Patients with primary carnitine deficiency treated with L‐carnitine are alive and doing well—A 10‐year follow‐up in the Faroe Islands

Abstract Primary carnitine deficiency (PCD) can be lethal. Carnitine is essential for the transfer of long‐chain fatty acids across the inner mitochondrial membrane for β‐oxidation. The reported prevalence of PCD in the Faroe Islands of 1:300 is the highest in the world. The Faroese PCD patient cohort has been closely monitored and we now report results from a 10‐year follow‐up study of 139 PCD patients. Four patients have died of natural causes since diagnosis. There were no signs of cardiac complications related to PCD. 70.5% reported an effect of L‐carnitine treatment. 33.7% reported current symptoms with fatigue and low stamina being the most common. 65.1% had experienced side effects during L‐carnitine treatment. Most common side effects were fish odor, abdominal pain, and diarrhea. The overall mean L‐carnitine dosage was 66.3 mg/kg/day. Free p‐carnitine was similar between male and female patients on L‐carnitine—18.6 and 18.8 μmol/L, respectively. L‐carnitine supplementation seems to be a safe and effective treatment when suffering from PCD. PCD patients in the Faroe Islands are alive and doing well more than 10 years after diagnosis.


| INTRODUCTION
Primary carnitine deficiency (PCD) can be lethal. 1The sudden death of a young Faroese female in 2009, who had just been diagnosed with PCD, but had not yet received supplementary L-carnitine treatment, sparked a public outcry for preventive measures.A voluntary screening program revealed a high prevalence of 1:300 in the Faroe Islands, a small island community in the North Atlantic with a population of only 54 000 inhabitants. 2 In a postmortem study among Faroese individuals under the age of 45, who had died suddenly, a total of 13 were found with genetic analysis to have suffered from PCD. 1 As a result, the Faroe Islands began to screen all newborns for PCD by measuring the carnitine level in their blood, 2-3 days after birth.Previous research has shown a direct correlation between the maternal and fetal carnitine levels. 3In order to accurately assess the infants carnitine level without any potential false elevation or reduction, a follow-up blood test is offered 4 weeks after birth.
Carnitine is a quaternary amine and is essential for the transfer of long-chain fatty acids across the inner mitochondrial membrane for β-oxidation, and the export of acetyl-and chain-shortened acyl products contributing to a stabile cell environment.][6] PCD is an autosomal recessive disorder that affects the function of the organic cation transporter 2 (OCTN2) high-affinity carnitine transporters, that localizes to the cell membrane and transport carnitine actively inside the cell.Without proper functioning OCTN2 carnitine transporters, the renal reabsorption of carnitine is impaired, and as a consequence, patients suffering from PCD have low plasma levels of carnitine.[9][10] Patients diagnosed with PCD in the Faroe Islands are treated with oral L-carnitine supplementation.The screening efforts, which culminated in 2010, revealed a cohort of Faroese adults, which had not been diagnosed with the disorder prior to the screening program and thus had not received L-carnitine supplementation.A study describing the health and cardiac status of 76 adult patients diagnosed with PCD in the Faroe Islands was published in 2014.It showed a marked improvement in self-reported fatigue following the initiation of L-carnitine treatment.But otherwise, most patients were asymptomatic. 11ore than 10 years have now passed since the largest part of the PCD cohort was diagnosed with the disorder.Children diagnosed then have now become adults.Patients have been rigorously followed in an outpatient setting and the aim of this article is to report an 10-year follow-up of the unique Faroese PCD patient cohort.

| MATERIALS AND METHODS
All registered PCD patients born prior to 2014 and living in the Faroe Islands at the time of the follow-up study were included.Furthermore, in patients who had emigrated from the Faroe Islands, we included data from their electronic health record, if they had been in contact with the Faroese health care system within 2 years of the study period from 2021 to 2022.

| Questionnaire
All patients living in the Faroe Islands, above 18 and below 75 years of age and not suffering from other serious sickness, were contacted by phone if possible and invited to answer the following questions by the principle investigator: Symptoms before diagnosis.If they had experienced improvements in their symptoms following treatment.Current symptoms.If they had experienced side effects from the L-carnitine treatment.Possible current side effects.Current dosage of L-carnitine supplementation.Height and weight.

| Blood tests
Hemoglobin, creatinine, Alanine Aminotransferase (ALAT) taken at least 5 years from the time of diagnosis.Mean plasma carnitine was calculated from the five most recently registered test results, though at least 5 years from diagnosis.

