2',5' PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES AND ANTIVIRAL USES THEREOF

Abstract: Optically active antiviral compounds having formula (I), wherein m is 0, 1, 2 or 3; n and q are selected from the group of 0 and 1, provided that n and q may not both be zero; R, R1 and R2 are independently of each other selected from the group consisting of oxygen and sulfur, provided that all R, R1 and R2, may not be oxygen, and further provided that all R, R1, and R2 may not be sulfur. The compounds possess increased antiviral activity and/or metabolic stability.

Description: 2 ' .5 ' PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES AND ANTIVIRAL USES THEREOF Field of the Invention This invention relates to synthetic analogues of naturally occurring antiviral 2 ', 5 ' -oligoadenylates wherein at least one of the internucleotide phosphodiester linkages is replaced with optically active phosphorothioate groups. The compounds with selected internucleotide phosphodiester linkages have antiviral activity and increased metabolic stability. Background of the Invention The full nomenclature of the subject matter of the present invention involves lengthy terms. It is customary for those skilled in the art to abbreviate oligoadenylate analogues and related terms in a manner well-known to the art. These general and customary abbreviations are set forth herein below and may be utilized in the text of this specification. Abbreviations : 2-5A, 2 ', 5 ' -oligoadenylate or p 3 A_: Oligomer of adenylic acid with 2 ', 5 ' -phosphodiester linkages and a 5 ' -terminal triphosphate group. A 2 , A 3 and A 4 : Dimer, trimer and tetramer of adenylic acid with 2 ', 5 ' -phosphodiester linkages. pA 3 , ppA 3 (or p 2 A 3 ) , pppA 3 (or p 3 A 3 ) : 5'- terminal mono-, di- and triphosphates of A 3 . pA 4 , ppA 4 (or p 2 A 4 ) , pppA 4 (or p 3 A 4 ) : 5 '-terminal mono-, di- and triphosphates of A 4 . ApA: Dimer of adenylic acid with 2'-5'- phosphodiester linkage. Ap * A: Dimer of adenylic acid with 2 '-5 '-phos¬ phorothioate linkage. PR: The R stereoconfiguration about a chiral phosphorous atom in a phosphorothioate internucleotide linkage. PS: The S stereoconfiguration about a chiral phosphorous atom in a phosphorothioate internucleotide linkage. A Rp * ApA: (PR) -P-thioadenylyl- (2 ' , 5 ' ) -adenylyl- (2 ' , 5 ' ) -adenosine . A Sp * ApA: (PS) -P-thioadenylyl- (2 ', 5 ') -adenylyl- (2 ' , 5 ' ) -adenosine . ApA j 'A: Adenylyl- (2 ' , 5 ' ) - (PR) -P-thioadenylyl- (2 ' , 5 ' ) -adenosine . ApA Sp * A: Adenylyl- (2 ' , 5 ' ) - (PS) -P-thioadenylyl- (2 ' , 5 ' ) -adenosine. pA Rp * ApA, ppA Rp * ApA, pppA Rp * ApA, pA Sp * ApA, ppA Sp * ApA, pppA Sp * ApA, pApA Rp * A, ppApA Rp * A, pppApA Rp * A, pApA Sp * A, ppApA Sp * A, pppApA Sp * A: 5 '-mono-, di- and triphosphates of A Rp * ApA, A Sp * ApA, ApA Rp * A, and ApA Sp * A. A Rp * ApApA: (PR) -P-thioadenylyl- (2 ' , 5 ' ) -adenyl¬ yl- (2 ' ,5' ) -adenylyl- (2 ' ,5' ) -adenosine. A sp * ApApA: (PS) -P-thioadenylyl- (2 ' , 5 ' ) -adenyl¬ yl- (2 ' ,5 ' ) -adenylyl- (2 ' , 5 ' ) -adenosine. Id.l ApA j ^ApA: Adenylyl- (2 ' , 5 ' ) - (PR) -P-thioadenyl- yl- (2' ,5' ) -adenylyl- (2 • ,5' ) -adenosine. ApA Sp * ApA: Adenylyl- (2 ' , 5 ' ) - (PS) -P-thioadenyl¬ yl- (2 ', 5 ' ) -adenylyl- (2 ' , 5 ' ) -adenosine . ApApAκ p * A: Adenylyl- (2 ' , 5 ' ) - (PR) -P-thioadenyl- yl- (2 ' ,5' ) -adenylyl- (2 ' , 5 ' ) -adenosine. ApApA Sp * A: Adenylyl- (2 ' , 5 ' ) -adenylyl- (2 ' , 5 ' ) - (PS) -P-thioadenylyl- (2 ' , 5 ' ) -adenosine . pA Rp * ApApA, ppA Rp * ApApA, pppA Rp * ApApA, pA Sp * ApApA, ppA Sp * ApApA, pppA Sp * ApApA, pApA Rp * ApA, ppApA Rp "ApA, pppApA Rp * ApA, pApA Sp * ApA, ppApA Sp * ApA, pppApA Sp * ApA, pApApA Rp * A, ppApApA Rp * A, pppApApA Rp * A, pApApA^'A, ppApApA Sp * A, pppApApA Sp * A: 5 '-mono-, di- and triphosphates of the above tetramers. bz: benzoyl ce : cyanoethyl CFS: Chronic Fatigue Syndrome DEAE: 2- (diethylamino) ethyl DBU: 1.8 diazabicyclo [5.3.0] undec-7-enc HIV: Human Immunodeficiency Virus MeOTr: monomethoxytrityl M.O.I: multiplicity of infection mRNA: Messenger RNA npe 2- (4-niytophenyl) ethyl PBL: Peripheral blood lymphocytes pCp: Cytidine 3 ' -5 ' -bisphosphate (PS) - ATP-alpha-S: Adenosine 5'0-(PS)-(l- thiotriphosphate) . RNase L 2-5A-dependent endoribonuclease . rRNA: Ribosomal RNA RT: Reverse transcriptase SCP: Specific cleavage products tbds (tert-butyl) dimethylsilyl Tris tris (hydroxymethyl) aminomethane tRNA: Transfer RNA It is generally regarded that activation of RNase L by 2-5A is key to the antiviral defense mecha¬ nisms. Interferon induces transcription of the enzyme 2-5A synthetase which produces 2', 5' linked oligoadeny- lates upon activation of double-stranded RNA. Previously, the only known biochemical effect of 2-5A is activation of RNase L. This enzyme hydrolyses mRNA and rRNA, thereby resulting in inhibition of protein synthesis. The activation of RNase L is transient unless 2-5A is continuously synthesized, since 2-5A is rapidly degraded. RNase L activation thus plays a critical role in inhibiting replication, and therefore in defending against infection by viruses. A correlation has also been established between 2-5A metabolism and the growth cycle of HIV-1, i.e. , high levels of 2-5A and activated RNase L correlate with failure of infected cells to release HIV-1, Schroder et al.. , J. Biol . Chem. 264: 5669-5673 (1989) . Conversely, when the intracellular 2-5A pool decreases, RNase L can not be activated and HIV -1 production increases. A role for 2-5A cores as inhibitors of HIV-1 replication has been established with reports that 2-5A trimer and tetramer cores, 5 ' -monophosphates and 5 ' -triphosphates inhibit HIV-l reverse transcriptase/primer complex formation, Montefiori et a_l . , Proc. Natl . Acad. Sci . USA 86 : 7191-7194 (1989) ; Mϋller et al., Biochemistry 30 : 2027-2033 (1991) ; Sobol et . al., Biochemistry 32 : 1211- 12118 (1993) . The introduction of the phosphorothioate group in the 2 ', 5 ' -internucleotide linkages of 2-5A, induces metabolic stability greater than authentic 2--5A and resulted in the first 2-5A cores (i.e. 2-5A lacking 5'- phosphate moieties) able to activate RNase L (Kariko et al . Biochemistry 26 : 7136-7142 (1987) ; Charachor. et al . Biochemistry 29: 2550-2556 (1990)) . Further, RNase L is a functionally stereoselective enzyme and 2-5A trimers and tetramers having at least one of the internucleotide phosphorothioate 2 ', 5 ' -linkages of the PS configuration have greatly enhanced metabolic stability. The chemical synthesis of the fully resolved 2', 5' -phosphorothioate adenylate trimer and tetramer cores has been reported, Suhadolnik et al . , U.S. Patent No. 4,924,624. Prepara- tion of the stereoisomers via enzymatic synthesis is not possible due to the sterospecificity of 2-5A synthetase for the substrate (PS) -ATP-alpha-S, which yields trimer and tetramer products of the PR configuration exclu¬ sively. Further, while Lebleu et al., U.S. Pat. No. 4,981,957, discloses the enzymatic synthesis of a phosphorothioate-substituted derivative of 2 ',5' oligoadenylate, the compounds disclosed are not stereo- specific. Summary of the Invention Compounds of the present invention useful in inhibiting viral infections in plants and mammals have increased metabolic stability and/or antiviral activity. The compounds and the water-soluble salts thereof are of the formula wherein m is zero, 1, 2 or 3 ; n and q are selected from the group of zero and 1, provided that n and q may not both be zero; and R, R α and R 2 are independently selected from the group of oxygen and sulfur, provided that all R, R x and R 2 , may not be oxygen, and further provided that all R, R x and R 2 may not be sulfur. The invention also comprises a method of in¬ hibiting viral infection in mammals or plants by admin¬ istering an antivirally effective amount of a compound according to the above formula, or a water-soluble salt thereof, and antiviral compositions containing such compounds with a carrier. Compounds according to the formula wherein n is 1 and q is 1 may be utilized to form oligoadenylate conjugates with the macromolecular carrier poly(L-lysine) for intracellular transport . Such poly(L-lysine) /2 ' , 5 ' - phosphorothioate/phosphodiesteroligoadenylate conjugates have the formula NH 2 NH 2 I I (CH 2 ) 4 (CH 2 ) 4 I I NH 2 CHCO-R q -NHCHCOOH wherein q is an integer from about 60 to about 70, and R is randomly R' or . From about five to about ten of the R groups comprise R' . R' has the following formula wherein m is 0, 1, 2 or 3; and where each R 3 , R 4 , or R 5 are independently selected from the group of oxygen and sulfur; provided that all R 3 , R 4 or R 5 may not be oxygen; and further provided that all R 3 , R 4 or R 5 , may not be sulfur. NH, I (CH 2 ) 4 I -NH— CH— CO— Preferably, at least one of the internucleotide phosphorothioate groups of the poly(L-lysine) /2 ', 5 ' -phosphorothioate/phospho- diester oligoadenylate conjugates is of the PR configu¬ ration. Description of the Figures Figure 1A represents the results of a radiobinding assay indicating the ability of 2 ' , 5 ' -phosphorothioate/- phosphodiester trimer core 2-5A derivatives to activate partially-purified RNase L from L929 cell extracts to hydrolyze the substrate poly(U) [ 32 P]pCp, but not poly(C) . Activation of RNase L was determined by the conversion of poly(I) [ 32 P] pCp to acid soluble fragments. 100% represents 25,000 dpm of poly(U) [ 32 P]pCp bound to glass fiber filters. 2-5A oligomer (phosphodiester internucleotide linkage) is included for comparison. The curves are labeled as follows: p 3 A 3 (•) ; A 3 (♦) ; A Rp * ApA (D) ; A Sp * ApA (■) ; ApA Rp * A (Δ) ; and ApA Sp 'A (A) . Figure IB represents the results of a radiobinding assay performed according to the method of Figure 1A for 2 ' , 5 ' -phosphorothioate/phosphodiestertrimer5 ' -monophos- phate 2-5A derivatives. The curves are labeled as follows: p 3 A 3 (•) ; pA 3 (O) ; pA Rp * ApA (□) ; pA Sp * ApA (■) ; pApA Rp * A (Δ) ; and pApA Sp * A (A) . Figure 1C represents the results of a radiobinding assay performed according to the method of Figure 1A for 2 ' , 5 ' -phosphorothioate/phosphodiester tetramer core _-5A derivatives. The curves are labeled as follows: p 3 A 4 (•) ; A Rp * ApApA (.) ; A Sp * ApApA (τ) ; ApA Rp * ApA (O) ; ApA Sp * ApA (■) ; ApApA Rp * A (Δ) ; and ApApA Sp * A (A) . Figure ID represents the results of a radiobinding assay performed according to the method of Figure 1A for 2 ' , 5 ' -phosphorothioate/phosphodiester tetramer 5'- monophosphate 2-5A derivatives. The curves are labeled as follows: p 3 A 4 (•) ; pA 4 (O) ; pA Rp * ApApA (v) ; pA Sp * ApApA (▼) ; pApA Rp * ApA (□) ; pApA Sp * ApA (■) ; pApApA Rp * A (Δ) ; and pApApA Sp * A (A) . Figure 2A represents the results of a ribosomal RNA cleavage assay with 2 ' , 5 ' -phosphorothioate/phosphodiester trimer core 2-5A derivatives. The procedure was performed according to the method of Kariko et a_l . , Biochemistry 26 : 7127-7135 (1987) . 2-5A oligomer (phosphodiester internucleotide linkage) is also included for comparison. L929 cell extracts were incubated in the absence (lane 1) or presence of p 3 A 3 at 10 "8 M (lane 2) , A Rp * ApA at 10 "6 M (lane 3) , A Sp * ApA at 10 "6 M (lane 4) , ApA Rp * A at 10 '6 M (lane 5) , ApA Sp * A at 10 "6 M (lane 6) or A 3 at 10 "6 M (lane 7) . The positions of 28S and 18S rRNA are shown; the arrows indicate the positions of the well- characterized specific cleavage products (SCP) of RNase L. Figure 2B represents the results of a ribosomal RNA cleavage assay performed according to the method of Figure 2A with 2 ' , 5 ' -phosphorothioate/phosphodiester trimer 5 ' -monophosphate derivatives. L929 cell extracts were incubated in the absence (lane 1) or presence of p 3 A 3 at 2 x 10 "9 M (lane 2) , pA 3 at 10 "6 M (lane 3) , pA Rp * ApA at 10 ~7 M (lane 4) , pA Sp * ApA at 10 "7 M (lane 5) , pApA Rp * A at 2 x 1M 9 M (Lane 6) , pApA Sp * A at 10 "7 M (lane 7) . Figure 3A represents the results of a ribosomal RNA cleavage assay performed according to the method of Figure 2A with 2 ' , 5 ' -phosphorothioate/phosphodiester tetramer core 2-5A derivatives . 2-5A oligomer (phosphod- iester internucleotide linkage) is also included for comparison. L929 cell extracts were incubated in the absence (lane 1) or presence of p 3 A 4 at 10 "8 M (lane 2) , A 4 at 10 "5 M (lane 3) , A Rp * ApApA at 10 "5 M (lane 4) , A Sp "ApApA at 10 -5 M (lane 5) , ApA Rp * ApA at 10 "5 M (lane 6) , ApA Sp * ApA at 10 "5 M (lane 7) , ApApA Rp * A at 10 "5 M (lane 8) or ApApA Sp * A at 10 "5 M (lane 9) . Figure 3B represents the results of a ribosomal RNA cleavage assay performed according to the method of Figure 2A with 2 ' , 5 ' -phosphorothioate/phosphodiester tetramer 5 ' -monophosphate derivatives. L929 cell extracts were incubated in the absence (lane 1) or presence of p 3 A 4 at 10-8 M (lane 2) , pA 4 at 10 "6 M (lane 3) , pA Rp * ApApA at 10-6 M (lane 4) , pA Sp * ApApA at 10 "6 M (lane 5) , pApA Rp * ApA at 10 "8 M (lane 6) , pApA Sp * ApA at 10 "5 M (lane 7) , pApApA Rp * A at 10 "7 M (lane 8) or pApApA Sp "A at 10 "7 M (lane 9) . Figure 4A represents the results of a ribosomal cleavage assay performed according to the method of Figure 2A, indicating the inhibition of activation of RNase L by pApA Sp * A in L929 cell extracts and by partial¬ ly-purified RNase L. L929 cell extracts were incubated in the presence of p 3 A 3 at 10 "9 M (lanes 1 and 2) , p 3 A, at 10 "8 M (lanes 3 and 4) pApA Rp * A, at 10 "9 M (lanes 5 and 6) , pApA Rp * A at 10 "8 M (lanes 7 and 8) , and pApA Sp * A at 10 "6 M (lanes 2 , 4 , 6 and 8) . Figure 4B represents the results of a radiooinding assay indicating the activation of RNase L partially- purified from L929 cell extracts. Activation of RNase L was determined by the conversion of poly(U) [ 32 P] pCp to acid-soluble fragments by immobilization on 2-5A 4 core-cellulose. 