Severe Antenatal Hypertrophic Cardiomyopathy Secondary to ACAD9-Related Mitochondrial Complex I Deficiency

Abstract

Introduction

Antenatal presentation of hypertrophic cardiomyopathy (HCM) is rare. We describe familial recurrence of antenatal HCM associated with intrauterine growth restriction and the diagnostic process undertaken.

Methods

Two pregnancies with antenatal HCM were followed up. Biological assessment including metabolic analyses, genetic analyses, and respiratory chain study was performed. We describe the clinical course of these two pregnancies, antenatal manifestations as well as specific histopathological findings, and review the literature.

Results

The assessment revealed a deficiency in complex I of the respiratory chain and two likely pathogenic variations in the ACAD9 gene.

Discussion and Conclusion

Antenatal HCM is rare and a diagnosis is not always made. In pregnancies presenting with cardiomyopathy and intrauterine growth restriction, ACAD9 deficiency should be considered as one of the potential underlying diagnoses, and ACAD9 molecular testing should be included among other prenatal investigations.

Introduction

Antenatal presentation of hypertrophic cardiomyopathy (HCM) is rare. This finding has been associated with various etiologies including environmental or obstetric factors, chromosomal anomalies, as well as syndromic or metabolic conditions (Table 1) [Clayton et al., 1992; Sonesson et al., 1992; Cincotta et al., 1996; Bonnet et al., 1998; Yunis et al., 1999; Narchi and Kulaylat, 2000; von Kleist-Retzow et al., 2003; Boldt et al., 2004; Schafer-Graf and Wockel, 2006; Lo et al., 2008; Malhotra et al., 2009; Hamdan et al., 2010; Sanchez Andres et al., 2011; Weber et al., 2014; Alston et al., 2015; Touraine et al., 2017; Mendez-Abad and Zafra-Rodriguez, 2018]. Nearly half of the antenatal HCM cases are described as idiopathic [Weber et al., 2014].

Table 1.

Known etiologies of antenatal hypertrophic cardiomyopathy

Etiology	Type	Reference
Maternal pathology	Gestational diabetes	Clayton et al., 1992; Sonesson et al., 1992; Cincotta et al., 1996
Iatrogenic	Infants exposed to prenatal corticosteroids or tacrolimus	Bonnet et al., 1998; Yunis et al., 1999
Twin pregnancies	When a twin-twin transfusion complication occurs	Cincotta et al., 1996; Narchi and Kulaylat, 2000
Inborn errors of metabolism	ATPase deficiency	Malhotra et al., 2009
	Pompe disease	Boldt et al., 2004
	Hurler syndrome	Malhotra et al., 2009
	Congenital disorder of glycosylation; type 1a and type 1L	Schafer-Graf and Wockel, 2006; Lo et al., 2008; Malhotra et al., 2009
Genetic syndromes	RASopathy (Noonan, Costello)	Malhotra et al., 2009; Hamdan et al., 2010; Sanchez Andres et al., 2011; Weber et al., 2014
	Trisomy 13	Malhotra et al., 2009
	Vici syndrome	von Kleist-Retzow et al., 2003
	Alpha thalassemia	Malhotra et al., 2009
Mitochondrial respiratory chain deficiency	Complex II deficiency due to SDHD mutation	Alston et al., 2015; Touraine et al., 2017; Mendez-Abad and Zafra-Rodriguez, 2018
Idiopathic		Malhotra et al., 2009

We depict herein familial recurrence of severe antenatal HCM beginning in the second trimester associated with intrauterine growth restriction (IUGR). Metabolic and molecular investigations uncovered respiratory chain complex I deficiency secondary to ACAD9 pathogenic variants. We describe the clinical course of these 2 pregnancies, antenatal manifestations, as well as specific histopathological findings and make a literature update.

Case Presentation

The 2 pregnancies were those of an unrelated healthy couple of Northern European descent. The mother was known for untreated junctional tachycardia, mostly asymptomatic.

