Benzazolides and their employment in phosphite ester oligonucleotide synthesis processes

Claim 2

A benzotriazolide of the formula ##STR12## wherein A, B, C, and D are independently selected from among hydrogen, chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulfonato amino and cyano, R is hydrogen or a base labile phosphite protecting group, P g is a selectively removable protecting group selected from levulinyl and acid labile protecting groups and B* is a base selected from 1-thyminyl, 1-(N-protected)cytosinyl, 9-(N-protected)adeninyl and 9-(N-protected)guaninyl.

Claim 3

The benzotriazolide of claim 1 wherein B, C and D are hydrogens.

Claim 4

The benzotriazolide of claim 2 wherein R is a base labile phosphite protecting group.

Claim 5

The benzotriazolide of claim 3 wherein R is selected from among 1 to 4 carbon alkyls, 6 to 12 carbon aryls, alkaryls and aralkyls and halo-substituted 6 to 12 carbon aryls, alkaryls and aralkyls.

Claim 6

The benzotriazolide of claim 1 wherein the N-protected base is protected with a group selected from among benzoyl, isobutyrl and anisoyl.

Claim 7

A phosphitylated oligonucleotide of the formula: ##STR13## wherein X is N or CH, A, B, C and D are each selected from hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulonato amino and cyano, R is a base labile phosphite protecting group, P g is a protecting group selected from levulinyl and acid-labile protecting groups and B* and B*' are independently selected from 1-thyminyl, 1-(N-protected)cytosinyl, 9-(N-protected)adeninyl and 9-(N-protected)guaninyl, and n is an integer 1 or greater.

Claim 8

A phosphitylated oligonucleotide of the formula: ##STR13## wherein X is N or CH, A, B, C and D are each selected from hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulonato amino and cyano, R is a base labile phosphite protecting group, P g is a protecting group selected from levulinyl and acid-labile protecting groups and B* and B*' are independently selected from 1-thyminyl, 1-(N-protected)cytosinyl, 9-(N-protected)adeninyl and 9-(N-protected)guaninyl, and n is an integer 1 or greater.

Claim 9

In a process of oligonucleotide synthesis wherein a first nucleoside optionally affixed to a solid support is reacted with a phosphitylated second nucleoside or nucleotide under nucleotide coupling conditions in the presence of an activating agent, the improvement comprising employing as said activating agent a benzotriazole of the formula ##STR14## wherein A, B, C and D are independently selected from among hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulfonato amino and cyano, and Y is hydrogen or a leaving group.

Claim 10

In a process of oligonucleotide synthesis wherein a first nucleoside optionally affixed to a solid support is reacted with a phosphitylated second nucleoside or nucleotide under nucleotide coupling conditions in the presence of an activating agent, the improvement comprising employing as said activating agent a benzotriazole of the formula ##STR14## wherein A, B, C and D are independently selected from among hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulfonato amino and cyano, and Y is hydrogen or a leaving group.

Claim 11

The process of claim 7 being further characterized as a flow process in which the first nucleotide is affixed to a solid support in a reaction zone and controlled amounts of solutions of said second nucleoside or nucleotide and said activating agent are serially fed for controlled periods into said reaction zone for a reaction period.

Claim 12

The process of claim 8 wherein the controlled periods are for from 50 microseconds to 5 seconds and the reaction period is at least 50 times a controlled period.

Claim 13

The process of claim 9 being further characterized as a flow process in which the first nucleotide is affixed to a solid support and controlled amounts of solutions of said second nucleoside or nucleotide and said activating agent are passed over said solid support immediately after being admixed together.

Claim 14

A process for synthesizing an oligonucleotide comprising (a) reacting a nucleotide or nucleoside with a phosphinedibenzazolide of the formula ##STR15## wherein X is N or CH, A, B, C and D are independently selected from hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulfonato amino and cyano, and R is a base-labile phosphite-protecting group, to yield a phosphitylated nucleotide, and (b) coupling said phosphitylated nucleotide with a second nucleotide or nucleoside.

Claim 15

A process for synthesizing an oligonucleotide comprising (a) reacting a nucleotide or nucleoside with a phosphinedibenzazolide of the formula ##STR15## wherein X is N or CH, A, B, C and D are independently selected from hydrogen and chloro, bromo, iodo, fluoro, lower alkyls of 1 to 4 carbon atoms, substituted lower alkyls, benzyl, phenyl, substituted benzyl, substituted phenyl, lower alkoxies of 1 to 4 carbon atoms, nitro, nitroso, sulfonato amino and cyano, and R is a base-labile phosphite-protecting group, to yield a phosphitylated nucleotide, and (b) coupling said phosphitylated nucleotide with a second nucleotide or nucleoside.

Claim 16

A process for synthesizing an oligonucleotide of claim 11 wherein X is N.

Claim 17

A process for synthesizing an oligonucleotide of claim 11 wherein X is CH.