100% represents 25,000 dpm of poly(U) [32P]pCp bound to glass fiber filters. 2-5 A oligomer (phosphodiester internucleotide linkage) is also included for comparison. The curves are labeled as follows: p 3 A 3 (•) ; p 3 A 3 + pApA Sp * A at 10 "6 M (O) . Figure 5A represents the results of an assay performed according to the method of Henderson et aj.. , Virology 182 : 186-198 (1991) , indicating the inhibition of HIV-1 (IIIB) -induced syncytia formation by adenosine, 2-5A trimer or tetramer core or 2 ' , 5 ' - phosphorothioate/- phosphodiester trimer derivatives: A Rp * ApA, A Sp * ApA, ApA Rp * A, ApA Sp * A, A Rp * ApApA, A Sp * ApApA, ApA Rp * ApA, ApA Sp * ApA, ApApA Rp * A, ApApA Sp * A. Figure 5B is the results of an assay performed according to the method of Figure 5A with a 2 ' , 5 ' - phosphorothioate/phosphodiester tetramer derivative of 2-5A core. Detailed Description of the Invention The individual nucleotide linkages of the trimer and tetramer derivatives of 2 ', 5 ' -oligoadenylate (2-5A) were stereochemically modified via phosphorothiate substitution by phosphotriester and phosphoramidite chemical synthesis. The approach described herein utilizes fully protected monomeric building blocks (Schemes 1 and 2) below, which can be individually manipulated. The protecting groups remain in place during the chemical synthesis of the oligonucleotide chain and are removed at the end of the sequence by ,_- elimination. The phosphorothioate/phosphodiester trimer cores, A Rp * ApA .10 . , A Sp * ApA i , ApA Rp * A .23 . , and ApA Sp * A 24/ were che- mically synthesized and separated by preparative thin layer chromatography on silica gel, deblocked and purified by applying the residue on a DEAE Sephadex column. The four trimer cores are prepared from phosphoramidite intermediates 4 . and JL5 . . The synthesis relies on separation of fully resolved protected inter¬ mediates, 8 . , 3_, 2 and . 22 . followed by removal of all blocking groups to yield the individually substituted 2 ' , 5 ' -phosphorothioate/phosphodiester trimer adenylate cores. While not part of the invention, the preparation of the dimer core 6 . is included for completeness. The selectively substituted tetramer cores, 3_8. a " 39 , 51 and 5_2 and 5_7 and 5_8, were derived from the fully protected _rimer cores, 3_0 and . 3_1, and subsequently subjected to detrilylation and condensation to add the tetramer moiety. The compounds of the present invention comprise 2-5A derivatives that are (i) nuclease-resistant , (ii) non-toxic, (iii) able to activate or inactivate RNase L and (iv) able to inhibit HIV-l replication. The inventive compounds are chemically synthesized phosphoro- thioate/phosphodiester trimer and tetramer 2-5A deriva¬ tives in which at least one 2 ', 5 ' -phosphodiester bond has been selectively replaced with a 2 ', 5 ' -pho¬ sphorothioate bond. The chemical synthesis of these phosphorothioate/phosphodiester derivatives utilizes the phosphotriester and phosphoramidite approach in which reactive functional groups are protected by blocking groups which can be individually manipulated. The phosphorothioate/phosphodiester trimer and tetramer 2-5A derivatives reveal heretofore unknown aspects of the stereochemical requirements for activation of RNase L, namely, that activation of RNase L requires PR chirality in the second internucleotide linkage from the 5'- terminus of the 2-5A molecule and that PS chirality in the second internucleotide linkage results in 2-5A derivatives that are antagonists of RNase L activation. Without wishing to be bound by any theory, it appears that PR chirality in the second internucleotide linkage from the 5 '-terminus may serve to facilitate formation of a productive complex between RNase L, the allosteric activator (ApA Rp * A or ApA Rp * ApA) and the RNA substrate such that hydrolysis of HIV-l RNA can occur. Phosphorothioate substitution of individual inter¬ nucleotide linkages in the 2-5A molecule has revealed that inhibition of HIV-l replication is influenced by the location and stereoconfiguration of the chiral phosphorothioate group in the phosophorothioate/phos- phodiester derivatives. Of the four phosphoro- thioate/phosphodiester trimer core derivatives, ApA Rp * A and ApA Sp "A were the most efficient inhibitors of HIV-l induced syncytia formation (Figure 4A) . Of the six phosphorothioate/phosphodiester tetramer core der¬ ivatives, ApApA Rp * A and ApApA Sp * A were the most efficient inhibitors (Figure 4B) . A Rp * A, A Sp * A, 3',5'-A 4 , adenosine and adenine did not inhibit HIV-l RT activity. Whereas ApA Rp * A and ApA Sp * A are both phosphodiesterase-resistant and inhibit HIV-l RT, the ApA Rp * A enantiomer (but not the ApA Sp * A enantiomer) can also activate RNase L. In this regard it appears, again, without wishing to be bound by any theory, that the relative differences in the inhibition of HIV-l replication by the phosphorot- hioate/phospohodiester trimer and tetramer core deriva¬ tives may be explained by their resistance to hydrolysis by serum phosphodiesterases (see Table 1, infra. ) . In contrast to 2 ', 5 ' -phosphodiester bonds in authentic A 2 and A 3 which are totally hydrolyzed in serum-containing medium in 20 minutes, both PR and PS 2 ', 5 ' -phosphorothi¬ oate bonds are more stable to hydrolysis by phosphodie¬ sterases. The phosphorothioate/phosphodiester tetramer core derivatives which are stereochemically modified at the 5 '-terminus (A Rp * ApApA and A Sp * ApApA) are rapidly hydrolyzed from the 2 ', 3 ' -terminus to their respective dimers, A Rp * A and A Sp * A. These dimers, although resistant to further hydrolysis, can neither activate RNase L nor inhibit HIV-l replication. The remaining four phosphoro- thioate/phosphodiester tetramer core derivatives (ApA Rp * ApA, ApA Sp * ApA, ApApA Rp * A and ApApA Sp * A) are hydro¬ lyzed from the 5 '-terminus to form their respective trimer cores, A Rp * ApA, A Sp * ApA, ApA Rp * A and ApA Sp * A, respec- _. tively. Because ApA Rp * A and ApA Sp * A are efficient inhibitors of HIV-l replication, this hydrolysis most likely accounts for the antiviral action of the tetramer derivatives, ApApA Rp * A and ApApA Sp * A. Therefore, the decreased anti-HIV-l activity observed with ApA Rp * ApA and ApA Sp * ApA (relative to ApApA Rp "A and ApApAp Sp * A) is likely due to hydrolysis from the 5 ' -terminus to form ApA Rp * A and ApA Sp * A, which are very efficient inhibitors of HIV-l induced syncytia formation (compare Figs. 4A and 4B) . In preliminary experiments, all phosphoro- thioate/phosphodiester trimer and tetramer 2-5A core derivatives of the present invention have been shown to inhibit HIV-l RT. Inhibition ranges from 22% to 70%. A Rp * A, A Sp * A, 3',5'-A 4 , adenosine and adenine did not inhibit HIV-l RT activity. Whereas ApA Rp * A and ApA Sp * A are both phosphodiesterase-resistant and inhibit HIV-l RT, the ApA Rp * A enantiomer (but not the ApA Sp * A enantiomer) can also activate RNase L. These three biological properties (i.e. , resistance to hydrolysis by phospohodiesterases, inhibition of reverse transcriptase and activation of RNase L) may account for the 100% inhibition of HIV-l replication observed with ApA Rp * A. The compounds of the invention are advantageously prepared as soluble salts of sodium, ammonium or po¬ tassium. The preparative scheme begins with 6-N-benzoyl- 3 ' -O-tert-butyldimethylsilyl-5 ' -O-monomethoxy-tri- tyladenosine 1 , which is advantageously prepared from adenosine according to the procedure of Flockerzi e aM , Liebig's Ann. Chem., 1568-1585 (1981) . Preparation of the compounds of the present invention is illustrated in more detail by reference to the following non-limiting examples. 3-Nitro-l,2,4-triazole andp-nitrophenylethanol used in the examples may be prepared advantageously from published procedures: Chattopadhyaya et a_l . , Nucleic Acids Res. 8:2039-2053 (1980); Schwarz et al. , Tetrahe- dron Lett . , 5513-5516 (1984); Uhlmann et al., Helv. Chim. Acta 4:1688-1703 (1981) . These compounds are also available commercially in the United States. 3-Nitro-l, 2, 4-triazole is available from Aldrich Chemical Co., P.O. Box 355, Milwaukee, WI 53201 (1986-1987 cat. no. 24,179.2) . p-Nitrophenylethanol is available from Fluka Chemical Corp. (cat. no. 73,610) . Pyridine and triethylamine used in the examples were purified by distillation over KOH, tosyl chloride and calcium hydride. Dichloromethane was distilled over calcium chloride and then passed through basic alumina. Pure acetonitrile was obtained by distillation over calcium hydride. Purification of the protected nucleotides was achieved by preparative column chromatography on silica gel 60 (0.063-0.2 mesh, Merck) and by preparative thick layer chromatography on silica gel 60 PF 254 (Merck) . Thin layer chromatography ("TLC") was carried out on precoated thin layer sheets F 1500 LS 254 and cellulose thin layer sheets F 1440 from Schleicher _ Scheull. Scheme 1 is the reaction scheme for the preparation of the fully resolved trimers, having phosphorothioate substitution of the first internucleotide linkage A Rp * ApA 10 and A Sp * ApA . 11 . , from the protected intermediates, 8_ and 9., wherein "bz" denotes the benzoyl radical, "tbds" denotes the tert-butyldimethylsilyl radical, "ce" denotes the cyanoethyl radical, "npe" denotes the nitrophenyleth- oxy radical and "MeOTr" represents the monomethoxytrityl radical. The preparation of the trimer cores, . 10 . and 11 is set forth in Preparations 1 through 4 and Example 1. Preparation 4 illustrates the fully protected trimer core, while Example 1 illustrates the removal of the blocking groups and the chemical purification of the fully resolved isomers. I to ι- » _ι O o LΠ σ t to o ι_π O <_π LΠ - 17 - Preparation 1 a. Bis- (diisopropylamino) - ( β -cyanoethoxy) - phosphane 3 . : Preparation of the titled compound was in accord with the procedure of Kraszewski _ Norris, Nucleic Acids Research Sump. Ser. 18 : 177-80 (1987) . β-Cyanoethanol (7 g; 0.1 mole) in absolute CH 3 CN (40 ml was added dropwise within 30 min to a solution of freshly distilled PCl j (40 ml; 0.4 mole) at room temperature ("r.t.") and under nitrogen atmosphere. After stirring for 3.5 h, the solvent and excess PCl 3 were removed in high vacuum, the residue was dissolved in 450 ml of absolute ether and at -10°C reacted with N,N-diisopropylamine (127 ml; 0.9 mole) by dropwise addition within 1 h under nitrogen atmosphere. The reaction mixture was stirred at -10°C for 30 min and at r.t. for 15 h. The precipitate was filtered under nitrogen and the solvent was removed in vacuo. The yellow crude product was fractionally distilled over CaH 2 to give 14.7 g (49%) of pure 2 of b.p. 114-118°C. This reagent was stored at -20°C under nitrogen. ^-N R (CDC1 3 ) : 3.75 (s, 2H, CH 2 ) ; 3.52 (m, 4H, 4 N-CH) ; 2.60 (t, 2H, j8-CH 2 ) ; 1.17 + 1.14 (2d, 24H, 4 N- C(CH 3 ) 2 ) . 31 P-NMR (CDC1 3 ) : 124.6 ppm. b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] - 5' -O- (monomethoxytrityl) -adenosine-2 ' -O- [ (β - cyanoethyl) -N,N-diisopropylaπ_ino] - phosphoramidite 4 : METHOD A The preparation of the titled compound was in accord with the procedures of Sinha et. al . , Nucleic Acids Res. 12: 4539-4557 (1984) wherein compound 1 , Flockerzie et. _l. (1981) , supra, (3.79 g; 5 mmole) and diisopropyl-ethylamine (3.5 ml) were dissolved in dry CH 2 C1 2 (20 ml) and chloro-N,N-diisopropylamino- cyanoethoxy phosphane (2.37 g; 10 mmole) was added. After 1.5 h stirring under nitrogen at r.t. , the reaction mixture was diluted with EtOAc (100 ml) and the organic - 18 - phase was washed with a saturated NaHC0 3 /NaCl solution (2 x 80 ml) . The organic layer was dried over Na 2 SO, , filtered and evaporated to dryness . The residue was dissolved in CH 2 C1 2 (10 ml) and added dropwise to n- hexane (200 ml) at -60°C. The product was collected and evaporated to dryness in high vacuum for 8 h to give 4.3 g (89%) of a colorless amorphous solid. METHOD B Alternatively, the titled compound was prepared according to the procedure of Kraszewski and Norris (1987) , supra. In this method compound 1 (3.79 g; 5 mmole) and tetrazole (0.175 g; 2.5 mmole) were dissolved in dry CH 2 C1 2 (20 ml) and then bis- (diisopropy- lamino) - ( β-cyanoethoxy) phosphane 3 . (3 g; 10 mmole) was added. After stirring at r.t. under argon for 17 h, the reaction mixture was extracted with EtOAc (100 ml) and washed with saturated NaHC0 3 /NaCl solution (80 ml) . This was repeated twice and work-up was performed analogous to method A to give 4.49 g (94%) of a colorless amorphous powder. Anal . calc. for C 52 H 64 N 7 0 7 PSi x 2 H 2 0 (994.2) : C 62.82, H 6.89, N 9.86. Found: C 62.52, H 7.08, N 10.35. UV (MeOH) : λ max (log e) 279 nm (4.33) ; 229 n (4.43) . R f on silica gel with toluol/EtOAc (1/1, v/v) : 0.64, 0.61 (diastereo ers) . 31 P-NMR (CDC1 3 ) : 150.98, 151.34. Preparation 2 6-N-Benzoyl-3 ' -0- [ (tert-butvl)dimethylsilyl] -5 ' -O- (monomethoxytrityl) -adenylyl-2 ' - [O p - (2-cyanoethyl) - 5 < _] -6-N-benzoyl-2 ' ,3'-di-0- [(tert-butyl)dimethyls- ilyl] -adenosine .6: The phosphoramidite 4 . (2.88 g; 3 mmole) and 6-N- benzoyl-2 ' , 3 ' -di-O- [ (ter -butyl)dimethyl-silyl] adenosine 5., Flockerzie et al. (1981) , supra, (1.2 g; 2 mmole) were dried at r.t. in high vacuum for 24 h and dissolved in dry CH 2 C1 2 (30 ml) . Tetrazole (0.5 g; 8 mmole) was added _ and after 3 h stirring at r.t. under argon, a solution of I 2 [0.5 g H 2 0/pyridine/ CH 2 C1 2 (l/3/l, v/v/v) ] was added dropwise until the brown color does not disappear. - 19 - The mixture was stirred for 15 min, then diluted with CHC1 3 (300 ml) . The organic phase was saturated with Na 2 S 2 0 3 /NaCl (3 x 80 ml) , dried over Na 2 S0 4 and evaporated to dryness. Final coevaporation was done with toluene (3 x 20 ml) . The crude product was purified by silica gel column chromatography (15 x 2.5 cm) using CHC1 3 (100 ml) , CHCl 3 /MeOH (100/0.5, v/v; 1.5 L) and CHCl 3 /MeOH: (100/1, v/v) to elute the product. Product fractions were collected and evaporated to dryness to give 2.33 g (79%) of the dimer 6 . in the form of a solid foam. Anal . calc. for C 75 H 94 N 11 0 13 PSi 3 (1490.9) : C 60.42, H 6.49, N 10.33. Found: C 60.50, H 6.49, N 10.22. UV (MeOH) : λ max (log e) 278 nm (4.62) , 230 nm (4.62) . R f on silica gel with CHCl 3 /MeOH (95/5, v/v) = 0.56. 