Purification of oligonucleotides
AssigneeIsis Pharmaceuticals, Inc.
InventorsAchim H. Krotz, Vasulinga T. Ravikumar
Life of this patent
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Abstract
Methods and compounds useful for the purification of oligonucleotides and their analogs are provided wherein the oligonucleotides are contaminated with at least one oligonucleotide having at least one abasic site by the formation of imines at the aldehyde moiety of the abasic site and subsequent separation based on extractions, precipitations or chromatography.
Claims59
A method of purifying an oligonucleotide from a mixture in solution, said mixture including said oligonucleotide and at least one contaminant comprising:
- treating said mixture with an amino reactive reagent for a time and under conditions effective to form a linkage of formula
- with each said contaminant;
- and
- separating said oligonucleotide from said linked contaminants;
- wherein the amino reactive reagent is selected from the group consisting of a polymeric support modified by attachment thereto of an amino reactive group Z and a compound of the formula
- wherein:
- x is from 0-20;
- y is from 0 to 5;
- n is from 0 to 150;
- and
- Z is —NH2, —NH—NH2, —O—NH2, —NH—C(O)—NH—NH2,
- or NH—C(S)—NH—NH2.
A method of purifying an oligonucleotide from a mixture, said mixture including said oligonucleotide and at least one contaminant, comprising:
- treating said mixture with a compound of formula I:
- wherein:
- x is from 0 to about 20,
- y is from 0 to about 5,
- n is from 0 to about 150,
- Z is a reactive nitrogenous moiety capable of reacting with an aldehyde to form a compound having a linkage of formula:
- for a time and under conditions effective to form said linkage with each contaminant;
- and
- separating said oligonucleotide from said linked contaminants.
A method for separating an abasic oligonucleotide from a mixture comprising said abasic oligonucleotide and one or more oligonucleotides having no abasic sites, said method comprising:
- providing a mixture containing at least one abasic oligonucleotide and one or more oligonucleotides having no abasic sites;
- treating said mixture with an amino reactive reagent for a time and under conditions effective to form a linkage of formula
- between said abasic oligonucleotide and said amino reactive reagent at said abasic site, thereby modifying the solubility of said abasic oligonucleotide;
- and
- separating said abasic oligonucleotide from said oligonucleotide or oligonucleotides having no abasic sites based on the solubility difference therebetween;
- wherein the amino reactive reagent is selected from the group consisting of a polymeric support modified by attachment thereto of an amino reactive group Z and a compound of the formula
- wherein:
- x is from 0-20;
- y is from 0 to 5;
- n is from 0 to 150;
- and
- Z is —NH2, NH—NH2, —O—NH2, —NH—C(O)—NH—NH2, or NH—C(S)—NH—NH2.
A compound of Formula I,
- wherein:
- x is from 0 to 20,
- y is from 0 to 5,
- n is from 0 to 150;
- and
- Z is —NH2, —NHNH2, —ONH2, —NHC(O)NHNH2 or —NHC(S)NHNH2.
Dependencies are inferred from the claim text.