50
MA 10 Deletion of exon 52
AN 10 Deletion of exons 3–44
MNA 8 Deletion of exons 47–50
E 8 Deletion of exons 44–50
R 4 Deletion of exons 44–50
RA 9 Deletion of exon 44
MNH 11 Duplication of exons 2–44
AJ 8 Duplication of exons 44–51
MNA 12 Duplication of exons 44–48
IAP 13 Duplication of exons 51–62
MFH 14 Duplication of exon 40
RD 7 Duplication of exons 12–44
RA 17 Duplication of exons 2–44
EM 12 NM 004006.2:c.6614+2T>C
IA 9 NM_004006.3:c.8086del
AS 10 NM_004006.3:c.9224+5G>A
HMS 14 NM_004006.3:c.5363C>A
Dp71 (N = 4)
NF 10 Deletion of exons 56–74
RR 12 Duplication of exons 63
GF 9 Duplication of exons 53–63
MYS 7 Duplication of exons 53–63
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There was no significant correlation between dystrophin disruption groups and the patients’ characteristics (Table 2).

Table 2. Patients’ characteristics.

Characteristics Dystrophin disruption groups p value
Dp427 (N = 10) Dp140 (N = 37) Dp71 (N = 4)
Age 10.9 ± 2.6 11.1 ± 3.1 9.5 ± 2.08 0.572
Disease onset 6.7 ± 2.9 5.8 ± 2.3 5.8 ±4.3 0.639
Ambulatory 0.349
    Early ambulatory 4 (36%) 4 (10.3%) 0 (0%)
    Late ambulatory 0 (0%) 2 (5.1%) 1 (25%)
    Non-ambulatory 5 (45%) 24 (61.5%) 2 (50%)
Family history of disease 2 (20%) 10 (27%) 1 (25%) 0.923
Physical Examination
    Pseudohypertrophy 4 (36%) 19 (51.4%) 1 (35%) 0.420
    Gower sign 5 (45%) 18 (48.6%) 3 (75%) 0.759
Laboratories result
    CK (U/L) 4314 (1,077–17,238) 6,788 (444–16,550) 6,681 (3,335–7,875) 0.604
    CKMB (U/L) 145 (91–399) 288 (49–831) 222 (173–311) 0.384
    Positive biopsy 3 (27%) 11 (29.7%) 3 (75%) 0.365
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Note: significance set as p<0.05; CK, creatine kinase; CKMB, creatine kinase myocardial band.

The average IQ test results for all patients were 80.6 ± 22.0 for total IQ, 82.9 ± 23.6 for verbal abilities, and 75.8 ± 19.7 for performance abilities. The Dp71 group significantly had the lowest cognitive function with total IQ score of 46.0 ± 24.8 compared to the other two groups (p = 0.003) accompanied with scores for verbal abilities of 49.75 ± 26.73 (p = 0.009), and for performance abilities of 45.3 ± 24.8 (p = 0.002). There were no significant differences of adaptive function profiles in the three groups (Table 3).

Table 3. Association between cognitive profile and dystrophin disruption group.

Dystrophin disruption group p value
Dp427 Dp140 Dp71
(N = 10) (N = 37) (N = 4)
WISC
Total IQ Score 83.0 ± 24.6 84.2 ± 17.5 46 ± 24.8 0.003*
Verbal 85.7 ± 28.4 87.0 ± 17.5 49.8 ± 26.7 0.009*
Performance 82.6 ± 18.6 78.2 ± 15.3 45.3 ± 24.8 0.002*
VABS
ABC 47.5 ± 22.3 46.6 ± 15.4 40.67 ± 26.35 0.850
Communication 43 (33–84) 59 (20–91) 40 (19–67) 0.683
Daily Living 69.0 ± 20.6 42.6 ± 23.1 39.3 ± 34.4 0.063
Socialization 65.5 (45–86) 61 (20–99) 53 (19–87) 0.491
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Note: significance set as p<0.05; ABC, Adaptive Behavior Composite; WISC, Wechsler Intelligence Scale for Children; VABS, Vineland Adaptive Behavior Scales.

The incidence of intellectual disability differed significantly between the Dp71 group and the other two groups (p = 0.009). All patients within the Dp71 group had intellectual disabilities. No significant findings of ADHD nor any patients with ASD were found among the patients with DMD in our study (Table 4).

Table 4. Prevalence of intellectual disability, ADHD, and ASD based in dystrophin disruption groups.

Dystrophin disruption groups Total N = 51 p value
Dp427 (n,%) Dp140 (n,%) Dp71 (n,%)
Intellectual disability 3 (30%) 8 (21.6%) 4 (100%) 15 (29.4%) 0.009*
ADHD 1 (10%) 3 (8.1%) 0 (0) 4 (7.8%) 0.819
ASD 0 (0) 0 (0) 0 (0) 0 (0) -
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Note: significance set as p<0.05; ADHD, attention deficit disorder; ASD, autism spectrum disorder.

No significant association was found between IQ test results and prevalence of intellectual disability, ADHD, and ASD with mutation type in each dystrophin disruption group. However, there was a significant difference in adaptive function, especially in ABC and communication in the Dp427 group (Table 5).

Table 5. Association between cognitive profile and prevalence of intellectual disability, ADHD, and ASD with mutation type in each dystrophin disruption groups.