The mother was 26 years old at the time of the couple's first pregnancy. First trimester course was unremarkable, without teratogenic exposure or maternal complications. Nuchal translucency was measured at 1.29 mm at 12+6 weeks' gestation. Second trimester fetal anatomical survey at 19+6 weeks' gestation commented on possible IUGR. Re-evaluation at 22+3 weeks' gestation reported all growth parameters just below the 5th centile, with heart appearing large for the thorax and outflow tracts not clearly seen. Fetal echocardiogram at 23 weeks' gestation identified prominent cardiac size, normal cardiac anatomy, moderate right ventricular hypertrophy, and borderline left ventricular hypertrophy. Follow-up fetal echocardiogram at 26 weeks' gestation reported signs compatible with an evolving cardiomyopathy (Fig. 1a, b) with moderately decreased biventricular systolic function.

Fig. 1.

Antenatal presentation of heart involvement in the two pregnancies. a, b First pregnancy. Four-chamber view at 23 (a) and 27 (b) weeks' gestation. Note the progression of the fetal cardiomegaly and degree of ventricular hypertrophy from the initial echocardiogram to the final follow-up. c–e Second pregnancy. Four-chamber view at 22 (c), 24 (d), and 27 (e) weeks' gestation with progressive biventricular hypertrophy and dilatation. RA, right atrium; RV, right ventricle; LA, left atrium; LV, left ventricle; LSVC, left superior caval vein to coronary sinus.

Investigations undertaken at that time included infectious studies (TORCH screen) and amniocentesis for the purpose of fetal chromosome studies. All were reported negative, with normal female karyotype.

Subsequent imaging at 28+2 weeks' gestation revealed fetal scalp skin edema, without other signs of developing hydrops or heart failure.

Delivery was induced at 32+4 weeks' gestation in the context of deteriorating fetal wellbeing. The infant girl was stillborn. Autopsy confirmed mild growth restriction, absence of hydrops, and severe cardiac hypertrophy (heart weight 18.3 g for an expected weight of 9.3 g ± 2.2 g for body weight, and 10.1 g ± 4.4 g for gestational age [GA]). Cardiac anatomy and histology were unremarkable. Pulmonary hypoplasia by weight (combined weight 10.9 g for an expected weight for body weight of 26.9 ± 14.6 g) was noted.

The remainder of the pathological findings were notable for occasional bilateral renal tubule interstitial microcalcifications of undetermined etiology, with focal tubular ectasia in deep cortex and medulla (Fig. 2a–c). Neuropathology examination showed a brain GA-appropriate in terms of weight and gyration. There were no malformations. On histologic examination, maturation was also GA-appropriate. The only abnormality was the presence of microvascular calcifications in the basal ganglia and thalami (Fig. 3e). Placenta evaluation was unremarkable.

Fig. 2.

Kidney pathology. a–c The first baby's kidney. a Low-power image showing cortex and medulla. This photo is representative of the pathological process showing full thickness fetal renal cortex with patchy tubular ectasia (more pronounced in deep cortex, marked by asterisks), with microcalcification (arrow). b Dilated tubules cortex (*) with calcifications (arrow). c Medullary focal ectasia. d, e Kidney histology of the second baby at low power (d) shows normal architecture with tubular microcysts occasionally surrounded by hyaline cylinders. Glomerular calcifications (arrows) are also well seen at low (d) and higher magnification (e).

Fig. 3.

Fetal cerebral anomalies in the two pregnancies. a−d, f, g Second pregnancy. Fetal MRI at 31 weeks' gestation shows cerebral gyration delay, polymicrogyria, and unilateral frontal paraventricular cyst (a–c). Neuropathology examination shows microcystic rearrangements (d) and microcalcifications and necrosis with reactive gliosis (f, g). e First pregnancy. Neuropathology examination of the baby shows the presence of microvascular calcifications in the basal ganglia and thalami.

The couple subsequently had a healthy term girl, as well as a first trimester miscarriage. During their fourth pregnancy (pedigree shown in Fig. 4), recurrence of significant growth restriction was noted at 20 weeks' gestation, with fetal growth below the 3rd percentile. No fetal anomalies were initially detected. Right ventricle hypertrophy was first noted at 23 weeks' gestation, with biventricular hypertrophy and hypokinesia of the right ventricle appearing at 25 weeks' gestation (Fig. 1c–e). Cardio-thoracic ratio was estimated at 50–55% (expected 30–35%). Bicuspid aortic valve and slight hypoplastic arch were also noted. Fetal MRI at 31 weeks' gestation showed cerebral gyration delay, possible polymicrogyria, and unilateral frontal paraventricular cyst (Fig. 3a–c).

Fig. 4.

