3-Hydroxy-3-methylglutaryl-CoA lyase deficiency, also known as HMG-CoA lyase deficiency (HMGCLD; OMIM 246450), is a rare inherited autosomal-recessive metabolic disorder caused by disease-causing homozygous or compound heterozygous variants in the HMGCL. This gene is located in the short arm of chromosome 1p36.11 (Pié et al., 2007; Menao et al., 2009), which encodes the enzyme 3-hydroxymethyl-3-methylglutaryl-coenzyme A lyase (HMG-CoA lyase). This mitochondrial enzyme has a role in dietary protein metabolism, which forms acetyl-CoA and acetoacetate from HMG-CoA, known as the final step of leucine degradation and fat breakdown for energy (Puisac et al., 2010; Václavík et al., 2020) (Figure 1). Therefore, the body cannot synthesize ketone bodies, which are essential for energy production during the fasting period (Václavík et al., 2020). The prevalence of this disorder is less than 1/100,000 in live borns (Santarelli et al., 2013), while in Saudi Arabia, the incidence is 1: 55,357 (Alfadhel et al., 2017) which is approximately twice the world incidence.

HMGCLD clinically manifests early during the neonatal period (Santarelli et al., 2013) or later within the first year of life. Acute presentations include seizures, vomiting, metabolic acidosis, lethargy, and hypoketotic hypoglycemia (Pié et al., 2007; Menao et al., 2009). These symptoms sometimes progress to coma and death (Robinson, 2006).

HMGCLD can be diagnosed through newborn screening by measuring 3-hydroxy isovaleryl carnitine (C5-OH) via the tandem mass spectrometry methodology, which is usually elevated in this disease, and subsequently, the diagnosis is confirmed by the measurement of organic acids which showed a characteristic pattern of increased 3-methylglutaric, 3-hydroxy-3-methylglutaric, and 3-hydroxyisovaleric acids. In addition, urinary levels of 3-methyl-crotonyl glycine may also be increased (Alfadhel et al., 2017).

Also, it can be detected by low enzymatic activity testing using lymphocytes, fibroblasts, or immortalized lymphoblastoid cells. The molecular diagnosis is achieved if it shows homozygous or compound heterozygous pathogenic variants in the HMGCL gene using Sanger sequencing. Despite the high incidence in Saudi Arabia, a few studies are associated with natural history, epidemiological, clinical, and molecular data related to HMGCLD pathogenesis.

In the present study, we report the largest cohort of HMGCLD from Saudi Arabia. In addition, we provide clinical and molecular data associated with HMG-CoA lyase deficiency. Also, we provided the psychological evaluation of some patients to give an insight into long-term outcomes.

In this study, we retrospectively reviewed clinical, molecular, biochemical, and radiological findings in 62 patients with HGMCLD. The clinical presentation of HGMCLD is highly heterogeneous but it is usually evident in the neonatal and first year of life. Most affected individuals in our study became symptomatic within the first year after birth (Grunert et al., 2017; Václavík et al., 2020; Grünert and Sass, 2020). HMGCLD patients typically present features such as acute metabolic decompensation that may be life-threatening (Grünert and Sass, 2020).

The biochemical findings include acidosis, hypoglycemia, hyperammonemia, increased plasma/serum transaminase activities, and an increased anion gap. As ketogenesis is vital for maintaining glucose homeostasis, some patients might have hypoglycemia with or without metabolic acidosis during the initial episode that might be fatal due to the accumulation of organic acids. Similarly, ketone body accumulation reduces cerebral and peripheral glucose utilization and decreases muscular proteolysis (Evans et al., 2017). Therefore, for patients with HMGCLD who are unable to synthesize ketone bodies, their glucose utilization will be high, and gluconeogenesis will increase to uphold normoglycemia (Evans et al., 2017; Grunert et al., 2017). Hyperammonemia is also commonly reported; however, five (8.064%) patients showed hyperammonemia in the presented cohort, while all had abnormal urine organic acid profiles. Thus, the pathogenesis mechanism underlying hyperammonemia in HMGCLD patients is possibly multifactorial and still not completely understood.

HMGCLD diagnosis can be achieved using metabolite screening. Previous studies suggested that patients display a distinguishing biochemical profile with typical metabolites in the blood acylcarnitine and/or a urinary organic acid pattern (Václavík et al., 2020). Therefore, the first step in the diagnosis is the MS/MS-based acylcarnitine profiling in dried blood spots/plasma and the GC/MS-based organic acid analysis in urine. Then, molecular diagnosis using next-generation sequencing should be followed to pinpoint precisely the genetic cause of the disease (Ismail et al., 2019; Alfadhel et al., 2021). Similarly, targeted newborn screening should also be initiated to eradicate this severe disorder (Alfadhel et al., 2019). Similarly, Reimão et al. described late-onset diseases where a 29-year-old man with no prior medical history presented sudden coma, profound hypoglycemia, hyperammonemia, and metabolic acidosis without ketosis (Reimão et al., 2009). A 36-year-old woman with a late onset of an acute episode of hyperproteinorachia (0.73 g/L), hypoglycemia, and generalized seizures has also been reported (Bischof et al., 2004). Patients showing different levels of severity in the phenotypic presentation highlight clinical heterogeneity of the disorder. Still, it is difficult to establish a proper genotype–phenotype correlation in HMGCLD (Grunert et al., 2017; Alfadhel et al., 2017).