Dp427 (N = 10) Dp140 (N = 37) Dp71 (N = 4)
Deletion (N = 9) Duplication (N = 1) Point Mutation (N = 0) p value Deletion (N = 26) Duplication (N = 7) Point Mutation (N = 4) p value Deletion (N = 1) Duplication (N = 3) Point Mutation (N = 0) p value
WISC
Total IQ Score 81.1 ± 25.6 98.00 N/A 0.553 82.3 ± 17.7 86.3 ± 19.0 93.7 ± 13.6 0.547 37.0 49.0 ± 29.5 N/A 0.758
Verbal 83.4 ± 29.5 104.00 N/A 0.531 84.3 ± 17.1 91.8 ± 8.5 97.3 ± 22.0 0.425 35.0 54.7 ± 30.4 N/A 0.632
Performance 81.4 ± 19.5 92.00 N/A 0.624 75.7 ± 17.0 81.0 ± 5.2 90.0 ± 3.6 0.314 35.0 48.7 ± 29.2 N/A 0.724
VABS
ABC 36.6 ± 9.8 76.00 N/A 0.022* 45.5 ± 15.5 56.0 ± 2.8 53.5 ± 24.4 0.584 33.0 43.0 ± 26.9 N/A 0.778
Communication 43.4 ± 10.0 84.00 N/A 0.021* 52.5 ± 18.5 64.5 ± 14.9 57.5 ± 25.7 0.702 37.0 43.0 ± 24.0 N/A 0.849
Daily Living 58.2 ± 23.0 70.00 N/A 0.664 42.3 ± 23.2 48.5 ± 14.9 57.0 ± 38.3 0.623 20.0 42.3 ± 33.7 N/A 0.624
Socialization 61.8 ± 13.4 86.00 N/A 0.176 59.5 ± 21.3 61.0 ± 1.4 58.0 ±26.2 0.986 51.0 53.7 ± 34.0 N/A 0.952
Prevalence of intellectual disability, ADHD, and ASD
Intellectual disability 3 (33.3%) 0 0 1.000 5 (19.2%) 1 (14.3%) 1 (25%) 0.892 1 (100%) 3 (100%) 0 -
ADHD 1 (11.1%) 0 0 1.000 2 (7.7%) 1 (14.3%) 0 0.705 0 0 0 -
ASD 0 0 0 - 0 0 0 - 0 0 0 -
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Note: significance set as p<0.05; ABC, Adaptive Behavior Composite; WISC, Wechsler Intelligence Scale for Children; VABS, Vineland Adaptive Behavior Scales; ADHD, attention deficit disorder; ASD, autism spectrum disorder.

Overall, no significant association was found between the severity of symptoms with IQ test results and adaptive function profiles and the prevalence of intellectual disability, ADHD, and ASD in each dystrophin disruption group (Table 6)

Table 6. Association between cognitive profile and prevalence of the intellectual disorder, ADHD, and ASD with ambulatory status in each dystrophin disruption groups.

Dp427 (N = 10) Dp140 (N = 37) Dp71 (N = 4)
Ambulatory (N = 5) Non ambulatory (N = 5) P value Ambulatory (N = 13) Non ambulatory (N = 24) P value Ambulatory (N = 2) Non ambulatory (N = 2) P value
WISC
Total IQ Score 96.0 ± 32.2 72.6 ± 11.2 0.168 85.0 ± 17.7 83.7 ± 17.8 0.847 52.5 ± 21.9 39.5 ± 34.7 0.698
Verbal 104.3 ± 34.2 70.8 ± 10.8 0.075 91.6 ± 20.2 84.5 ± 15.9 0.339 56.0 ± 29.7 42.5 ± 33.2 0.730
Performance 87.0 ± 24.7 79.0 ± 14.2 0.558 85.8 ± 13.9 74.2 ± 14.9 0.065 49.5 ± 20.5 41.0 ± 36.8 0.802
VABS
ABC 53.5 ± 31.8 38.0 ± 10.8 0.387 56.1 ± 9.2 43.8 ± 18.7 0.121 52.5 ± 27.6 28.5 ± 12.4 0.384
Communication 63.5 ± 26.2 42.5 ± 11.4 0.180 61.1 ± 13.4 51.2 ± 21.4 0.285 52.0 ± 21.2 31.0 ± 17.0 0.388
Daily Living 76.0 ± 8.5 52.3 ± 21.7 0.228 59.4 ± 23.0 38.3 ± 24.5 0.080 50.5 ± 43.1 23.0 ± 5.7 0.466
Socialization 69.5 ± 23.3 64.0 ± 14.5 0.729 68.6 ± 9.4 54.6 ± 23.8 0.189 69.0 ± 25.5 37.0 ± 25.5 0.336
Prevalence of intellectual disability, ADHD, and ASD
Intellectual disability 1 (16.7%) 2 (40%) 1.000 2 (15.4%) 5 (20.8%) 1.000 2 (100%) 2 (100%) -
ADHD 0 1 (20%) 1.000 1 (7.7%) 2 (8.3%) 1.000 0 0 -
ASD 0 0 - 0 0 - 0 0 -
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Note: significance set as p<0.05; ABC, Adaptive Behavior Composite; WISC, Wechsler Intelligence Scale for Children; VABS, Vineland Adaptive Behavior Scales; ADHD, attention deficit disorder; ASD, autism spectrum disorder.

