Clinical and genetic characteristics and prenatal diagnosis of patients presented GDD/ID with rare monogenic causes

Global developmental delay/intellectual disability (GDD/ID), used to be named as mental retardation (MR) is one of the most common phenotypes in neurogenetic diseases. Global developmental delay (GDD), is characterized by a delay in achieving developmental milestones in at least two of the following domains: motor skills, speech and language, cognitive skills, and social and emotional skills [1]. After growing up, many patients with GDD would demonstrate intellectual disability (ID), which is characterized by an intelligence quotient below 70 and limitations on adaptability [1, 2]. The prevalence of GDD/ID in the world population is estimated to be 1–3% [3], and the average lifetime costs (direct and indirect) to support an individual with ID have reached $1 million [4, 5].

Various environmental and genetic factors can result in GDD/ID [6]. Genetic reasons, including aneuploidy, copy number variants and single gene variants, account for 30–50% of cases [7], and Down syndrome, MECP2-related Rett syndrome and fragile X syndrome are the most common forms of genetic GDD/ID [6]. The modes of inheritance of genes related to GDD/ID include autosomal recessive (AR), autosomal dominant (AD), X-linked (XL) and mitochondrial [8]. On the basis of inherited patterns, some scholars divided those genes into ARID (autosomal recessive intellectual disability), ADID (autosomal dominant intellectual disability) and XLID (X-linked intellectual disability) [9‐11]. With the improvement of next-generation sequencing (NGS) [12], monogenic genetic causes are being found more frequently in previously unexplained or idiopathic cases. To date, nearly 1334 causative genes and 1159 candidate genes have been identified as related to GDD/ID [13], and the number continues to grow.

With increasing number of patients have obtained the genetic diagnoses by NGS, it poses great challenges on subsequential genetic counseling for family reproduction plan because of our limited knowledge on most rare monogenic diseases. As effective and specific treatments for most monogenic diseases are still in development, and prenatal molecular diagnosis is an important method to prevent recurrence. Previous studies mainly focused on comparing the diagnostic yields of different NGS methods [14] on unidentified GDD/ID or expanding the phenotype and genotype spectrum of a genetic disorder or specific gene. However, the general characteristics of patients presented GDD/ID with rare monogenic causes have not been well studied. Besides, few studies concerned the genetic counseling and prenatal diagnosis.

In this article, we collected a cohort of patients with GDD/ID caused by rare single gene variants. It is the first time, to the best of our knowledge, to observed these patients from the perspective of a genetic counseling and prenatal diagnostic center. The first aim of the study was to describe the diagnostic courses and clinical and genetic spectrum of these patients. The second aim was to report prenatal molecular diagnostic results of affected families, intending to raise awareness on this area.

As a tertiary genetic counseling and prenatal diagnosis center, our center served 290 families with individuals suspected of rare monogenic diseases during the 4-year study, and 142 (nearly 50%) of those patients had GDD/ID. After excluding 13 patients for missing information, 18 patients for uncertain diagnosis and 3 patients who had pathogenic variants along with atypical manifestations that could not be explained by the variants, we included a total of 108 subjects at preliminary screening. Further reviewing the case history, we excluded 16 patients who reached developmental lime stones normally in early stage and suffered GDD secondary to developmental regression, 5 patients that experienced abundant epileptiform activity in the neonatal period and then showed developmental arrest or regression and 7 patients for early death. Finally, we considered a total of 81 subjects and their core family members (Additional file 1: Fig. S1).

In this article, we analyzed the diagnostic courses, clinical and genetic characteristics, and prenatal diagnosis of 81 individuals with GDD/ID of monogenic origin. It often took 0.5–4.5 years and 2–8 referrals to obtain a genetic diagnosis after disease onset, reflecting the difficulty of diagnosis. Many factors are associated with this difficulty, including genetic heterogeneity, phenotype and penetrance variability, and shared signs and symptoms. Despite the great variability, when treated as a group of diseases, some features are noteworthy. The empirical findings regarding onset age, severity and coexisting symptoms can be summarized as follows.

