IP Library › Patent Application 18984508

Patent Application

App. No. 18/984,508

COMPOSITIONS AND METHODS FOR NON-GENOTOXIC CELL CONDITIONING

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App. No.: 18/984,508

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Abstract

Compositions and methods for non-genotoxic monoclonal antibody (mAb) conditioning, where the methods involve altering a cluster of differentiation 117 (CD117; c-KIT) polynucleotide sequence in a hematopoietic stem cell (HSC) or progenitor thereof to encode a CD117 polypeptide with reduced binding to the antibody. In various embodiments, the methods further include introducing a therapeutic alteration to a gene of the HSC or progenitor thereof for treatment of a hemoglobinopathy.

Claims (698)

1 . A method for hematopoietic stem cell transplantation in a subject, the method comprising:

(a) contacting an isolated hematopoietic stem cell or progenitor thereof with a guide polynucleotide and a base editor comprising a nucleic acid programmable DNA binding protein (napDNAbp) and an adenosine deaminase domain or a polynucleotide encoding the base editor, wherein the guide polynucleotide targets a nucleic acid molecule encoding a CD117 polypeptide, thereby generating an edited cell;

(b) administering the edited cell to the subject; and

(c) administering to the subject an antibody or antigen binding fragment thereof, wherein the antibody is selected from the group consisting of ABTx025, ABTx030, ABTx052, ABTx061, ABTx062, ABTx070, ABTx071, ABTx196, ABTx198, ABTx202, ABTx203, ABTx205, ABTx206, ABTx248, ABTx250, ABTx251, ABTx253, ABTx254, ABTx255, ABTx256, ABTx265, ABTx268, ABTx270, ABTx271, ABTx272, ABTx273, ABTx274, ABTx307, ABTx308, ABTx309, and ABTx313.

2 . The method of claim 1 , wherein in step (a) generating an edited cell comprises

i) introducing an alteration of a nucleobase in a codon encoding a serine at amino acid position 261, wherein the alteration of the nucleobase encoding the serine at amino acid position 261 results in the codon expressing a glycine,

ii) introducing an alteration of a nucleobase in a codon encoding a serine at amino acid position 251, and/or

iii) introducing an alteration of a nucleobase in a codon encoding an asparagine at amino acid position 260 and introducing an alteration of a nucleobase in a codon encoding a serine at amino acid position 261, or corresponding positions in another CD117 polypeptide, and generating an edited cell;

3 . The method of claim 1 , wherein the adenosine deaminase domain comprises a combination of alterations to TadA*7.10 selected from:

i) I76Y, V82T, Y123H, Y147R, F149Y, and Q154R; and

ii) I76Y, V82T, Y123H, Y147D, F149Y, Q154R, T166I, and D167N; wherein the adenosine deaminase domain has at least 85% sequence identity to TadA*7.10.

4 . A method for treating a hemoglobinopathy in a subject, the method comprising:

(a) contacting an isolated hematopoietic stem cell or progenitor thereof with two or more guide polynucleotides and a base editor comprising a nucleic acid programmable DNA binding protein (napDNAbp) and an adenosine deaminase domain or a polynucleotide encoding the base editor, wherein one guide polynucleotide targets a nucleic acid molecule encoding a CD117 polypeptide and another guide polynucleotide targets the base editor to effect a deamination of a nucleobase of a hemoglobin subunit gamma 1 and/or 2 (HBG1/2) promoter, thereby generating an edited cell;

(b) administering the edited cell to the subject; and

5 . The method of claim 4 , wherein in step (a),

1. one guide polynucleotide targets a nucleic acid molecule encoding a CD117 polypeptide, thereby

ii) introducing an alteration of a nucleobase in a codon encoding a serine at amino acid position 251, and/or

iii) introducing an alteration of a nucleobase in a codon encoding an asparagine at amino acid position 260 and introducing an alteration of a nucleobase in a codon encoding a serine at amino acid position 261 or corresponding positions in another CD117 polypeptide; and

2) another guide polynucleotide targets the base editor to effect an alteration to a beta globin polynucleotide (HBB) that results in expression of a beta globin polypeptide having an alanine at position 6 (Hb G-Makassar), thereby generating an edited cell;

(b) administering the edited cell to the subject; and

(c) administering to the subject an antibody or antigen binding fragment thereof, antibody drug conjugate, or a chimeric antigen receptor T (CAR-T) cell, each of which selectively binds a wild type CD117 polypeptide.