Discussion

Exon deletions were found in 36 out of 55 patients (70.6%), higher than the result from a previous study in Indonesia, in which DMD patients had 44.1% deletions detected by multiplex PCR [10]. This result was similar to a study conducted by Aartsma-Rus et al., which found ~68% deletion. We found 21.6% for duplications and 7.8% for point mutations, in which one patient (1.9%) had intron mutation. A previous study reported duplications and point mutations were ~11% and ~20% of patients, respectively [11].

Dystrophin loss is associated with anatomic and physiological adaptations that contribute to the cognitive deficits present in patients with DMD. Dystrophin is one of the proteins that build up the Dystrophin-Glycoprotein Complex (DGC). DGCs play a role in the organization of the gamma-aminobutyric acid (GABA) receptors on neurons and the aquaporin-4 protein complex (AQP4) on glial cells. Accordingly, defects in DGC processing and formation are associated with brain abnormalities across a wide spectrum, ranging from mild cognitive impairment to brain migration disorders, apart from muscle disorders [12]. There is evidence of impaired central nervous system (CNS) architecture, dendritic abnormalities, and neuronal loss in patients with DMD. Electroencephalogram (EEG) abnormalities, brain atrophy on computerized tomography (CT) scan and magnetic resonance imagery (MRI) in patients with DMD have also been reported previously [13, 14].

Assessment of patients’ intellectual ability was done by measuring cognitive domains using the WISC test and the adaptive scale using the VABS test. The average IQ score in the patients with DMD in this study was 80.6 ± 22.0. This is lower than the average IQ score in the general population, which was 100 ± 15. This result is in accordance with research results in a multicenter meta-analysis of patients with DMD in Europe, which previously reported the average IQ score of 81.3 ± 5.1 [5, 13]. This is also in line with study results from Asian populations in Japan and India [15, 16]. Studies in various populations show that the mean IQ score in patients with DMD is 1–1.5 SD below the normal mean [7, 17]. In the present study, verbal IQ was higher than performance in all three dystrophin isoform disruption groups. This finding is in contrast to a previous study which found verbal IQ was lower than performance IQ in patients with DMD. They suggested the impairment in verbal abilities was caused by weaknesses in sequential auditory and visual information processing that reduced both attention and memory [13].

In this research, intellectual disability was found in 15 children (29.4%). Previous studies reported the frequency of intellectual disability appeared in 18.6–27% of patients with DMD in the Caucasian race [6, 12]. Cognitive function in the Dp71 group was significantly lower (p<0.05), both in total IQ scores and verbal and performance abilities. The total IQ score in the Dp71 group corresponds to intellectual disability, which is <70. Ricotti et al. reported that there was an increased risk of cognitive impairment in boys with mutations at the 3’ end of the DMD gene. This DMD gene mutation, which is located downstream of exon 63, encodes an isoform of the dystrophin protein called Dp71, which is the most expressed dystrophin isoform in the brain [4, 18].

The adaptive function of patients with DMD in this study was measured by VABS. The mean of adaptive function in children with DMD was also low in this study, which was 46.2 ± 17.4. There was no significant correlation between dystrophin disruption groups and adaptive function. Nevertheless, patients with DMD were reported to have a significant delay in adaptive function profiles compared with unaffected family members. Adaptive function profiles in patients with DMD were 1 SD lower than controls, with the most significant differences observed in communication and motoric skills [17].

In this study, there were no patients with suspected ASD based on the results of screening using the Autism Mental Status Exam. This measuring instrument is used for children over five years, with a specificity of 90.5% and a sensitivity of 81.2% [19]. Other different diagnostic methods that are more sensible and more specific could have been used (i.e., ADI-ADOS-2). ASD is reported to occur in 4–37% of patients with DMD. However, the measurement tools for screening and diagnosis of ASD vary between reported studies. Simone et al. reported that 19% of patients with DMD met the ASD criteria using the Autism Diagnostic Interview-Revised tool [20]. In comparison, Ricotti et al. reported a figure of 21% using the 3D diagnostic interview, which is a high sensitivity test tool considered to be more specific and reliable by taking into account the possibility of overestimation [15].

Dystrophin has an essential role in brain development. Lack of dystrophin and its mutations disrupt brain isoforms and are more likely to be associated with neurodevelopmental disorders [18]. The exact mechanism between ASD and DMD remains unclear. The previous study found that mutations in the distal dystrophin gene, which is associated with the production of short isoforms such as Dp71, are more likely to be associated with ASD compared to mutations of upstream parts of the gene [21]. However, this contradicts the findings of a previous study, which found that upstream gene mutations are also associated with the severity of autistic disorders [15]. This might be due to the loss of shorter isoforms progressively affecting the dystrophin as a whole [20].