31 P-NMR (CDC1 3 ) : -0.74 and -1.07 ppm (diastereomers) . Preparation 3 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - adenylyl-2 ' - [O p - (2-cyanoethy1) -5 ' ] -6-N-benzoyl- 2 ' ,3 ' -O-di- [ (tert-butyl)dimethylsilyl] adenosine 2: Compound 6 . (2.22 g; 1.51 mmole) was stirred with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v; 30 ml) at. r.t. for 30 min. The reaction mixture was diluted with CH 2 C1 2 (300 ml) , washed with phosphate buffer, pH 7.0 (2 x 100 ml) , dried over Na 2 S0 4 and evaporated to dryness. The residue was applied to a silica gel column (9 x 4.5 cm) , washed with CHC1 3 (0.7 L) and CHCl 3 /MeOH (100/1, v/v; 300 ml) . The product was eluted with CHCl 3 /MeOH (50/1, v/v; 300 ml and 100/3, v/v; 300 ml) . The combined product fractions were evaporated to dryness in high vacuum to give 11.65 g (90%) of 5 ' -hydroxy dimer 1_ as an amorphous solid. Anal . calc. for C 55 H 78 N 11 0 12 PSi 3 x H 2 0 (1218.5) : C 54.21, H 6.62, N 12.64. Found: C 54.53, H 6.58, N 12.62. UV (MeOH) : λ^ (log e) 278 nm (4.60) , 232 nm (4.42) . * R f on silica gel with CHCl 3 /MeOH (95/5, v/v) = 0.36. 31 P-NMR (CDC1 3 ) : -0.77 and -1.30 ppm (diastereomers) . - 20 - Preparation 4 a . 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] - 5 ' -O- (mono ethoxytrityl) - (PR) -thioadenylyl-2 ' - [O p - (2 -cyanoethy1) -5 ' ] -6-N-benzoyl-3 ' -O- [ (tert.-butyl) dimethylsilyl] -adenylyl-2 ' - [O p - (2-cyanoethyl) -5 ' ] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert-butyl)dimethylsilyl] -adenosine A_. p * ApA ! b. 6-N-Benzovl-3'-0- [ (tert-butyl)dimethylsilyl] - 5' -O- (monomethoxytrityl) - (PS) -thioadenylyl-2 ' - [O p - (2-cyanoethyl) -5 '] -6-N-benzoyl-3 ' -O- t (tert-butyl) dimethylsilyl] -adenylyl-2 ' - [O p - (2-cyanoethyl) -5 '] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert-butyl)dimethylsilyl] -adenosine A Sp * ApA ! '• The phosphoramidite 4 . (2.79 g; 2.92 mmole) , the 5'- hydroxy dimer 1_ (1.94 g; 1.62 mmole) and tetrazole (0.567 g; 8.1 mmole) were dissolved in dry CH 3 CN (8.1 ml) and stirred at r.t. under nitrogen. After 3 h, S B (1.66; 6.48 mmole) and pyridine (7.8 ml) were added and stirred further for 20 h at r.t. The reaction mixture was then diluted with CH 2 C1 2 (300 ml) , washed with saturated NaCl (2 x 200 ml) , dried over Na 2 S0 4 and evaporated to dryness. Final coevaporation was with toluene (3 x 20 ml) . The crude diastereomeric mixture (A Rp * ApA 8 . + A Sp * ApA 9.) was dissolved in CH 2 C1 2 and applied to a silica gel column (21 x 3.5 cm) . The column was washed with CH 2 C1 2 (450 ml) and CH 2 Cl 2 /MeOH (99/1, v/v; 200 ml) and the product was eluted with CH 2 Cl 2 /MeOH (97/3, v/v; 400 ml) . The product fractions were collected and evaporated to dryness to give 3.16 g (93%) of an isomeric mixture of A Rp * ApA 8 . and A Sp * ApA 9.. Separation into the pure diastereoisomers was achieved by medium pressure chromatography as described above by elution with CHCl 3 /MeOH (99/1, v/v; 800 ml; 20 ml/fraction; fractions 1-40) followed by elution with CHCl 3 /MeOH (95/5, v/v; 800 ml; 20 ml/fraction; fractions 41-80) . The fully protected A Rp "ApA isomer 8. (0.287 g) was eluted in fractions 21-56 (20 ml/fraction) . Fractions 57-61 gave the isomer mixture (0.07 g) and the fully protected - 21 - A Sp * ApA isomer 9_ (0.132 g) was eluted in fractions 62-64. Chromatographic separation was repeated with each 0.5 g of the crude mixture to yield 1.62 g (51%) of A Rp * ApA 8. and 0.94 g (30%) of A Sp * ApA 9 . . Anal . calc. for A Rp * ApA - C ltu H 127 N 17 0 19 P 2 SSi 4 (2089.6) : C 58.05, H 6.13, H 11.40. Found: C 58.65, H 6.24, N 11.50. UV (MeOH) : λ max (log e) 279 nm (4.76) , 260 nm (4.56) , 236 nm (4.73) . R f on silica gel with CHCl 3 /MeOH (97/3, v/v) = 0.35. 31 P-NMR (CDC1 3 ) : 69.35 and -1.10 ppm. Anal . calc. fcr A Sp * ApA - C 101 H 127 N 17 O 19 P 2 SSi 4 (2089.6) : C 58.05, H 6.13, N 11.25. Found: C 57.03, H 6.33, N 11.14. UV (MeOH) : λ max (log e) 279 nm (4.77) , 260 nm (4.57) , 236 nm (4.73) . 31 P-NMR (CDCI 3 ) : 68.33 and -0.84 ppm. Example 1 a. (PR) -P-Thioadenylyl-2 ' -5 ' -adenylyl-2 ' -5 ' - adenosine A Rp * ApA 10; b. (PS) -P-Thioadenylyl-2 ' -5 ' -adenylyl-2 • -5 ' - adenosine A Sp * ApA 11; The corresponding fully protected trimers 8 . and 9 . , respectively, were separately deblocked by stirring the trimer (0.06 g; 0.029 mmole) with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v; 1.2 ml) for 1.5 h at r.t. The reaction mixture was diluted with CHC1 3 (50 ml) , washed with H 2 0 (2 x 25 ml) , dried and evaporated to dryness. The crude product was purified on preparative silica gel plates (20 x 20 x 0.2 cm) in CHCl 3 /MeOH (8/2, v/v) . The product bands were eluted with CHCl 3 /MeOH (4/1, v/v) and evaporated to a foam to give 0.04 g (84%) of the A Rp * ApA isomer JL0 . and 0.034 g (73%) of the A Sp * ApA isomer H. The 5'-hydroxy trimer (0.034 g; 0.08 mmole) was then stirred with 0.5 M DBU in pyridine (5.0 ml) and after stirring at r.t. for 20 h, the solution was neutralized with 1 M acetic acid in dry pyridine (2.5 ml) and evaporated to dryness. The residue was treated with methanolic ammonia (5 ml) and after 48 h stirring the solvents were - 22 - removed in vacuo. Desilylation was performed with 1 M tetrabutylammonium fluoride in THF (2 ml) . After 48 h stirring, the solvent was removed in vacuo and the residue was dissolved in H 2 0 (10 ml) and applied to a DEAE Sephadex A-25 column (60 x 1 cm) . The pure product was eluted with a linear gradient of 0.14 -0.17 M TEAB buffer, pH 7.5. After evaporation and coevaporation with water several times, the trimer was applied to four paper sheets (35 x 50 cm) and developed in i-PrOH/conc. ammonia/H 2 0 (6/1/3, v/v/v) . The product band was cut out, eluted with H 2 0, evaporated and lyophilized to give 500 O.D. 260 nm units (79%) of the A Rp * ApA isomer ljO and 410 °- D - 260 nm units (65%) of the A Sp * ApA isomer IX - UV λ„ ax in both cases was 258 nm in H 2 0. A Rp * ApA JUD: R f on cellulose in i-PrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.33. 'H-NMR (D 2 0) : 8.20; 8.19; 8.14 (3s, 3H, H-C(8)) ; 7.97 (IS, 2H, 2 H-C(2) ) and 7.76 (1 s, 1H, 1 H-C(2) ) ; 6.08; 5.93; 5.82 (3d, 3H, 3 H-C(l') ) . Retention time on reverse-phase HPLC was 5.60 min. A Sp * ApA 3_L: R f on cellulose in i-PrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.33. X H-NMR (D 2 0) : 8.14; 8.09; 8.02 (3s, 3H, H-C(8) M 7.94; 7.89: 7.80 (3s, 3H, 2 H-C(2)) ; 6.03; 5.92; 5.80 (3d, 3H, 3 H-C(l') ) . Retention time on reverse-phase HPLC was 6.51 min. Scheme 2 is the reaction scheme for the preparation of the remaining pair of trimer cores, ApA Rp * A 2 . 3 and ApA Sp * A 2 , from the protected intermediates 2 . 1 and 22 , and is outlined in detail in preparations 5 and 6 and Example 2, below. 23 - 10 5 - 23/1 o (X - 24 - Preparation 5 a . N,N-Diisopropyl- trimethylsilylamine: The preparation of the titled compound weis in accord with the procedure of Noth and Staudigl, Chem. Ber. 115 : 3011-3024 (1982) . Methyl iodide (37.6 ml; 0.6 mole) in absolute ether (50 ml) was added dropwise (over 90 min) to a suspension of 14.6 g (0.6 mole) of magnesium and a few crystals of iodine in absolute ether (100 ml) . The reaction was then stirred for 30 min until all the magnesium was dissolved. Subsequently, N,N-diisopropyl- amine (78 ml; 0.55 mole) was added within 10-15 min and the reaction was refluxed for 1 h. After cooling to 0°C, tri ethylsilyl chloride (76 ml; 0.6 mole) was added dropwise and the reaction mixture was again heated in an oil-bath with vigorous stirring to 80°C for 20 h. The supernatant liquid was decanted and the residue was extracted with ether (4 x 50 ml) . The supernatant and the ether extract were combined. The solvent, excess trimethylsilyl chloride and unreacted N,N-diisoproylamine were removed by distillation. The product was then isolated by distillation under vacuum at an oil-bath temperature of 60°C to yield 73 g (80%) , Kp 18 = 36-39°C. : H-NMR (CDC1 3 ) : 0.08 (s, 9H, SiCH 3 ) ; 1.04-1.07 (d, 12H, N-C-CH 3 ) , 3.2 (m, 2H, N-CH) . b. Chloro-N,N-diisopropylamino-2 - ( 4 - nitrophenyl) e thoxy -phosphane 14 ; p-Nitrophenylethanol (4.16 g; 25 mmole) was added portion wise to a solution of freshly distilled PC1 3 (14 ml; 0.16 mole) in absolute ether (40 ml) at -30°C under a nitrogen atmosphere within 45 min. The reaction mix¬ ture was stirred at r.t. for 1.5 h and the solvent and excess PC1 3 were then removed in vacuo at 0°C. The residue was treated with N,N-diisopropyl-trimethylsilyl- amine (Preparation 5a) (4.33 g; 25 mmole) at 0°C under a nitrogen atmosphere for 30 min and then at. r.t. for - 25 - 20h. The resulting trimethylsilyl chloride was removed under high vacuum at r.t. to yield a syrupy pale yellow product (7.1 g; 85%) which crystallized upon storage at -20°C. This material was then used for the subsequent phosphitylation reactions. α H-NMR (CDC1 3 ) : 8.1-8.2 ( , 2H, o to N0 2 ) ; 7.39-7.43 (m, 2H, m to N0 2 ) ; 4.04-4.18 (m, 2H, P-0-CH 2 ) ; 3.63-3.79 (m, 2H, N-CH) ; 3.07-3.13 (t, 2H, P-0-C-CH 2 ) ; 1.14-1.27 (2d, 12H, N-C-CH 3 ) . 31 P-NMR (CDC1 : 181.60 ppm. 6-N-Benzoyl-3 ' -O- t (tert-butyl)dimethylsilyl] - 5' -O- (monomethoxytrityl) -adenosine-2 ' -O- [ (4- nitrophenyl) ethyl) -N,N-diisopropylamino] - phosphoramidite 15: METHOD A. Compound 1 (3.79 g; 5 mmole) and diisopropylethylamine (3.5 ml) were dissolved in dry CH 2 C1 2 (20 ml) and then chloro-N,N-diisopropylamino-2- (4- nitrophenyl) -ethoxyphosphane .14 . (2.37 g; 10 mmole) was added dropwise under a nitrogen atmosphere. After stirring at r.t. for 2 h, the reaction mixture was di¬ luted with EtOAc (200 ml) , the organic phase was washed with a saturated NaHC0 3 /NaCl solution (3 x 80 ml) , dried over Na 2 S0 4 and evaporated to dryness . The crude product was dissolved in toluene/EtOAc (7/3, v/v) and chro- matographed on a silica gel column (12 x 2 cm) equil¬ ibrated with EtOAc/NEt 3 (95/5, v/v) . The product frac¬ tions were eluted with EtOAc/NEt 3 (95/5, v/v) , collected and evaporated to dryness, yielding 1 . 5 (5.28 g; 79%) as a colorless solid foam. Anal . calc. for C 57 H 68 N 7 0 9 PSi (1054.3) : C 64.94, H 6.50, N 9.30. Found: C 64.81, H 6.51, N 9.01. UV (MeOH) : λ^ (log e) 277 nm (4.50) , 229 nm (4.48) . 31 P-NMR (CDC1 3 ) : 150.27, 150.01 ppm. R f on silica gel in toluene/EtoAC (l/l, v/v) : 0.62 and 0.68 (diastereomers) . METHOD B. Alternatively, compound JL5 was synthe¬ sized using bis- (diisopropylamino) - [2- (4- nitrophenyl) ethoxy] -phosphane 2_7, infra. To a solution - 26 - of 1.52 g (2 mmole) of compound 1 in absolute CH 3 CN (10 ml) , bis- (diisopropylamino) - [2- (4-nitrophenyl) ethoxy] - phosphane 27 (1.59 g, 4 mmole) and tetrazole (0.07 g ; 1 mmole) were added under nitrogen atmosphere and the reaction mixture was stirred for 17 h at r.t. The re¬ action mixture was diluted with EtOAc (120 ml) and washed twice with saturated NaHC0 3 /NaCl solution (60 ml) , dried over Na 2 S0 4 and evaporated to dryness. The crude solid foam was applied onto a flash silica gel column (20 x 2.5 cm) and chromatographed with toluene/EtOAc (1/1, v/v; 250 ml) . The product fraction (90 ml) was evaporated to give 1^ . (1.9 g, 90%) as a colorless solid foam. c. Bis - ( di i sopropylamino ) - [ 2 - ( 4 - nitrophenyl) ethoxy] -phosphane 27 ; 2- (4-Nitrophenyl) ethanol (8.35 g, 50 mmole) was added in small portions over 30 min to a solution of distilled PC1 3 (28 ml; 280 mmole) in absolute ether (80 ml) at -5°C under a nitrogen atmosphere. After stirring for 15 min at -5°C and 1.5 h at r.t., the solvent and excess PC1 3 were removed under high vacuum. Then, the yellowish syrupy residue was dissolved in 200 ml of absolute ether and reacted at -10°C with N,N- diisopropylamine (64 ml, 450 mmole) by dropwise addition over 30 min under a nitrogen atmosphere. The reaction mixture was stirred at -10°C for 15 min and r.t. for 16 h. The voluminous precipitate of N,N-diisopropyl-amine hydrochloride was filtered under nitrogen and the solvent was removed in vacuo. The yellowish syrupy product (17.6 g; 89%) , which crystallized on storage at -20°C was pure enough to be used for phosphitylation reactions . -Η- MR (CDC1 3 ) : 8.10-8.13 (d, 2H, o to N0 2 ) ; 7.36-7.40 (d, 2H, m to N0 2 ) ; 3.75-3.82 (q, 2H, P-0-CH 2 ) ; 3.36-3.51 (m, 2H, N- CH) ; 2 . 95 - 3 . 00 ( t , 2H , P-0-C- CH 2 ) ; 1 . 05 - 1 . 12 ( 2d , _2H , N- C- CH 3 ) . 31 P-NMR ( CDC1 3 ) : 123 . 53 ppm . - 27 - Preparation 6 a . 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - 5 ' -O- (monomethoxytrityl) -adenylyl -2 ' - [ (0 p -2 - (4-nitrophenyl) ethyl-5 ' ] -6 -N-benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - (PR) -P- thioadenyl- yl-2 ' - [ (O p -2- (4-nitro-phenyl) ethyl-5 ' ] -6-N- benz oyl - 2 ' , 3 ' - di - O - [ ( t e r t - b u - tyl) dimethylsilyl] -adenosine ApA^'A 21 ; b. 6 -N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - 5 ' -O- (monomethoxytrityl) -adenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5 ' ] -6-N-benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - (PS) -P-thioadenyl- yl-2 ' - [ (O p -2- (4-nitro-phenyl) ethyl-5 ' ] -6-N- benz oyl - 2 ' , 3 ' - di - O - [ ( te r t - b u - tyl)dimethylsilyl] -adenosine ApA Sp * A 22 ; Triethylammonium 6-N-benzoyl-3 ' -O- [ (tert-butyl) - dimethylsilyl] -5 ' -O- (monomethoxytrityl) adenosine-2 ' - [2- (4-nitrophenyl) ethyl] -phosphate 20 (0.10 g; 0.1 mmole) , Charubala et al . , Liebig' s Ann. Chem.. 2392-2406 (1981) , and the respective 5 ' -hydroxy dimers PR 1_8 and PS 15 , (0.066 g; 0.05 mmole) , Charubala and Pfleiderer (1992) supra, were coevaporated with dry pyridine (3 x 5 ml) , dissolved in one ml dry pyridine and (2,4,6- riisopropyl) benzenesulfonyl chloride (0.062 g; 0.2 mmole) and 3-nitro-l, 2 , 4-triazole (0.068 g; 0.6 mmole) , Kroger and Mietchen, Z . Chem. .9: 378-379 (1969) ; Jones et al . , Tetrahedron 36 : 3075-3085 (1980) , were added. After suirring at r.t. for 20 h, the reaction mixture was diluted with CHC1 3 (100 ml) , washed with H 2 0 (2 x 50 ml) , dried and evaporated. Final evaporations were done with toluene (2 x 10 ml) to remove pyridine. The crude -rimers 2 . 