Oligonucleotides, reagents, and preparation thereof

A nucleotide or oligonucleotide represented by Formula III or IIIP,

• or a salt thereof, wherein
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is optionally protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently H, C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A nucleotide or oligonucleotide represented by Formula III′ or HIP′,

• or a salt thereof, wherein
• Q is a hydroxyl protecting group;
• is a nucleobase comprising a NH2 group which is modified by Z;
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is optionally protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently H, C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V),

• or a salt thereof, comprising the steps of:
• 1) deprotecting a compound of formula (VA):
• or a salt thereof, to form a compound of formula (VB):
• or a salt thereof;
• 2) reacting the compound of formula (VB), or a salt thereof, with a compound of formula (VC ):
• or a salt thereof, to form a compound of formula (VD),
• or a salt thereof;
• 3) sulfurizing or oxidizing the compound of formula (VD), or a salt thereof, with a sulfurization or oxidation agent to form a compound of formula (VE):
• or a salt thereof;
• 4) deprotecting the compound of formula (VE), or a salt thereof to form a compound of formula (VF):
• or a salt thereof;
• 5) when q is equal or greater than 2, starting with the compound of formula (VF), repeating steps 2), 3) and 4) for q-2 times, followed by steps 2) and 3) to yield the fragment of formula (V), or a salt thereof, wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V′),

• or a salt thereof, comprising the steps of:
• 1) deprotecting a compound of formula (VA):
• or a salt thereof, to form a compound of formula (VB):
• or a salt thereof;
• 2) reacting the compound of formula (VB), or a salt thereof, with a compound of formula (VC′):
• or a salt thereof, to form a compound of formula (VD′),
• or a salt thereof;
• 3) sulfurizing or oxidizing the compound of formula (VD′), or a salt thereof, with a sulfurization or oxidation agent to form a compound of formula (VE′):
• or a salt thereof;
• 4) deprotecting the compound of formula (VE′), or a salt thereof to form a compound of formula (VF′):
• or a salt thereof;
• 5) when q is equal or greater than 2, starting with the compound of formula (VF′), repeating steps 2), 3) and 4) for q-2 times, followed by steps 2) and 3) to yield the fragment of formula (V′), or a salt thereof, wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V-C1) or (V-C2),

• or a salt thereof, comprising the steps of:
• 1) reacting the compound of formula (VB),
• or a salt thereof, with a compound of formula (V-CR1) or (V-CR2),
• or a salt thereof, and a base, to form a compound of formula (V-C1) or (V-C2), wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S, provided when X is S, the phosphorothiolate group has S-configuration, R-configuration or a mixture thereof (e.g., a racemic mixture);
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V-C1) or (V-C2),

• or a salt thereof, comprising the steps of:
• 1) reacting the compound of formula (VB),
• or a salt thereof, with a reagent of formula (VR1) or (VR2),
• to form a compound of formula (V-CR3) or (V-CR4),
• or a salt thereof;
• 2) reacting the compound of formula (V-CR3) or (V-CR4), or a salt thereof, with a compound of formula (VG):
• or a salt thereof, and a base, to form a compound of formula (V-C1) or (V-C2), wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S, provided when X is S, the phosphorothiolate group has S-configuration, R-configuration or a mixture thereof (e.g., a racemic mixture);
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (VBZ),

• or a salt thereof, comprising the steps of:
• 1) reacting the compound of formula (VBZ-1),
• or a salt thereof, with a compound of formula (VBZ-2):
• or a salt thereof, to form a compound of formula (VBZ-3),
• or a salt thereof;
• 2) sulfurizing or oxidizing the compound of formula (VBZ-3), or a salt thereof, with a sulfurization or oxidation agent to form a compound of formula (VBZ), or a salt thereof;
• wherein:
• Q is a hydroxyl protecting group;
• is a nucleobase comprising a NH2 group which is modified by Z;
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V),

• or a salt thereof, comprising the steps of:
• c) coupling a nucleotide of formula (V-1):
• or a salt thereof, with an oligonucleotide fragment of formula (V-2):
• or a salt thereof, in a solution to form an oligonucleotide fragment of formula (V-3),
• or a salt thereof;
• and
• d) sulfurizing or oxidizing the oligonucleotide of formula (V-3), or a salt thereof, to form an oligonucleotide of formula (V):
• or a salt thereof;
• wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide fragment of formula (V*),