6 . A method of altering a nucleobase of a CD117 polynucleotide, the method comprising:

contacting the CD117 polynucleotide with a base editor polypeptide comprising a nucleic acid programmable DNA binding protein (napDNAbp) and an adenosine deaminase domain, and a guide polynucleotide that targets said base editor to

i) effect an alteration of a nucleobase in a codon encoding a serine at amino acid position 261, wherein the alteration of the nucleobase encoding the serine at amino acid position 261 results in the codon expressing a glycine,

ii) effect an alteration of a nucleobase in a codon encoding a serine at amino acid position 251, and/or

iii) effect an alteration of a nucleobase in a codon encoding an asparagine at amino acid position 260 and effect an alteration of a nucleobase in a codon encoding a serine at amino acid position 261, or corresponding positions in another CD117 polypeptide, thereby altering the nucleobase of the CD117 polynucleotide.

7 . The method of claim 6 , wherein the adenosine deaminase domain comprises a combination of alterations to TadA*7.10 selected from:

a) I76Y, V82T, Y123H, Y147R, F149Y, and Q154R; and

b) I76Y, V82T, Y123H, Y147D, F149Y, Q154R, T166I, and D167N; wherein the adenosine deaminase domain has at least 85% sequence identity to TadA*7.10, and a guide polynucleotide that targets the base editor to effect an alteration of a nucleobase in a polynucleotide encoding a CD117 polypeptide, thereby altering the nucleobase of the CD117 polynucleotide.

8 . A cell produced by the method of claim 1 .

9 . A pharmaceutical composition comprising an effective amount of the cell of claim 8 .

10 . A base editor system comprising a base editor comprising a nucleic acid programmable DNA binding protein (napDNAbp) and an adenosine deaminase domain or a polynucleotide encoding the base editor, and a guide polynucleotide that targets said base editor to

ii) effect an alteration of a nucleobase in a codon encoding a serine at amino acid position 251, and/or

11 . The base editor system of claim 10 , wherein the adenosine deaminase domain comprises a combination of alterations to TadA*7.10, wherein the combinations are selected from:

a) I76Y, V82T, Y123H, Y147R, F149Y, and Q154R; and

b) I76Y, V82T, Y123H, Y147D, F149Y, Q154R, T166I, and D167N; wherein the guide polynucleotide targets the base editor to effect an alteration of a nucleobase of a CD117 polynucleotide, wherein the adenosine deaminase domain has at least 85% sequence identity to TadA*7.10.

12 . The base editor system of claim 10 , wherein the guide polynucleotide comprises a polynucleotide sequence selected from the group consisting of: AUAAUAGCUGGCAUCACGGU (SEQ ID NO: 693; CC200);

(SEQ ID NO: 694; gRNA889)

CCACUAGCUUUCCAAACGGU;

(SEQ ID NO: 695; gRNA908)

GCUGAACUGAUAGUCAACGU;

(SEQ ID NO: 696; gRNA918)

UUUGACAAAGCCCGGAUCAG;

(SEQ ID NO: 697; gRNA923)

UGAAAGUGAGGCCAGGUACU;

(SEQ ID NO: 698; gRNA928)

AAACAGUCAGGUGAGUGAAU;

(SEQ ID NO: 699; gRNA929)

AACUACAGGAGAAAUAUAAU;

and

(SEQ ID NO: 700; gRNA944)

GAUUAAAAGGCACCGAAGGA.

13 . A polynucleotide encoding the base editor system of claim 10 .

14 . A guide polynucleotide comprising a spacer sequence selected from the group consisting of:

(SEQ ID NO: 693; gRNA931)

AUAAUAGCUGGCAUCACGGU;

(SEQ ID NO: 694; gRNA889)

CCACUAGCUUUCCAAACGGU;

(SEQ ID NO: 695; gRNA908)

GCUGAACUGAUAGUCAACGU;

(SEQ ID NO: 696; gRNA918)

UUUGACAAAGCCCGGAUCAG;

(SEQ ID NO: 697; gRNA923)

UGAAAGUGAGGCCAGGUACU;

(SEQ ID NO: 698; gRNA928)

AAACAGUCAGGUGAGUGAAU;

(SEQ ID NO: 699; gRNA929)

AACUACAGGAGAAAUAUAAU;

and

(SEQ ID NO: 700; gRNA944)

GAUUAAAAGGCACCGAAGGA.