1 and .22 . , respectively, were purified by silica gel column chromatography (15 x 2 cm) , using first CHC1 3 and then CHCl 3 /MeOH (100/1, v/v) as eluants. The product fraction was collected and evaporated to a solid foam, which was dried under high vacuum to give 0.08 g (70%) of 2 . 1. Anal, calc. for C 111 H 135 N 17 0 23 P 2 SSi 4 x 2 H 2 0 (2317 8) : C 57.52, H 5.95, N 10.27. Found: C 57.15, H 6.13, N 10.72. UV (MeOH) : λ max (log e) 276 nm (4.87) , 227 nm - 28 - ( 4 . 83 ) . R f on s i l ica ge l in CH 2 Cl 2 /EtOAc ( 1 / 1 ) = 0 . 6 3 . : P - NMR ( CDCl j ) : 6 9 . 88 and - 1 . 0 ppm . Example 2 a. Adenylyl- (2 '-5') -(PR) -P-thioadenylyl- (2 '-5' ) - adenosine ApA R * A 23 Adenylyl- (2 '-5')- (PS) -P-thioadenylyl- (2 '-5') adenosine and ApA Sp * A 24: The fully protected trimers, ApA Rp * A . 21 . and ApA Sp * A 22 , were separately deblocked by stirring the corres¬ ponding trimer (0.088 g; 0.037 mmole) with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v; 0.8 ml) . After 30 min stirring at r.t., the reaction mixture was diluted with CHC1 3 (50 ml) and washed with H 2 0 (2 x 25 ml) . The organic phase was dried over NaS0 4 and evaporated to dryness. The crude product was purified on a silica gel cclumn (5 x 2 cm) and the product eluted with CHCl 3 /MeOH (100/1, v/v) , evaporated and dried under high vacuum to give 0.073 g (94%) of the 5 ' -hydroxy trimer ApA Rp * A 2∑ a d 0.061 g (84%) of the 5 ' -hydroxy trimer ApA Sp * A 22 . . The resulting 5 ' -hydroxy trimer (0.04 g; 0.02 mmole) was then stirred with 10 ml of 0.5 M DBU in pyridine. After 24 h, the solution was neutralized with 1 M acetic acid in pyridine (10 ml) and evaporated to dryness. The residue was treated with saturated methanolic ammonia 16 ml) and after stirring at r.t. for 48 h, the solvent was removed in vacuo and the residue was desilylated with 1 M Bu 4 NF in THF (5 ml) for 48 h. The solvent was then removed in vacuo and the residue was dissolved in water (10 ml) and applied onto a DEAE Sephadex A-25 column (60 x 1 cm) . The product was eluted with a linear gradient of 0.14- 0.17 M TEAB buffer, pH 7.5. After evaporation and coevaporation with water several times, the trimer was applied to four paper sheets (35 x 50 cm) and developed in i-PrOH/conc. aπ_nonia/H 2 0 (6/1/3, v/v/v) . The product band was cut out, eluted with H 2 0, evaporated and - 29 - lyophilized to give 354 O.D. 260 nm units (79%) of the ApA Rp " A isomer _L and 410 O.D. 260 nm units (58%) of the ApA Sp * A isomer 2± . UV λ max in both cases was 258 nm in H 2 0. ApA Rp * A .23 . : R f on cellulose in iPrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.34. __-NMR (D 2 0) : 8.17; 8.16; 8.09 (3S, 3H, H-C(8) ) ; 7.90, 7.78 (2 s, 3H, 3 X H-C(2) ) ; 6.04; 5.96; 5.80 (3d, 3H, 3 H-C(l')) . Retention time on reverse-phase HPLC was 5.98 min. ApA Sp * A (24) : R f on cellulose in i-PrOH/ ammonia/H 2 0 (6/1/3, v/v/v) = 0.33. : H-NMR (D 2 0) : 8.17; 8.07; 8.04 (3s, 3H, 3 X H-C(8) ) ; 8.01; 7.92 : 7.72 (3s, 3H, 2 3 X H-C(2) ) ; 6.04; 5.92; 5.82 (3d, 3H, 3 x H-C(l')) . Retention time on reverse-phase HPLC was 7.23 min. Preparations 7 and 8 begin the preparation for the fully resolved tetramers, ApA Rp * ApA 3_8 and ApA Sp * ApA 39 , from their corresponding dimer .28. (Scheme 1) . The reaction scheme continues with the addition of the trimer moiety in Preparations 9 and 10 (Scheme 2) . Preparation 7 6-N-Benzoyl-3 ' -0- [ (tert-butyl) dimethylsilyl] -5 ' - (monomethoxytrityl) -adenylyl-2 ' - [ (O p -2- (4-nitro¬ phenyl) ethyl-5 ' ] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert- butyl) dimethylsilyl] -adenosine A (Rp Sp) * A 28 ; The phosphoramidite JL5. (1.41 g; 1.34 mmole) , 6-N- benzoyl-2 ' , 3 ' -di-O- [ (tert-butyl) dimethylsilyl] -adenosine 5. (0.36 g, 0.93 mmole) and tetrazole (0.188 g, 2.68 mmole) were stirred at r.t. in absolute CH 3 CN (9 ml) under a nitrogen atmosophere . After 4 h, a solution of I 2 [0.5 g in CH 2 Cl 2 /H 2 0/pyridine (1/1/3, v/v/v)] was added dropwise until the brown color did not disappear. The mixture was stirred was stirred for another 15 min, then extracted with CH 2 C1 2 (3 x 60 ml) and saturated Na 2 S 2 0 3 /NaCl solution (2 x 60 ml) . The CH 2 C1 2 phase was collected, dried over Na 2 S0 4 , evaporated and coevaporated with toluene (2 x 20 ml) to remove the pyridine. The - 3 0 - crude dimer (1.85 g) was dissolved in CH 2 C1 2 and applied onto a flash silica gel column (12 x 2.5 cm) and chromatographed using CH 2 Cl 2 /l% MeOH (400 ml) , 2% MeOH (200 ml) and 3% MeOH (200 ml) to elute the product (600 ml) . This fraction was evaporated to dryness to give 1.45 g (quant, yield) of the dimer 28 as a colorless amorphous solid. The identity of the isolated dimer 20 . was proven by comparison with authentic material by spectrophotometric comparison. The authentic material was synthesized by the phosphotriester method. Anal . calc. for ApA 2£ = C 80 H 98 N 11 O 15 PSi 3 (1569.0) : C 61.24, H 6.30, N 9.82. Found: C 61.24, H 6.24, N 9.65. UV (MeOH) : λ max (loge) 277 (4.69) ; [260 (4.54)] ; [231 (4.66)] . R £ on silica gel with CHCl 3 /MeOH (49/1, v/v) = 0.37. Preparation 8 6-N-Benzoyl-3 ' -O- (tert-butyl)dimethylsilyl-adenyl- yl-2' - [(O p -2- (4-nitrophenyl) ethyl-5 '] -6-N-benzoyl- 2 ' , 3 ' -di-O- t (tert-butyl) dimethylsilyl] -adenosine- 5'-OH-A (Rp>Sp) * A 29: The crude dimer mixture 2 . 8 (2.24 g, 1.43 mmole) was stirred with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v, 20 ml) at r.t. for 30 min. The reaction mixture was diluted with CH 2 C1 2 (200 ml) , washed with H 2 0 (2 x 80 ml) , dried over Na 2 S0 4 and evaporated to dryness . The colorless amorphous residue (2.0 g) was applied onto a flash silica gel column (21 x 2.5 cm) and chromatographed with CH 2 C1 2 (200 ml) , CH 2 Cl 2 /2% MeOH (400 ml) and the product was eluted with CH 2 Cl 2 /2% MeOH (500 ml) . The product fraction was evaporated and dried under high vacuum to give 1.2 g (75% calculated to compound 5_ over 2 steps) of 5 ' -OH dimer . 29. as an amorphous solid. The identity of the isolated dimer 2S_ with authentic material was proven by chromatographic and spectrophotometric comparison. Anal . calc. for 5 ' -OH-ApA 29 = C 60 H 82 N 11 O 14 PSi 3 (1296.6) : C 55.58, H 6.37, N 11.88. Found: C 55.33, H 6.38, N 11.78. UV (MeOH) : λ max (loge) 278 (4.68) ; [259 - 31 - (4.51)] ; 233 (4.46) . R f on silica gel with tcluene/EtOAc/MeOH (5:4:1) = 0.53. 31 P=NMR (CDC1 3 : -0.36 and -0.73 ppm. Preparation 9 a. 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -5 ' -O- (mono ethoxytrityl) - (PR) -P-thioadenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5'] -6-N-benzoyl-3 ' -0- [ (tert- butyl) dimethylsilyl] -adenylyl -2 ' - [ (O p - 2 - (4 - nitrophenyl) ethyl-5 ' -] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert-butyl) dimethylsilyl] -adenosine A Rp * ApA 30 : b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -5 ' -O- (mono ethoxytrityl) - (PS) -P-thioadenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5'] -6-N-benzoyl-3 ' -0- [ (tert- butyl) dimethylsilyl] -adenylyl -2 ' - [ (O p -2- (4- nitrophenyl) ethyl-5 ' -] -6-N-benzoyl-2 ' ,3 ' -di-O- [ (tert-butyl) dimethylsilyl] -adenosine A Sp * ApA 31: The phosphoramidite 15 . (0.59 g, 0.56 mmole) , the 5 '-hydroxy ApA dimer 29 . (0.52 g, 0.40 mmole) and tetrazole (0.079 g; 1.12 mmole) were dissolved in dry CH ; CN (4 ml) and stirred at r.t. under a nitrogen atmosphere. After 3 h, phosphoramidite .15 . (0.464, 0.44 mmole) and tetrazole (0.062 g, 0.88 mmole) were added again and the mixture was stirred for another 3 h. Then, oxidation with S e (0.257 g, 1 mmole) and pyridine (2.6 ml) was followed within 16 h at r.t. The reaction mixture was diluted with CH 2 C1 2 (200 ml) at r.t. The reaction mixture was diluted with CH 2 C1 2 (200 ml) , washed with a saturated NaCl solution (2 x 80 ml) , dried over Na 2 S0 4 and evaporated to dryness. Final coevaporation was done with toluene (3 x 20 ml) to remove pyridine. The crude diastereoisomeric mixture A( Rp , Sp ) * ApA 3JD + 3_1 was dissolved in CH 2 C1 2 (20 ml) , applied onto a flash silica gel column (11 x 2.5 cm) and chromatographed with CH 2 C1 2 (400 ml) , CH 2 Cl 2 /0.5% MeOH (200 ml) , 1% MeOH (200 ml) and the product was eluted with CH 2 Cl 2 /1.5% MeOH (200 ml) . The product fraction was evaporated to dryness to give 0.713 g (78%) of the isomeric mixture 3_0 + 31. Separation into the pure diastereomers was achieved by - 32 - application to preparative silica gel plates (40 x 20 x 0.2 cm, 8 plates) in toluene/EtOAc (l/l, v/v, 4 developments) . The isomeric products bands were separately eluted with CH 2 Cl 2 /MeOH (4/1, v/v) and evaporated to solid foams, which were dried under high vacuum to give 0.311 g (34%) of the fully protected A Rp * ApA isomer 3_0 and 0.245 g (27%) of the fully protected A Sp * ApA isomer 3JL. Anal . calc. for A Rp * ApA 3.0 = C 111 H 13 _ 5 N 17 0 23 P 2 SSi 4 X H 2 0 (2299.8) : C 57.97, H 6.00, N 10.35. Found: C 57.63, H 6.11, N 10.39. UV (MeOH) : λ max (loge) 278 (4.87) ; [260 (4.72)] ; [231 (4.75)] . R £ on silica gel with toluene/EtOAc (l/l, v/v, 2 developments) and toluene/EtOAc (1/2, v/v, 1 development) = 0.37. Anal . calc. for A Sp * ApA 3L = C 111 H 135 N 17 0 23 P 2 SSi 4 X 2 H 2 0 (2317.8) : C 57.52, H 6.05, N 10.27. Found: C 57.44, H 6.19, N 10.41. UV (MeOH) : λ max (loge) 277 (4.86) ; [260 (4.72)] ; [231 (4.75)] . R f on silica gel with toluene/EtOAc (1:1, 2 developments) and toluene/EtOAc (1:2, 1 development) = 0.27. Preparation 10 a. 6-N-Benzoyl-3 ' -0- [ (tert-butyl)dimethylsilyl] - (PR) - P-thioadenylyl-2'- [O p -2- (4-nitrophenyl) ethyl-5 '] -6- N-benzoyl-3 ' -O- [tert-butyl)dimethylsilyl] -adenylyl- 2'- [O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl-2' ,3 ' - di-O- (tert-butyl) dimethylsilyl] -adenosine 5 ' - hydroxy A Rp * ApA 32 ; b. 6-N-Benzoyl-3 ' -O- [ (tert-butvl)dimethylsilyl] - (PS) - P-thioadenylyl-2'- [O p -2- (4-nitrophenyl)ethyl-5' ] -6- N-benzoyl-3 ' -O- [tert-butyl)dimethylsilyl] -adenylyl- 2 ' - [O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl-2 ' ,3 ' - di-O- (tert-butyl)dimethylsilyl] -adenosine 5 ' - hydroxy A Sp * ApA 33 : The corresponding fully protected trimers 3_0 and 31, respectively, were separately detritylated by stirring the trimer (A Rp * ApA 3_0: 0.263 g, 0.115 mmole; A Sp * ApA 11: 0.21g, 0.92 mmole) with 2% p-TsOH % in CH 2 Cl 2 /MeOH (4/1, v/v, for 1 : 3.2 ml; for 3_1 : 2.6 ml) for 75 min at r.t. The reaction mixture was diluted with - 33 - Ch 2 Cl 2 (120 ml) , washed with H 2 0 (2 x 40 ml) , dried over Na 2 S0 4 and evaporated to dryness . The crude product was purified on preparative silica gel plates (40 x 20 x 0.2 cm) in toluene/EtOAc (3/7, v/v), the product bands were eluted with CH 2C1 2/MeOH (4/1, v/v) and evaporated to a solid foam to give 0.2 g (86%) of the 5 ' -hydroxy A Rp * ApA 32 and 0.121 g (66%) of the 5 ' -hydroxy A Sp * ApA H, respectively. Anal . Calc. for 5 ' -0H-A Rp * ApA 32= C 91 H 119 N 17 0 22 SSI 4 (2009.4: C 54.39, H 5.97, Nil.85. Found: C54.12, H 6.13, N 1174. UV (MeOH) : λ max (loge) 278 (4.86) ; [260 (4.71)] ; [233 (4.64] . R f on silica gel with toluene/EtOAc (3:7, 2 developments) = 0.35 (diastereomers) . 31 P-NMR: 69.60, 68.91, -0.36 and -0.56 ppm (diastereomers) . Anal . calc. for 5 ' -OH-A Sp * ApA ϋ = C 91 H 119 N 17 0 22 P 2 SSi 4 (2009.4) : C 54.39, H 5.97, N 11.85. Found: C54.29, H 6.23, N 11.51. UV (MeOH) ; λ max (loge) 277 (4.85; [260 (4.71)] ; [233 (4.63)] . R f silica gel with toluene/EtOAc (3:7, 2 developments = 0.42 (diastereomers) . 31 P-NMR: 69.28, 69.09, -0.33 and -0.56 ppm (diastereomers) . Scheme 3 is the reaction scheme for the preparation of the fully resolved tetra ers, ApA Rp * ApA 3_8 and ApA Sp * ApA 39, from the protected intermediates, . 34 . and 3_5. The preparation of these compounds is outlined in Prepara¬ tions 11 and 12 and Example 3. 34 - 35 - Preparation 11 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] -5 ' -O- (monomethoxytrityl) -adenylyl-2 ' - [O p -2- ( 4 - nitrophenyl) ethyl-5'] - 6 -N-benzoyl -3 ' -O- [ (.tert- butyl) dimethylsilyl] - (PR) -P-thioadenylyl -2 ' - [0 P - (2- (4 -nitrophenyl) ethyl) -5 '] -6 -N-benzoyl -3 ' -O- [ (tert- butyl) dimethylsilyl] -adenylyl -2 ' - [O p -2 - (4 - nitrophenyl) -ethyl-5 ' ] - 6 -N-benzoyl-2 ' , 3 ' -di-O- ( tert-butyl) dimethylsilyl -adenosine ApA Rp * ApA 34 . : b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] -5' -O- (monomethoxytrityl) -adenylyl-2 ' - [O p -2- (4- nitrophenyl) ethyl-5'] -6 -N-benzoyl -3 ' -0- [ (tert- butyl) dimethylsilyl] - (PS) -P-thioadenylyl -2 ' - [O p - (2- (4 -nitrophenyl) ethyl) -5'] -6 -N-benzoyl -3 ' -O- [ (tert- butyl) dimethylsilyl] -adenylyl -2 ' - [0 p -2- (4- nitrophenyl) -ethyl-5 ' ] - 6-N-benzoyl-2 ' , 3 ' -di-O- ( tert-butyl ) dimethylsilyl - adenos ine ApA Sp * ApA 35 : The condensation to the fully protected tetramers 34 and 3_5, respectively, were separately realized by coevaporatingtriethylammonium- 6 -N-benzoyl -3 ' -O- [ (tert- butyl) dimethylsilyl] -5 ' -O- (monomethoxytrityl) adenosine- 2 ' - [2- (4 -nitrophenyl) ethyl] -phosphate 20 (0.108 g, 0.1 mmole) and the 5 ' -hydroxy PR trimer H or PS trimer ϋ (0.1 g, 0.05 mmole) , respectively, with dry pyridine (3 x 2 ml) , dissolved in dry pyridine (0.5 ml) and then (2 , 4 , 6-triisopropyl) benzenesulf onyl chloride (0.061 g, 0.2 mmole) and 3 -nitro-1 , 2 , 4 -triazole (0.068 g, 0.6 mmole) were added. The solution was stirred at r.t. for 21 h, then extracted with CH 2 C1 2 (2 x 20 ml) and H 2 0 (3 x 20 ml) . The organic phase was collected, dried over Na 2 S0 4 , evaporated and coevaporated with toluene (3 x 20 ml) to remove pyridine. The crude tetramers 3_4 . and 35 , respectively, were separately purified on preparative silica gel plates (40 x 20 x 0.2 cm) with toluene/- EtOAc/MeOH (5/4/0.5, v/v/v/) , the product bands were eluted with CH 2 Cl 2 /MeOH (4/1, v/v/) and evaporated to solid foams, which were dried under high vacuum to give 0.116 g (78%) of ApA Rp * ApA 14 . and 0.12 g (81%) of -the ApA Sp * ApA 15. Anal . calc. for ApA Rp * ApA 3_4 = C 142 H 172 N 2 . 