Compounds for improved viral transduction
The present invention provides methods and compositions for improving efficacy of viral transduction of cells. More particularly, the present invention provides methods and materials useful for safely and reliably improving the efficiency of methods and materials useful for safely and reliably improving the efficiency of methods for transducing cells, such as human hematopoietic stem cells (HSC), with viruses and/or viral vectors. The compositions and methods are useful for therapeutic indications amenable to treatment with hematopoietic stem cell gene therapies.
1. A method for increasing the lentiviral vector transduction efficiency of human CD34 + hematopoietic stem or progenitor cells comprising:
contacting the human CD34 + hematopoietic stem or progenitor cells with a culture medium comprising a lentiviral vector and prostaglandin E 2 (PGE 2 ), 16,16-dimethyl PGE 2 , or an analogue thereof, ex vivo
wherein the lentiviral vector transduction efficiency is increased in the human CD34 + hematopoietic stem or progenitor cells contacted with the culture medium compared to the lentiviral vector transduction efficiency of human CD34 + hematopoietic stem or progenitor cells contacted with a culture medium comprising the lentiviral vector in the absence of PGE 2 , 16,16-dimethyl PGE 2 , or an analogue thereof.
2. The method of claim 1 , wherein the human CD34 + hematopoietic stem or progenitor cells are human CD34 + hematopoietic stem cells.
3. The method of claim 1 , wherein the human CD34 + hematopoietic stem or progenitor cells are human CD34 + hematopoietic progenitor cells.
4. The method of claim 1 , wherein
a) at least 50% of the human CD34 + hematopoietic stem or progenitor cells are transduced;
b) at least 75% of the human CD34 + hematopoietic stem or progenitor cells are transduced; or
c) at least 90% of the human CD34 + hematopoietic stem or progenitor cells are transduced.
5. The method of claim 1 , wherein the medium further comprises a histone deacetylase (HDAC) inhibitor.
6. The method of claim 5 , wherein the HDAC inhibitor is selected from the group consisting of: Trichostatin A (TSA), valproic acid (VPA), sodium butyrate, suberoylanilide hydroxamic acid (SAHA), sodium phenylbutyrate, depsipeptide, trapoxin (TPX), cyclic hydroxamic acid-containing peptide 1 (CHAP1), MS-275, LBH589, and PXD-101.
7. The method of claim 1 , wherein the lentiviral vector is derived from a Human immunodeficiency virus (HIV) virus.
8. The method of claim 1 , wherein the lentiviral vector is pseudotyped with a vesicular stomatitis virus G-protein (VSV-G) envelope protein.
9. The method of claim 1 , wherein the human CD34 + hematopoietic stem or progenitor cells are contacted with the culture medium comprising the lentiviral vector and PGE 2 , 16,16-dimethyl PGE 2 , or an analogue thereof for at least four hours or for at least twenty-four hours.
10. The method of claim 1 , wherein the lentiviral vector comprises:
a) a left (5′) lentiviral long terminal repeat (LTR) comprising a heterologous promoter;
b) an expression control sequence operably linked to a gene of interest; and
c) a right (3′) lentiviral SIN LTR.
11. The method of claim 1 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi packaging sequence (Ψ+);
c) an HIV-1 central polypurine tract/DNA flap (cPPT/FLAP);
d) a rev response element (RRE);
e) a β-globin promoter and a β-globin locus control region (LCR) operably linked to a gene of interest; and
f) a right (3′) lentiviral SIN LTR that comprises
i) one or more insulator elements, or
ii) a rabbit β-globin polyA (rβgpA) sequence.
12. The method of claim 1 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi (Ψ+) packaging signal;
c) a cPPT/FLAP;
d) an RRE;
e) a myeloproliferative sarcoma virus enhancer, negative control region deleted, d1587rev primer-binding site substituted (MND) promoter, operably linked to a polynucleotide encoding a human ATP-binding cassette, sub-family D, member 1 (ABCD1) polypeptide;
f) a right (3′) HIV-1 SIN LTR; and
g) a rβgpA sequence.