Pedigree of the family showing familial recurrence of antenatal hypertrophic cardiomyopathy in two of their baby girls presenting with 2 variants in the ACAD9 gene (c.1204G>A, maternally inherited and c.1628T>G, paternally inherited). SB, stillbirth.

A non-hydropic female was delivered at 36+6 weeks' gestation, with the following growth parameters: weight 1,800 g (below 3rd percentile), length of 43 cm (3rd percentile), and head circumference of 31 cm (10th percentile). The baby passed away a few hours after birth.

Autopsy confirmed HCM without specific histology findings. Liver showed normal architecture, discrete microvacuolar steatosis, some periportal oxyphilic hepatocytes, and hepatocyte ferric accumulation. Kidney histology (Fig. 2d) showed normal architecture with tubular microcysts occasionally surrounded by hyaline cylinders and microcalcifications as well (Fig. 2e). The kidney cortex was discreetly thinned with enlarged sinuses. In the cerebral cortex, microcystic rearrangements with spongious lesions associated with microcalcifications and necrosis with reactive gliosis were noted both at the supra- and subtentorial levels (Fig. 3d, f, g). No other histologic lesions were noted (lungs, thymus, pancreas, spleen). Muscle biopsy was unremarkable, without fiber hypertrophy and normal distribution of type 1 and 2 fibers. COX activity was normal.

Additional Investigations

Analysis of a targeted panel of genes involved in RASopathies as well as metabolic studies were conducted for the first pregnancy. Given familial recurrence, exome sequencing was then performed on the firstborn's preserved DNA sample. Simultaneously, mitochondrial studies were initiated on the cardiac and liver tissues of the second baby.

Results

Postnatal investigations performed on the first child included comprehensive molecular studies for RASopathies, lysosomal enzymatic studies including Pompe disease, acylcarnitine profile, and chromosome microarray, all of which were reported as negative.

Exome sequencing was performed because of the familial recurrence of antenatal HCM and identified 2 variants in the ACAD9 gene (MIM *611103): NM_014049: c.1204G>A; p.(Gly402Arg) (exon 12; maternally inherited) and NM_014049: c.1628T>G; p.(Met543Arg) (exon 16; paternally inherited). These 2 variants were subsequently confirmed to be present in the second affected child. The p.(Gly402Arg) was classified as a likely pathogenic variant according to ACMG guidelines [Richards et al., 2015] with the following criteria: PM1 (mutational hotspot); PM2 (present in controls at extremely low frequency, MAF: 0.000007074), using strength Strong because the position is strongly conserved; PM5 since p.(Gly402Trp) has previously been reported as pathogenic [Fragaki et al., 2017]; and PP3 (in silico prediction of pathogenicity). The p.(Met543Arg) was classified as a likely pathogenic variant as well, with the following criteria: PM2 (absent from controls, using strength Strong because the position is strongly conserved; PM3 (in trans with a pathogenic variant); and PP3 (in silico prediction of pathogenicity).

In addition, mitochondrial studies on the first baby's fibroblasts confirmed a profound isolated complex I deficiency on Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) (Fig. 5), with significant decrease of complex I to complex II enzymatic activity ratio (Table 2) [Medja et al., 2009].

Fig. 5.

Mitochondrial studies on the first baby's fibroblasts. BN-PAGE showing profound isolated complex I deficiency.

Table 2.

Results of spectrophotometric analysis of mitochondrial respiratory chain in both patients

Patient	Cells tested	Total activity of NADH ubiquinone oxidoreductasea, nmol/min/mg	Complex IV/complex I ratioa	Complex I/complex II ratioa
Patient 1	Fibroblasts	Not tested	Not tested	0.11 (N: 0.5–1.1)
Patient 2	Heart muscle cells	8 (N: 30–50)	34 (N: 4–6)	0.09 (N: 0.7–1.1)
	Liver cells	6 (N: 26–40)	36 (N: 3–4)	0.03 (N: 0.16–0.24)

^aReference measures were established based on studies of 144 (muscle), 51 (liver), and 50 (fibroblast) control samples [Medja et al., 2009]. N, normal range.

Discussion

ACAD9 (MIM *611103) pathogenic variants lead to an autosomal recessive multisystem disorder usually characterized by infantile onset of acute metabolic acidosis, HCM, and muscle weakness, associated with mitochondrial complex I deficiency [Haack et al., 2010]. Atypical presentation with dystonia has also been reported [Haack et al., 2010]. Pathogenic variants in ACAD9 typically cause mitochondrial complex I deficiency as well as a mild defect in long chain fatty acid metabolism [Haack et al., 2010; Nouws et al., 2010, 2014]. The clinical phenotype of ACAD9 deficiency and associated mitochondrial complex I dysfunction reflects this dual role: reported symptoms are variable in severity and onset, and it has been suggested that the ACAD9-related fatty acid oxidation defect contributes to the severity of the phenotype in ACAD9-deficient patients [Schiff et al., 2015].

A recent review of 70 cases described to date reported that ACAD9 pathogenic variants result in multiple phenotypes with a large clinical spectrum and inconstant signs, from a severe form beginning at birth to a moderate phenotype with later onset [Aintablian et al., 2017; Repp et al., 2018]. Of those 70 cases, 22 patients presented features in the neonatal period (birth to 28 days), and only 9 of them were reported with prenatal manifestations. All remaining cases developed symptoms after the age of 1 month. Of the cases for which prenatal manifestations were sought after postnatal diagnosis, oligohydramnios and IUGR were the most commonly reported clinical signs, in 4 and 6 cases, respectively. Those manifestations were considered nonspecific by the authors [Aintablian et al., 2017; Repp et al., 2018]. Our 2 cases presented as well with IUGR detectable in the second trimester, which makes this ultrasonographic finding relatively frequent in the severe form of ACAD9 deficiency.

Prenatal Cardiac Findings

The most common reported manifestation of ACAD9-related complex I deficiency is postnatal cardiac dysfunction secondary to HCM. The 2 cases reported herein presented prenatal ACAD9 dysfunction, with cardiac findings detectable as early as at 23 weeks' gestation (Fig. 1). A single case of type I mitochondrial complex deficiency has been reported with antenatal biventricular non-compaction associated with congenital complete heart block [Dhar et al., 2015], but the underlying genetic etiology for this case was not determined. Three other cases with pathogenic variants in ACAD9 had fetal cardiac findings, one of them cardiomegaly [Lagoutte-Renosi et al., 2015] and the other two dysrythmias [Repp et al., 2018]. Interestingly, the case with prenatal cardiomegaly also showed growth retardation detected as early as at 22 weeks' gestation. The cardiomegaly was detected later at 33 weeks' gestation by ultrasound. The progression of the fetal cardiomegaly led to fatal outcome shortly after birth. The diagnosis of HCM was made in the postnatal period by heart ultrasound examination.

Of note, HCM as presented by our patients was never reported in association with ACAD9 deficiency in the antenatal period [Collet et al., 2016; El-Hattab and Scaglia, 2016].

Prenatal Cerebral Findings

There are few reports of cerebral findings associated with ACAD9 deficiency. Here, both children presented with cerebral findings either on autopsy or on prenatal imaging (Fig. 3).

One previous patient was described with lissencephaly and agenesis of the corpus callosum in association with IUGR [Repp et al., 2018].

Neuropathology findings were seen in the 2 affected infants from this report, however it is interesting to see very different neuropathological findings in the 2 babies. One brain showed only microvascular calcifications, which were probably of no functional significance, while the other showed extensive ischemic damage.

Apart from the typical findings of Leigh syndrome, there are very few reports of the specific histopathological cerebral findings associated with complex I deficiency [Ruhoy and Saneto, 2014]. Association between abnormal neuronal migration and/or intracerebral calcifications and complex I deficiency is anecdotal [Lake et al., 2015]. Additional reports are needed to establish the possible specificity of these lesions in relation to mitochondrial dysfunction.

Kidney Findings

Renal autopsy findings in the 2 infants presented in this report are unusual, with interstitial microcalcifications and focal tubular ectasia (Fig. 2). Again, these findings are not specific, and the rarity of reported histopathological findings makes a correlation with the underlying genetic etiology difficult. Leslie et al. [2016] reported excessive cytoplasmic eosinophilic granularity in proximal tubules of neonatal patients with ACAD9 deficiency which can be an indication of mitochondrial hyperplasia.

