Compositions and methods for treating beta-hemoglobinopathies

Abstract

The present disclosure provides expression vectors comprising at least two nucleic acid sequences, namely a nucleic acid sequence encoding an anti-HPRT RNAi, and a nucleic acid sequence encoding a gamma globin gene. In some embodiments, the viral vector is a self-inactivating lentiviral vector. In some embodiments, the gamma-globin gene is used to genetically correct sickle cell disease or β-thalassemia or to reduce symptoms thereof.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]The present application claims the benefit of the filing date of U.S. Provisional Patent Application No. 62 / 653,913, filed on Apr. 6, 2018; the benefit of the filing date of U.S. Provisional Patent Application No. 62 / 541,931, filed on Aug. 7, 2017; and also, the benefit of the filing date of U.S. Provisional Patent Application No. 62 / 533,719 filed on Jul. 18, 2017, the disclosures of which are each hereby incorporated by reference herein in their entireties.FIELD OF DISCLOSURE[0002]This disclosure generally relates to the fields of molecular biology and, in particular, vectors and host cells transduced by vectors.BACKGROUND OF THE DISCLOSURE[0003]β-Hemoglobinopathies, including beta-thalassemia and sickle-cell disease (SCD), are a heterogeneous group of commonly inherited disorders affecting the function or levels of hemoglobin. SCD and β-thalassemia are the most common monogenic disorders in the world with approximately 400,000 affected ...

AI Technical Summary

Benefits of technology

[0009]Gene therapy strategies to modify human stem cells hold great promise for curing many human diseases, included hemoglobinopathies. It is believed that the engraftment of gene modified stem cells may be enhanced by engineering stem cells in which hypoxanthine guanine phosphoribosyitransferase (“HPRT”) expression is knocked down, thereby enabling the selection of genetically modified cells by conferring resistance to a guanine analog antimetabolite.

[0016]In some embodiments, the second nucleic acid encoding the therapeutic gene is one which may genetically correct sickle cell disease or β-thalassemia; or reduce symptoms thereof (including the symptoms of severe SCD). In other embodiments, the nucleic acid encoding the therapeutic gene is one which may genetically correct immune deficiencies, hereditary diseases, blood diseases (e.g. hemophilia, hemoglobin disorders), neurological diseases, and / or lysosomal storage diseases; or reduce symptoms thereof. In some embodiments, the vector is a lentiviral vector. In some embodiments, the therapeutic gene is gamma globin. In some embodiments, the second nucleic acid sequence encoding the therapeutic gene has a sequence having at least 80% identity to that of SEQ ID NO: 55. In some embodiments, the second nucleic acid sequence encoding the therapeutic gene has a sequence having at least 90% identity to that of SEQ ID NO: 55. In some embodiments, the second nucleic acid sequence encoding the therapeutic gene has a sequence having at least 95% identity to that of SEQ ID NO: 55. In some embodiments, the second nucleic acid sequence encoding the therapeutic gene has a sequence having at least 97% identity to that of SEQ ID NO: 55. In some embodiments, the second nucleic acid sequence encoding the therapeutic gene has a sequence of SEQ ID NO: 55.

[0032]It is believed that with a strategy of combined conditioning and chemoselection (such as with a purine analog), efficient and high engraftment of HPRT-deficient, gamma globin gene-containing hematopoietic stem cells can be achieved, and it is believed that such high engraftment may be accomplished with low overall toxicity. It is believed that the enhanced engraftment and chemoselection of the gene-modified HSCs, combined with lineage-specific expression of the gamma globin gene, may result in a sufficient frequency of red blood cells expressing the therapeutic gamma globin transgene, allowing for increased levels of fetal hemoglobin formation to correct for SCD and / or beta thalassemia. As a safety measure, HPRT-deficient cells can be negatively selected, such as by introducing MTX or MPA, to inhibit the enzyme dihydrofolate reductase (DHFR) in the purine de novo synthetic pathway, thus killing HPRT deficient cells.

[0033]It is further believed that HPRT-deficient HSCs can be selected in vivo using a regimen of a purine analog (e.g. 6TG) to enhance engraftment. It is also believed that the expanded gene-modified HSCs can differentiate into erythrocytes expressing the therapeutic gamma globin transgene. The gene therapy compositions described herein have the potential to not only correct SCD and beta thalassemia, but also to greatly improve on the current “gold standards” for autologous hematopoietic stem cell transplantation. Improvements may allow for (i) out-patient procedures using the gene-modified HSCs; (ii) low adverse events (AEs), including avoiding infertility associated with other clinical therapies; (iii) low dose oral administration for conditioning (as compared with high-dose IV conditioning); (iv) in vivo selection of gene-modified cells; and / or (v) low procedure mortality rate related to transplantation and conditioning.

Problems solved by technology

Clinical manifestations typically appear several months after birth during the switch from fetal hemoglobin (HbF) to adult β-globin (HbA) and can be severe with substantial morbidity and mortality.

Allogenic bone marrow transplantation is curative but limited to those patients with an appropriately matched donor.

It is further believed that the alpha-globin aggregates cause cell membrane damage and lead to early erythroid precursor death.

The resultant ineffective erythropoiesis found in patients, if severe, may necessitate frequent blood transfusions.

A matched allogeneic hematopoietic stem cell (HSC) transplantation is believed to be curative but restricted by the availability of matched related donors and has potential serious complications.

However, after birth, the expression of fetal gamma-globin gene decreases to negligible levels, with a concomitant increase in beta-globin expression.

Their high prevalence, significant morbidity and mortality, and the resulting high cost of lifelong palliative medical care portends that a curative therapy can greatly improve patient outcomes and significantly reduce associated medical costs.

Despite promising results, the majority of subjects in these trials failed to achieve levels of engraftment of gene-corrected autologous HSPC or reach a threshold level of expression of the therapeutic protein associated with clinical benefit.

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