Milestones in treatments for inborn errors of metabolism: Reflections on Where chemistry and medicine meet.

From Sir Archibald Garrod's initial description of the tetrad of albinism, alkaptonuria, cystinuria, and pentosuria to today, the field of medicine dedicated to inborn errors of metabolism has evolved from disease identification and mechanistic discovery to the development of therapies designed to subvert biochemical defects. In this review, we highlight major milestones in the treatment and diagnosis of inborn errors of metabolism, starting with dietary therapy for phenylketonuria in the 1950s and 1960s, and ending with current approaches in genetic manipulation.

[Clinical practice guidelines for albinism].

Albinism is an autosomal or X-linked recessive Mendelian trait in man, which mainly manifests as hypopigmentation and related lesions of eye, skin and hair. At least 18 genes have so far been identified as causative genes for albinism. The mutational spectrum is population-specific. Molecular genotyping of albinism is important for genetic and prenatal diagnosis, and is a prerequisite for the practice of precision medicine. Based on long-term study of albinism in Chinese population, a guideline for the clinical management of albinism is provided.

The color of skin: white diseases of the skin, nails, and mucosa.

White diseases are a heterogenous group characterized by hypopigmentation or depigmentation. Skin and eye color are determined by the number and size of melanosomes present. Melanin is produced by melanosomes in the melanocytes present within the epidermis of the skin, uvea, and retinal pigmented epithelium (RPE). Conditions altering the number of melanocytes or concentration of melanin result in a lack of pigmentation, appearing as "white diseases" ranging from the well-known albinism and vitiligo to more esoteric white hand syndrome and Degos disease.

Adenine base editing in mouse embryos and an adult mouse model of Duchenne muscular dystrophy.

Adenine base editors (ABEs) composed of an engineered adenine deaminase and the Streptococcus pyogenes Cas9 nickase enable adenine-to-guanine (A-to-G) single-nucleotide substitutions in a guide RNA (gRNA)-dependent manner. Here we demonstrate application of this technology in mouse embryos and adult mice. We also show that long gRNAs enable adenine editing at positions one or two bases upstream of the window that is accessible with standard single guide RNAs (sgRNAs). We introduced the Himalayan point mutation in the Tyr gene by microinjecting ABE mRNA and an extended gRNA into mouse embryos, obtaining Tyr mutant mice with an albino phenotype. Furthermore, we delivered the split ABE gene, using trans-splicing adeno-associated viral vectors, to muscle cells in a mouse model of Duchenne muscular dystrophy to correct a nonsense mutation in the Dmd gene, demonstrating the therapeutic potential of base editing in adult animals.

Hemophagocytic lymphohistiocytosis: pathogenesis and treatment.

Hemophagocytic lymphohistiocytosis (HLH) is not an independent disease but rather a life-threatening clinical syndrome that occurs in many underlying conditions and in all age groups. HLH is the consequence of a severe, uncontrolled hyperinflammatory reaction that in most cases is triggered by an infectious agent. Persistent stimulation of lymphocytes and histiocytes results in hypercytokinemia, leading to the characteristic symptoms of HLH. Genetic defects in familial HLH and in immunodeficiency syndromes associated with albinism affect the transport, processing, and function of cytotoxic granules in natural killer cells and cytotoxic T lymphocytes. This leads to defective killing of target cells and a failure to contract the immune response. The defects are increasingly found also in adolescents and adults. Acquired HLH occurs in autoinflammatory and autoimmune diseases (macrophage activation syndrome) and in patients with iatrogenic immunosuppression or with malignancies, but also in otherwise healthy persons with infections. Treatment of HLH aims at suppressing hypercytokinemia and eliminating the activated and infected cells. In genetic HLH, hematopoietic stem cell transplantation (HSCT) is needed for the correction of the immune defect. Treatment modalities include immunosuppressive, immunomodulatory, and cytostatic drugs; T-cell antibodies; and anticytokine agents. Using immunochemotherapy, familial HLH, which had been invariably fatal, has become a curable disease with more than 50% survivors. Reduced intensity conditioning for HSCT, which is associated with less transplantation-related mortality, will further improve cure rates.

Albinism in Europe.