3 0 32 P 3 SSi s X 2 H 2 0 (3014.5) : C 56.58, H 5.89, N 10.69; - 36 - found: C 56.22, H 6.07, N 10.57. UV (MeOH) : λ max (loge) 277 (4.99) : [260 (4.85)] ; [231 (4.85)] . R f silica gel with toluene/EtOAc/MeOH (5:4:0.5) = 0.63 (diastereomers) . Anal . calc. for ApA Sp * ApA 5 = C 142 H 172 N 23 0 32 P 3 SSi 5 x H 2 0 (2996.5) : C56.92, H 5.85, N 10.75; found: C 56.40, H 5.89, N 10.61. UV (MeOH) : λ max (loge) 277 (5.01) : [260 (4.88)] ; [232 (4.89)] . R f silica gel with toluene/EtOAc/MeOH (5/4/0.5, v/v/v) = 0.62 (diastereomers) . Preparation 12 a. 6-N-Benzoyl-3 ' -0- [ (tert-butyl)dimethylsilyl] -adeny- lyl-2'- [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl- 3 ' -O- [ (tert-butyl)dimethylsilyl] - (PR) -P-thioadenyl- yl-2 ' - [(O p -2- (4-nitrophenyl) ethyl-5'] -6-N-benzoyl- 3 ' -O- [ (tert-butyl)dimethylsilyl] -adenylyl-2' - [(O p -2- (4-nitrophenyl) -ethyl-5'] -6-N-benzoyl-2 ' ,3 ' -di-O- [ (tert-butyl) dimethylsilyl] -adenosine 5 ' -OH- ApA Rp * ApA 36 ; b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adeny- lyl-2' - [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl- 3' -O- [ (tert-butyl)dimethylsilyl] - (PS) -P-thioadenyl- yl-2'- [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl- 3 ' -O- [ (tert-butyl)dimethylsilyl] -adenylyl-2' - [(O p -2- (4-nitrophenyl) -ethyl-5 ' ] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert-butyl)dimethylsilyl] -adenosine 5 ' -OH- ApA Sp * ApA 37 . : The fully protected tetramers (0.105 g; 0.035 mmole) 34 and 3_5, respectively, were separately detritylated by treatment with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v, 1.2 ml) for 1 h at r.t. The reaction mixture was extracted with CH 2 C1 2 (3 x 40 ml) and washed with H 2 0 (3 x 30 ml) . The organic phase was collected, dried over Na 2 S0 4 and evaporated to dryness. The resulting residue was purified on preparative silica gel plates (20 x 20 x 0.2 cm) in toluene/EtOAc/MeOH (5/4/0.5, v/v/v) . The product bands were eluted with CH 2 Cl 2 /MeOH (4:1) and evaporated to a solid foam, which was dried in high vacuum to give 0.064 g (67%) of the 5 ' -hydroxy ApA Rp * ApA 36. and 0.057 g (60%) of the corresponding PS tetramer 3_7. Anal . calc. - 37 - for 5 ' -OH-ApA Rp * ApA 3_6 = C 122 H 156 N 23 0 31 P 3 SSi 5 x H 2 0 (2724.1) : C 53.79, H 5.85, N 11.83; found: C 53.62, H 5.87, N 11.51. UV (MeOH) : λ max (loge) 277 (5.00) ; [260 (4.87)] ; 92.35 (4.81)] . R f silica gel with toluene/EtOAc/MeOH (5:4:0.5) = 0.41. Anal, calc. for 5 ' -OH-ApA Sp * ApA 17 = c i 22 H 156 N 23 0 31 P 3 SSi 5 x HO (2724.1) : C 53.69, H 5.85, N 11.83; found: C 53.58, H 5,97, N 11.32. UV (MeOH) : λ max (loge) 277 (4.96) : [260 (4.82)] ; 234 (4.74)] . R f silica gel with toluene/EtOAc/MeOH (5/4/0.5, v/v/v) = 0.39. Example 3 a. Adenylyl (2 ' -5 ')- (PR) -P-thioadenylyl- (2 ' -5 ') - adenylyl-2 (2 ' -5 ') -adenosine ApA Rp * ApA 38: b. Adenylyl (2 ' -5 ' ) - (PS) -P-thioadenylyl- (2 ' -5 ' ) - adenylyl-2 (2 ' -5' ) -adenosine ApA Sp * ApA 39 : The corresponding 5 ' -hydroxy tetramers 3_6 and 37 , respectively, were separately deblocked by stirring each 5 ' -hydroxy tetramer (0.056 g; 0.021 mmole) with 0.5 M DBU in absolute CH 3 CN (2.5 ml) at r.t. and after 22 h, the solution was neutralized with 1 M AcOH in absolute CH 3 CN (1.25 ml) and evaporated to dryness. The residual mixture was then treated with methanolic ammonia and after stirring at r.t. for 60 h, the solvent was removed in vacuum and finally the residue was disilylated with 1 M Bu 4 NF in THF (5 ml) for three days. The solvent was then removed, the residue was dissolved in HO (10 ml) , applied onto a DEAE Sephadex column A-25 (30 x 2 cm) and chromatographed first with H 2 0 (200 ml) and then with a linear gradient of 0-0.04 ml TEAB buffer, pH 7.5, within 3000 ml (flow rate 2 ml/min) . Under this condition, the ApA Rp * ApA tetramer 3_8 was eluted with a 0.23-0.28 M TEAB buffer and ApA Sp * ApA tetramer 19 . with 0.245-0.305 M TEAB buffer, respectively. The product fractions were collected, evaporated and coevaporated several times with MeOH. For further purification, paper chromatography was performed using a system of i-PrOH/ammonia H 2 0 - 3 8 - (55/10/35, v/v/v) . The product band was cut out, eluted with H 2 0, concentrated to a smaller volume and finally lyophilized to give 728 O.D. 260 nm units (73%) of the ApA Rp * ApA isomer 18 and 686 O.D. 260 nm units (69%) of the ApA Sp * ApA isomer 3_9, respectively. ApA Rp * ApA 18: R f on cellulose in i-PrOH/ammonia/H 2 0 (55:10:35) = 0.36. UV (H 2 0) : λ max 257 nm. X H-NMR (D 2 0) : 8.15, 8.07, 8.06, 7.93 (4s, 4H, 4xH-C(8)) ; 7.92 (s, 2H, 2xH-C(2)) ; 6.03, 5.89, 5.86, 5.79 (4d, 4xH-C(l')) . HPLC: on PR-18, A: 50 mM NH 4 H 2 P0 4 (pH 7.24) . B: MeOH/H 2 0 (l/l, v/v) ; gradient: 0-1 min, 80% A, 20% B; 1-31 min, 30% A, 70% B; retention time: 9.55 min. ApA Sp * ApA 19: R f on cellulose in i- PrOH/ammonia/H 2 0 (55/10/35, v/v/v) = 0.40. UV (H 2 0) : λ max 257 nm. HPLC: PR-18, A: 50 mM NH 4 H 2 P0 4 (pH 7.24) . B: MeOH/H 2 0 (1/1, v/v) ; gradient: 0-1 min, 80% A, 20% B; 1- 31 min, 30% A, 70% B; retention time: 10.37 min. Scheme 4 is the beginning of the reaction scheme for the preparation of the remaining tetramers from their intermediates, A Rp * Ap-diester 4 . 5 and the A Sp * Ap-diester 46 , with the blocking groups in place. The preparation of the diesters is outlined in Preparations 13 and 14. - 39 NH-bz 15 20 30 41 5 42 43 45 - 40 - Preparation 13 a. 6-N-Benzoyl-3 ' -0- [ (tert-butyl)dimethylsilyl] - 5 ' ) - (monomethoxytrityl) -adenylyl-2 ' - [ (O p -2- (4- nitrophenyl)ethyl-5'] -6-N-benzoyl-3 ' -0- [ (tert- butyl) dimethylsilyl] -adenosine-2 ' - [2, 5- dichlorophenyl, 2- (4-nitrophenyl) - ethylphosphate] ApAp-triester 4_1 and 41a; 6-N-Benzoyl-3 ' -0- [ (tert-butyl)dimethylsilyl. -adeno- sine-2 ' - [2 , 5-dichlorophenyl , 2- (4 -nitrophenyl) ethyl¬ phosphate] 5 ' -OH-Ap-triester 4_0 (0.43 g, 0.5 mmole) and the phosphoramidite JL5 . (0.735 g, 0.7 mmole) were dissolved in absolute CH 3 CN (5 ml) in the presence of tetrazole (0.098 g, 1.5 mmole) under a nitrogen atmosphere. After stirring for 3.5 h at r.t., phosphoramidite (0.2 g, 0.19 mmole) and tetrazole (0.026 g, 0.37 mmole) were added again and the reaction mixture was stirred for another 30 min. A solution of I 2 [0.5 g in Ch 2 Cl 2 /H 2 0/pyridine (1/1/3, v/v/v)] was added dropwise until the brown color did not disappear. The mixture was stirred for another 10 min, diluted with CH 2 C1 2 (20 ml) and washed with saturated Na 2 S 2 0 3 NaCl solution (2 x 80 ml) . The organic phase was collected, dried over Na 2 S0 4 , evaporated and coevaporated with toluene (3 x 30 ml) to remove the pyridine. The crude product was purified by flash silica gel chromatography (15 x 2 cm) , using toluene/EtOAc (l/l, v/v) , EtOAc and EtOAc/2-4% MeOH as eluants. The product fraction was evaporated to a solid foam, which was dried in high vac- uum at 30°C to give 0.610 g (67%) of the ApAp-triester 41 and 41a. The identity of the isolated compound with authentic material was proved by spectrophotometric comparison. The authentic material was synthesized from the Ap-diester 0 (Charubala et al . , 1981) with the 5'- hydroxy P-triester 40 . by the phosphotriester method. Anal . calc. for ApAp-triester = C 88 H 94 N 12 O :;0 Cl 2 P 2 SSi 2 (1828.8) : C57.80, H 5.18, N 9.9. Found: C 57.77, H 5.20, N 9.02. UV (MeOH) : λ^ (loge) : 277 (4.75) ; [260 (4.62)] ; - 4 1 - [228 (4.72)] . R f on silica gel with toluene/EtOAc/MeOH (5/4/1, v/v/v) _ 0.78. b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] - 5 ' ) - (monomethoxytrityl) - (PR, PS) -P-thioad- enylyl-2 '- [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N- benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - adenosine-2 ' - [2, 5-dichlorophenyl-2- (4-nitr- ophenylethyl) -phosphate] A( Rp , Sp ) * Ap-triester 4 + 14: 6-N-Benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adeno- sine-2 ' - [2, 5-dichlorophenyl , 2- (4-nitrophenyl) - ethylphosphate] 5 ' -OH-Ap-triester 40 (0.52 g; 0.6 mmole) and the phosphoramidite 15 . (0.95 g; 0.9 mmole) were dissolved in absolute CH 3 CN (6.5 ml) in the presence of tetrazole (0.126 g, 1.8 mmole) and under nitrogen atmosphere. After stirring for 3 h at r.t., S 8 (0.39 g, 1.51 mmole) and absolute pyridine (3.9 ml) were added and the reaction mixture was further stirred for 20 h, then extracted with CH 2 C1 2 (2 x 80 ml) and H 2 0 (2 x 80 ml) . The organic phase was collected, dried over Na 2 S0 4 and evaporated to dryness. Final coevaporation was done with toluene (4 x 20 ml) to remove the pyridine. The crude diastereomeric mixture A( Rp , Sp ) * Ap-triester 4! + 44 . was purified by flash silica gel column chromatography (14 x 2.5 cm) , using 200 ml CH 2 C1 2 , CH 2 C1 2 /1% MeOH, 2% MeOH and finally 200 ml CH 2 Cl 2 /l% MeOH, 2% MeOH and finally CH 2 Cl 2 /3% MeOH as eluants. The product fraction (150 ml) was evaporated to a solid foam, which was dried in high vacuum to give 0.975 g (88%) of (43) and (44) as a diastereomeric mixture. Anal. calc. for A( Rp , Sp ) * Ap- triester 4 + ___ = C 88 H 94 N 12 0 19 Cl 2 P 2 SSi 2 (1844.9) : C 57.29, H 5.14, N 9.11. Found: C 56.96, H 5.16, N 9.09. UV (MeOH) : λ^ (loge) : 277 (4.75) ; [228 (4.72)] . R f on silica gel with toluene/EtOAc/CHCl 3 (l/l/l, v/v/v) = 0.21. 31 P-NMR (CDC1 3 ) 69.87, 69.25, -6.89, -7.22 and - 7.31 ppm. - 42 - Preparation 14 a . Triethylammonium-N-6-benzoyl-3 ' -O- [ (tert-butyl) - dimethylsilyl] -5 ' ) - (monomethoxytrityl) - (PR) -P- thioadenylyl-2 ' - [ (O p -2- (4-nitrophenyl) -ethyl-5 '] -N- 6-benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adeno- sine-2 ' - [2- (4-nitrophenylethyl) -phosphate] A Rp * Ap- diester 45: b. Triethylammoni_m-N-6-benzoyl-3 ' -O- [ (tert-butyl) - dimethylsilyl] -5 ' ) - (monomethoxytrityl) - (PS) -P- thioadenylyl-2 ' - [ (O p -2- (4-nitrophenyl) -ethyl-5 '] -N- 6-benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adeno- sine-2 ' - [2- (4-nitrophenylethyl) -phosphate] A Sp * Ap- diester 46 : The solution of 0.558 g (3.36 mmole) of 4-nitro- benzaldehyde oxime in 15 ml of H 2 0/dioxane/Et 3 N (1:1:1) was stirred for 30 min at r.t. Then, 0.62 g (0.336 mmole) of the diastereomeric mixture of the A( Rp , Sp ) * Ap- triester 43_ + 44 . was added and stirred for 2.5 h at r.t. The mixture was evaporated, then coevaporated with pyridine (3 x 15 ml) , toluene (3 x 15 ml) and finally with CH 2 C1 2 (3 x 15 ml) . The residue was dissolved in a small amount of CHC1 3 and chromatographed on a flash silica gel column (15 x 2.5 cm) with CHC1 3 (150 ml) , CHCl 3 /2% MeOH (200 ml) , 4% MeOH (100 ml) , 6% MeOH (200 ml) , CHCl 3 /6% MeOH/0.5% Et 3 N (300 ml) and CHCl 3 /6_ MeOH/2% Et 3 N (250 ml) . The product fraction (600 ml) was evaporated to a solid foam, which was dried under high vacuum to give 0.55 g (91%) of the isomeric mixture 4_5 + 4_6. Separation into the pure diastereomers was achieved by chromatography on preparative silica gel plates (7 plates, 40 x 20 x 0.2 cm) and three develop¬ ments in CHCl 3 /MeOH (9/1, v/v) . The product bands were eluted with CHCl 3 /MeOH (4/1, v/v) containing 1% Et 3 N and evaporated to a solid foam to give 0.262 g (43%) of A Rp * Ap-diester 45 . , 0.144 (24%) of A Sp * Ap-diester 4 . 6 and 0.045 g (7%) of A( Rp , Sp ) * Ap-diester 45 . + 4j5- Anal, calc. for A Rp * Ap-diester 45 . = C 88 H 107 N 13 O 19 P 2 SSi 2 x 2 H 2 0 (1837Tl) : C 57.53, H 6.09, N 9.91. Found: C 57.30, H 6.7C, N 9.75. UV (MeOH) : λ max (loge) ; 277 (4.69) ; [260 (4.56)] ; [231 - 43 - (4.61)] . R £ on silica gel with CHCl 3 /MeOH (9/1, v/v) = 0.37. 31 P-NMR (CDC1 3 ) : 69.66 and -0.09 ppm. Anal . calc. for A Sp * Ap-diester 4_6 = C 88 H 107 N 13 O 19 P 2 SSi 2 x 2 H 2 0 (1837.1) : C 57.53, H 6.09, N 9.91. Found: C 56.44, H 7.15, N 8.61. UV (MeOH) : λ max (loge) ; 276 (4.60) ; [260 (4.49)] ; [232 (4.52)] . R f on silica gel with CHCl 3 /MeOH (9/1, v/v) = 0.28. 31 P-NMR (CDCI 3 ) : 68.96 and -0.06 ppm. Scheme 5 is the reaction scheme for the preparation of the remaining fully resolved tetramers, ApApA Rp * A 51 , ApApA Sp * A 5_2, A Rp * ApApA 5_7, and A Sp * ApApA 5_8, from their respective protected intermediates, 4J7, 4_8, 5_3 and 54. The corresponding preparations are outlined in Prepara¬ tion 15, 16, and 17 and Examples 4 and 5. 44 - - 45 - Preparation 15 a . 