Methods of making conjugated 4' desmethyl nucleoside analog compounds

• US 5,688,941 A
• Filed: 12/05/1996
• Issued: 11/18/1997
• Est. Priority Date: 07/27/1990
• Status: Expired due to Fees

• Current Assignee: ISIS Pharmaceuticals Incorporated
• Original Assignee: ISIS Pharmaceuticals Incorporated
• Inventors: Teng, Kelly, Cook, Phillip Dan
• Primary Examiner(s): Elliott, George C.
• Assistant Examiner(s): LARSON, THOMAS GEORGE
• Application Number:
  US08/760,848
• Time in Patent Office: 348 Days
• Field of Search: 536/22.1, 536/23.1, 536/24.3, 536/24.5, 536/25.3, 536/25.32, 536/25.6, 536/26.1, 536/26.6, 536/27.1, 435/6, 435/375, 514/44
• US Class Current: 536/25.3
• CPC Class Codes: A61K 31/70 Carbohydrates; Sugars; Derivatives thereof sorbitol A61K31/047
• CPC Class Codes: A61K 47/59 obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: C07D 405/04 directly linked by a ring-member-to-ring-member bond
• CPC Class Codes: C07D 405/14 containing three or more hetero rings
• CPC Class Codes: C07F 7/1804 Compounds having Si-O-C linkages Si-O-acyl linkages C07F7/1896
• CPC Class Codes: C07H 19/04 Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
• CPC Class Codes: C07H 19/06 Pyrimidine radicals
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/16 Purine radicals
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids

Abstract

Tetrahydrofuranyl compounds are provided that are functionalized to include pendant conjugate groups, and which are useful in diagnostic assays and as research reagents. Novel intermediates for the synthesis of the compounds are also provided.

Claims

1. A method for forming a 4'"'"'-desmethyl conjugated monomer having structure:

##STR13## comprising the steps of;

(a) providing a synthon having structure;

##STR14## and (b) contacting this synthon with an activated conjugating group;

the contacting being for a time and under reaction conditions sufficient to form the conjugated monomer;

wherein;

R_L is a group of formula;

space="preserve" listing-type="equation">R.sub.C -- Y!.sub.C --Z--

R_C is O-alkyl, O-alkenyl, O-alkynyl, a polyamine, a polyether, a steroid molecule, a reporter molecule, an aromatic lipophilic molecule, a non-aromatic lipophilic molecule, a reporter enzyme, a peptide, a protein, a water soluble vitamin, a lipid soluble vitamin, a carbohydrate, a terpene molecule, a phospholipid, an intercalator, a cell receptor binding molecule, a crosslinking agent, of a porphyrin;

Z is O, S or HN;

Y is a bivalent linker;

e is 0 or 1;

B_x is a nucleobase;

X is H, OH, O-alkyl, O-alkoxylalkyl, O-alkylamino, or F;

J is a leaving group;

Q is O, S, CH₂, CHF or CF₂ ; and

R_D is H, hydroxyl, an activated phosphorous group, a nucleoside, an activated nucleotide, a nucleotide, an oligonucleotide, an oligonucleoside or a protected derivative thereof.

2. The method of claim 1 wherein J is OH, SH, NH₂, trifluoromethylsulfonyl, methylsulfonyl, halogen, O-trichloroacetimidate, acyloxy, dialkyl phosphite, 2,4,6-trichlorophenyl, p-toluenesulfonyl, 4-dimethylaminoazobenzenesulfonyl or 5-dimethylaminonaphthalenesulfonyl.

3. The method of claim 1 wherein J is trifluoromethylsulfonyl, methylsulfonyl, halogen or acyloxy.

4. A method for forming a 4'"'"'-desmethyl conjugate oligomer having structure:

##STR15## comprising the steps of;

(a) providing a first synthon having structure;

##STR16## and (b) contacting the first synthon with a second synthon having structure;

##STR17## the contacting being for a time and under reaction conditions sufficient to form the conjugated oligomer;

wherein;

R_L is a group of formula;

space="preserve" listing-type="equation">R.sub.C -- Y!.sub.e --Z--

R_C is O-alkyl, O-alkynyl, a polyamine, a polyether, a steroid molecule, a reporter molecule, an aromatic lipophilic molecule, a non-aromatic lipophilic molecule, a reporter enzyme, a peptide, a protein, a water soluble vitamin, a lipid soluble vitamin, a carbohydrate, a terpene molecule, a phospholipid, an intercalator, a cell receptor binding molecule, a crosslinking agent, or a porphyrin;

Z is O, S or HN;

Y is a bivalent linker;

e is 0 or 1;

E is a phosphodiester, phosphorothioate, phosphorodithioate, phosphamidate, alkyl phosphonate or phosphotriester phosphorous linking group;

R_E is an activated phosphorous group;

W is H, a hydroxyl protecting group or a solid state support;

k is an integer from 0 to 50;

B_x is a nucleobase;

X is H, OH, O-alkyl, O-alkoxylalkyl, O-alkylamino, or F; and

Q is O, S, CH₂, CMF or CF₂.

5. A method for forming an oligonucleotide having a conjugate group thereon comprising the steps

(a) selecting an oligonucleotide having a 4'"'"'-desmethyl nucleoside at the 5'"'"'-terminal position of the oligonucleotide;

and

(b) linking a conjugate group to said 4'"'"'-desmethyl nucleoside, to form said oligonucleotide having said conjugate group thereon.