Albinism is a rare genetic condition associated with a variable hypopigmentation phenotype, which can affect the pigmentation of only the eyes or both the eyes and the skin/hair, resulting in ocular (OA) or oculocutaneous albinism (OCA), respectively. At least four forms of OCA and one of OA are known, associated with TYR (OCA1), OCA2 (OCA2), TYRP1 (OCA3), SLC45A2 (OCA4) and GPR143 (OA1) loci, respectively. Additionally, the rarest syndromic forms of albinism, affecting the normal function of other organs, can be grouped in Hermansky-Pudlak syndrome (HPS1-9) and the Chediak-Higashi syndrome (CHS1). In summary, a total of 15 genes are currently associated with various types of albinism. However, new genes have been recently described, associated with autosomal recessive oculocutaneous albinism with highly similar phenotypes but diverse molecular origin, indicating that there are likely to be more than 15 genes whose mutations will be associated with albinism. In this review, we will describe the different types of albinism and comment on its prevalence in European countries. Some preclinical attempts for innovative therapeutic approaches of different types of albinism will be also discussed.

Albinism: classification, clinical characteristics, and recent findings.

PURPOSE: To describe the clinical characteristics and recent findings in the heterogeneous group of inherited disorders of melanin biosynthesis grouped as "albinism." METHODS: The current classification of albinism, and the cutaneous, ocular, and central nervous system characteristics are presented. Recent clinical findings are summarized. RESULTS: Albinism is now classified based on genes known to be responsible for albinism. Foveal hypoplasia is invariably present and individuals with albinism often have delayed visual development, reduced vision, nystagmus, a positive angle kappa, strabismus, iris transillumination, and absent or reduced melanin pigment in the fundi. A visual-evoked potential can document the excessive retinostriate decussation seen in albinism. Grating acuity can be used to document delayed visual development in preverbal children. Glasses are often needed to improve visual acuity and binocular alignment. CONCLUSIONS: Albinism is caused by several different genes. Heterogeneity in clinical phenotype indicates that expressivity is variable.

A novel approach to gene therapy of albino hair in histoculture with a retroviral streptomyces tyrosinase gene.

In order to induce melanin production in mammalian cells with pigment disorders such as albino hair, a recombinant retrovirus containing the mel locus of Streptomyces antibioticus was constructed. The S. antibioticus mel locus, which consists of the open reading frame (ORF)-438 and the tyrosinase gene, was specifically derived by polymerase chain reaction (PCR) from Streptomyces plasmid pIJ702. The ORF-438 is required for the transfer of copper to apotyrosinase, which is essential for tyrosinase enzymatic activity. The tyrosinase gene was inserted into the XhoI/BamHI cloning site of the pLXSN retroviral vector to obtain pLtyrSN. An internal ribosome entry site (IRES) suitable for mammalian cell expression was obtained from the pLXIN retroviral vector by PCR. The ORF-438 and IRES DNA fragments were inserted into the pLtyrSN vector to obtain the tyrosinase-expression retroviral vector pLmelSN. The expression vector was amplified in murine PT67 packaging cells, where the ORF-438 and tyrosinase genes were also co-expressed as determined by reverse transcription-PCR. In order to evaluate the vector's ability to restore pigment production in cells with a pigment disorder, albino-mouse skins were histocultured and then infected with the pLmelSN retrovirus. Six days after infection, melanin granules were observed in approximately 60% of albino-mouse hair follicles in the histocultured skin. These results demonstrated that the S. antibioticus mel operon could express an active tyrosinase and produce melanin in the albino-mouse hair follicles. This novel gene therapy approach, using a small and simple tyrosinase operon in a high-expression vector, has a potentially wide application for therapy of pigment disorders in hair follicles.

Gene correction by RNA-DNA oligonucleotides.

An oligonucleotide composed of a contiguous stretch of RNA and DNA residues has been developed to facilitate the correction of single-base mutations of episomal and chromosomal targets in mammalian cells. The design of the oligonucleotide exploited the highly recombinogenic RNA-DNA hybrids and featured hairpin capped ends avoiding destruction by cellular helicases or exonucleases. The RNA-DNA oligonucleotide (RDO) was designed to correct a point mutation in the tyrosinase gene and caused a permanent gene correction in mouse albino melanocytes, determined by clonal analysis at the level of genomic sequence, protein and phenotypic change. Recently, we demonstrated correction of the tyrosinase gene using the same RDO in vivo, as detected by dark pigmentation of several hairs and DOPA staining of hair follicles in the treated skin of albino mice. Such RDOs might hold a promise as a therapeutic method for the treatment of skin diseases. However, the frequency of gene correction varies among different cells, indicating that cellular activities, such as recombination and repair, may be important for gene conversion by RDOs. As this technology becomes more widely utilized in the scientific community, it will be important to understand the mechanism and to optimize the design of RDOs to improve their efficiency and general applicability.

Localized in vivo genotypic and phenotypic correction of the albino mutation in skin by RNA-DNA oligonucleotide.