There were four patients (7.8%) diagnosed with ADHD based on the DSM-5 diagnostic criteria according to the screening result score using the Abbreviated Conner Rating Scale 13. Another instrument for diagnosing ADHD is available, for example, Nepsy II in its dedicated subparts. There was no significant relationship between the location of the DMD gene mutation with the incidence of ADHD in this study. In a cohort study, ADHD was confirmed in 32%, compared to ADHD in general populations, which was 3–7% [22]. Previously, one study reported a range between 12–50% [7]. Another study by Ricotti et al. reported that 44% of patients with DMD presented with attention deficit disorder [15]. ASD and ADHD were found to be frequently associated with mutations affecting Dp140 and dystrophin short isoforms, including Dp71, between exons 62 and 63. Mutations on the middle and 3’ end of the gene may have a higher risk of manifesting attention disorders in children, where 25% of DMD patients with ADHD have mutations of the upstream exon 44 [7]. Another theory proposed that steroid therapy may cause behavioral problems in children. Long-term use of corticosteroid therapy may result in memory and attention problems [23]. However, this result is still controversial [7].

Correlation between types of mutation (deletion, duplication, and point mutation) and severity of symptoms (ambulatory and non-ambulatory) with cognitive profiles and prevalence of intellectual disability and ADHD in each dystrophin disruption group were also analyzed. No significant difference was found in the IQ mean and the prevalence of intellectual disability and ADHD in each type of mutation. At the same time, there is a significant correlation between types of mutation and adaptive and communication functions in the Dp427 group. Previous studies also reported no significant difference in mean IQ in each type of mutation [24]. Furthermore, the significant difference in cognitive profiles, adaptive profiles, and the prevalence of intellectual disability and ADHD was not observed for the severity of symptoms in each group.

This study is a cross-sectional study conducted with a valid assessment to measure neurodevelopment in male children with DMD with a broad spectrum of mutations, using genetic testing diagnostic techniques according to international guidelines over five years. This is the first study ever reported in Indonesia which examined the neurobehavior profiles of patients with DMD concerning the genotypes of the DMD gene mutations. As a single-center study, limits exist in its ability to depict the condition of the entire Indonesian population.

Conclusions

Boys with DMD are at higher risk of intellectual disability. The risk appears to increase with mutations at the 3’ end of the gene (Dp71 disruption). However, dystrophin disruption might not be associated with ADHD and ASD in patients with DMD. Further multicenter studies with larger sample populations are needed to confirm our findings.

Supporting information

S1 Data

(XLSX)

Click here for additional data file. (15.5KB, xlsx)

Acknowledgments

We are sincerely grateful to our patients and patients’ families who were willing to participate in this research. We would also like to thank Dwi Susilawati, MA, S.Psi., Psychology, Melina Dian Kusumadewi, S.Psi., M.A., Psychology, and all the staff and nurses who helped in caring for the patients’ wellbeing.

Data Availability

All relevant data are within the manuscript and its Supporting Information files.

Funding Statement

This research is funded by Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Indonesia and 3 billion, Inc, South Korea. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