One of the distinguishing features is the early age of onset. In our study, all individuals presented developmental delay before 3 years of age, and 80% of them showed abnormal symptoms in the first year of life. Nearly 10% of patients had abnormal during the prenatal stage. This finding is in accordance with previous studies showing that in monogenic forms of ID, the time of onset ranges from the 12th week after conception to early childhood [8, 47]. It also implies that future efforts should be made using NGS in the prenatal stage to detect abnormal prenatal ultrasound findings available and affordable [48, 49].

The severity of GDD/ID ranged from mild to profound in our study, and about 80% of patients had severe to profound disability. This finding is consistent with previous reports that GDD/ID caused by genetic factors could be more severe than those resulting from environmental factors, as the latter are usually mild [50, 51]. Previous studies concluded that de novo variants in ADID genes are the major causes of severe ID, and ARID and XLID are rare in outbred European or Korean populations [41‐44], and ARID with homozygous variants is most prevalent in consanguineous populations [45]. In this outbred Chinese cohort, we found that ARID, ADID and XLID had similar rates of severe cases (81%, 80%, and 60%). In addition, ARID with compound heterozygous variants and ADID accounted for approximately 32% and 35.8% of severe cases. Therefore, our results suggested that ARID with compound heterozygous variants also plays an important role in patients with GDD/ID caused by monogenic origin in outbred population. The inconsistency could be partially explained by the difference in study design and population. Another possible explanation is that each person carries 100 to 200 heterozygous private variants that are potentially deleterious [52], and when asymptomatic and unrelated parents carry such a variant in the same ARID gene, their offspring have a 25% chance of illness.

Most patients (over 95%) had other symptoms besides GDD/ID. Approximately 58% of subjects encompassed epilepsy and 10% had ASD. This finding supports the theory that GDD/IDs share a common etiology with other cognitive and neurological disorders including ASD and seizures [50]. In addition, 43.2% (35/81) of patients manifested abnormalities in appearance, such as microcephalus (21%), macrocephalus (8.6%), facial anomalies (17.2%), short stature (10.4%), and changes in skin or hair (3.8%). The percentage of patients in our cohort who had involvement in other organs, including the heart, kidney, liver, bone and endocrine, was under 16%, which might be lower than the rate in GDD/ID caused by aneuploid.

Genetic heterogeneity was prominent among these patients. Patients with different variants in the same gene could have different manifestations, while patients could have similar phenotype even with different causal genes. For example, the clinical presentation of three patients (No. 88, 89, 90) who were diagnosed with Phelan-McDermid syndrome and carried different frameshift variants in SHANK3 were not exactly similar. Patient 88 and 89 presented profound speech delay and ASD, while facial dysmorphism was found only in patient 88 and epilepsy was found only in patient 89. Patient 90 showed moderate developmental delay and facial dysmorphism but did not have epilepsy or ASD. This finding is consistent with previous studies intended to discover the phenotype-genotype correlation of Phelan-McDermid syndrome [53, 54].

Although experts recommended that when a genetic disorder was highly suspected, the first tier genetic testing is to detect the specific gene directly. We observed that most of our patients were diagnosed with the help of NGS rather than specific gene detection. It suggested the strong power of NGS in diagnosing patients with GDD/ID. We also observed that the percentages of patients diagnosed by WES or panel were similar in this cohort, while WGS is currently not widely used in domestic clinical situations. The best NGS technique for screening patients with GDD/ID remained controversial. It was reported that the diagnostic yields of panel, WES, and WGS in unexplained GDD/ID were up to 11–32%, 40%, and 42% respectively [43, 44, 55‐57]. Previously opinions considered that targeted NGS has the priorities of deeper coverage depth and lower cost than WES, however, hardly any targeted ID-panel could cover a great number of ID genes and chase up to the speed of the discovering new candidate genes, as it is estimated that the number of ID genes is over 1000 and still uprising. Therefore, the choice of personalized diagnostic method relies on the clinicians’ experience and varies from person to person.

