Case Report: Novel pathogenic variant in NFIX in two sisters with Malan syndrome due to germline mosaicism

Abstract

Malan syndrome is an autosomal dominant disorder caused by pathogenic variants in NFIX with less than 100 cases reported thus far. NFIX is important for stem cell proliferation, quiescence, and differentiation during development and its protein plays a role in replication, signal transduction, and transcription. As a result of pathogenic variants, symptoms of Malan syndrome include overgrowth, intellectual disability, speech delay, and dysmorphic features. Currently, the recurrence risk for this disorder is indicated at less than 1%, standard for de novo autosomal dominant disorders. Herein, we report an additional set of sisters with the same novel pathogenic variant in NFIX and clinical features consistent with Malan syndrome providing evidence of germline mosaicism. Considering the rarity of this condition in conjunction with three previous reports of germline mosaicism, it is worthwhile to investigate and re-evaluate the proper recurrence risk for this condition. This discovery would be paramount for family planning and genetic counseling practices in families with affected individuals.

Full text

Case Report: Novel pathogenic
variant in NFIX in two sisters with
Malan syndrome due to germline
mosaicism
Elizabeth Langley, Laura S. Farach and Kate Mowrey*
Department of Pediatrics, Division of Medical Genetics, McGovern Medical School at the University of
Texas Health Science Center at Houston (UTHealth Houston) and Children’s Memorial Hermann
Hospital, Houston, TX, United States
Malan syndrome is an autosomal dominant disorder caused by pathogenic
variants in NFIX with less than 100 cases reported thus far. NFIX is important for
stem cell proliferation, quiescence, and differentiation during development and
its protein plays a role in replication, signal transduction, and transcription. As a
result of pathogenic variants, symptoms of Malan syndrome include
overgrowth, intellectual disability, speech delay, and dysmorphic features.
Currently, the recurrence risk for this disorder is indicated at less than 1%,
standard for de novo autosomal dominant disorders. Herein, we report an
additional set of sisters with the same novel pathogenic variant in NFIX and
clinical features consistent with Malan syndrome providing evidence of
germline mosaicism. Considering the rarity of this condition in conjunction
with three previous reports of germline mosaicism, it is worthwhile to
investigate and re-evaluate the proper recurrence risk for this condition. This
discovery would be paramount for family planning and genetic counseling
practices in families with affected individuals.
KEYWORDS
NFIX, malan syndrome, germline mosaicism, recurrence risk, genetic counseling,
intellectual disability, overgrowth
Introduction
Malan syndrome, also known as Sotos syndrome 2, is an autosomal dominant
overgrowth disorder characterized by mild to moderate overgrowth, delayed speech,
intellectual disability (ID), and dysmorphic features (Priolo et al., 2018). Malan syndrome
is caused by pathogenic variants in nuclear factor I X (NFIX) and is located at
19p13.2 [OMIM: 164005] (Malan et al., 2010). Malan syndrome was first described as
Sotos-like syndrome, as its presentation is similar to Sotos syndrome that presents with
overgrowth, difficulties with learning, cardiac and renal anomalies, seizures, as well as
dysmorphic features. However, Sotos syndrome is caused by haploinsufficiency of the
NSD1 gene, rather than pathogenic variants in NFIX (Klaasens et al., 2015). NFIX is
important for stem cell proliferation, quiescence, and differentiation during development
and its proteins play a role in replication, signal transduction, and transcription (Malan
OPEN ACCESS
EDITED BY
Jennifer Melissa Kalish,
Children’s Hospital of Philadelphia,
United States
REVIEWED BY
Tri Indah Winarni,
Diponegoro University, Indonesia
Marcella Zollino,
Catholic University of the Sacred Heart,
Italy
*CORRESPONDENCE
Kate Mowrey,
Kate.Mowrey@uth.tmc.edu
SPECIALTY SECTION
This article was submitted to Human
and Medical Genomics,
a section of the journal
Frontiers in Genetics
RECEIVED 14 September 2022
ACCEPTED 31 October 2022
PUBLISHED 09 November 2022
CITATION
Langley E, Farach LS and Mowrey K
(2022), Case Report: Novel pathogenic
variant in NFIX in two sisters with Malan
syndrome due to germline mosaicism.
Front. Genet. 13:1044660.
doi: 10.3389/fgene.2022.1044660
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© 2022 Langley, Farach and Mowrey.