References

  • 1.Sheikh O, Yokota T. Advances in genetic characterization and genotype-phenotype correlation of Duchenne and Becker muscular dystrophy in the personalized medicine era. J Pers Med. 2020;10(3):111. doi: 10.3390/jpm10030111 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2.Nizamis K, Schutte W, Grutters JJ, Goseling J, Rijken NHM, Koopman BFJM. Evaluation of the cognitive-motor performance of adults with Duchenne muscular dystrophy in a hand-related task. PLoS ONE. 2020;15(1):e0228128. doi: 10.1371/journal.pone.0228128 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Mento G, Tarantino V, Bisiacchi PS. The neuropsychological profile of infantile Duchenne muscular dystrophy. Clin Neuropsychol. 2011;25(8):1359–77. doi: 10.1080/13854046.2011.617782 [DOI] [PubMed] [Google Scholar]
  • 4.Naidoo M, Anthony K. Dystrophin Dp71 and the neurophysiology of Duchenne muscular dystrophy. Mol Neurobiol. 2020;57:1748–1767. doi: 10.1007/s12035-019-01845-w [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Doorenweerd N. Combining genetics, neuropsychology and neuroimaging to improve understanding of brain involvement in Duchenne muscular dystrophy: a narrative review. Neuromuscul Disord. 2020;30(6):437–42. doi: 10.1016/j.nmd.2020.05.001 [DOI] [PubMed] [Google Scholar]
  • 6.Banihani R, Smile S, Yoon G, Dupuis A, Mosleh M, Snider A, et al. Cognitive and neurobehavioral profile in boys with Duchenne muscular dystrophy. J Child Neurol. 2015;30(11):1472–82. doi: 10.1177/0883073815570154 [DOI] [PubMed] [Google Scholar]
  • 7.Pane M, Lombardo ME, Alfieri P, D’Amico A, Bianco F, Vasco G, et al. Attention deficit hyperactivity disorder and cognitive function in Duchenne muscular dystrophy: phenotype-genotype correlation. J Pediatr. 2012;161(4):705–709.e1. doi: 10.1016/j.jpeds.2012.03.020 [DOI] [PubMed] [Google Scholar]
  • 8.Fujino H, Saito T, Matsumura T, Shibata S, Iwata Y, Fujimura H, et al. Autism spectrum disorders are prevalent among patients with dystrophinopathies. Neurol Sci. 2018;39(7):1279–82. doi: 10.1007/s10072-018-3341-2 [DOI] [PubMed] [Google Scholar]
  • 9.Iskandar K, Triono A, Herini ES. Profil kognitif dan neurobehavior berdasarkan genotipe mutasi isoform distrofin pada pasien Duchenne muscular dystrophy (DMD) di Yogyakarta. [Thesis] [Bahasa]. Yogyakarta [Indonesia]: Universitas Gadjah Mada; (2021). [Google Scholar]
  • 10.Iskandar K, Dwianingsih EK, Pratiwi L, Kalim AS, Mardhiah H, Putranti AH, et al. The analysis of DMD gene deletions by multiplex PCR in Indonesian DMD/BMD patients: the era of personalized medicine. BMC Res Notes. 2019;12(1):1–6. doi: 10.1186/s13104-019-4730-1 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Aartsma-Rus A, Ginjaar IB, Bushby K. The importance of genetic diagnosis for Duchenne muscular dystrophy. J Med Genet. 2016;53(3):145–151. doi: 10.1136/jmedgenet-2015-103387 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Waite A, Brown S, Blake D. The dystrophin-glycoprotein complex in brain development and disease. Trends Neurosci. 2012;35(8):487–96. doi: 10.1016/j.tins.2012.04.004 [DOI] [PubMed] [Google Scholar]
  • 13.Ueda Y, Suwazono S, Maedo S, Higuchi I. Profile of cognitive function in adults with Duchenne muscular dystrophy. Brain Dev. 2017;39(3):225–30. doi: 10.1016/j.braindev.2016.10.005 [DOI] [PubMed] [Google Scholar]
  • 14.Anderson JL, Head SI, Rae C, Morley JW. Brain function in Duchenne muscular dystrophy. Brain. 2002;125(1):4–13. doi: 10.1093/brain/awf012 [DOI] [PubMed] [Google Scholar]
  • 15.Ricotti V, Mandy WPL, Scoto M, Pane M, Deconinck N, Messina S, et al. Neurodevelopmental, emotional, and behavioural problems in Duchenne muscular dystrophy in relation to underlying dystrophin gene mutations. Dev Med Child Neurol. 2016;58(1):77–84. doi: 10.1111/dmcn.12922 [DOI] [PubMed] [Google Scholar]
  • 16.Perumal AR, Rajeswaran J, Nalini A. Neuropsychological profile of Duchenne muscular dystrophy. Appl Neuropsychol Child. 2015;4(1):49–57. doi: 10.1080/21622965.2013.802649 [DOI] [PubMed] [Google Scholar]
  • 17.Cyrulnik S, Fee R, Batchelder A, Kiefel J, Goldstein E, Hinton V. Cognitive and adaptive deficits in young children with Duchenne muscular dystrophy (DMD). JINS. 2008;14(5):853–61. doi: 10.1017/S135561770808106X [DOI] [PubMed] [Google Scholar]
  • 18.Ricotti V, Roberts RG, Muntoni F. Dystrophin and the brain. Dev Med Child Neurol. 2011;53(1):12–12. doi: 10.1111/j.1469-8749.2010.03836.x [DOI] [PubMed] [Google Scholar]
  • 19.Betz E, Hackman NM, Mayes S, Chin EM, Ivy JW, Tierney C. Validity of the Autism Mental Status Exam in developmental pediatrics and primary care settings. Glob Pediatr Health. 2019;6:1–6. doi: 10.1177/2333794X19847905 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Simone M, Margari L, Pompamea F, De Giacomo A, Gabellone A, Marzulli L, et al. Autism spectrum disorder and Duchenne muscular dystrophy: a clinical case on the potential role of the dystrophin in autism neurobiology. J Clin Med. 2021;10(19):4–11. doi: 10.3390/jcm10194370 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Hendriksen RGF, Hoogland G, Schipper S, Hendriksen JGM, Vles JSH, Aalbers MW. A possible role of dystrophin in neuronal excitability: a review of the current literature. Neurosci Biobehav Rev. 2015;51:255–62. doi: 10.1016/j.neubiorev.2015.01.023 [DOI] [PubMed] [Google Scholar]
  • 22.Polanczyk G, de Lima MS, Horta BL, Biederman J, Rohde LA. The worldwide prevalence of ADHD: a systematic review and metaregression analysis. Am J Psychiatry 2007;164:942–8. doi: 10.1176/ajp.2007.164.6.942 [DOI] [PubMed] [Google Scholar]
  • 23.Brown ES, Vera E, Frol AB, Woolston DJ, Johnson B. Effects of chronic prednisone therapy on mood and memory. J Affect Disord. 2007;99(1–3):279–83. doi: 10.1016/j.jad.2006.09.004 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Taylor PJ, Betts GA, Maroulis S, et al. Dystrophin gene mutation location and the risk of cognitive impairment in Duchenne muscular dystrophy. PLoS One. 2010;5(1):e8803. doi: 10.1371/journal.pone.0008803 [DOI] [PMC free article] [PubMed] [Google Scholar]

Decision Letter 0

Giulio Piluso

12 May 2022

PONE-D-22-04415Dp71 and intellectual disability in Indonesian patients with Duchenne muscular dystrophyPLOS ONE

Dear Dr. Iskandar,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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Additional Editor Comments (if provided):

Both reviewers, who are expert in the field, have raised some criticism that authors have to address. In my opinion, the manuscript would furtherly benefit by adding a table detailing all mutations identified in DMD patients included in the study. This table would help the reader in interpreting the impact of the variants on the different Dp isoforms.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: N/A