13. A method for increasing the lentiviral transduction efficiency of human CD34 + hematopoietic stem or progenitor cells comprising:
contacting the human CD34 + hematopoietic stem or progenitor cells with a culture medium comprising a lentiviral vector and PGE 2 , ex vivo wherein the lentiviral transduction efficiency is increased in the human CD34 + hematopoietic stem or progenitor cells contacted with the culture medium compared to the lentiviral transduction efficiency of human CD34 + hematopoietic stem or progenitor cells contacted with a culture medium comprising the lentiviral vector in the absence of PGE 2 .
14. The method of claim 13 , wherein
a) at least 50% of the human CD34 + hematopoietic stem or progenitor cells are transduced;
b) at least 75% of the human CD34 + hematopoietic stem or progenitor cells are transduced; or
c) at least 90% of the human CD34 + hematopoietic stem or progenitor cells are transduced.
15. The method of claim 13 , wherein the lentiviral vector is derived from an HIV virus.
16. The method of claim 13 , wherein the lentiviral vector is pseudotyped with a VSV-G envelope protein.
17. The method of claim 13 , wherein the human CD34 + hematopoietic stem or progenitor cells are contacted with the culture medium comprising the lentiviral vector and PGE 2 for at least four hours or for at least twenty-four hours.
18. The method of claim 13 , wherein the lentiviral vector comprises:
a) a left (5′) lentiviral LTR comprising a heterologous promoter;
b) an expression control sequence operably linked to a gene of interest; and
c) a right (3′) lentiviral SIN LTR.
19. The method of claim 13 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi packaging sequence (Ψ+);
c) an HIV-1 cPPT/FLAP;
d) an RRE;
e) a β-globin promoter and a β-globin LCR operably linked to a gene of interest; and
f) a right (3′) lentiviral SIN LTR that comprises
i) one or more insulator elements, or
ii) a rβgpA sequence.
20. The method of claim 13 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi (Ψ+) packaging signal;
c) a cPPT/FLAP;
d) an RRE;
e) an MND promoter, operably linked to a polynucleotide encoding a human ABCD1 polypeptide;
f) a right (3′) HIV-1 SIN LTR; and
g) a rβgpA sequence.
21. A method for increasing the lentiviral transduction efficiency of human CD34 + hematopoietic stem or progenitor cells comprising:
contacting the human CD34 + hematopoietic stem or progenitor cells with a culture medium comprising a lentiviral vector and 16,16-dimethyl PGE 2 , ex vivo
wherein the lentiviral transduction efficiency is increased in the human CD34 + hematopoietic stem or progenitor cells contacted with the culture medium compared to the lentiviral transduction efficiency of human CD34 + hematopoietic stem or progenitor cells contacted with a culture medium comprising the lentiviral vector in the absence of 16,16-dimethyl PGE 2 .
22. The method of claim 21 , wherein
a) at least 50% of the human CD34 + hematopoietic stem or progenitor cells are transduced;
b) at least 75% of the human CD34 + hematopoietic stem or progenitor cells are transduced; or
c) at least 90% of the human CD34 + hematopoietic stem or progenitor cells are transduced.
23. The method of claim 21 , wherein the lentiviral vector is derived from an HIV-1 virus.
24. The method of claim 21 , wherein the lentiviral vector is pseudotyped with a VSV-G envelope protein.
25. The method of claim 21 , wherein the human CD34 + hematopoietic stem or progenitor cells are contacted with the culture medium comprising the lentiviral vector and 16,16-dimethyl PGE 2 for at least four hours or for at least twenty-four hours.
26. The method of claim 21 , wherein the lentiviral vector comprises:
a) a left (5′) lentiviral LTR comprising a heterologous promoter;
b) an expression control sequence operably linked to a gene of interest; and
c) a right (3′) lentiviral SIN LTR.
27. The method of claim 21 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi packaging sequence Ψ+);
c) an HIV-1 cPPT/FLAP;
d) an RRE;
e) a β-globin promoter and a β-globin LCR operably linked to a gene of interest; and
f) a right (3′) lentiviral SIN LTR that comprises
i) one or more insulator elements, or
ii) a rβgpA sequence.
28. The method of claim 21 , wherein the lentiviral vector comprises:
a) a left (5′) HIV-1 LTR comprising a CMV promoter;
b) a Psi (Ψ+) packaging signal;
c) a cPPT/FLAP;
d) an RRE;
e) an MND promoter, operably linked to a polynucleotide encoding a human ABCD1 polypeptide;
f) a right (3′) HIV-1 SIN LTR; and
g) a rβgpA sequence.