15 . A kit comprising base editor system of claim 10 .

16 . An anti-CD117 antibody or antigen-binding portion thereof comprising one or more complementarity determining regions (CDRs) which comprise or consist of heavy chain variable region (VH) CDRs and/or light chain variable region (VL) CDRs selected from the following:

VL CDR1:

(SEQ ID NO: 394)

QSVSSSY;

VL CDR2:

GAS;

VL CDR3:

(SEQ ID NO: 395)

QQYGTSLT;

VH CDR1:

(SEQ ID NO: 391)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 392)

ISWNSGTI;

VH CDR3:

(SEQ ID NO: 393)

AKDSPPGYCSSASCYGAFDI (ABTx025);

VL CDR1:

(SEQ ID NO: 409)

QSISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 410)

QQSYSTPLT;

VH CDR1:

(SEQ ID NO: 406)

GFTFSSYS;

VH CDR2:

(SEQ ID NO: 407)

IGTISSYI;

VH CDR3:

(SEQ ID NO: 408)

ARDYYGGLFDY (ABTx030);

VL CDR1:

(SEQ ID NO: 439)

QSVSSSY;

VL CDR2:

GAS;

VL CDR3:

(SEQ ID NO: 440)

QQYGSSPLT;

VH CDR1:

(SEQ ID NO: 436)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 437)

ISWNSGSI;

VH CDR3:

(SEQ ID NO: 438)

AKDTPLGYCSTTSCYGAFDI (ABTx061);

VL CDR1:

(SEQ ID NO: 454)

QSISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 455)

QQSYSTPFT;

VH CDR1:

(SEQ ID NO: 451)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 452)

ISWNSGTI;

VH CDR3:

(SEQ ID NO: 453)

AKDSPPGYCSSASCYGAFDI (ABTx062);

VL CDR1:

(SEQ ID NO: 469)

QGISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 470)

QQSYSTPIT;

VH CDR1:

(SEQ ID NO: 466)

GFTFDDYG;

VH CDR2:

(SEQ ID NO: 467)

INWNGGST;

VH CDR3:

(SEQ ID NO: 468)

ARESWDGSGIYYMDV (ABTx070);

VL CDR1:

(SEQ ID NO: 484)

QGISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 485)

QQLNSYPYT;

VH CDR1:

(SEQ ID NO:481)

GFTFDDYG;

VH CDR2:

(SEQ ID NO: 482)

INWNGGST;

VH CDR3:

(SEQ ID NO: 483)

ARESWNYEGYYYMDV (ABTx071);

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 425)

QQFNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 425)

QQFNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 954)

QQFYSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

TYPGDSDTR;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1020)

QQFSSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1023)

QQTNSHPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1025)

QQVRSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1029)

QQTYSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1027)

ARHGRGYDAYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1036)

QQTRSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 952)

QQVNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1025)

QQVRSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1027)

ARHGRGYDAYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1044)

QQTNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1032)

ARHGRGYDAYDGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1025)

QQVRSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

IYPGDSDTK;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 953)

QQFRSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1044)

QQTNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

TYPGDSDTK;

VH CDR3:

(SEQ ID NO: 948)

ARHGRGYDGYEGAFDI;

VL CDR1:

(SEQ ID NO: 394)

QSVSSSY;

VL CDR2:

GAS;

VL CDR3:

(SEQ ID NO: 1074)

QQSETCLT;

VH CDR1:

(SEQ ID NO: 391)

GFTFDDYA

VH CDR2:

(SEQ ID NO: 1070)

ISWNSGTIG;

VH CDR3:

(SEQ ID NO: 393)

AKDSPPGYCSSASCYGAFDI;

VL CDR1:

(SEQ ID NO: 394)

QSVSSSY;

VL CDR2:

GAS;

VL CDR3:

(SEQ ID NO: 1074)

QQDSLGLT;

VH CDR1:

(SEQ ID NO: 391)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 1070)

ISWNSGTIG;

VH CDR3:

(SEQ ID NO: 1090)

AKDSPPGYCASASCYGAFDI;

VL CDR1:

(SEQ ID NO: 394)

QSVSSSY;

VL CDR2:

GSS;

VL CDR3:

(SEQ ID NO: 1084)

QQYNFWPYT;

VH CDR1:

(SEQ ID NO: 391)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 1070)

ISWNSGTIG;

VH CDR3:

(SEQ ID NO: 393)