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

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Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder which causes progressive muscle weakness leading to loss of ambulation in the mid-adolescent years. Affected males generally present in the first few years of life with motor symptoms and enlarged calves. However, neurodevelopmental disorders are increasingly recognized features and can be the initial presenting symptoms. The Authors report the results of a cross-sectional study aiming to characterize cognitive and neurodevelopmental profiles in a cohort of 48 patients with molecular diagnosis of DMD, aged between 5 and 18 years. Cognitive function was measured by intelligence quotient testing using the Wechsler Intelligence Scale for Children, and adaptive function tests with Vineland Adaptive Behavior Scales. The Autism Mental Status Exam and Abbreviated Conner's Rating Scale were used to screen autism spectrum disorder (ASD) and attention deficit and hyperactivity disorder (ADHD), respectively.

They found that patients with DMD have in general an IQ score lower compared to their peers and present with intellectual disability in about 20% of cases. No case of ASD, and only two boys with ADHD are reported. When trying a genotype-phenotype correlation, they conclude that DMD patients with mutation affecting Dp71 isoform of dystrophin are at higher risk to develop intellectual disability, but not ASD or ADHD.

The study is not original, as the knowledge that mutations disrupting the dystrophin brain isoforms Dp140 and Dp71 are more frequently associated with lower IQ scores in DMD patients, is not new. However, some results, such as the absence of patients with ASD and the low rate of patients with ADHD are interesting as they differ from what has been reported so far.

Critical points

- In tables there is no mention of the number of patients included in each dystrophin disruption group (Dp427, Dp140 e Dp71);

- Furthermore, in the tables it should be better clarified the difference between which groups the significance is referred;

- It would also be interesting to subdivide the patients within the same group according to the type of mutation - deletions vs duplications - and evaluate, if any, differences in these subgroups.

- In my opinion, the absence of patients with ASD and the low rate of patients with ADHD should be underlined and widely discussed as they differ markedly from what has been reported so far.

Reviewer #2: The article is well done and the topic is very interesting.

Anyway, data presented, even of interesting, appear partially in line with what available in literature. This could be due to a too small cohort or due to the application of evaluation methods not as sensitive as needed to analyze diseases object of investigation.

As said by the Author themselves, infact, the "Autism mental status exam" used by them, has a sensitivity of 81.2%.

Therefore, it could have been used a different diagnostic methods, more sensible and specific than previous

(i.e ADI-ADOS-2).

Another diagnostic method could also be used for the diagnosis of attention deficit disorder (Nepsy II in its dedicated subparts).

Moreover, since the age of the sample is very large and Duchenne muscular dystrophy is a progressive pathology, it could also be useful to differentiate the cohort according to the level of severity of symptoms (for example ambulatory and non-ambulatory).

Concerning IQ score much lower in dp71 group, this data is partially confirmed in literature but, in my opinion, this group should be enlarge to provide a wider benchmark and compare it.

For these findings, the paper need major work with a further evaluation for publication

**********

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Reviewer #1: No

Reviewer #2: No

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PLoS One. 2022 Oct 31;17(10):e0276640. doi: 10.1371/journal.pone.0276640.r002

Author response to Decision Letter 0

12 Jul 2022

Journal Requirements:

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Thank you, we have provided details regarding participant consent in the Methods section and Ethics Statement field:

“Patients older than 12 years old and/or their parents or guardians (for patients <12 years old) signed a written informed consent form to be included in this study. The Institutional Review Board of the Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada/Dr. Sardjito General Hospital approved this study (KE/FK/0894/EC/2020 and KE/FK/0180/EC/2021).”

3. Thank you for stating the following in the Funding Section of your manuscript:

“This work was supported by a grant from the Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Indonesia and 3 billion, Inc, South Korea.”

Please note that funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form.

Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows:

"This research is funded by the Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Indonesia and 3 billion, Inc, South Korea. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.”

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Please include your amended statements within your cover letter; we will change the online submission form on your behalf.

We have included amended statements in the cover letter. Thank you for your kind help.

Additional Editor Comments (if provided):

Both reviewers, who are expert in the field, have raised some criticism that authors have to address. In my opinion, the manuscript would furtherly benefit by adding a table detailing all mutations identified in DMD patients included in the study. This Table would help the reader in interpreting the impact of the variants on the different Dp isoforms.

Thank you for your suggestion. We have added a table detailing all mutations identified in DMD patients included in the study (Table 1).

Reviewers' comments:

Comments to the Author

Reviewer #1:

Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder which causes progressive muscle weakness leading to loss of ambulation in the mid-adolescent years. Affected males generally present in the first few years of life with motor symptoms and enlarged calves. However, neurodevelopmental disorders are increasingly recognized features and can be the initial presenting symptoms. The Authors report the results of a cross-sectional study aiming to characterize cognitive and neurodevelopmental profiles in a cohort of 48 patients with molecular diagnosis of DMD, aged between 5 and 18 years. Cognitive function was measured by intelligence quotient testing using the Wechsler Intelligence Scale for Children, and adaptive function tests with Vineland Adaptive Behavior Scales. The Autism Mental Status Exam and Abbreviated Conner's Rating Scale were used to screen autism spectrum disorder (ASD) and attention deficit and hyperactivity disorder (ADHD), respectively.