| Echocardiography and electrocardiogram
Results from follow-up echocardiography and electrocardiograms were only included if performed more than 5 years after diagnosis.

| RESULTS
We included 139 PCD patients in the study-74 males and 65 females (Table 1).The mean age was 37.8 years with most patients between 30 and 45 years of age (n = 33).Height, weight, and body mass index for those above and below the age of 18 and 75, respectively were unremarkable and as expected in the Faroese population (Table 1).The overall mean L-carnitine dosage was 66.3 mg/kg/day, with females tending to be on a higher dosage (68.7 mg/kg/day) than men (63.9 mg/kg/day; Table 1).
Free p-carnitine was similar between male and female patients on L-carnitine-18.6 and 18.8 μmol/L respectively.Hemoglobin, creatinine, and ALAT were normal (Table 1).
Follow-up echocardiography and electrocardiogram data were available in 100 patients with a mean followup time of 8.7 years (Table 1).All echocardiography parameters were reported normal in 91%, while there

Synopsis
Patients with primary carnitine deficiency treated with L-carnitine are alive and doing well more than 10 years after diagnosis.
were reported major and minor changes 2% and 7%, respectively.The major changes were signs of mild to moderate left ventricular hypertrophy and a mildly reduced left ventricular ejection fraction (LVEF) of 45% in two middle-aged male patients.Both had left bundle branch block.The minor changes ranged from mild valvular regurgitation to slight hypertrophy (Patient Table S1).Electrocardiograms were normal in 83% while there were signs of major and minor changes in 15% and 2% of the patients, respectively.Most major changes were atrial fibrillation and bundle branch block (Patient Table S1).
Four patients, two males and two females, have died since being diagnosed with PCD.They were 86, 81, 77, and 48 years of age and died of chronic obstructive pulmonary disease, ischemic cardiomyopathy and heart failure, lung cancer, and breast cancer.
Of the patients included in the questionnaire study (n = 83), 70.5% reported symptoms prior to treatment-and 70.5% reported an effect of L-carnitine treatment (Table 2).A third of the patients (33.7%) reported current symptoms with fatigue and low stamina being the most common-especially females reported current symptoms (51.2%) compared with 15% in males (Table 2).Two thirds of the patients (65.1%) had experienced side effects during L-carnitine treatment while a third (33.8%) was currently suffering from side effects (Table 2).Most common side effects were fish odor, abdominal pain, and diarrhea (Table 2).
When stratified in relation to genotypes most patients were compound heterozygous for the c.95A > G (p.Asn32Ser) and the c.-149G > A (n = 58), while 48 patients were homozygous for the c.95A > G mutation (Table 3).The c.95A > G homozygous group had a lower mean age than the other genotypes (27.6 years).And, the amount of daily L-carnitine ingested was higher among c.95A > G homozygous patients (86.7 mg/kg/day) compared with the other genotypes.Major changes (%) 2 Minor changes (%) 7 Electrocardiogram c n = 100 Normal (%) 83 Major changes (%) 15 Minor changes (%) 2 therapeutic reference was 15-25 μmol/L for all patients, and the free p-carnitine during L-carnitine treatment was kept in the similar range between the genotypes ranging from 18.0 to 18.8 μmol/L (Table 3).
There was no marked deviation between the genotypes regarding hemoglobin, creatinine, and ALAT (Table 3).
Almost 90% in the c.95A > G homozygous group reported a beneficial effect of L-carnitine treatment, T A B L E 3 Genotype follow-up characteristics, blood samples, mean (SD).which also was the genotype with lowest free carnitine prior to treatment (Table 3).Forty-eight patients had been admitted since being diagnosed (Patient Table S1).In the adult patients, the causes of admission to hospital do not seem to be related to PCD.In the children, it seems that they have been admitted quite often as a precautionary measure when experiencing signs of gastroenteritis and vomiting because their ability to adhere to the L-carnitine treatment was affected.

