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Background: Erythropoietic protoporphyria (EPP) is a rare inherited disease associated with heme metabolism, characterized by severe life-long photosensitivity and liver involvement., Objective: To provide epidemiological data of EPP in Italy., Materials & Methods: Prospective/retrospective data of EPP patients were collected by an Italian network of porphyria specialist centres (Gruppo Italiano Porfiria, GrIP) over a 20-year period (1996-2017)., Results: In total, 179 patients (79 females) with a clinical and biochemical diagnosis of EPP were assessed, revealing a prevalence of 3.15 cases per million persons and an incidence of 0.13 cases per million persons/year. Incidence significantly increased after 2009 (due to the availability of alfa-melanotide, which effectively limits skin photosensitivity). Mean age at diagnosis was 28 years, with only 22 patients (12.2%) diagnosed ≤10 years old. Gene mutations were assessed in 173 (96.6%) patients; most (164; 91.3%) were FECH mutations on one allele in association with the hypomorphic variant, c.315-48C, on the other (classic EPP), and nine (5.2%) were ALAS2 mutations (X-linked EPP). Only one case of autosomal recessive EPP was observed. Of the 42 different FECH mutations, 15 are novel, three mutations collectively accounted for 45.9% (75/164) of the mutations (c.215dupT [27.2%], c.901_902delTG [11.5%] and c.67 + 5G > A [7.2%]), and frameshift mutations were prevalent (33.3%). A form of light protection was used by 109/179 (60.8%) patients, and 100 (56%) had at least one α-melanotide implant. Three cases of severe acute liver involvement, requiring OLT, were observed., Conclusion: These data define, for the first time, the clinical and molecular epidemiology of EPP in Italy.

Alkaptonuria (AKU) is an autosomal recessive disorder caused by mutations in homogentisate-1,2-dioxygenase (HGD) gene leading to the deficiency of HGD enzyme activity. The DevelopAKUre project is underway to test nitisinone as a specific treatment to counteract this derangement of the phenylalanine-tyrosine catabolic pathway. We analysed DNA of 40 AKU patients enrolled for SONIA1, the first study in DevelopAKUre, and of 59 other AKU patients sent to our laboratory for molecular diagnostics. We identified 12 novel DNA variants: one was identified in patients from Brazil (c.557T>A), Slovakia (c.500C>T) and France (c.440T>C), three in patients from India (c.469+6T>C, c.650-85A>G, c.158G>A), and six in patients from Italy (c.742A>G, c.614G>A, c.1057A>C, c.752G>A, c.119A>C, c.926G>T). Thus, the total number of potential AKU-causing variants found in 380 patients reported in the HGD mutation database is now 129. Using mCSM and DUET, computational approaches based on the protein 3D structure, the novel missense variants are predicted to affect the activity of the enzyme by three mechanisms: decrease of stability of individual protomers, disruption of protomer-protomer interactions or modification of residues in the region of the active site. We also present an overview of AKU in Italy, where so far about 60 AKU cases are known and DNA analysis has been reported for 34 of them. In this rather small group, 26 different HGD variants affecting function were described, indicating rather high heterogeneity. Twelve of these variants seem to be specific for Italy.

Mutations and a low-expressed allele IVS3-48c (in trans to the mutation) of the ferrochelatase (FECH) gene are responsible for erythropoietic protoporphyria (EPP) which is characterized clinically by cutaneous photosensitivity. In this study of 15 Italian EPP families, we identified 10 different FECH gene mutations, six of them were novel mutations (Q32X; IVS2-2, a-->g; IVS3-67, g-->a; 488-501del 14bp; IVS5-3, c-->t and 757-761delAGAAG). Four were known mutations (213insT; R115X; S264L and 899-900delTG). Real-time PCR quantification of FECH mRNA from a patient carrying mutation IVS-67, g-->a revealed a 51% reduction in the normal FECH transcript. This mutation generated an abnormal mRNA species with a 63-bp intron 3 retention--the same mRNA species which the low-expressed FECH allele IVS3-48c also produces. In addition, a double intronic deletion (IVS8-61 del ag /IVS9-58 del a) was found in some patients as well as their healthy relatives of this cohort. Sequencing of 50 Italian control subjects revealed an 11% prevalence of the double deletion. However, the double intronic deletion is not a disease-causing mutation as evidenced by a normal amount of FECH mRNA measured from a non-porphyric individual who carried the double deletion. Another interesting finding of this study was a 1% prevalence of IVS3-48c allele in the white Italian population, comparing to 11% in the French population. Further epidemiological studies are needed to test whether the 1% frequency of the low-expressed IVS3-48c allele is associated with a low prevalence of EPP in the white Italian population.