They found that patients with DMD have in general an IQ score lower compared to their peers and present with intellectual disability in about 20% of cases. No case of ASD, and only two boys with ADHD are reported. When trying a genotype-phenotype correlation, they conclude that DMD patients with mutation affecting Dp71 isoform of dystrophin are at higher risk to develop intellectual disability, but not ASD or ADHD.

The study is not original, as the knowledge that mutations disrupting the dystrophin brain isoforms Dp140 and Dp71 are more frequently associated with lower IQ scores in DMD patients, is not new. However, some results, such as the absence of patients with ASD and the low rate of patients with ADHD are interesting as they differ from what has been reported so far.

Thank you very much for these encouraging comments from this reviewer. We have emphasized our findings from previous studies in the Discussion section, particularly the absence of patients with ASD and the low rate of patients with ADHD compared to the previous result.

Critical points

- In tables there is no mention of the number of patients included in each dystrophin disruption group (Dp427, Dp140 e Dp71);

Thank you for your suggestions. We have added the number of patients included in each dystrophin disruption group in table 2 and table 3.

- Furthermore, in the tables it should be better clarified the difference between which groups the significance is referred.

We have clarified the difference between which groups the significance is referred to. We have added, "The Dp71 group significantly had the lowest cognitive function with a total IQ score of 46 ± 24.83 compared to dp427 and dp140 group (p=0.005 and 0.001, consecutively) accompanied with scores for verbal abilities of 49.75 ± 26.73 (p=0.013 and 0.005, consecutively), and for performance abilities of 45.25 ± 24.80 (p=0.001 and 0.002, consecutively)” at result section.

- It would also be interesting to subdivide the patients within the same group according to the type of mutation - deletions vs duplications - and evaluate, if any, differences in these subgroups.

We have subdivided the patient within the same group according to the type of mutation (deletion, duplication, and point mutation) and evaluated the difference in cognitive profile, the prevalence of intellectual disability, ADHD, and ASD between the group (Table 5).

- In my opinion, the absence of patients with ASD and the low rate of patients with ADHD should be underlined and widely discussed as they differ markedly from what has been reported so far.

Thank you very much for your opinion. We have underlined and added discussion about the absence of patients with ASD, and the low rate of patients with ADHD: “Correlation between types of mutation (deletion, duplication, and point mutation) and severity of symptoms (ambulatory and non-ambulatory) with cognitive profiles and prevalence of intellectual disability and ADHD in each dystrophin disruption group were also analyzed. No significant difference was found in the IQ mean and the prevalence of intellectual disability and ADHD in each type of mutation. At the same time, there is a significant correlation between mutation types and adaptive and communication functions in the Dp427 group. Previous studies also reported no significant difference in mean IQ in each type of mutation [23]. Furthermore, the significant difference in cognitive profiles, adaptive profiles, and the prevalence of intellectual disability and ADHD was not observed for the severity of symptoms in each group.”

Reviewer #2:

The article is well done and the topic is very interesting.

Anyway, data presented, even of interesting, appear partially in line with what available in literature. This could be due to a too small cohort or due to the application of evaluation methods not as sensitive as needed to analyze diseases object of investigation.

As said by the Author themselves, infact, the "Autism mental status exam" used by them, has a sensitivity of 81.2%.

Therefore, it could have been used a different diagnostic methods, more sensible and specific than previous (i.e., ADI-ADOS-2).

Thank you very much for your kind comments. We have added “Other different diagnostic methods that are more sensible and more specific could have been used (i.e., ADI-ADOS-2)” in the Discussion section: “Other different diagnostic methods that are more sensible and more specific could have been used (i.e., ADI-ADOS-2).”

Another diagnostic method could also be used for the diagnosis of attention deficit disorder (Nepsy II in its dedicated subparts).

We have added “Another instrument to diagnose ADHD also available, for example Nepsy II in its dedicated subparts” in the Discussion section.

Moreover, since the age of the sample is very large and Duchenne muscular dystrophy is a progressive pathology, it could also be useful to differentiate the cohort according to the level of severity of symptoms (for example ambulatory and non-ambulatory).

We added a table differentiating the cohort according to the severity of symptoms. There is no difference in IQ, ASD, and ADHD according to the severity of symptoms (Table 6).

Concerning IQ score much lower in dp71 group, this data is partially confirmed in literature but, in my opinion, this group should be enlarge to provide a wider benchmark and compare it.

As suggested by the reviewer, we have already added more patients to this study. The new Table has been inserted in the text accordingly. Unfortunately, there were no additional patients with Dp71 mutation.

For these findings, the paper need major work with a further evaluation for publication

We have added additional DMD patients for further analysis and expanded the Discussion section.

Attachment

Submitted filename: Response to reviewers_Kristy_rev.edited.docx

Click here for additional data file. (19.8KB, docx)

Decision Letter 1

Giulio Piluso

18 Aug 2022

PONE-D-22-04415R1Dp71 and intellectual disability in Indonesian patients with Duchenne muscular dystrophyPLOS ONE

Dear Dr. Iskandar,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Oct 02 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Giulio Piluso, M.Sc.