AKDSPPGYCSSASCYGAFDI;

VL CDR1:

(SEQ ID NO: 469)

QGISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 1097)

QQSYSTPYT;

VH CDR1:

(SEQ ID NO: 391)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 1070)

ISWNSGTIG;

VH CDR3:

(SEQ ID NO: 1094)

AKDWPSGFCSSAYCYGAFDI;

VL CDR1:

(SEQ ID NO: 469)

QGISSY;

VL CDR2:

AAS;

VL CDR3:

(SEQ ID NO: 1097)

QQSYSTPYT;

VH CDR1:

(SEQ ID NO: 1071)

GFTFDDYA;

VH CDR2:

(SEQ ID NO: 1070)

ISWNSGTIG;

VH CDR3:

(SEQ ID NO: 393)

AKDSPPGYCSSASCYGAFDI;

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 425)

QQFNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI;

AA)

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1020)

QQFSSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 972)

TYPGDSDTK;

VH CDR3:

(SEQ ID NO: 1032)

ARHGRGYDAYDGAFDI;

AB)

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 1044)

QQTNSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1032)

ARHGRGYDAYDGAFDI;

and

AC)

VL CDR1:

(SEQ ID NO: 424)

QGISSA;

VL CDR2:

DAS;

VL CDR3:

(SEQ ID NO: 425)

QQFSSYPLT;

VH CDR1:

(SEQ ID NO: 421)

GYRFTSYW;

VH CDR2:

(SEQ ID NO: 958)

IYPGDSDTR;

VH CDR3:

(SEQ ID NO: 1012)

ARHGRGYDGYDGAFDI.

17 . An isolated polynucleotide that encodes the antibody of claim 16 .

18 . An anti-CD117 antibody or antigen-binding portion thereof comprising complementarity determining regions (CDRs) that comprise or consist of the following heavy chain variable region (VH) CDR and light chain variable region (VL) CDR amino acid sequences:

VH CDR1: GX 1 X 2 FX 3 X 4 YX 5 , wherein

X 1 is F or Y,

X 2 is R or T,

X 3 is D, S, or T,

X 4 is D or S, and

X 5 is A, G, S, or W;

VH CDR2 is IX 6 X 7 X 8 X 9 X 10 X 11 X 12 X 13 , wherein

X 6 is G, N, S, or Y,

X 7 is P, T, or W,

X 8 is G, I, or N,

X 9 is D, G, or S,

X 10 is Gor S,

X 11 is D, S, T, or Y,

X 12 is I or T, and

X 13 is G, K, R, or Y;

VH CDR3 is selected from the group consisting of ARHGRGYDX 14 YDGAFDI (SEQ ID NO: 1105), ARDYYGGLFDY (SEQ ID NO: 1106), ARESWX 15 X 16 X 17 GX 18 YYMDV (SEQ ID NO: 1107), and AKDX 19 PX 20 GX 21 CX 22 X 23 X 24 X 25 CYGAFDI (SEQ ID NO: 1108), wherein

X 14 is A or G,

X 15 is D or N,

X 16 is Gor Y,

X 17 is E or S,

X 18 is I or Y,

X 19 is S, T, or W,

X 20 is L, P, or S,

X 21 is F or Y,

X 22 is A or S,

X 23 is S or T,

X 24 is A or T, and

X 25 is S or Y;

VL CDR1 is QSX 26 SSX 27 (SEQ ID NO: 1109) or QSVSSSY (SEQ ID NO: 1110), wherein

X 26 is Gor S, and

X 27 is A or Y;

VL CDR2: X 28 X 29 S, wherein,

X 28 is A, D, or G, and

X 29 is A or S; and

VL CDR3 is QQX 30 X 31 X 32 X 33 PX 34 T (SEQ ID NO: 1111) or QQX 35 X 36 X 37 X 38 LT (SEQ ID NO: 1112), wherein

X 30 is F, L, S, T, or Y,

X 31 G, N, S, or Y,

X 32 is S or F,

X 33 S, T, W, or Y,

X 34 F, I, L, or Y,

X 35 is D, S, or Y,

X 36 is E, G, or S,

X 37 is L or T, and

X 38 is C, G, or S;

and wherein the anti-CD117 antibody comprises at least one amino acid alteration relative to the amino acid sequence of ABTx052.