The mechanism of neurological pathophysiology in HGMCLD is still poorly understood; however, possible mechanisms include secondary carnitine deficiency, hypoketotic hypoglycemia, and intracellular toxic organic acid accumulation (Puisac et al., 2010). Furthermore, MRI shows a unique combination of T2-weighted diffuse mild signal changes with multiple foci of a more severe signal abnormality (Legault et al., 2015). Studies using rats revealed that the accumulation of the metabolite in HMGCLD results in oxidative stress in the developing rat’s striatum that disrupts bioenergetics dynamics, signaling pathways, and ER–mitochondria communication, which might explain the HMGCLD disease pathogenesis (da Rosa et al., 2015). Therefore, it might be concluded that these events cause the onset and progression of symptoms during decompensation in the neonatal period (da Rosa et al., 2020).

There have been notable highlights of some of the complications of the disease in recent years as HMCLD causes varying damages to the brain, liver, pancreas, and a very rare heart defect. Seizure is the most common complication in 27.14% (n: 17) of the presented patients. An MRI of the brain exhibited nonspecific periventricular and deep white matter hyperintense signal changes in 16 patients and cerebral atrophy was also found in one patient. Practically the same as the previously described marked lesions lie in periventricular white matter, arcuate fibers, and most prominently in the frontal or periarterial region, and slightly hyperintense subcortical white matter signals were also found (Yalçinkaya et al., 1999). Developmentally affected patients exhibit a delay in motor, speech, adaptive, and communication skills ranging from mild to moderate delay, found in 9.67% (n: 6) to profound intellectual disabilities found in one patient. However, in the present cohort, five patients had normal development despite the same MRI findings as previously described.

As HMGCL-deficient patients are prone to metabolic decompensation during illness, the presented patients demonstrated the same. Out of 37 patients reported previously with HMGCL variants, 10 had no metabolic decompensations after diagnosis, while 22 had at least one metabolic crisis, primarily associated with infections (Grunert et al., 2017). Similarly, one of the present patients had 10 metabolic decompensations, while several others had never had a hospital admission before. This signifies the heterogeneity of the disorder and the involvement of additional vital players defining the pathogenicity of the disease. In addition, about 15% of patients in the study reported by Grunert et al. (2017) died before the time of data collection. However, none of the patients reported in the present study died.

An 8-month-old infant with HMG-CoA lyase deficiency revealed recurrent pulmonary infections, left ventricular noncompaction, macrocephaly, nonketotic hypoglycemia, metabolic acidosis, and seizure (Köksal et al., 2015). However, in the present cohort, left ventricular noncompaction, macrocephaly, and recurrent pulmonary infections were not observed. Similarly, 10 novel HMGCL variants were reported in 24 patients of different origins suffering from 3-hydroxy-3-methyl-glutaric aciduria (Menao et al., 2009). The main complications in these patients were hepatic involvement with hepatomegaly and elevation of transaminases, with nine patients showing these symptoms during the neonatal period (Menao et al., 2009). However, in our cohort, 12 patients (19.35%) revealed hepatomegaly, and 27 patients (43.54%) were diagnosed during the neonatal period. Furthermore, Ozand et al. (1991) have reported overlapping clinical presentations, where 11 Saudi infants suffered from HMG-CoA lyase deficiency, with clinical presentation of acidotic attack including hyperpnea, lethargy, seizures, and tachypnea, at either birth or refusal of food in later infancy (Ozand et al., 1991).

Based on our data, no special diet or carnitine supplementation can be suggested to the patients. However, most of the patients were on a protein and/or leucine-restricted diet. Based on clinical reports and patho-biochemical considerations, the avoidance of fasting seems to be the mainstay of therapy for ketogenesis. In addition, administration of l-carnitine may have detoxifying effects and help to avoid secondary l-carnitine deficiency and intracellular depletion of free coenzyme A (Mitchell et al., 2008, Grünert and Sass, 2020).

The molecular study revealed that missense homozygous variants in the HMGCL gene 9c.122G>A; p.(Arg41Gln)] were identified in 48 patients (77.41%). Previously, the c.122G>A; p.(Arg41Gln) variant was reported as a founder variant and accounted for >94% of the pathogenic variants underlying HMGCLD in Saudi Arabia (Mitchell et al., 1998; Al-Sayed et al., 2006; Pie et al., 2007). Furthermore, we identified two previously reported variants (c.206_207del; p(Ser69Cysfs*11), c.914_915delTT; p(Phe305Tyrfs*10)) in additional seven affected individuals (Mitchell et al., 1998; Al-Sayed et al., 2006; Menao et al., 2009; Alfadhel et al., 2016).

To reach the diagnosis, some accumulated toxic metabolites should be tested in urine as the disease is considered one of the organic academic disorders. Several studies demonstrated that the patient excreted abnormal amounts of leucine catabolites such as 3-methyl glutaconic, 3-hydroxy-3-methyl glutaric, 3-methyl glutaric, 3-hydroxy isovaleric, and glutaric acids in the urine, which were found in all our patients, and we believe it is the leading cause of metabolic acidosis and the cause of an apparently secondary carnitine deficiency (Alfadhel et al., 2017).

4In conclusion, our data demonstrate that HMGCLD is a pan-ethnic disease, although some variants are clustered in some geographic regions with close connections throughout history. Our study also provides the most extensive genotype analysis of HMGCLD patients from Saudi Arabia. In addition, we conclude that HMGCLD is a heterogeneous disease with a variable clinical presentation and outcome; thus, early detection of the disease contributes to proper intervention, minimizing recurrent metabolic decompensation, and therefore, a normal cognitive outcome. HMGCLD patients suffer from avoidable neurologically devastating metabolic decompensations, and based on our results, we recommend newborn screening (NBS) of Saudi ethnicity for the HMGCL [p.(Arg41Gln)] variant.