• or a salt thereof, comprising the steps of:
• a) coupling a nucleotide of formula (V-1):
• or a salt thereof, with an oligonucleotide fragment of formula (V-2′):
• in a solution to form an oligonucleotide fragment of formula (V-3′),
• or a salt thereof;
• and
• b) sulfurizing or oxidizing the oligonucleotide of formula (V-3′), or a salt thereof, to form the oligonucleotide of formula (V*) or a salt thereof;
• wherein:
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently H, C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• q is an integer from 1 to 20;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide of formula (VI) or (VI-1),

• or a salt thereof, comprising
• a) coupling an oligonucleotide fragment of formula (F1) or (F1-1):
• or a salt thereof, with an oligonucleotide fragment of formula (F2):
• or a salt thereof, in a solution to form an oligonucleotide fragment of formula (F3) or (F3-1),
• or a salt thereof;
• and
• b) sulfurizing or oxidizing the oligonucleotide fragment of formula (F3) or (F3-1), or a salt thereof, to form the oligonucleotide of formula (VI) or (VI-1) or a salt thereof,
• wherein:
• Q is a hydroxyl protecting group;
• is a nucleobase comprising a NH2 group which is modified by Z;
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• p is an integer from 2 to 20;
• o is an integer from 1 to 200;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.

A process for preparing an oligonucleotide of formula (VI′) or (VI′-1),

• or a salt thereof, comprising
• a) coupling an oligonucleotide fragment of formula (F1) or (F1-1):
• or a salt thereof, with an oligonucleotide fragment of formula (F2′):
• or a salt thereof, in a solution to form an oligonucleotide fragment of formula (F3′) or (F3′-1),
• or a salt thereof;
• and
• b) sulfurizing or oxidizing the oligonucleotide fragment of formula (F3′) or (F3′-1), or a salt thereof, to form the oligonucleotide of formula (VI′) or (VI′-1) or a salt thereof,
• wherein:
• Q is a hydroxyl protecting group;
• is a nucleobase comprising a NH2 group which is modified by Z;
• R31, for each occurrence, is independently a nucleobase, wherein the NH2 of the nucleobase, if present, is protected by an amine protecting group;
• R32, for each occurrence, is independently selected from the group consisting of H, halo, OH, and C1-6alkoxy optionally substituted with C1-6alkoxy;
• wherein the OH group is optionally protected by a hydroxyl protecting group;
• R34, for each occurrence, is independently H or forms a ring with the alkoxy group of R32;
• R35 is a hydroxyl protecting group;
• R36, for each occurrence, is independently C1-6alkyl group, C2-6alkenyl group, phenyl or benzyl group, each of which is optionally substituted with —CN, —NO2 or halogen;
• or
• R36 is
• R37a and R37b are independently C1-6alkyl;
• p is an integer from 2 to 20;
• o is an integer from 1 to 200;
• X, for each occurrence, is independently O or S;
• Z is a group represented by Formula I* or B*,
• wherein
• —#represents the point of attachment for Z;
• one of A1, A2 and A3 is YA and the others are H;
• is a single bond or a double bond;
• YA is Y—(CH2)a1CH2O(CH2)a2—, wherein a1 and a2 are each independently 0 or an integer from 1 to 10;
• ring A is phenyl, 8- to 10-membered bicyclic aryl, 5- to 6-membered heteroaryl having 1 to 3 heteroatoms independently selected from oxygen, nitrogen, and sulfur, or 7- to 10-membered bicyclic heteroaryl having 1-4 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• Y is H, halogen, OR1A, NR2AR3A, SR4A, CR5AR6AR7A, or a hydrophobic group comprising one or more aliphatic hydrocarbon group having 10 or more carbon atoms;
• wherein R1A, R2A, R3A, R4A, R5A, R6A, and R7A are each independently C1-6alkyl, C1-6alkenyl, C1-6alkynyl, phenyl, OR8A, —OC(O)R8A, —C(O)OR8A, NR8AR9A, —NR8ACOR9A, —CONR8AR9A, 3- to 7-membered saturated or partially unsaturated monocyclic carbocyclyl, or 3- to 7-membered saturated or partially unsaturated monocyclic heterocyclyl having 1-2 heteroatoms independently selected from oxygen, nitrogen, and sulfur;
• wherein R8A and R9A, for each occurrence, is independently H or C1-6alkyl;
• P1 is NO2 or a silyl hydroxyl protecting group;
• R1 and R2 are independently H, C1-6alkyl, or phenyl;
• wherein C1-6alkyl and phenyl are optionally substituted by 1-3 R3;
• R3 is C1-30alkoxy;
• e is an integer from 0 to 6;
• and
• f is an integer from 0 to 6.