US 11,898,179 B2
Suppression of pain by gene editing
Juan Pablo Maianti, Revere, MA (US); and David R. Liu, Lexington, MA (US)
Assigned to President and Fellows of Harvard College, Cambridge, MA (US)
Appl. No. 16/492,548
Filed by President and Fellows of Harvard College, Cambridge, MA (US)
PCT Filed Mar. 9, 2018, PCT No. PCT/US2018/021664
§ 371(c)(1), (2) Date Sep. 9, 2019,
PCT Pub. No. WO2018/165504, PCT Pub. Date Sep. 13, 2018.
Claims priority of provisional application 62/469,408, filed on Mar. 9, 2017.
Prior Publication US 2021/0115428 A1, Apr. 22, 2021
Int. Cl. C12N 9/78 (2006.01); A61K 9/00 (2006.01); A61K 31/7088 (2006.01); A61K 38/46 (2006.01); C12N 9/22 (2006.01); C12N 9/96 (2006.01); A61K 48/00 (2006.01); C12N 15/11 (2006.01); A61K 38/50 (2006.01)
CPC C12N 9/78 (2013.01) [A61K 9/0029 (2013.01); A61K 9/0053 (2013.01); A61K 31/7088 (2013.01); A61K 38/465 (2013.01); A61K 38/50 (2013.01); C12N 9/22 (2013.01); C12N 9/96 (2013.01); C12N 15/11 (2013.01); C12Y 305/04 (2013.01); A61K 48/00 (2013.01); C07K 2319/00 (2013.01); C12N 2310/20 (2017.05); C12N 2320/31 (2013.01); C12N 2320/32 (2013.01); C12N 2800/80 (2013.01)] 28 Claims
 
1. A method of editing a polynucleotide encoding an ion channel in a dorsal root ganglion (DRG) neuron, the method comprising contacting the ion channel-encoding polynucleotide with:
(i) a fusion protein comprising: (a) a guide nucleotide sequence-programmable DNA binding protein domain; and (b) a cytosine deaminase domain, wherein the fusion protein further comprises a uracil glycosylase inhibitor (UGI) domain; and
(ii) a guide nucleic acid molecule targeting the fusion protein of (i) to a target cytosine (C) base in the ion channel-encoding polynucleotide;
whereby the contacting results in deamination of the target C base by the fusion protein, resulting in a cytosine (C) to thymine (T) change in the ion channel-encoding polynucleotide; and
wherein the C to T change leads to a mutation in the ion channel that either introduces a premature stop codon in the ion channel-coding polynucleotide that leads to a truncated or non-functional ion channel or destabilizes ion-channel protein folding, or both.