6-N-Benzoyl-3 ' -O- [ ( . tert-butyl) dimethylsilyl] - 5 ' -O- (monomethoxytrityl) -adenylyl-2 ' - [ (O p -2- (4 -nitrophenyl) ethyl-5 '] -6 -N-benzoyl -3 ' -O- [ (tert-butyl) -dimethylsilyl] -adenylyl-2 ' - [ (O p - 2- (4 -nitrophenyl) ethyl-5 ' ] -6-N-benzoyl-3 ' -0- [ ( tert-butvl ) dimethylsilyl] - (PR) -P- thioadenylyl-2 ' - [ (O p -2- (4 -nitrophenyl) ethyl - 5 ' ] -6-N-benzoyl-2 ' , 3 ' -di-O- [ (tert-butvl) - dimethylsilyl] -adenosine ApApA-. p * A 47 : Triethylammonium 6-N-benzoyl-3 ' -O- [ (tert-butyl) - dimethylsilyl] -5 ' -O- (monomethoxytrityl) -adenylyl -2 ' - [ (O p - 2- (4 -nitrophenyl) ethyl-5 ' ] -6 -N-benzoyl -3 ' -O- [ (tert- butyl ) dimethylsilyl] -adenosine-2 ' - [2 - ( 4 - nitrophenyl) ethyl -phosphate] 4_2 (0.14 g; 0.078 mmole) and the 5 ' -hydroxy PR dimer 18 . (0.08 g, 0.06 mole) were coevaporated with dry pyridine (4 x 0.5 ml) , dissolved in dry pyridine (0.6 ml) and then (2 , 4 , 6-triisopropyl ) - benzenesulfonyl chloride (0.047 mg, 0.156 mmole) and 3- nitro-1, 2 , 4-triazole (0.053 mg, 0.47 mmole) were added. The solution was stirred at r.t. for 22 h, extracted with CH 2 C1 2 (2 x 30 ml) , washed with H 2 0 (2 x 20 ml) , dried over Na 2 S0 4 and evaporated to dryness. Pyridine was re- moved by coevaporation with toluene (3 x 20 ml) . The crude tetramer 4_7 was purified by flash silica gel column chromatography (15 x 1 cm) and eluted first with CH 2 C1 2 (50 ml) , then with CH 2 C1 2 /1% MeOH (100 ml) , 2% MeOH (50 ml) and finally with CH 2 Cl 2 /3% MeOH (100 ml) . The product fraction (80 ml) was evaporated to dryness to give 0.11 g (62%) of the fully protected tetramer ApApA Rp * A 4_7 as a colorless foam after drying under high vacuum at 35°C. Anal . calc. for C 142 H 172 N 23 0 32 P 3 SSi s x H 2 0 (2996.5) : C 56.92, H 5.85, N 10.75. Found: C 56.51, H 5.91, N 10.37. UV (MeOH) : λ^ (loge) 277 (4.99) ; [259 (4.84)] ; [233 (4.85)] . R f on silica gel with CHCl 3 /MeOH (19/1, v/v) = 0.46. - 46 - 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - 5 ' -O- (monomethoxytrityl) -adenylyl-2 ' - [ (O p -2- (4 -nitrophenyl) ethyl-5 ' ] -6 -N-benzoyl -3 ' -O- [ (tert-butvl) dimethylsilyl] -adenylyl-2 ' - [ (O p - 2- (4 -nitrophenyl) -ethyl-5'] -6 -N-benzoyl -3 ' -0- [ (tert-butyl) dimethylsilyl] - (PS) -P-thioade- nylyl-2'- [ (0 p -2- (4 -nitrophenyl) ethyl-5 ' ] -6-N- benzoyl-2 ' , 3 ' -di-0- [ ( t er t - butyl) - dimethylsilyl] -adenosine ApApA Sp * A 48: ApAp-diester 4_1 (0.14 g, 0.078 mmole) and the 5'- hydroxy PS dimer 9 (0.08 g, 0.06 mmole) were coevapor¬ ated with dry pyridine (4 x 0.5 ml), dissolved in dry pyridine (0.6 ml) and then (2,4, 6-triisopropyl) - benzenesulfonyl chloride (0.47 mg, 0.156 mmole) and 3- nitro-1, 2 , 4-triazole (0.053 mg, 0.46 mmole) were added. After stirring for 4.5 h at r.t., (2,4,6- triisopropyl)benzenesulfonyl chloride (0.024 g, 0.078 mmole) and 3-nitro-1,2, 4-triazole (0.027 g, 0.234 mmole) were added again. The solution was stirred at r.t. for 16.5 h, then extracted with CH 2 C1 2 (4 x 20 ml) and added with H 2 0 (3 x 20 ml), dried over Na 2 S0 4 and evaporated. Final coevaporations were done with toluene (4 x 15 ml) to remove pyridine. The crude tetramer 4_8 was purified by flash silica gel column chromatography (15 x 1 cm) and eluted analogous to tetramer 4_7 with CH 2 C1 2 aned CH 2 Cl 2 /l-3% MeOH to give 0.107 g (60%) of the fully protected tetramer ApApA Sp * A 48 as a colorless foam after drying under high vacuum at 35°C. Anal . calc. for C 142 H 172 N 23 0 32 P 3 SSi s x H 2 0 (2996.5) : C 56.92, H 5.85, 10.75. Found: C 56.51, H 5.91, N 10.85. UV (MeOH) : λ„ x (loge) 277 (4.99) ; [259 (4.85)] ; [231 (4.86)] . R f on silica gel with CHCl 3 /MeOH (19/1, v/v) = 0.46. - 47 - Preparation 16 a . 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - adenylyl -2 ' - [ (O p -2- (4 -nitrophenyl) ethyl-5 '] -6-N- benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] -adenylyl - 2 ' - [ (O p -2- (4 -nitrophenyl) ethyl-5'] - 6 -N-benzoyl - 3 ' -O- [ (tert-butyl) dimethylsilyl] - (PR) -P-thioadenylyl -2 ' - [(O p -2- (4 -nitrophenyl) ethyl-5'] -6 -N-benzoyl -2 ' ,3 ' - di-O- [ (tert-butyl) dimethylsilyl] -adenosine 5 ' -OH- ApApA- p * A 41: b. 6-N-Benzoyl-3 ' -O- [ (tert-butyl) dimethylsilyl] - adenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5 ' ] -6-N- benzoyl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adenylyl- 2 ' - [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzoyl-3 ' -O- [ (tert-butvl)dimethylsilyl] - (PS) -P-thioadenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5 ' ] -6-N-benzoyl-2 ' , 3 ' - di-O- [ (tert-butyl) dimethylsilyl] -adenosine 5 ' -OH- ApApA Sp * A : The fully protected tetramer ApApA Rp * A 4J7 (0.104 g, 0.035 mmole) was stirred with 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v, 1.4 ml) at r.t. After 1.5 h, the reaction mixture was extracted with CH 2 C1 2 (3 x 40 ml) and H 2 0 (2 x 40 ml) . The combined organic phase was dried over Na 2 S0 4 and evaporated to dryness. The crude product was purified on a flash silica gel column (11 x 1 cm) and _he product eluted with 20 ml CH 2 C1 2 and 50 ml CH 2 C1 2 /1% MeOH to 5% MeOH. The product fraction (100 ml) was evaporated and dried under high vacuum to give 0.075 g (80%) of the hydroxy tetramer ApApA Rp * A 4_9. Anal . calc. for C 122 H 156 N 23 0 31 P 3 SSi s x H 2 0 (2706.5) : C 54.15, H 5.81, N 11.90. Found: C 53.97, H 6.02, N 11.65. UV (MeOH) : λ max (loge) 277 (5.00) ; [260 (4.85)] ; [234 (4.77)] . R f on silica gel with CHCl 3 /MeOH (19/1, v/v) = 0.43. The fully protected tetramer ApApA Sp * A 4 . was treated in an analogous manner through the purification stage. The crude product . 50 . was purified on two preparative silica gel plates (20 x 20 x 0.2 cm) in ___Cl 3 /MeOH (19/1, v/v) , the product band was eluted with CH 2 Cl 2 /MeOH (4/1, v/v) and evaporated to a solid foam to give 0.068 g (72%) of the 5 ' -hydroxy tetramer ApApA Sp * A 50. - 48 - Example 4 a. Adenylyl- (2 ' -5') -adenylyl- (2' -5' ) - (PR) -P-thioad¬ enylyl- (2 ' -5 ' ) -adenosine ApApA j ^'A 51: b. Adenylyl- (2 ' -5') -adenylyl- (2 ' -5' )- (PS) -P-thioad¬ enylyl- (2 ' -5 ' ) -adenosine ApApA Sp * A 52 : The corresponding 5 ' -hydroxy tetramers 4 . 9 and 50 , respectively, were deblocked separately by stirring the 5 '-hydroxy tetramer (0.067 g, 0.025 mmole) with 0.5 M DBU in absolute CH 3 CN (3 ml) at r.t. for 20 h, the solu¬ tion was neutralized with 1 M AcOH in absolute CH 3 CN (1.5 ml) and evaporated to dryness. [R f on silica gel with {EtOAc/i-PrOH/ ammonia/H 2 0, 7/1/2, v/v/v) 7/3, v/v: ApApA Rp * A = 0.58; ApApA Sp * A = 0.66. The residue was then treated with methanolic ammonia (10 ml) and after 3 days reaction time, the solvent was removed under vacuum. [R f on silica gel with EtOAc/i-PrOH/ammonia/H 2 0, 7/1/2, v/v/v) 1/1, v/v} : ApApA Rp * A = 0.38; ApApA Sp * A = 0.36] . Desilylation was done with 1 M Bu 4 NF in THF (5 ml) . The reaction mixture was stirred at r.t. for 48 h and then the solvent was evaporated in vacuum. The residue was taken up in H 2 0 (10 ml) and applied to a DEAi: Sephadex A-25 column (30 x 2 cm) . With flow rates of 2 ml/min, the pure tetramer ApApA Rp * A was eluted with 0.15-0.20 I. TEAB buffer, pH 7.5, and in the case of the tetramer ApApA Sp * A with 0.24-0.32 M TEAB buffer, pH 7.5. After evaporation and coevaporation with MeOH several times, the tetramer was applied onto eight paper sheets (25 x 50 cm) and developed in i-PrOH/ammonia/H 2 0 (6/1/3, v/v/v) . The product band was cut out, eluted with H 2 0, concentrated to a smaller volume and finally lyophilized to give 675 O.D. 260 nm units (57%) of ApApA Rp * A isomer H and 753 O.D. 260 nm (65%) of ApApA Sp * A isomer 5_2 . . ApApA Rp * A 5L: R f on cellulose in i-PrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.33. UV (H 2 0) : λ max 258 nm. HPLC: PR-18, A: 50 mM NH 4 H 2 P0 4 , pH 7.2. B: MeOH/H 2 0 (l/l, v/v) , gradient: 0-1 min, 80% A, 20% B; 1-31 min, 30% A, 70% B; retention - 49 - time: 9.70 min. ApApA Sp * A 5.2: R f on cellulose in i- PrOH/ ammonia/ H 2 0 (6/1/3, v/v/v) = 0.21. UV (H 2 0) : λ max 258 r_n. HPLC: PR-18, A: 50 mM NH 4 H 2 P0 4 , pH 7.2. B: MeOH/H 2 0 (1/1, v/v) , gradient: 0-1 min, 80% A, 20% B; 1- 31 min, 30% A, 70% B ; retention time: 13.49 min. Preparation 17 a. 6-N-Benzoyl-3' -O- [(tert-butyl)dimethylsilyl] - 5 ' -O- (monomethoxytrityl) - (PR) -P-thioadenylyl- 2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5 ' ] -6-N-benzo- yl-3 ' -O- [ (tert-butyl)dimethylsilyl] -adenylyl- 2'- [(O p -2-(4-nitrophenyl) ethyl-5'] -6-N-benzo- yl-3 ' -O- [ (tert-butvl)dimethylsilyl] -adenylyl- 2'- [(O p -2-(4-nitrophenyl) ethyl-5'] -6-N-benzo- yl-2 ' , 3 ' -di-O- [ (tert-butvl) dimethylsilyl] - adenosine A Rp * ApApA 53 : Triethylammonium 6-N-benzoyl-3 ' -O- [ (tert-butyl) - dimethylsilyl] -5 ' -O- (monomethoxytrityl) - (PR) -P-thio- adenylyl-2 ' - [ (O p -2- (4-nitrophenyl) ethyl-5 ' ] -6-N-benzoyl- 3 ' -O- [ (tert-butyl) dimethylsilyl] -adenosine-2 ' - [ (O p -2- (4- nitrophenyl) ethyl-phosphate] A Pp Αp-diester 45 (0.141 g, 0.078 mmole) and the 5' -hydroxy dimer 2S_ (0.078 g, 0.06 mmole) were coevaporated with dry pyridine (4 x 0.5 ml) and finally dissolved in dry pyridine (0.6 ml) . Then (2, 4, 6-triisopropyl) -benzenesulfonyl chloride (0.047 mg, 0.156 mmole) and 3-nitro-1, 2, 4-triazole (0.053 mg, 0.47 mmole) were added and stirred at r.t. for 21 h. The reaction mixture was diluted with CH 2 C1 2 (60 ml) and washed with H 2 0 (2 x 30 ml) , dried over Na 2 0 4 and evaporated to dryness . Pyridine was removed by coevaporation with toluene (3 x 20 ml) . The crude tetramer H was purified by flash silica gel column chromatograpy (11 x 1 cm) and eluted first with CH 2 C1 2 (50 ml) , then with CH 2 Cl 2 /l% MeOH (100 ml) , 2% MeOH (200 ml) , 3% MeOH (50 ml) and finally with CH 2 Cl 2 /5% MeOH (50 ml) . The product fraction (200 ml) was evaporated to dryness. The residue was chromatographed again on two preparative silica gel plates (20 x 20 x 0.2 cm) in toluene/EtOAc/MeOH (5/4/0.5, v/v/v) to remove small - 50 - amount of 5 ' -hydroxy dimer. The tetramer product band was eluted with CH 2 Cl 2 /MeOH (4/1, v/v) and evaporated to a solid foam to give 0.053 g (30%) of the tetramer A Rp * ApApA 51 after drying in high vacuum at 35°C. Anal . calc. for C 142 H 172 N 23 0 32 P 3 SSi 5 x 3 H 2 0 (3032.5) : C 56.24, H 5.92, N 10.62. Found: C 55.75, H. 5.71, N 9.83. UV (MeOH) : λ max (loge) 277 (4.96) ; [260 (4.82)] ; [232 (4.82)] . R f on silica gel with toluene/EtOAc/MeOH (5:4:1) = 0.78. b. 6-N-Benzoyl-3 ' -O- [(tert-butyl)dimethylsilyl] - 5 ' -O- (monomethoxytrityl) - (PS) -P-thioadenylyl- 2 ' - [(O p -2- (4-nitrophenyl) ethyl-5 '] -6-N-benzo- yl-3 ' -O- [ (tert-butvl)dimethylsilyl] -adenylyl- 2' - [(O p -2- (4-nitrophenyl)ethyl-5'] -6-N-benzo- yl-3 ' -O- [ (tert-butvl)dimethylsilyl] -adenylyl- 2 '- [(O p -2- (4-nitrophenyl) ethyl-5'] -6-N-benzo- yl-2 ' , 3 ' -di-O- [ (tert-butvl)dimethylsilyl] - adenosine A Sp * ApApA 54: A Sp * Ap-diester 46 (0.141 g, 0.078 mmole) and the 5'- hydroxy dimer 2j9 (0.078 g, 0.06 mmole) were coevaporated with dry pyridine (4 x 0.5 ml) and dissolved in dry pyridine (0.6ml) . (2,4, 6-Triisopropyl) -benzenesulfonyl chloride (0.047 mg, 0.156 mmole) and 3-nitro-l, 2 , 4- triazole (0.053 mg, 0.47 mmole) were added and the mixture was stirred at r.t. After 21 h, (2,4,6- triisopropyl)benzenesulfonyl chloride (0.024 mg, 0.079 mmole) and 3-nitro-1,2, -triazole (0.027 mg, C.24 mmole) were added again. The reaction mixture was stirred for another hour, then diluted with CH 2 C1 2 (60 ml) and washed with H 2 0 (2 x 30 ml) , dried over Na 2 S0 4 and evaporated to dryness . Further work-up was performed analagous to that described for tetramer 5! to give 43 mg (24%) of A Sp * ApApA 54 in the form of a solid foam. Anal. calc. for C 142 H 172 N 23 0 32 P 3 SSi 5 X H 2 0 (2996.5) : C 56.92, H 5.85, N 10.75. Found: C 56.63, H 6.08, N 10.18. UV (MeOH) : λ max (loge) 277 (4.96); [260 (4.84)] ; [232 (4.85)] . R f on silica ^ gel with toluene/EtOAc/MeOH (5/4/1, v/v/v) = 0.78. - 51 - Example 5 a. (PR) -P-Thioadenylyl- (2 ' -5 ' ) -adenylyl- (2 ' -5 ' ) - adenylyl- (2 ' -5 ') -adenosine A Rp * ApApA 57 : b. (PS) -P-Thioadenylyl- (2 ' -5 ') -adenylyl- (2 ' -5 ') - adenylyl- (2 ' -5 ') -adenosine A Sp * ApApA 58 : The corresponding fully protected tetramers 5! and 54 were deblocked by stirring a solution of 0.047 g (0.016 mmole) of PR tetramer 5_1 (PS tetramer 5_4= 0.032 g, 0.012 mmole) in 2% p-TsOH in CH 2 Cl 2 /MeOH (4/1, v/v; for PR: 0.5 ml; for PS: 0.38 ml) for 1 h at r.t. The reaction mixture was diluted with CH 2 C1 2 (60 ml) , washed with H 2 0 (2 x 30 ml) , dried over Na 2 S0 4 and evaporated to dryness. The crude products were purified on preparative silica gel plates (20 x 20 x 0.2 cm) in CHCl 3 /MeOH (19/1, v/v) , the product bands were eluted with CH 2 Cl 2 /MeOH (4/1, v/v) and evaporated to solid foams to give 0.034 g (80%) of the 5 ' -hydroxy A Rp * ApApA isomer 5.5 and 0.02 g (68%) of the 5 ' -hydroxy A Sp * ApApA isomer 56. The solution of the 5 '-hydroxy tetramers 5_5 (0.034 g, 0.013 mmole) and 5_6 . (0.02 g, 0.007 mmole) , respectively, were separately stirred with 0.5 M DBU in absolute CH.CN [(55: 1.5 ml; 56: 0.9 ml)] for 18 h at r.t., then neutralized by addition of 1 M AcOH [(55: 0.75 ml; 56: 0.45 ml)] and evaporated. The residue was treated with 10 ml of saturated methanolic ammonia and the solution, after stirring at r.t. for 60 h, was evaporated to dryness. Desilylation was done by treatment with 1 M Bu 4 NF in THF (2.5 ml) . After stirring at r.t. for 60 h, the solvent was removed under vacuum. Some H 2 0 (10 ml) was added to the resulting residue and applied to a DEAE Sephadex column A-25 (32 x 2 cm) and eluted with 0-0.5 M TEAB buffer, pH 7.5. The fractions of the main peak were collected, evaporated and coevaporated several times with MeOH. Further purification by paper chromatography v(i- PrOH/ammonia/H 2 0, 55/10/35, v/v/v) gave, after lyophili- zation, 347 O.D. 260 nm units (58%) of A Rp * ApApA 57 and 111 - 52 - °-D. 260 nm units (31%) of A Sp * ApApA 5_8, respectively. A Rp * ApApA 5J : UV (H 2 0) = 257 nm. R f on cellulose in i- PrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.21. HPLC: PR-18, A: 50 mM NH 4 H 2 P0 4 (pH 7.2) . B: MeOH/H 2 0 (1/1, v/v) , gradient: 0-1 min, 80% A, 20% B; 1-31 min, 30% A, 70% B; retention time: 7.47 min. A Sp * ApApA 5_8 : UV (H 2 0) = 257 nm. R f on cellulose in iPrOH/ammonia/H 2 0 (6/1/3, v/v/v) = 0.32. HPLC: PR-18, A: 50 mM NH 4 H 2 P0 4 , pH 7.2. B: MeOH/H 2 0 (1/1, v/v) , gradient: 0-1 min, 80% A, 20% B; 1- 31 min, 30% A, 70% B; retention time: 9.84 min. PREPARATION OF 2 ' ,5 ' -PHOSPHOROTHIOATE/PHOSPHO- DIESTER OLIGOADENYLATE 5 ' -MONOPHOSPHATES The phosphorothioate/phosphodiester trimer and tetramer cores were synthesized as described above in Examples 1, 2, 3, 4, and 5. The trimer and tetramer 5'- monophosphates were enzymatically synthesized according to the procedure of Sambrook et al . , Molecular Cloning - A Laboratory Manual , 2 ed. , Cold Spring Harbor Laborato¬ ry Press, pp. 5.68-5.71 (1989) from their corresponding cores and ATP with T4 polynucleotide kinase. 