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Additional Editor Comments (if provided):

Concerning table 1, I believe the identified mutations should be reported according to the HGVS nomenclature. It's useful indicate the interval of exons deleted or duplicated but authors should complete table also reporting the expected phenotype DMD/BMD. Similarly, the indication "point mutation" for some patients seems very incomplete. These changes and that suggested by reviewers are mandatory.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The present version of the manuscript is much improved compared to the previous one. The Authors adressed all my suggestions.

I still have a few comments:

- Table 1. In the deletion group Dp427, the deletion 3-7 (patient NRR, 15 years) is listed: this is reported among the Malhotra’s exceptions to the reading frame rule. In fact, it is associated to a BMD severe phenotype.

Similarly in the deletion group Dp140, the deletions 48-49 (patient MA, 8 years) and 45-49 (patient AS, 11 years) are listed. Both these deletions are in frame and usually result in a BMD phenotype. These three patients must therefore be eliminated from the study.

- Table 2. The Measure Unit for CK and CKMB should be indicated;

- Table 3. WISC group. Specify better that the significance of “p” is between the Dp71 group and the other two groups, as I understand or not;

- Table 4. Specify better the significance for the intellectual disability to which group it refers;

- pag.15, line 17. I would say “motor” skills rather than “motoric” skills.

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Reviewer #1: Yes: Luisa Politano

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PLoS One. 2022 Oct 31;17(10):e0276640. doi: 10.1371/journal.pone.0276640.r004

Author response to Decision Letter 1

3 Oct 2022

RESPONSES TO REVIEWER AND EDITOR:

The editor and reviewer comments are in italics in the material below, and our responses are regular.

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Our manuscript has not cited papers that have been retracted.

Additional Editor Comments (if provided):

Concerning table 1, I believe the identified mutations should be reported according to the HGVS nomenclature. It's useful indicate the interval of exons deleted or duplicated but authors should complete table also reporting the expected phenotype DMD/BMD. Similarly, the indication "point mutation" for some patients seems very incomplete. These changes and that suggested by reviewers are mandatory.

Thank you for your suggestions. We have already specified the mutation detail as for point mutation according to HGVS nomenclature. Moreover, we have excluded three patients with the expected BMD phenotype as per reviewer #1 suggestion, so all patients have DMD phenotype.

Reviewers' comments:

Review Comments to the Author

Reviewer #1: The present version of the manuscript is much improved compared to the previous one. The Authors adressed all my suggestions.

I still have a few comments:

- Table 1. In the deletion group Dp427, the deletion 3-7 (patient NRR, 15 years) is listed: this is reported among the Malhotra’s exceptions to the reading frame rule. In fact, it is associated to a BMD severe phenotype.

Similarly in the deletion group Dp140, the deletions 48-49 (patient MA, 8 years) and 45-49 (patient AS, 11 years) are listed. Both these deletions are in frame and usually result in a BMD phenotype. These three patients must therefore be eliminated from the study.

Thank you for your comments. We have already excluded the deletions 3-7 (patient NRR, 15 years) from the deletion group Dp427 also the deletions 48-49 (patient MA, 8 years) and 45-49 (patient AS, 11 years) from the deletion group Dp140.

- Table 2. The Measure Unit for CK and CKMB should be indicated;

Thank you for your suggestion. We have already added the measuring unit for CK (U/L) and CKMB (U/L) in Table 2.

- Table 3. WISC group. Specify better that the significance of “p” is between the Dp71 group and the other two groups, as I understand or not;

Thank you for your suggestions. We have already added the following sentences: “The Dp71 group significantly had the lowest cognitive function with total IQ score of 46.0 ± 24.8 compared to the other two groups (p=0.003) accompanied with scores for verbal abilities of 49.75 ± 26.73 (p=0.009), and for performance abilities of 45.3 ± 24.8 (p=0.002).”

- Table 4. Specify better the significance for the intellectual disability to which group it refers;

Thank you for your suggestions. We have already added the following sentences: “The incidence of intellectual disability differed significantly between the Dp71 group and the other two groups (p=0.009). All patients within the Dp71 group had intellectual disabilities.”

- pag.15, line 17. I would say “motor” skills rather than “motoric” skills.

Thank you for your suggestion. We have already replaced “motoric” skills with “motor” skills.

Attachment

Submitted filename: Responses to reviewer and editor_rev2.docx

Click here for additional data file. (17.4KB, docx)

Decision Letter 2

Giulio Piluso

11 Oct 2022

Dp71 and intellectual disability in Indonesian patients with Duchenne muscular dystrophy

PONE-D-22-04415R2

Dear Dr. Iskandar,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Giulio Piluso, M.Sc.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

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4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

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PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

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6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The present revision of the manuscript clarified the points I underlined. All my comments have been accepted and the manuscript changed accordingly.

I have no further requests.

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Reviewer #1: Yes: Luisa Politano

**********

Acceptance letter

Giulio Piluso

21 Oct 2022

PONE-D-22-04415R2

Dp71 and intellectual disability in Indonesian patients with Duchenne muscular dystrophy

Dear Dr. Iskandar:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

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Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Professor Giulio Piluso

Academic Editor

PLOS ONE

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    Attachment

    Submitted filename: Responses to reviewer and editor_rev2.docx

    Click here for additional data file. (17.4KB, docx)

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