Engineered primate L-methioninase for therapeutic purposes
Methods and compositions relating to the engineering of an improved protein with methionine-γ-lyase enzyme activity are described. For example, in certain aspects there may be disclosed a modified cystathionine-γ-lyase (CGL) comprising one or more amino acid substitutions and capable of degrading methionine. Furthermore, certain aspects of the invention provide compositions and methods for the treatment of cancer with methionine depletion using the disclosed proteins or nucleic acids.
1. A polypeptide comprising a variant of a native primate cystathionine-γ-lyase (CGL) enzyme, wherein the variant CGL has methionine-γ-lyase activity, comprises a sequence at least 95% identical to SEQ ID NO: 1, and comprises amino acid substitutions comprising E59N, R119L, and E339V of SEQ ID NO: 1, and at least one of the following substitutions: S63L, L91M, K268R, T311G, and I353S.
2. The polypeptide of claim 1 , wherein the variant CGL has at least 98% sequence identity to SEQ ID NO: 1.
3. The polypeptide of claim 1 , wherein the variant CGL is human CGL comprising SEQ ID NO: 1 having the E59N, S63L, L91M, R119L, K268R, T311G, E339V, and I353S substitutions.
4. The polypeptide of claim 1 , further comprising a heterologous peptide segment.
5. The polypeptide of claim 4 , wherein the heterologous peptide segment is an XTEN peptide, an IgG Fc, an albumin, or an albumin binding peptide.
6. The polypeptide of claim 1 , wherein the polypeptide is coupled to polyethylene glycol (PEG).
7. The polypeptide of claim 6 , wherein the polypeptide is coupled to PEG via one or more Lys or Cys residues.
8. A pharmaceutical formulation comprising the polypeptide of claim 1 in a pharmaceutically acceptable carrier.
9. A method of treating a tumor cell or subject having a tumor cell comprising administering to the tumor cell or the subject the formulation of claim 8 .
10. The method of claim 9 , wherein the subject is maintained on a methionine restricted diet.
11. The method of claim 9 , wherein the subject is maintained on a normal diet.
12. The method of claim 9 , wherein the subject is a human patient.
13. The method of claim 9 , wherein the formulation is administered intravenously, intradermally, intraarterially, intraperitoneally, intralesionally, intracranially, intraarticularly, intraprostaticaly, intrapleurally, intratracheally, intraocularly, intranasally, intravitreally, intravaginally, intrarectally, intramuscularly, subcutaneously, subconjunctival, intravesicularlly, mucosally, intrapericardially, intraumbilically, orally, by inhalation, by injection, by infusion, by continuous infusion, by localized perfusion bathing target cells directly, via a catheter, or via a lavage.
14. The method of claim 9 , wherein the formulation is administered to a nutrient medium of the tumor cell.
15. The method of claim 14 , wherein the nutrient medium is blood, lymphatic fluid, or spinal fluid.
16. The method of claim 9 , further comprising administering at least a second anticancer therapy to the subject.
17. The method of claim 16 , wherein the second anticancer therapy is a surgical therapy, chemotherapy, radiation therapy, cryotherapy, hormone therapy, immunotherapy or cytokine therapy.
18. The method of claim 9 , wherein the tumor cell is a cell of breast cancer, prostate cancer, neuroblastoma, or pancreatic carcinoma.