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Frontiers in Genetics frontiersin.org01
TYPE Case Report
PUBLISHED 09 November 2022
DOI 10.3389/fgene.2022.1044660

et al., 2010). Notably, Marshall-Smith syndrome is another rare
genetic condition that occurs only from frameshift and splice site
variants in exons six to eight of NFIX and presents with advanced
skeletal maturation, respiratory compromise, and failure to
thrive (Klaassens et al., 2015). Malan syndrome was named a
distinct entity from both Sotos syndrome and Marshall-Smith
syndrome due to its differing clinical presentation and genetic
etiology. Since first described in Malan et al., 2010, it is estimated
that less than 100 cases of Malan syndrome have been reported.
Although the majority of the prior reports of Malan
syndrome are the result of de novo pathogenic variants,
germline mosaicism was observed in eight individuals from a
total of three families (Nimmakayalu et al., 2013;Hancarova
et al., 2019;Sihombing et al., 2020). Since rare disorders are
underreported, the incidence of Malan syndrome as well as the
presence of germline mosaicism may be more prevalent than
previously estimated. In general, the recurrence risk for de novo
variants in most autosomal dominant genetic disorders are
estimated to be less than 1% (Rahbari et al., 2016). With the
increased availability of next-generation sequencing, it is
recognized that there are other factors that can influence the
accuracy of this recurrence risk, including the presence of
parental somatic mosaicism, mutation rate during
embryogenesis, and the gene of interest (Veltman and
Brunner, 2012;Samocha et al., 2014;Rahbari et al., 2016). For
genetic counseling, it is prudent to highlight the incidence of
germline mosaicism in families with Malan syndrome to educate
families more precisely on the recurrence risk of this disorder.
Here, we present a case of two full sisters with genetically
confirmed Malan syndrome due to a novel frameshift variant in
NFIX. Both sisters have phenotypes in line with previously
described cases and provide further evidence that the
incidence of germline mosaicism in Malan syndrome is higher
than the standardly quoted less than 1% for other autosomal
dominant genetic disorders.
Case presentation
Patient 1
Patient 1 is a 9-year-old Hispanic female born vaginally at
37 weeks gestation to non-consanguineous parents after an
uncomplicated pregnancy. After birth, she was in the neonatal
intensive care unit for a total of 11 days due to hyperventilation,
feeding issues, and jaundice. Motor and speech delays were noted
as early as 9 months. Specifically, she pulled to stand at 1 year,
cruised at 14 months, and walked at 18 months. At 4 years, she
put two words together, and at 9 years, she can speak in two-to-
three-word phrases. Due to her global developmental delay, she
receives occupational and speech therapy and is enrolled in
special education classes. At 4 years, neurology evaluations
occurred following a single episode of a febrile seizure. At that
time, anti-epileptic drugs were not recommended. At 6 years, she
had surgery for bilateral strabismus which was unsuccessful due
to optic nerve hypoplasia. A brain MRI at age 7 years
demonstrated decreased volume of the bilateral anterior optic
pathway. She was 8 years old at her first genetics evaluation and
at that time her dysmorphic features included macrocephaly,
upslanting palpebral fissures, almond shaped eyes, bitemporal
narrowing, flat feet bilaterally low-set, posteriorly rotated ears, a
straight border on the inside of her lower legs, and deep, hockey
stick hand creases bilaterally (Figures 1A–C). In addition to her
overgrowth, she had ID, acquired acanthosis nigricans, and
autistic-like features including difficulty communicating, nail
biting, and rocking. After evaluation by pediatric neurology,
she was diagnosed with autism spectrum disorder of major
severity due to persistent deficits in social communication and
interaction across multiple contexts, stereotypic behaviors,
hyperreactivity to sensory input, and clinically significant
impairment in social settings as outlined in the Diagnostic
and Statistical Manual 5 (DSM-5). Additionally, these findings
were accompanied by language impairment and not better
explained by her ID. At her follow up genetics evaluation, she
was noted to have an abnormal EEG with continuous generalized
slowing, consistent with mild encephalopathy. At this time, anti-
epileptics drugs are being considered. At 9 years, her weight is
62 kg (+4 SD) height is 151.6 cm (+2.8 SD) and her head
circumference is 58.5 cm (+4.9 SD).
