REFERENCE TO A SEQUENCE LISTING

The Sequence Listing written in file SequenceListing_077867-1162156-652100US.txt, created on Mar. 16, 2020, 15,771 bytes, machine format IBM-PC, MS-Windows operating system, is hereby incorporated by reference in its entirety for all purposes.

BACKGROUND OF THE INVENTION

Disorders of adult beta-globin (HBB) synthesis, or beta-globinopathies, are the most common monogenic disorders in the world. Two best-known examples include sickle cell disease and beta-thalassemia.

Sickle cell disease is caused by a single point mutation known as HbS in beta-globin, where a hydrophilic amino acid glutamic acid is substituted with a hydrophobic amino acid valine at the sixth position due to codon GAG being replaced by GTG. This substitution creates a hydrophobic spot on the outside of the protein, which tends to stick to the hydrophobic region of an adjacent hemoglobin molecule's beta chain. This further causes clumping of hemoglobin molecules into rigid fibers, causing “sickling” of the entire red blood cells in the HbS/HbS homozygotes. While the homozygous HbS/HbS alleles lead to a severe and potentially fatal genetic disorder, heterozygous HbS carriers are resistant to malaria and are minimally affected by the symptoms of anemia. Beta-thalassemia is a genetic disease caused by total or partial loss of beta-globin in a patient. Total loss, termed thalassemia major or beta-0-thalassemia, is due to mutation in both alleles of the beta-globin genes leading to failure to form beta chain of hemoglobin. It prevents oxygen supply in the tissues and can therefore be highly lethal. Symptoms, such as severe anemia and heart attack, often appear within two years after birth. Reduced beta-globin function termed thalassemia minor or beta+thalassemia is due to mutation in only one of the alleles. It is less severe, but patients are prone to other diseases such as asthma and liver problems.

Beta-globinopathies can be treated by lifelong blood transfusion and bone marrow transplantation. Although previous studies have demonstrated the curative potential of bone marrow transplantation, this approach is limited to a small fraction of affected patients due to the requirement for an HLA-matched donor, the highly specialized approach that requires critical infrastructure, and the high cost. In a more recent study, transcription factor Pogo transposable element with Zinc finger domain (POGZ) has been shown to negatively regulate beta-globin synthesis, suggesting its suppression as a possible means for treating beta-globinopathies. Targeting a transcription factor, however, tends to present many practical difficulties in the context of developing therapeutic agents. Therefore, there exists a distinct need to develop new and cost-effective treatment methods, especially by increasing fetal hemoglobin expression as a potential cure for beta-globinopathies. This invention fulfills this and other related needs.

BRIEF SUMMARY OF THE INVENTION

The application provides the first disclosure of RIO kinase 3, a serine/threonine kinase and a member of the right open reading frame (RIO) kinase family, playing a role as a downstream effector of transcription factor Pogo transposable element with Zinc finger domain (POGZ) in the negative regulation of beta-globin synthesis. New compositions and methods for treating diseases and disorders involving suppressed beta-globin synthesis are therefore devised from this discovery.

As such, in a first aspect, this invention provides a method for promoting beta-globin synthesis in a cell. The method includes a step of contacting the cell with an effective amount of an inhibitor of RIOK3. In some embodiments, the cell is an erythroid cell, e.g., a primary erythroid progenitor cell or a CD34+ erythroid cell. In some embodiments, the cell is with the body of a patient, especially a human patient.

In some embodiments, the method comprises administering an effective amount of an inhibitor of RIOK3 to the patient. In some embodiments, the inhibitor is a neutralizing antibody against RIOK3 (e.g., a monoclonal or polyclonal antibody that specifically binds to RIOK3 protein and interferes with its activity), an siRNA, a microRNA, a miniRNA, a lncRNA, an antisense oligonucleotide, or a small molecule (e.g., Midostaurin, Axitinib, Bosutinib, or Ruxolitinib). In some embodiments, the inhibitor is administered by subcutaneous, intramuscular, intravenous, intraperitoneal, or oral administration. In some embodiments, the inhibitor is administered in the form of a solution, a powder, a paste, a tablet, or a capsule. In some embodiments, the patient receiving treatment by the claimed method has been diagnosed with a beta-globinopathy, such as sickle cell disease or beta-thalassemia. The patient optionally is receiving concurrently another therapy for the beta-globinopathy, such as blood transfusion.

In a second aspect, the present invention provides a composition comprising an effective amount of an inhibitor of RIOK3 and a physiologically acceptable excipient. In some embodiments, the composition is formulated for subcutaneous, intramuscular, intravenous, intraperitoneal, or oral administration. For example, the composition may be in form of a solution, a powder, a paste, a tablet, or a capsule. In some embodiments, the composition comprises one or more of the RIOK3 inhibitor. In some embodiments, the inhibitor is a neutralizing antibody against RIOK3 (e.g., a monoclonal or polyclonal antibody that specifically binds to RIOK3 protein and interferes with its activity), an siRNA, a microRNA, a miniRNA, a lncRNA, an antisense oligonucleotide, or a small molecule (e.g., Midostaurin, Axitinib, Bosutinib, or Ruxolitinib). In some embodiments, the inhibitor is Midostaurin, Axitinib, Bosutinib, or Ruxolitinib, or any combinations thereof.

In a third aspect, the present invention provides a kit for promoting beta-globin synthesis. The kit includes a container containing a composition comprising an effective amount of an inhibitor of RIOK3. Optionally, a second container containing a second RIOK3 inhibitor in an effective amount is also included in the kit. In some embodiments, the RIOK3 inhibitor-containing composition is formulated for subcutaneous, intramuscular, intravenous, intraperitoneal, topical, or oral administration. For example, the composition may be in form of a solution, a powder, a paste, a tablet, or a capsule. In some embodiments, the inhibitor is a neutralizing antibody against RIOK3 (e.g., a monoclonal or polyclonal antibody that specifically binds to RIOK3 protein and interferes with its activity), an siRNA, a microRNA, a miniRNA, a lncRNA, an antisense oligonucleotide, or a small molecule (e.g., Midostaurin, Axitinib, Bosutinib, or Ruxolitinib). In some embodiments, the inhibitor is Midostaurin, Axitinib, Bosutinib, or Ruxolitinib, or any combinations thereof. In some embodiments, the kit further comprises an instruction manual for administration of the composition comprising the RIOK3 inhibitor(s).

Related to this aspect of the present invention, a use of a RIOK3 inhibitor is further provided for the manufacturing of (1) a medicament for treating beta-globinopathies; or (2) a kit containing the medicament for treating beta-globinopathies. In some embodiments, the medicament comprising an effective amount of one or more RIOK3 inhibitors is formulated for subcutaneous, intramuscular, intravenous, intraperitoneal, topical, or oral administration. For example, the medicament may be in form of a solution, a powder, a paste, a tablet, or a capsule. In some embodiments, the inhibitor is a neutralizing antibody against RIOK3 (e.g., a monoclonal or polyclonal antibody that specifically binds to RIOK3 protein and interferes with its activity), an siRNA, a microRNA, a miniRNA, a lncRNA, an antisense oligonucleotide, or a small molecule (e.g., Midostaurin, Axitinib, Bosutinib, or Ruxolitinib). In some embodiments, the inhibitor used for manufacturing the medicament or kit for treating a beta-globinopathy is Midostaurin, Axitinib, Bosutinib, or Ruxolitinib, or any combinations thereof.

