Materials and methods for treatment of pain related disorders
The present application provides materials and methods for treating a patient with one or more conditions associated with SCN10A whether ex vivo or in vivo. In addition, the present application provides materials and methods for editing and/or modulating the expression of SCN10A gene in a cell by genome editing.
1. A method for editing the Sodium Channel, Voltage Gated, Type X Alpha Subunit (SCN10A) gene in a cell by genome editing comprising: introducing into the cell an S. pyogenes Cas9 endonuclease or one or more polynucleotide encoding the S. pyogenes Cas9 endonuclease and one or more gRNA or sgRNA or one or more nucleic acid encoding the one or more gRNA or sgRNA, wherein the one or more gRNA or sgRNA comprises a spacer sequence that targets a sequence within or near the SCN10A gene to effect one or more single-strand breaks (SSBs) or double-strand breaks (DSBs) within or near the SCN10A gene or SCN10A regulatory elements that results in one or more permanent insertions, deletions or mutations of at least one nucleotide within or near the SCN10A gene, thereby reducing or eliminating the expression or function of SCN10A gene products, wherein the sequence within or near the SCN10A gene that is targeted by the spacer sequence of the one or more gRNA or sgRNA is SEQ ID NO: 21893.
2. An in vivo method for treating a patient with an SCN10A related disorder comprising editing the SCN10A-gene in a cell of the patient according to the method of claim 1 .
3. The method of claim 1 , wherein the one or more polynucleotide encoding the S. pyogenes Cas9 endonuclease is DNA, wherein the DNA is codon optimized.
4. The method of claim 1 , wherein the one or more polynucleotide encoding the S. pyogenes Cas9 endonuclease is RNA, wherein the RNA is:
chemically modified, optionally in the coding region;
codon optimized; or
a combination thereof.
5. The method of claim 1 , wherein the one or more gRNA or sgRNA is:
chemically modified;
pre-complexed with the S. pyogenes Cas9 endonuclease; or
a combination thereof.
6. The method of claim 1 , wherein:
the S. pyogenes Cas9 endonuclease or the one or more polynucleotide encoding the S. pyogenes Cas9 endonuclease is formulated in a liposome or lipid nanoparticle which also comprises the one or more gRNA or sgRNA;
or
the S. pyogenes Cas9 endonuclease and the one or more gRNA or sgRNA are encoded by one or more AAV vector particle.
7. The method of claim 1 , wherein the S. pyogenes Cas9 endonuclease comprises one or more nuclear localization signals (NLSs).
8. The method of claim 1 , wherein the cell is a stem cell or somatic cell.
9. The method of claim 8 , wherein the stem cell is a pluripotent stem cell, an induced pluripotent stem cell (iPSC), or a mesenchymal stem cell.
10. The method of claim 2 , wherein the cell is a neuron of the peripheral nervous system.