We recently demonstrated that an RNA-DNA oligonucleotide corrected a point mutation in the mouse tyrosinase gene, resulting in permanent and inheritable restoration of tyrosinase enzymatic activity, melanin synthesis, and pigmentation changes in cultured melanocytes. In this study, we extended gene correction of melanocytes from tissue culture to live animals, using a chimeric oligonucleotide designed to correct a point mutation in the tyrosinase gene. Both topical application and intradermal injection of this oligonucleotide to albino BALB/c mouse skin resulted in dark pigmentation of several hairs in a localized area. The restored tyrosinase enzymatic activity was detected by dihydroxyphenylacetic acid (DOPA) staining of hair follicles in the treated skin. Tyrosinase gene correction was also confirmed by restriction fragment length polymorphism analysis and DNA sequencing from skin that was positive for DOPA staining and melanin synthesis. Localized gene correction was maintained three months after the last application of the chimeric oligonucleotides. These results demonstrated correction of the tyrosinase gene point mutation by chimeric oligonucleotides in vivo.

[Spectral filters as a method of therapeutic correction]. / Spektral'nye fil'try kak vid lechebnoi korrektsii.

Spectral filters were used for additional correction of vision in 67 patients: in 15 adults with initial cataracts (intensive yellow filter), 26 children with albinism (yellow-brown filter), 14 children with macular hypoplasia (orange filter), and 12 children with aphakia after removal of congenital cataracts (yellow filter). Selection of the filter density is carried out using a special method including visocontrastometry, examination of sensitivity to lateral light, and study of color perception thresholds. Use of filters resulted in improvement of the vision acuity by 43.5% in patients with initial cataracts, by 10% in those with albinism, by 20% in those with macular hypoplasia, and by 22% in those with aphakia; moreover, an improvement of the frequency-contrast characteristics was observed, as well as a reduction of photophobia, and a reduction of vision amplitude in patients with nystagmus. The possible applications of spectral correction are discussed.

Partial albinism with immunodeficiency (Griscelli syndrome).

Partial albinism with immunodeficiency is a rare and fatal immunologic disorder characterized by pigmentary dilution and variable cellular immunodeficiency. To define the phenotype, therapy, and outcome, we retrospectively analyzed seven consecutive patients. Primary abnormalities included a silvery-grayish sheen to the hair, large pigment agglomerations in hair shafts, and an abundance of mature melanosomes in melanocytes, with reduced pigmentation of adjacent keratinocytes. Clinical onset occurred between the ages of 4 months and 4 years and was characterized by accelerated phases (lymphohistiocytic infiltration of multiple organs, including the brain and the meninges), triggered by viral and bacterial infections. Characteristic laboratory features included pancytopenia, hypofibrinogenemia, hypertriglyceridemia, and hypoproteinemia. Consistent immunologic abnormalities were characterized by absent delayed-type cutaneous hypersensitivity and impaired natural killer cell function. Some patients had secondary hypogammaglobulinemia, impaired major histocompatibility complex-mediated cytotoxic effects, a decreased capacity of lymphocytes to trigger a mixed lymphocyte reaction, or various functional granulocytic abnormalities. The disease seems to be invariably lethal without bone marrow transplantation; the mean age at the time of death was 5 years. Bone marrow transplantation has been performed in three cases; two patients died in the immediate posttransplantation period of infectious complications, but one patient is cured after a follow-up of 5 years. We conclude that partial albinism with immunodeficiency (Griscelli syndrome) can be differentiated from Chédiak-Higashi syndrome by pathognomonic histologic features. One of the underlying immunologic defects may be a defective function of natural killer cells, predisposing the patient to virus-associated hemophagocytic syndrome or accelerated phases. The prognosis is very poor unless early bone marrow transplantation is carried out.

Uso de Clofazimina em pacientes com albinismo óculo-cutâneo portador de Hanseníase na forma Virchowiana / Use of clofazimina in patients with oculo-cutaneous albinism in Bearer's of Virchow's form o f Hansen disease

O presente trabalho mostra um caso de paciente com Albinismo óculo-cutâneo, tirosinase +, portador de Hanseníase na forma V., submetido à MDT. O paciente foi medicado por um período de 2 anos, mas näo se notou qualquer pigmentaçäo de pele e mucosas pela Clofazimina um dos componentes da multidrogaterapia (MDT). Já que o Albinismo é uma doença congênita, devido à deficiência na produçäo de Melanina, estaria o metabolismo da Clofazimina relacionado ao da Melaniana, explicando a falta de pigmentaçäo neste paciente?