NEWBORN SCREENING OF RARE GENETIC DISEASES: VARIANT DISTRIBUTION IN CYSTIC FIBROSIS AND MEDIUM-CHAIN ACYL-CoA DEHYDROGENASE DECIFIENCY

Introduction: Newborn Screening (NBS) is one of the most advanced tools in precision medicine because it enables the early diagnosis of selected inherited disorders so that an effective treatment can be started before irreversible organ damage occurs. Italian legislation (Law 104/1992 and Law 167/2016) states that more than 40 diseases are mandatory on Italian newborns. Cystic Fibrosis (CF) is the most common autosomal recessive genetic disorder with a poor prognosis in populations of European descent. Medium-Chain Acyl-CoA Dehydrogenase Deficiency (MCADD) is the most frequent autosomal recessive fatty acid oxidation defect. For both conditions, early detection by NBS reduces morbidity and mortality. The aim of this thesis was to evaluate incidence and variant distribution of CF and MCADD in the Piedmont and Aosta Valley screening program over a 6-year period. Materials and methods: Between 2019 and 2024, a total of 163,997 newborns were screened. Capillary blood was collected on Dried Blood Spot cards (DBSs) between 48 and 72 hours after birth. CF screening involved a second-level genetic testing strategy (Second Tier Test, 2TT), so followed an IRT/DNA – IRT algorithm (IRT measured by DELFIA®, CFTR variants detected by targeted NGS). MCADD screening relied on tandem mass spectrometry (MS/MS) for the Acylcarnitines measurements. Results: For CF screening, of the screened newborns, 1,844 (1.12%) were investigated for 420 CFTR variants and 290 (0.18%) newborns were referred to Clinical Center; of these, 40 CF (63.5% of cases, incidence of 1:4,099) and 23 CFTR-Related Disorders (36.5% of cases, incidence of 1:7,130) affected individuals were found. In addition, 227 CFTR carriers were identified (incidence rate 1:722). The most frequent CFTR variant was the loss of phenylalanine-508 (F508del) and it was present in 18.6% of disease-associated alleles (9 out of 40 patients with classic-CF were homozygous for this variant), and in 16.5% of alleles in the carrier group. For MCADD screening, 11 newborns were confirmed as affected cases with an incidence of 1 in 12,436 live births. Molecular testing identified c.985A>G (p.Lys329Glu) as the most frequent pathogenic ACADM variant; it was observed in 4 patients (1 homozygous and 3 compound heterozygotes). Based on the genotype distribution among affected individuals, the allele frequency of c.985A>G was estimated at 23% (5 of 22 alleles) in the affected cohort. Conclusions: The regional NBS program effectively detected both CF and MCADD patients. The relatively high frequency of F508del among CF alleles is a common finding in several European published data and it is known that of the 2% of chromosomes which carry a CF mutation, 70% carry F508del, so this suggests that there is a carrier advantage at the individual, gamete or gene level. A strong founder effect has also been postulated to explain the high frequency of this variant but has been discarded or excluded. Instead, the high prevalence of p.Lys329Glu in Southern European populations has be explained by a common ancestral origin and a founder effect