| DISCUSSION
The major finding in this study is, that almost all patients diagnosed with PCD more than 10 years ago and receiving L-carnitine treatment are still alive and doing well, with the oldest female and male patients aged 89 and 81 years old, respectively.The four patients who have died since diagnosis died of natural causes.At least 13 sudden deaths among young Faroese individuals have been attributed to untreated PCD. 1 PCD seems especially to have been a major cause of sudden unexplained death among young Faroese women, but the rigorous efforts to uncover undiagnosed PCD in the Faroe Islands might potentially prevent half of the sudden unexplained deaths in young Faroese females in the future. 1 PCD patients seem to adhere well to L-carnitine treatment, even though they have to ingest L-carnitine tablets at least three times a day.The treatment ensures far higher levels of L-carnitine in the blood compared with when not receiving L-carnitine supplementation.This again leads to higher intracellular carnitine and stabilizes and protects against cellular dysfunction for example erratic cardiomyocytes and malignant cardiac arrhythmia. 12ore than two-thirds of the patients (70.5%) reported having had symptoms prior to initiation of treatment, while only a third (33.7) reported still suffering from especially fatigue and low stamina when on L-carnitine.There can be little doubt, that L-carnitine supplementation, as well as protecting against serious symptoms, also alleviates subtler symptoms of especially fatigue.But, it comes at a cost, because almost twothirds (65.1%) of the patients have experienced side effects.Although abdominal pain and diarrhea are a nuisance the more common side effect of bodily fish odor experienced by patients on L-carnitine is potentially stigmatizing. 13Especially among the younger patients.Intestinal bacterial degradation of carnitine into trimethylamine (TMA), a nontoxic chemical with an unpleasant odor, is the cause of the unfortunate side effect.Advice given for reducing the odor includes frequent washing, use of antiperspirants, probiotic food supplements, and only rarely oral metronidazole. 13MA is absorbed through the intestinal mucosa into the bloodstream and converted to trimethylamine-N-oxide (TMAO) in the liver.14 This leads to elevated levels of TMAO in patients supplemented with L-carnitine.15 Increased levels of TMAO have been linked to an increased risk of atherosclerosis and cardiac disease.16,17 We have not observed that Faroese PCD patients on high-dose L-carnitine supplementation have an increased risk of ischemic events-which also seems to be supported by previous reports.15,18 We though need to investigate the Faroese PCD cohort more closely with regards to the association between elevated TMAO and atherosclerosis in order to rule it out.
There seems to be a gender difference in self-reported current symptoms, as half (51.2%) of the females still report symptoms of especially fatigue compared with only 15% of the males-and females report less side effects (Table 2).Of the 13 reported sudden deaths in the Faroe Islands 10 were female. 1 But, whether females really are at an increased physiological risk of adverse effects from PCD compared with males will be difficult to answer.We can at least see from our data, that the mean free p-carnitine in both male and female PCD patients treated with L-carnitine is almost identical (18.6 vs. 18.8 μmol/L).
Two males had mild left ventricular hypertrophy and mildly reduced LVEF.But both also had LBBB, which often causes a mild reduction in LVEF because of left ventricular dyssynchrony.They both suffered from hypertension and did not seem to benefit from L-carnitine with regards to the mild hypertrophy, which likely is not related to PCD.
The PCD genotypes differ with regards to residual OCTN2 transporter activity and carnitine levels when untreated. 8Among the Faroese patients those homozygous for the c.95A > G mutation are especially vulnerable, as their residual OCTN2 transporter activity and free carnitine levels without treatment are the lowest among the different genotypes in the Faroe Islands. 8Furthermore, the 13 sudden death subjects were all homozygous for the c.95A > G mutation. 1 Patients homozygous for the c.95A > G mutation receive more L-carnitine supplement than the other genotypes in order to maintain p-carnitine in the therapeutic range (Table 3).But, they also have the highest selfreported benefit of L-carnitine treatment (87.5%;Table 3).
Antibiotics containing pivalic acid have either been banned or are in strict limited use in the Faroe Islands since the discovery of a high prevalence of PCD.0][21] Several of the subjects who died suddenly of PCD had taken antibiotics containing pivalic acid shortly before their death. 9The far more cautious use antibiotics containing pivalic acid in the Faroe Islands has and will continue to limit the risk of hazardous exposure to PCD patients-but only in the Faroe Islands.PCD patients traveling abroad are informed to be vigilant when being prescribed antibiotics in order to avoid being unnecessarily exposed to pivalic acid.

| CONCLUSION
L-carnitine supplementation seems to be a safe and effective treatment when suffering from PCD. PCD patients from the large Faroese PCD cohort treated with L-carnitine are alive and doing well more than 10 years after diagnosis.A continued effort to diagnose PCD as early as possible in order to treat patients with L-carnitine must be considered highly relevant.
T A B L E 1 19