Oligonucleotide compositions and methods of making the same

1. A method of synthesizing a polynucleotide of the formula:

wherein:

each B is independently a purine, a protected purine, a pyrimidine or a protected pyrimidine, or an analog thereof;

each X is independently oxygen or sulfur;

each R 3 is hydrogen, fluoro, or hydroxyl, an alkoxy, a substituted alkoxy or a protected hydroxyl;

L is an optional linker;

Z is H, a lipid, a support, a carrier, an oligonucleotide, a PEG, a polypeptide, a detectable label, or a tag;

R 6 is amino, hydroxyl, a protected amino, a protected hydroxy, —O-L-Z or NH-L-Z;

R is hydrogen, an alkyl, a substituted alkyl, an aryl, a substituted aryl, or a phosphate protecting group; and

n is an integer of 1 to 1000; or a salt thereof;

the method comprising the steps of:

(a) deprotecting a protected 3′ amino group of a terminal nucleoside attached to a solid phase support, said deprotecting forming a free 3′ amino group;

(b) reacting the free 3′ amino group with either:

(i) a 3′-protected amino-dinucleotide phosphoramidate-5′-phosphoramidite dimer; or

(ii) a 3′-protected aminonucleoside-5′-phosphoramidite monomer;

in the presence of a nucleophilic catalyst to form an internucleoside N3′→P5′ phosphoramidite linkage;

(c) oxidizing the linkage; and

(d) repeating steps (a) through (c) until the polynucleotide is synthesized, wherein the repeating steps (a) through (c) comprises performing step (b)(i) at least once.

2. The method of claim 1 , wherein oxidizing the linkage comprises sulfurization to produce a thiophosphoramidate linkage.

3. The method of claim 1 , wherein oxidizing the linkage produces an oxophosphoramidate linkage.

4. The method of claim 1 , wherein the polynucleotide comprises a sequence of nucleoside subunits complementary to the RNA component of human telomerase, and wherein at least two of the nucleoside subunits are joined by a N3′→P5′ phosphoramidate inter-subunit linkage.

5. The method of claim 4 , wherein the N3′→P5′ phosphoramidate inter-subunit linkage is a N3′→P5′ thiophosphoramidate inter-subunit linkage having the structure:

3′-NH—P(S)(OR)—O-5′

wherein R is selected from the group consisting of hydrogen, an alkyl, a substituted alkyl, an aryl, a substituted aryl and a phosphate protecting group, or a salt thereof.

6. The method of claim 1 , wherein the polynucleotide comprises the sequence TAGGGTTAGACAA.

7. The method of claim 6 , wherein all of the internucleotide inter-subunit linkages of the TAGGGTTAGACAA sequence are N3′→ P5′ phosphoramidate inter-subunit linkages.

8. The method of claim 7 , wherein the polynucleotide has the structure:

or a salt thereof;

wherein “nps” represents a thiophosphoramidate linkage —NH—P(═O)(SH)—O—, connecting the 3′-carbon of one nucleoside to the 5′-carbon of the adjacent nucleoside.

9. The method of claim 1 , wherein the polynucleotide has the structure:

wherein each M x+ is independently hydrogen or a counterion of a pharmaceutically acceptable salt, each x of M x+ is independently 1, 2 or 3 and n is an integer from 5 to 13.

10. The method of claim 9 , wherein the polynucleotide has the structure:

11. The method of claim 6 , wherein the C11 nucleotide residue of the TAGGGTTAGACAA sequence derives from a 3′-protected aminonucleoside-5′-phosphoramidite monomer.

12. The method of claim 6 , wherein the method comprises sequential coupling of the following 3′-protected amino-dinucleotide thiophosphoramidate-5′-phosphoramidite dimers TA, GG, GT, TA, GA and AA and 3′-protected aminonucleoside-5′-phosphoramidite monomer C to the solid phase support.