The second aim of our study was to analyze the prenatal diagnostic situation of these groups of patients. Among the 33 families experienced prenatal diagnosis, the recurrence rates are 26.7%, 15.4% and 20% for ARID, ADID, and XLID, respectively. For GDD/ID caused by variants in autosomal genes, the recurrence rate is determined by whether the origin of the variants is inherited from parents or occurring de novo. For XLID, the recurrence rate is related not only to the originality of the variants but also to the sex of the fetus, which should be taken into consideration in specific situations. The necessity for prenatal diagnosis of variants inherited from parents has reached consensus in ARID and recessive XLID.

However, with an estimated recurrence rate less than 1%, the question of whether it is necessary to perform prenatal diagnosis for de novo variants in ADID and XLID remains controversial. In this cohort, 2 families with ADID tested positive in prenatal diagnosis, suggesting the high possibility of parental mosaicism in these cases. However, the mosaicism was confirmed in only one (50%) AD family via Sanger sequencing with peripheral blood. Therefore, the recurrence rate of de novo variant in AD families reached 8.3% (1/12), much higher than 1%.

Multiple lines of evidence suggested that the occurrence of parental germline mosaicism is underestimated. Firstly, while previous studies suggested that both Sanger sequencing and exome sequencing have the ability to detect somatic mosaicism [44, 51]. Due to the limitation of sequencing depth and difficulty in specimen acquisition, the existence of mosaicism in asymptomatic parents, which results in an increasing recurrence rate, is usually undetectable at present. Studies using deep amplicon sequencing and digital PCR methods to detect multiple samples found that the proportion of parental mosaicism in some AD or XLID genes reached 5–20% [58‐60]. Secoundly, nearly half of the ADID and XLID genes in this study have related case reports on germline mosaicism (Additional file 7: S5). Therefore, whether confirmation of parental mosaicism or not, we recommend that prenatal diagnosis is also necessary for de novo variants in ADID and XLID situations. In conclusion, genetic counseling and prenatal diagnostic services are important for families with any ARID, ADID and XLID probands.

Another important observation is that at present, genetic counseling and prenatal diagnostic services are not timely for nearly 50% of families, who were first referred for genetic counseling only after conceiving again. This might result from the lack of awareness and limitation of resources in this field. With the improvement and availability of genetic testing technology, an increasing number of individuals will obtain accurate genetic diagnoses. Additionally, with the implementation of a universal two-child policy [61], the need for genetic counseling and prenatal diagnosis is bound to increase. Therefore, more attention should be paid to this area.

The strengths of our study could be summarized as following. Firstly, it is the first article, to report the general characteristic of a group of patients presented GDD/ID with rare monogenic causes. Secondly, our observations on diagnostic time and methods could be set as baseline data and compared with the that obtained 5–10 years later. Thirdly, via calculating the recurrence rate in a group of families affected by AR, AD and XL genes, our research revealed that the recurrence rate of de novo variant might be underestimated in real world practice. Finally, it is the first retrospective cohort study observed from the perspective of genetic counseling and prenatal diagnostic center, and our result reflected the lack of awareness and shortage of resources in this field.

There are several limitations in our study. One is that rare monogenic neurogenetic diseases with GDD/ID consist of a group of different disorders, and while analyzing it in a cohort, we failed to perform genotype–phenotype correlation of a single syndrome or gene. However, to delineate the relationship in detail, a group of individuals with variants in the same gene or diagnosed with the same syndrome are needed. This kind of study is restricted by sporadic cases. Additonally, our understanding of these rare monogenic diseases is insufficient, and regular follow-up observations of such patients might provide additional clinical information. Futhermore, as it is a single center study, the results would be influenced by selection bias.

In summary, individuals with GDD/ID caused by rare monogenic variants are characterized by early onset, relatively severe phenotype as well as great clinical variability and genetic heterogeneity. Patients or pedigrees with such features should be considered to undergo appropriate NGS as early as possible. The spectrum of causal genes and pathogenic variants has not yet been fully discovered. Therefore, clinicians, genetic counselors and genetic laboratories should collaborate tightly to address the problems of diagnosis posed by the bewildering clinical and genetic heterogeneity. Moreover, obtaining clinical and genetic diagnosis is not the final step; timely referral to genetic counseling and prenatal diagnostic laboratories are important for families that plan to have additional children.