19 . A method for hematopoietic stem cell transplantation in a subject, the method comprising:

(a) administering a hematopoietic stem cell or progenitor thereof to the subject, wherein the hematopoietic stem cell or progenitor thereof expresses a CD117 variant comprising an S261G amino acid alteration, or a CD117 variant comprising Y259C and N260D amino acid alterations; and

(b) administering to the subject an antibody or antigen binding fragment thereof that selectively binds a wild type CD117 polypeptide.

20 . The method of claim 19 , wherein the hematopoietic stem cell or progenitor thereof:

i) either expresses a CD117 variant comprising an S261G amino acid alteration or a CD117 variant comprising Y259C and N260D amino acid alterations, and

ii) comprises a nucleobase alteration to the HBG1/2 promoter that effects an increase in gamma globin expression and/or expresses an HBB polypeptide comprising an alanine at position 6.

21 . A hematopoietic stem cell or progenitor thereof expressing a CD117 polypeptide comprising

i) an S261G alteration,

ii) alterations at amino acid positions 260 and 261, and/or

ii) an alteration at amino acid position 251, relative to the following amino acid sequence,

wherein the CD117 polypeptide has at least 85% sequence identity to the following amino acid sequence:

Wild Type CD117

(SEQ ID NO: 499)

MRGARGAWDFLCVLLLLLRVQTGSSQPSVSPGEPSPPSIHPGKSDLIVRVGDEIRLLCTDPGFV

KWTFEILDETNENKQNEWITEKAEATNTGKYTCTNKHGLSNSIYVFVRDPAKLFLVDRSLYGKE

DNDTLVRCPLTDPEVTNYSLKGCQGKPLPKDLRFIPDPKAGIMIKSVKRAYHRLCLHCSVDQEG

KSVLSEKFILKVRPAFKAVPVVSVSKASYLLREGEEFTVTCTIKDVSSSVYSTWKRENSQTKLQ

EKYNSWHHGDFNYERQATLTISSARVNDSGVFMCYANNTFGSANVTTTLEVVDKGFINIFPMIN

TTVFVNDGENVDLIVEYEAFPKPEHQQWIYMNRTFTDKWEDYPKSENESNIRYVSELHLTRLKG

TEGGTYTFLVSNSDVNAAIAFNVYVNTKPEILTYDRLVNGMLQCVAAGFPEPTIDWYFCPGTEQ

RCSASVLPVDVQTLNSSGPPFGKLVVQSSIDSSAFKHNGTVECKAYNDVGKTSAYFNFAFKGNN

KEQIHPHTLFTPLLIGFVIVAGMMCIIVMILTYKYLQKPMYEVQWKVVEEINGNNYVYIDPTQL

PYDHKWEFPRNRLSFGKTLGAGAFGKVVEATAYGLIKSDAAMTVAVKMLKPSAHLTEREALMSE

LKVLSYLGNHMNIVNLLGACTIGGPTLVITEYCCYGDLLNFLRRKRDSFICSKQEDHAEAALYK

NLLHSKESSCSDSTNEYMDMKPGVSYVVPTKADKRRSVRIGSYIERDVTPAIMEDDELALDLED

LLSFSYQVAKGMAFLASKNCIHRDLAARNILLTHGRITKICDFGLARDIKNDSNYVVKGNARLP

VKWMAPESIFNCVYTFESDVWSYGIFLWELFSLGSSPYPGMPVDSKFYKMIKEGFRMLSPEHAP

AEMYDIMKTCWDADPLKRPTFKQIVQLIEKQISESTNHIYSNLANCSPNRQKPVVDHSVRINSV

GSTASSSQPLLVHDDV

22 . The hematopoietic stem cell or progenitor thereof expressing a CD117 polypeptide, the CD117 polypeptide comprising a sequence comprising 10, 20, 30, or 40 consecutive amino acids, wherein the sequence of consecutive amino acids comprises amino acid position 260, amino acid positions 261, and/or amino acid position 251 relative to the following amino acid sequence, wherein:

i) an amino acid corresponding to amino acid position 261 is substituted with a glycine,

ii) the amino acids corresponding to amino acid positions 260 and 261 are altered, and/or

iii) an amino acid corresponding to amino acid position 251 is altered relative to the following amino acid sequence in the sequence of consecutive amino acids, wherein the sequence of consecutive amino acids has at least 85% sequence identity to a fragment of the following amino acid sequence that has the same length as the sequence of consecutive amino acids, and wherein the CD117 polypeptide is capable of binding a stem cell factor (SCF) polypeptide: