Curing hemoglobinopathies : challenges and advances of conventional and new gene therapy approaches

Inherited hemoglobin disorders, including beta-thalassemia (BT) and sickle-cell disease (SCD), are the most common monogenic diseases worldwide, with a global carrier frequency of over 5%.1 With migration, they are becoming more common worldwide, making their management and care an increasing concern for health care systems. BT is characterized by an imbalance in the \u3b1/\u3b2-globin chain ratio, ineffective erythropoiesis, chronic hemolytic anemia, and compensatory hemopoietic expansion.1 Globally, there are over 25,000 births each year with transfusion-dependent thalassemia (TDT). The currently available treatment for TDT is lifelong transfusions and iron chelation therapy or allogenic bone marrow transplantation as a curative option. SCD affects 300 million people worldwide2 and severely impacts the quality of life of patients who experience unpredictable, recurrent acute and chronic severe pain, stroke, infections, pulmonary disease, kidney disease, retinopathy, and other complications. While survival has been dramatically extended, quality of life is markedly reduced by disease- and treatment-associated morbidity. The development of safe, tissue-specific and efficient vectors, and efficient gene-editing technologies has led to the development of several gene therapy trials for BT and SCD. However, the complexity of the approach presents its hurdles. Fundamental factors at play include the requirement for myeloablation on a patient with benign disease, the age of the patient, and the consequent bone marrow microenvironment. A successful path from proof-of-concept studies to commercialization must render gene therapy a sustainable and accessible approach for a large number of patients. Furthermore, the cost of these therapies is a considerable challenge for the health care system. While new promising therapeutic options are emerging,3,4 and many others are on the pipeline,5 gene therapy can potentially cure patients. We herein provide an overview of the most recent, likely potentially curative therapies for hemoglobinopathies and a summary of the challenges that these approaches entail

2'-O-methoxyethyl splice-switching oligos correct splicing from IVS2-745 β-thalassemia patient cells restoring HbA production and chain rebalance

\u3b2-thalassemia is a disorder caused by altered hemoglobin protein synthesis and affects individuals worldwide. Severe forms of the disease, left untreated, can result in death before the age of 3 years (1). The standard of care consists of chronic and costly palliative treatment by blood transfusion combined with iron chelation. This dual approach suppresses anemia and reduces iron-related toxicities in patients. Allogeneic bone marrow transplant is an option, but limited by the availability of a highly compatible HSC donor. While gene therapy is been explored in several trials, its use is highly limited to developed regions with centers of excellence and well-established healthcare systems (2). Hence, there remains a tremendous unmet medical need to develop alternative treatment strategies for \u3b2-thalassemia (3). Occurrence of aberrant splicing is one of the processes that affects \u3b2-globin synthesis in \u3b2-thalassemia. The (C>G) IVS-2-745 is a splicing mutation within intron 2 of the \u3b2-globin gene. It leads to an aberrantly spliced mRNA that incorporates an intron fragment. This results in an in-frame premature termination codon that inhibits \u3b2-globin production. Here, we propose the use of uniform 2'-O-methoxyethyl (2'-MOE) splice switching oligos (SSOs) to reverse this aberrant splicing in the pre-mRNA. With these lead SSOs we show aberrant to wild type splice switching. This switching leads to an increase of adult hemoglobin (HbA) up to 80% in erythroid cells from patients with the IVS-2-745 mutation. Furthermore, we demonstrate a restoration of the balance between \u3b2-like- and \u3b1-globin chains, and up to an 87% reduction in toxic \u3b1-heme aggregates. While examining the potential benefit of 2'-MOE-SSOs in a mixed sickle-thalassemic phenotypic setting, we found reduced HbS synthesis and sickle cell formation due to HbA induction. In summary, 2'-MOE-SSOs are a promising therapy for forms of \u3b2-thalassemia caused by mutations leading to aberrant splicing

Combined Inclusion of Former Foodstuff and Distiller Grains in Dairy Cows Ration: Effect on Milk Production, Rumen Environment, and Fiber Digestibility

Simple Summary One-third of the global food produced for human consumption is wasted every year. This leads to the wasting of economic, environmental, and social resources. The reallocation of some unconventional agro-industrial by-products, such as former foods or distiller grains, into the animal feed chain, can contribute to increasing the sustainability of livestock production, reducing the need for natural resources such as arable soil, water, fertilisers, and fuels, and consequentially reduce the impact of animal requirements. Many agro-food industrial by-products are already used in animal feeding, but the potential of these feed ingredients has not yet been fully investigated, especially in ruminant nutrition. The aim of the present study was to investigate the effect of the substitution, in dairy cow rations, of traditional protein and starch sources with more sustainable "circular" feeds to increase the sustainability of dairy production. For this purpose, eight multiparous mid-lactating cows were blocked and assigned to one of four treatments and were used in a replicated 4 x 4 Latin squares design with 21-days periods (14 days of adaptation and 7 of data collection). Two different circular feedstuffs were tested: a bakery's former foodstuff (FF) and a wheat distiller's grain with solubles (WDGS). These ingredients were used, alone and in combination, in three experimental diets (FF, WDGS; FF + WDGS) and compared to a standard ration (CTR). Dry matter intake and rumination time were not influenced by these diets. Conversely, dietary treatments partially influenced the milk yield, rumen pH, Volatile Fatty Acids (VFA) production, and fibre digestibility. In particular, the combined inclusion of FF and WDGS increased milk production (37.39 vs. 36.92, 35.48, 35.71 kg/day, for FF, WDGS and CTR diets, respectively) and reduced milk urea content (13.14 vs. 16.19, 15.58, 16.95 mg/dL for FF, WDGS, and CTR diets, respectively). No effects of this association were found in the milk composition, acetic and propionic production, and fibre digestibility. These results suggest that the association of former foodstuff and wheat distillers' grains could be safely included in dairy cow rations to increase the sustainability of cow nutrition and improve milk production without impairing animal health, dry matter intake, and fibre digestibility