Patient 2
Patient 2 is an 8-year-old Hispanic female born at term after
an uncomplicated pregnancy. She is the full sister of Patient 1.
She had delayed speech and did not put 2 words together until
5 years. Her motor milestones were developmentally
appropriate. She presents with ID and is enrolled in special
education classes. At 6 years, she was diagnosed with paralytic
strabismus and did not require surgical intervention. At 7 years,
a 23-h EEG was performed and exhibited right focal temporal
slow waves without evidence of epileptic or epileptogenic
events. Patient 2 was 6 years at her first genetics evaluation
and at that time, she could speak in two-to-three-word phrases
and received occupational and speech therapy twice a week. Her
dysmorphic features included epicanthal folds, hypertelorism,
retrognathia, small and low-set, posteriorly rotated ears, and a
thick upper lip (Figures 2A–C). At her follow up genetics
evaluation, she was noted to have an abnormal EEG due to
continuous generalized slowing, consistent with mild
encephalopathy, more prominent on the left than the right
suggestive of cortical dysfunction. Like Patient 1, anti-epileptic
drugs are being considered, but have not been started at this
time.At8years,herweightis30.8kg(+0.77SD),heightis
131.2 cm (+0.56 SD) and her head circumference is 56.8 cm
(+3.9 SD).
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Testing
Chromosome Microarray (CMA) was normal for both Patient
1 and Patient 2. Prader-Willi methylation analysis was found to be
normal in Patient 1. Patient 2 was identified as a premutation carrier
for Fragile X syndrome through Fragile X testing. Lastly, both sisters
along with their biological unaffected mother and unaffected father
underwent a Quad whole exome sequencing + mitochondrial
sequencing (Quad, WES + Mito) analysis. Patient 1 acted as the
proband and Patient 2 and their parents were used for segregation
analysis. The methodology of the Quad, WES + Mito includes next
generation sequencing with copy number variant calling. All variants
identified on the testing were confirmed via Sanger sequencing.
Results revealed that both Patient 1 and Patient 2 had the same novel
de novo heterozygous pathogenic variant located at
c.170_177dupCGAAGGAC (p.E60RfsX8l) [Clinvar ID: 1708174]
in NFIX [NM_001365902.3]. Specifically, parents were evaluated
forthisspecific genetic change via next generation sequencing and
were determined to not harbor this pathogenic variant, suggesting
that one parent has germline mosaicism. No further haplotyping or
additional genetic testing was utilized to elucidate which parent may
have germline mosaicism. In addition, a heterozygous known
pathogenic variant in PKP2 [NM_004572.3] located at c.235C>T
(p.R79X) [Clinvar ID: 6754] was identified in Patient 1, Patient 2, as
well as their mother via next generation sequencing leading to an
additional diagnosis of arrhythmogenic right ventricular
cardiomyopathy (ARVC). To our knowledge, ARVC has not been
found to be related to Malan syndrome in the literature. Due to this
additional diagnosis, both Patient 1 and Patient 2 were referred to
cardiology and have not had any cardiac issues to date.
Discussion
Here, we present a case of two full sisters with evidence of
germline mosaicism leading to Malan syndrome. Of the less than
100 cases presented in the literature, this set of siblings brings the
total number of cases of gonadal mosaicism in Malan syndrome to
four families and 10 individuals. Given the rarity of this condition
and the number of cases of germline mosaicism in Malan syndrome
thus far, the recurrence risk may be higher than the standardly
quoted less than 1% for this disorder in genetic counseling practices.
While recurrence risk is generally less than 1% for the majority
de novo genetic diseases, some disorders found to be de novo such as
Duchenne muscular dystrophy (DMD), Becker muscular dystrophy
FIGURE 1
Front (A), profile (B), and full (C) views of patient 1 at 9 years
demonstrate overgrowth, macrocephaly, upslanting palpebral
fissures, almond shaped eyes, low-set, posteriorly rotated ears,
dental crowding, and bitemporal narrowing.
FIGURE 2
Patients 1 and 2 at ages 9 and 8 years, respectively
demonstrate similar facial features (A). Profile (B) and full (C) views
of patient 2 at age 8 demonstrating almond-shaped eyes, small,
low-set, posteriorly rotated ears, retrognathia, dental
crowding, and bi-temporal narrowing.