5'- Monophosphorylation was determined by reverse-phase HPLC analysis and confirmed by the subsequent hydrolysis of each 5 ' -monophosphate derivative by 5 ' -nucleotidase (data not shown) . Yields of phosphorylation ranged from 15% to 68%. PREPARATION OF 2 ' , 5 ' -PHOSPHOROTHIOATE/PHOSPHO- DIESTER OLIGOADENYLATE 5 ' -DIPHOSPHATE AND 5'- TRIPHOSPHATE The 5 ' -diphosphate and 5-triphosphate of the 2 ' , 5 ' -phosphorothioate/phosphodiester oligoadenylates may be prepared from the 5 ' -monophosphate by following the procedure of Example 6. - 53 - Example 6 All reactions are performed in glassware oven-dried at 125° C for 18-24 hr. A 2 ' , 5 ' -phosphorothioate/phos- phodiester oligoadenylate stereoisomer (trimer or tetramer, 400 OD units at 260 nm) is dissolved in 500 microliters of dry dimethylformamide ("DMF") and dried in vacuo in a 10 ml conical flask at 35° C. This process is repeated three times. To the dry residue, 50 micromoles of triphenylphosphine, 100 micromoles of imidazole and 50 micromoles of dipyridinyl disulfide are added. The mixture is dissolved in 500 microliters dry DMF plus 50 microliters of dry dimethylsulfoxide . The solution is stirred with a stirring bar for 2 hr at room temperature. After 2 hr the solution is homogeneous (after 30 minutes, the solution begins to change to yellow) . The solution is transferred dropwise to 10 ml of a 1% Nal/dry acetone (w/v) solution. The clear colorless precipitate which forms is the sodium salt of the 5 ' -phosphoroimidazolidate. The precipitate is centrifuged at room temperature, the supernatant is decanted, and the precipitate is washed three times with 10 ml dry acetone. The centrifuging is repeated. The precitipate is dried over P 2 0 5 in vacuo for 2 hr. The precipitate is dissolved in 200 microliters of freshly prepared 0.5 M tributylammonium pyrophosphate in dry DMF. The solution is maintained at room temperature for 18 hr after which time the DMF is removed in vacuo. The residue is dissolved in 0.25 M triethylammonium bicarbon¬ ate buffer ("TEAB") (pH 7.5) . The 5'-di and 5'-triphos- phate products are separated using a DEAE-Sephadex A25 column (HC0 3 -form; 1x20 cm) with a linear gradient of 0.25 M to 0.75 M TEAB. Fractions (10 ml) are collected. The product is observed by ultraviolet spectroscopy at 254 nm. The fractions containing the 5 ' -di and^δ'- triphosphates are separately pooled and dried in vacuo. The TEAB is removed by repeated additon of water followed - 54 - by lyophilization. The yield of the 5 ' -diphosphate is about 5%; the yield of the 5 ' -triphosphate is about 60%. STABILITY OF THE PHOSPHOROTHIOATE/PHOSPHODIESTER TRIMER AND TETRAMER CORE DERIVATIVES TO SERUM PHOSPHODIESTERASE The stability of authentic 2-5A and phosphoro- thioate/phosphodiester trimer and tetramer core deriv- atives (300 μM) was determined by incubaticn in 200 μL of RPMI-1640 medium supplemented with 10% fetal calf serum in 5% C0 2 -in-air at 37°C. Aliquots (30 μL) were removed at time zero and 6 hours. The hydrolysis products were identified by HPLC as described in Kariko e_t al., Biochemistry 26: 7127-7135 (1987) Under the conditions described therein, authentic A 2 and A 3 were completely hydrolyzed to inosine and hypoxanthine in 20 min (Table 1) , while A Rp * A and A Sp * A were not hydrolyzed. No hydrolysis of the phosphorothioate/phosphodiester trimer core derivatives was observed. However, the phosphorothioate/ phosphodiester tetramer core deriva¬ tives were hydrolyzed from the 5'- or 2 ', 3 ' -terminus, depending on the location of the phosphorothioate- substituted internucleotide linkage. For example, A Rp * ApApA and A Sp * ApApA were 50% degraded to their respective dimer cores, A Rp * A and A Sp * A, whereas ApA Rp * ApA and ApA Sp * ApA are degraded from the 2 ' , 3 ' -terminus to form the trimer cores, ApA Rp * A and ApA Sp * A. ApApA Rp * A and ApApA Sp * A are degraded from the 5 '-terminus to yield ApA Rp * A and ApA Sp * A, respectively. - 55 Table 1. Hydrolysis of Phosphorothioate/Phosphodiester Trimer and Tetramer 2-5A Core Derivatives by Serum Phosphodiesterases a Incubations were for 6 h as described in text . Number in parentheses indicates the time at which 100% hydrol¬ ysis was observed. b Identified as described by Kariko et al . . , Biochemistry 26 : 7127-7135 (1987) . - 56 - BINDING OF 2 ' ,5 ' -PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES TO RNase L The affinity of the phosphorothioate/ phosphodiester 2-5A derivatives for RNase L was determined in radiobinding assays according to the method of Knight et al., Meth. Enzymol 79: 216-227 (1981) . Authentic A 3 , pA 3 and p 3 A 3 bind to RNase L with IC 50 values of 1 x 10 "6 M, 1 x 10 "9 M and 1 x 10 "9 M, respectively. According to the invention, the binding of the phos- phorothioate/phosphodiester core and their 5 ' -mono- phosphates to RNase L was equivalent to or slightly better than the corresponding authentic 2-5A cores and 5 ' -monophosphates, with IC 50 values from 8 x 10 "7 M to 8 x 10 "6 M for the cores and from 1 x 10 "8 M to 1 x 10 "9 M for the 5 ' -monophosphates. The trimer and tetramer 2-5A core derivatives with phosphorothioate substitution in the first internucleotide linkage from the 5 '-terminus exhibited lower affinity compared to those with phosphorothioate substitution in the second or third internucleotide linkage. ACTIVATION OF RNASE L BY PHOSPHOROTHIOATE/ PHOSPHODIESTER DERIVATIVES OF 2-5A Correlation of biological properties with absolute configuration has only been possible with the preparation of the fully resolved 2 ' , 5 ' -phosphorothioate/phosphodies- ter adenylate trimer cores. However, the trimer core compounds have been found to bind and/or activate RNase L only modestly. RNase L activation by the 2 ', 5 ' -phos¬ phorothioate core molecules is significantly enhanced by 5 ' -phosphorylation. Core-cellulose assays were performed according to the method of Silverman, Anal. Biochem. 144 : 450-460 (1985) and Kariko et. al . . (1987) , supra, in which RNase L was partially purified from L929 cell extracts by immobilization on 2-5A 4 core-cellulose. Activation of RNase L was measured by the conversion of poly(U) - 57 - [ 32 p]pCp to acid soluble fragments. The results indicate that authentic p 3 A 3 , p 3 A 4 , pA 3 and pA 4 have IC 50 values of 5 x 10 "10 M, 5 x 10 "10 M, 2 x 10 "7 M and 2 x 10 "8 M, respectively, while surprisingly, of the phosphor- othioate/phosphodiester trimer core derivatives, only ApAMA can activate RNase L (IC 50 of 5 x 10 "7 M) (Figure 1A, Δ) . Three of the phosphorothioate/phosphodiester trimer 5 ' -monophosphates can activate RNase L, with pApA Hp * A being the most potent activator of RNase L (IC 50 of 1 x 10 "9 M) (Figure IB, Δ) . pA Rp * ApA (□) and pA Sp * ApA (■) are 100-fold less potent activators of RNase L. Of the six phosphorothioate/phosphodiester tetramer core derivatives, only ApA Rp * ApA (α) and ApApA Rp * A (Δ) can activate RNase L (IC 50 of 5 x 10 "7 M and 5 x 10 "7 M, respectively) (Figure 1C) . Five of the six phosphorothioate/phosphodiester tetramer 5 ' - monophosphates activate RNase L (IC 50 values >6 x 10 "7 M to 8 x 10 "10 M) . The pApA Sp * ApA enantiomer did not activate RNase L, even at concentrations as high as 1 x 10 "5 M (Figure ID, ■) . Activation of RNase L by the phosphorothioate/phos- phodiester trimer and tetramer 2-5A derivatives was also measured in a rRNA cleavage assay using L929 cell extracts according to the method of Kariko e_t aJL . (1987) , supra, in which extracts of L929 cells were incubated for 1 h at 30°C in the presence or absence of 2-5A or 2-5A derivative. Consistent with the results from the core-cellulose assays (Figure 1A) , ApA Rp * A (1 x 10 "6 M) was the only trimer core able to activate RNase L to cleave rRNA to the well-characterized specific cleavage products (SCP) of RNase L (Figure 2A, lane 5) . A Rp * ApA, A Sp * ApA and ApA Sp * A, as well as authentic A 3 , did not activate RNase L at concentrations as high as 1 x 10 "6 M (Figure 2A, lanes 3, 4, 6, 7) . Authentic p 3 A 3 "was active at 1 x 10 "8 M (Figure 2A, lane 2) . The corres¬ ponding 5 ' -monophosphates, pA Rp * ApA, pA Sp * ApA and pApA Rp * A, - 58 - activated at 1 x 10 "7 M, 1 x 10 "7 M, and 2 x 10 "9 M, respectively (Figure 2B, lanes 4-6) , as compared with pA 3 which was active at 1 x 10" 6 M (lane 3) . Incubation with pApA Sp * A, even at concentrations as high as 5 x 10 6 M, did not result in detectable rRNA degradation (data not shown) . Comparable degradation of rRNA was observed with two of the six phosphorothioate/phosphodiester tetramer core derivatives relative to the authentic p 3 A 4 control. ApA Rp * ApA and ApApA Rp * A activated RNase L at 1 x 10 "5 M (Figure 3A, lanes 6 and 8) . As was observed in the core- cellulose assays (Figure ID) , five of the six phosphorothioate/phosphodiester tetramer 5 '-mono¬ phosphates were able to activate RNase L (pA Rp * ApApA, pA Sp * ApApA, pApA Rp * ApA, pApApA Rp * A, and pApApA Sp * A) (Figure 3B, lanes 4, 5, 6, 8, 9, respectively) . The most efficient activator or RNase L was pApA Rp * ApA (1 x 10 "8 M) (Figure 3B, lane 6) , while pApA Sp * ApA was an antagonist of RNase L activation and was unable to activate RNase L even at concentrations as high as 1 x 10 "5 M (Figure 3B, lane 7) . INHIBITION OF RNASE L ACTIVATION BY pApA ?p * A. The high affinity of pApA Sp * A for RNase L and the observation that pApA Sp * A does not activate RNase L, suggests that it might be a specific inhibitor of RNase L. Indeed, pApA Sp * A inhibits the activation of RNase L by p 3 A 3 or pApA Rp * A (Figure 4A) . Authentic p 3 A 3 activates RNase L to hydrolyze 28S and 18S rRNA to SCP at 10 "9 M or 10 "8 M (lanes 1 and 3) . However, addition of pApA Sp * A (lO "6 M) results in the inhibition of RNase L-catalyzed hydrolysis of rRNA (lanes 2 and 4) . Similarly, whereas pApA Rp * A activates RNase L at 10 "9 M or 10 "8 M (lanes 5 and 7) , the addition of pApA Sp * A (10 "6 M) inhibits this activation (lanes 6 and 8) . The inhibitory activity of pApA Sp * A was also observed with partially-purified RNase - 59 - 1 (Figure 4B) . p 3 A 3 activates RNase L with an IC 50 value of 5 x 10 "1C M (•) ; however, upon addition of pApA Sp * A (1 x 10 "; M) , the observed IC 50 value shifts to 1 x 10 "8 M ;o) , demonstrating specific inhibition of p 3 A 3 -mediated activation of RNase L by pApA Sp * A. Notwithstanding, pApA Sp * A is useful as a probe in the evaluation of the role of RNase L in the interferon- induced biological cascade. Most importantly, pApA Sp * A selectively inhibits activation of RNase L at physio- logical concentrations, and is metabolically stable to specific and non-specific phosphodiesterases. The molecule provides the means to selectively inhibit RNase 1 activation. Moreover, it is expected that pApA Sp * A has therapeu- _ic activity. Individuals afflicted with chronic —yelogenous leukemia ("CML") display a highly elevated RNase L activity, as evidenced by novel rRNA CML-specific cleavage products. Thus, pApA Sp * A, which is a metaboli¬ cally stable inhibitor of RNase L, has potential utility in treating chronic myelogenous leukemia. Additionally, individuals afflicted with chronic fatigue syndrome ("CFS") (also known as myalgic encepha- lomyelitiε (ME) or low natural killer ("NK") cell disease) and other HHV-6 related disorders also display a highly elevated RNase L activity compared to controls [mean basal level = 466 ± 23 compared to 123 ± 12 in controls; p < .0001) , Suhadolnik et. al., Clinical Infectious Disease 18 (SUPPL. 1) : 96-104 (1994) . In experiments performed using extracts of peripheral blood mononuclear cells ("PBMC") from individuals with CFS before and during therapy with a biological response modifier, poly (I) -poly (C 12 U) (mismatched dsRNS, Ampligen ® ) , as compared to healthy individuals, the mean basal latent 2-5A synthetase level in PMBC extracts'was significantly decreased following therapy (610 ± 220 picomoles 2-5A/mg protein/hour) compared to controls - 60 - (2035 ± 325 picomoles 2-5A/mg protein/hour, P< 0001) . Id. Further, all pretherapy PBMC extracts tested were positive for human herpes virus-6 (HHV-6) replication. Therapy resulted in a significant decrease in HHV-6 activity (p <.01) and down regulation of the 2-5A synthetase/RNase L pathway in temporal association with clinical and neuropsychological improvement. Without wishing to be bound by any theory, it appears that the upregulated 2-5A pathway observed in CFS pretherapy is consistent with a hypothesis that an activated immune state and persistent viral infection may play a pathogen- esis of CFS. Thus, pApA Sp * A, which, as stated above, is a metabolically stable inhibitor of RNase L, also has potential utility in treating CFS. INHIBITION OF CELL GROWTH BY PHOSPHOROTHIOATE/PHOS- PHODIESTER 2-5A CORE DERIVATIVES Cell viability was determined by Trypan blue exclusion. Post-treatment colony forming ability was determined by growth in microtiter wells as outlined in the procedure of Kraemer et. a . Mutation Res. 72 : 285- 292 (1980) . The results indicate that no decrease in survival or inhibition of Sup Tl cell growth in microtiter plates was observed with any of the dimer, trimer or tetramer phosphorothioate/phosphodiester 2-5A core derivatives. On the basis of the lack of cyto- toxicity and estimated uptake of 1% on a previous report with the cordycepin derivative of 2-5A, Suhadolnik et . al., Nucleosides and Nucleotides 2: 351-366 (1983), 3 x 10 "4 M was chosen as the concentration at which to screen the phosphorothioate/phosphodiester derivatives for anti-HIV-1 activity. - 61 - EFFECT OF 2 ' ,5' -PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES ON INHIBITION OF HIV-l-INDUCED SYNCYTIA FORMATION The infected