Tubulopathy (including Fanconi syndrome) and renal cysts are part of the spectrum of mitochondrial disorder manifestations [Lee et al., 2001]. The proximal tubulopathy is often moderate, and several investigators have reported isolated hyperaminoaciduria. Other patients may have acidosis, hypophosphatemic rickets, hypercalciuria, glycosuria, and tubular proteinuria. Renal biopsy shows nonspecific abnormalities of the tubular epithelium with dilatation and obstruction by casts, dedifferentiation, or atrophy. Giant mitochondria are often observed [Niaudet, 1998]. In a recent review which summarizes the renal manifestations of primary mitochondrial disorders, authors report that these manifestations can include renal insufficiency, nephrolithiasis, nephrotic syndrome, renal cysts, renal tubular acidosis, Bartter-like syndrome, Fanconi syndrome, focal segmental glomerulosclerosis, tubulointerstitial nephritis, nephrocalcinosis, and even benign or malignant neoplasms [Finsterer and Scorza, 2017].

Genotype-Phenotype Correlation

The 2 siblings of this case report presented with similar age of onset and severity of symptoms. Previously published familial cases also seem to indicate a certain correlation of degree and age of manifestation of symptoms among family members. Repp et al. [2018] reported 7 families with multiple affected sibs. In 6 out of 7 families, clinical presentation was very similar (symptoms and age of onset). In one family, the presenting symptom was HCM in both sibs but the age of onset differed dramatically: 2 months for the first affected child and 10 years for the second, who had received riboflavin supplementation as soon as the diagnosis of ACAD9 deficiency had been made in the neonatal period, which may explain the difference in age of presentation in this family. However, in some other families, riboflavin treatment did not influence the course of disease or outcome.

Despite this apparent concordance in clinical presentation between sibs, no obvious genotype-phenotype correlation could be established by Repp et al. [2018]. These authors also commented that no patients harboring biallelic loss-of-function mutations were documented in their cohort, indicating that this combination of alleles may be incompatible with life. Similarly, no correlation could be established between level of protein expression and degree of complex I activity. Nevertheless, it has been previously shown that residual ACAD9 enzyme activity, and not complex I activity, correlates with the severity of clinical symptoms in ACAD9-deficient patients [Schiff et al., 2015].

Treatment

Some ACAD9-deficient patients have undergone cardiac transplantation, but this treatment, while improving heart function, did not impact the other systems (e.g., muscular, neurologic, renal systems, etc.) that can be affected to various degrees in ACAD9 deficiency [Collet et al., 2016].

Some patients have also shown improvement of their cardiac function with riboflavin therapy. Recently, improvement in complex I activity with riboflavin supplementation was demonstrated in a majority of patient-derived fibroblast cultures [Scholte et al., 1995; Gerards et al., 2011; Repp et al., 2018]. While these results are encouraging, particularly for patients presenting with precocious forms of this condition, more cases are needed to evaluate the actual long-term benefit of this treatment.

Conclusion

Cardiomyopathy is one of the leading manifestations of a wide range of inherited disorders of energy metabolism, including mitochondrial disorders [Sidi et al., 1992; Kelly and Strauss, 1994; Zhuge et al., 2017]. While myocardial dysfunction often develops over a period in patients affected with these conditions, cardiomyopathy can also be the presenting sign, particularly in the antenatal period.

In pregnancies presenting with cardiomyopathy and IUGR in which other etiologies (infections, maternal illnesses) have been excluded, ACAD9 deficiency should be considered as one of the potential underlying diagnoses. Maternal riboflavin supplementation could be considered for pregnancies for which this diagnosis has been established.

Statement of Ethics

Toulouse University Hospital (TUH) has pledged to uphold France's CNIL MR-004 standards, which govern the use of data for medical research not directly involving human subjects. The TUH Data Protection Officer found this study compliant with the EU General Data Protection Regulation. It was included in both the TUH retrospective study registry (as RnIPH, 2021-102) and the CNIL MR-004 registry (as #2206723 v 0). Informed consent was obtained from our patients included in the study.

Conflict of Interest Statement

The authors have no conflicts of interest to declare.

Funding Sources

This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.

Author Contributions

C. Dubucs: conception of the work, data collection, drafting the article. N. Chassaing and I. De Bie: critical revision of the article, final approval of the version to be published. J. Aziza, A. Sartor, F. Heitz, A. Sevely, D. Sternberg, C. Jardel, T. Cavallé-Garrido, S. Albrecht, C. Bernard: revision of the article and the diagnosis procedure.

Data Availability Statement

Data sharing is not applicable to this article as no new data were created or analyzed in this study.

Funding Statement

This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.