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(BMD), osteogenesis imperfecta type II, Rett syndrome, and
Hemophilia A carry a higher recurrence risk of 4–20%, 5–7%,
11%, and 13%, respectively (Byers et al., 1988;Helderman-van den
Enden et al., 2009;Mari et al., 2005;Leuer et al., 2001). Generally,
germline mosaicism linked to dominantly inherited conditions is
rare and is likely due to genes undergoing variations that are not
repaired in germ cells (Byers et al., 1988). Like Malan syndrome,
osteogenesis imperfecta type II is an autosomal dominant disorder
where germline mosaicism has been observed. Byers et al., 1988
determined a recurrence risk of approximately 6% in families with
new dominant pathogenic variants due to gonadal mosaicism (Byers
et al., 1988). In another study, germline mosaicism was determined
by DNA analysis in blood and fibroblast DNA for 13 individuals,
while 2 parents were determined to be mosaic based on family
structure and having affected children with different partners (Pyott
et al., 2014). While the underlying mechanism of recurrence risk in
osteogenesis imperfecta type II is not fully understood, this
condition provides an example of increased risk for germline
mosaicism in an autosomal dominant disorder.
Furthermore, DMD/BMD is an X-linked disorder where
germline mosaicism has been well studied and better
understood. In Helderman-van den Enden et al., 2009,19cases
out of the 318 families identified with DMD/BMD with de novo
pathogenic variants were noted to have to have a second affected
male suggesting the presenceof germline mosaicism. This study set
out to determine the difference between true germline mosaicism
and somatic mosaicism. By utilizinghaplotyping, the authors were
able to determine if the de novo pathogenic variant was from the
maternal, grandmaternal, and/or grandpaternal X chromosome.
The results of the study indicated that in families with no
information about the risk haplotype, the recurrence risk was
approximately 4.3%. In the families with haplotype information
available, the recurrence risk varied from 14 to 20% depending on
the risk haplotype that is transmitted (Helderman-van den Enden
et al., 2009). This study identified that the recurrence risk for DMD
and BMD did not align with the less than 1% risk for de novo
conditions. As a result, genetic counseling practices have reflected
this by educating families with de novo cases about the likelihood
of having another affected child being higher at 4.3%. Given the
increased risk for siblings of Malan syndrome, targeted genetic
testing, whether prenatal or postnatally, should be offered. The
framework for genetic counseling in DMD and BMD cases may
provide a solid foundation to open discussions of how to
appropriately modify the recurrence risk for Malan syndrome,
so that families can be accurately informed during family planning.
Lastly, regarding clinical features of our patients, they
present similarly to other patients with Malan syndrome
while harboring a novel pathogenic variant. Although Malan
syndrome has variable expressivity, the literature describes core
features including global developmental delay, overgrowth,
dysmorphic features, strabismus, and ID (Sihombing et al.,
2020). This is consistent with the presentation of our two
patients.
As previously stated, the pathogenic variant found in both
Patient 1 and 2 is novel and not previously reported in the
literature to our knowledge. According to ClinVar, 30 pathogenic
variants in NFIX are associated with Malan syndrome in addition
to 11 variants noted to be likely pathogenic (Landrum et al.,
2018).
Overall, this sibling set adds another case to the medical
literature of germline mosaicism in Malan syndrome and
highlights an increasing number of cases of gonadal
mosaicism. It appears that germline mosaicism may be more
common in Malan syndrome than previously estimated based on
the inheritance pattern. This finding may indicate that a higher
recurrence risk should be cited for parents of children with Malan
syndrome. This topic is essential for genetic counseling and
family planning in families of affected individuals.
Data availability statement
The original contributions presented in the study are
included in the article/supplementary material, further
inquiries can be directed to the corresponding author.
Ethics statement
Ethical review and approval was not required for the study on
human participants in accordance with the local legislation and
institutional requirements. Written informed consent to
participate in this study was provided by the participants’legal
guardian/next of kin. Written informed consent was obtained
from the minor(s)’legalguardian/next of kin for the publication of
any potentially identifiable images or data included in this article.
Author contributions
EL drafted the manuscript. KM and LF both assisted in
project conceptualization and revised the manuscript. All
authors contributed and approved the submitted manuscript.
Acknowledgments
We thank the family for allowing us to publish this report.
Consent from all patients was obtained prior to this publication.
Conflict of interest
The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could
be construed as a potential conflict of interest.
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