centers assay as described by Henderson et al . . , Virology 182 : 186-198 (1991) , was used to measure the ability of the phosphorothioate/phosphodiester derivatives of 2-5A trimer and tetramer cores to inhibit HIV-l induced syncytia formation, an indicator of HIV-l replication in T cells. Freshly isolated peripheral blood lymphocytes (PBL) were treated with 2-5A or derivatives for 2 h and infected with HIV-l strain IIIB at a m.o.i. of approximately 0.1. The infected PBL were maintained in RPMI-1640 medium supplemented with 10% (v/v) heat-inactivated fetal calf serum at 37°C in a humidified 5% C0 2 in air atmosphere. After 48 h, the cells were washed twice in Hank's balaned salt solution, serially diluted and seeded into multiple wells of a 96- well microtiter plate. Immediately, 2 x 10 5 exponentially growing Sup Tl cells were added to each well; Sup Tl cells readily form a syncytium with a cell which is productively infected with HIV-l. The wells were examined daily for the presence of syncytia, using a tissue culture microscope. The first signs of syncytia formation can be seen in 12 h, with some complete syncytia developing by 24 h. Final results were read at 72 h. Each syncytium was counted as a single infected cell . The number of syncytia per seeded cell is deter- mined and expressed as an infected center per infected cell. In the control (no 2-5A derivative added) , 100% syncytia formation was equivalent to 12 ± 3 syncytia per 200 HIV-l infected cells. The data is shown in Figures 5A and 5B. As shown in Figure 5A, ApA Rp * A was a highly efficient inhibitor of syncytia formation, with 100% inhibition observed at 3 x 10 "4 M. Its PS enantiomer, ApA Sp * A inhibited syncytia formation 78%. A Rp * ApA and A Sp * ApA inhibited syncytia - 62 - formation only 10% and 15%, respectively. Authentic A, and A 4 (3 x 10 "4 M) inhibited syncytia formation 21% and 15%, respectively while adenosine (9 x 10 "4 M) did not inhibit syncytia formation. Of the six phosphoro- thioate/phosphodiester tetramer core derivatives, ApApA Rp * A and ApApA Sp * A were the most inhibitory (90% and 76% inhibition, respectively) (Figure 5B) . ApA Rp * ApA, ApA Sp * ApA, A Rp * ApApA and A Sp * ApApA inhibited syncytia for¬ mation 26%, 32%, 16% and 18%, respectively. Adenosine and A 4 (Figure 5B) , as well as the A Rp * A and A Sp * A dimers, adenine or 3 ' , 5 ' -A 4 (data not shown) , were not able to inhibit syncytia formation. EFFECT OF 2 ' , 5 ' -PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES ON HIV-l REVERSE TRANSCRIPTASE ACTIVITY Sup Tl cells were treated with 2-5A or a phosphoro- thioate/phosphodiester derivative at 300 μM for 6 hours and then infected with HIV-l at a multiplicity of infection (M.O.I) of approximately 0.1. Adenosine and adenine were tested at 900 μM. At 96 hours post- infection, culture supernatant was removed and HIV-l RT activity was assayed in triplicate as described by Henderson e_t al. , Virology 182:186-198 (1991) . Briefly in this method, 25 μl of culture supernatant was added to a 50 μl cocktail containing 50 mM Tris (pH 8.0) , 20 mM dithiothreitol, 10 mM MgCl 2 , 60 mM NaCl, 0.05 Nonidet p-40, 5 μg/ml oligodeoxythymidylic acid, 10 μg/ml polyriboadenylic acid, 10 μM deoxythymidine triphosphate and lmCi [ 3 P] thymidine 5 ' -triphosphate. The mixture was incubated at 37°C for 2 hours. Fifty microliters of the cocktail were then spotted onto diethylaminoethyl (DEAE) paper, dried, washed with 2X SSC solution (three times for 10 minutes each time) and 95% ethanol (two times for 5 minutes each time) , dried and exposed to radiographic film for 18 to 24 hours at -80°C. The filters were cut 63 - and final quantitation was determined by scintillation spectrometry. The data for the HIV-l RT activity is shown in Table 2. As indicated, the trimer ApA Sp * A was the most effi¬ cient inhibitor of HIV-l RT activity (78%) . On the contrary, its PR enantiomer, ApA Rp * A inhibited HIV-l reverse transcription by 31%. Similarly, the tetramer with the PS phosphorothioate/phosphodiester linkage adjacent to the 2' terminal linkage, ApApA Sp * A, was able co suppress RT activity 62% while its PR counterpart was only 38% effective in inhibiting this activity. Table 2. Inhibition of HIV-l Reverse Transcriptase Activity by Phosphorothioate/Phosphodiester 2-5A 2-5A or Percent Inhibition of HIV-l Derivative Reverse Transcriptase 1 32 56 31 78 57 54 52 42 38 62 26 8 0 4 \Average of triplicate determinations, Intra-assay variation for replicates was <10%. - 64 - The compounds of the present invention may be combined with appropriate pharmaceutical or agricultural carriers to form an antiviral composition. For pharmaceutical use, the compounds of the inven- tion may be taken up in pharmaceutically acceptable carriers, such as, solutions, suspensions, tablets, capsules, ointments, elixirs and injectable composition and the like. They are administered to subjects suf¬ fering from viral infection. The dosage administered depends upon the nature and severity of the infection, the disease stage, and, when administered systematically, the size and weight of the infected subject . The compounds are generally administered in the form of water-soluble salts. Pharmaceutically acceptable water soluble salts include, for example, the sodium, potassium or ammonium salts of the active compounds. They are readily dissolved in water or saline solution. Thus, the preferred formulation for pharmacological use comprises a saline solution of the desired compound in salt form. The formulation may further contain an agent, such as a sugar or protein, to maintain osmotic balance. The salt form of the compound is preferred owing to the relatively high acidity (about pH 3) of the acid form of the compounds. The compounds of the invention may be used as a treatment or prophylactically for humans and animals from viral infectives such as Herpes simplex, rhinovirus, hepatitis and other infections of the hepatitis virus family, Epstein Barr virus, measles virus, multiple sclerosis (which may be caused by a viral agent) and the various Human Immunodeficiency Viruses ("HIV") , such as HIV-l, which causes cutaneous T cell lymphoma, HIV-2, which causes Sezary lymphoma, and HIV-3, which is responsible for Acquired Immune Deficiency Syndrome ("AIDS") . The compounds of the invention inhibit the HIV-l Induced Syncytia formation. - 65 - The compounds may be applied topically to treat skin cancers caused by radiation, carcinogens or viral agents . Such skin cancers include cutaneous T-cell lymphoma, Sezary lymphoma, Xeroderma pigmentosium, ataxia telangiectasia and Bloom' s syndrome. A sufficient amount of a preparation containing a compound of the invention is applied to cover the lesion or affected area. An effective concentration of active agent is between about 10 "3 M and 10 "5 M, with 10 "4 M being preferred. The compounds of the present invention may also be used to treat plant-infecting virus, particularly tobacco mosaic virus, and other viruses which cause necrosis in turnips, cucumber, orchids and in other plants. Such viruses include, but are not limited to, tobacco vein mottling virus, vesicular stomatitis virus, vaccinia virus, turnip necrosis virus, and cymbidium orchid virus . The compounds may be administered effectively to plants by topical application by abrasion of the leaf surface, aerosol spray, treatment of the soil, spraying, or dusting. An effective antiviral composition may be formed by combining one or more of the compounds of the inven¬ tion with a carrier material suitable for agricultural use. While the individual stereoisomers are preferred for pharmaceutical use, mixtures of one or more of stereoisomers may be employed in agricultural applica¬ tions. The active compound may also be administered by spraying insect vectors such as aphids, thrips and whiteflies which carry virus to plants. The dosage administered depends upon the severity of the infection. The compounds of the invention may be applied to plant seeds prior to germination to control viruses contained in the germ plasm. The seeds may be soaked in a solution of polyethylene glycol ("PEG") containing one or more of the compounds. PEG brings the seeds to physiological activity and arrest. The relative con- - 66 - centration of active compound to PEG depends upon the type of seed under treatment. Plants may be effectively treated with an aqueous formulation containing from about 10 "1 to about 10 "2 M concentration of active ingredient. The compounds of the invention may be applied at very low concentrations. An effective amount of active ingredient on the plant surface is from about 10 "8 to about 10 "12 mole per cm 2 of plant surface area, with about 10 "10 mole to about 10 "12 mole per cm 2 being preferred. For the typical tobacco plant of 1,000 cm 2 , 10 "5 M of compound is effective. At this rate, one pound of active ingredient is sufficient to treat 2 x 10 8 tobacco plants. For agricultural application, the compounds are advantageously administered in the form of water-soluble salts, e.g. ammonium or potassium salts. Sodium salts are generally avoided in treating edible plants. The compounds of the invention are readily dissolved in water, particularly at such low concentrations. Aqueous formulations for agricultural use may optionally contain a sticker and/or a UV-stabilizer. Such agents are well-known to those skilled in the art. Fatty acids (1%) are useful as spreader sticker agents. Effective UV-stabilizers include, for example, p-aminobenzoic acid. For antiviral use in mammals, the compounds of the invention are administered parenterally, such as intravenously, intraarterially, intramuscularly, subcutaneously or when administered as an anti-cancer agent, intratumorally. The preferred route of adminis- tration for antiviral therapy is intravenous injection. The compounds of the invention may be administered to mammals at very low concentrations. The actual dosage administered may take into account the size and weight of the patient, whether the nature of the treatment is prophylactic or therapeutic in nature, the age, health and sex of the patient, the route of administration, the - 67 - nature and stage of the affliction, and other factors. An effective daily dosage of active ingredient, based upon m vivo studies involving other 2-5A analogues, is from about 0.25 g per 70 kg of body weight (approximately 152 lbs) to about 2.5 g per 70 kg of body weight. The preferred daily dosage is about 0.5 g per 70 kg of body weight. Those skilled in the art should readily be able to derive appropriate dosages and schedules of aministration to suit the specific circumstance and needs of the patient. It is expected that an effective treatment regimen includes administration of the daily dosage for two days. Treatment is continued at least until the disease condition is substantially abated. Preferably, the therapeutic end point is determined by testing for the continued presence of viral DNA. Such testing can be done by polymerase chain reaction (PCR) in which the presence of viral DNA is assayed according to convential PCR. PCR primers of appropriate nucleotide sequences for amplification of viral DNA can be prepared from known viral nucleotide sequences. To obtain DNA for testing, patient peripheral blood mononuclear cells are lysed with an appropriate lysing agent, such as NP- 40. Alternatively, testing for the continued presence of the virus can be performed by an antigen-antibody assay using any of the known monoclonal or polyclonal antisera against a protein antigen of the target virus' protein coat. For example, an antigen-antibody assay may be employed to detect any of the protein antigen in the antigens HIV protein coat, for example, the gpl20, pl7 or p24. Moreover, the target antigen is not limited merely to coat protein antigens. Antisera can be targeted against a suitable non-coat protein antigen, such as the HIV reverse transcriptase (RT) molecule. - 68 - Monoclonal antibodies to HIV RT are known. Sobol et al . , Biochemistry 30 : 10623-10631 (1991) . Additionally, testing for the presence of the infecting virus during or post-treatment could be accom- plished by an assay which assesses the viral load in the patient's blood stream. This can be done by determining the level of syncytia formation, i.e., by measuring the formation of viral particles. See procedure outlined in Henderson et . aJL., Virology 182 : 186-198 (1994) . In addition to administration with conventional carriers, the compounds of the present invention may be administered by a variety of specialized oligonucleotide or nucleic acid delivery techniques. 2-5A and its analogues have been successfully encapsulated in various encapsulating materials, such as in unilamellar liposomes and delivered with the aid of monoclonal antibodies to cells, Bayard et. al. , Eur. J. Biochem. 151 :319-325 (1985) . Reconstituted Sendai virus envelopes have been successfully used to deliver RNA and DNA to cells, Arad et. al., Biochem. Biophvs. Acta. 859: 88-94 (1986) . Moreover, the virus envelope is not limited to Sendai virus, but could include encapsulation in any retroviral amphotrophic particle. For example, an HIV envelope could be formed from any part or all of the outer protein coat of a non-infectious HIV particle. Such particles as gp 120 can be cloned by known recombinant techniques. These techniques may be utilized for introduction of the present 2 ' , 5 ' -phosphorothioate/phosphodiesteroligoadeny- lates into cells. It is further contemplated that the compounds of the invention may be administered in the form of prodrugs in which lipophilic groups are attached to, for example, the 5 '-terminal hydroxyl group of the core compound. - 69 - CONJUGATION OF 2 ' , 5 ' -PHOSPHOROTHIOATE TETRAMER ADENYLATES The 2 ' , 5 ' -phosphorothioate/phosphodiester tetramers of the invention may be conjugated with the carrier (poly) L-lysine. (Poly) L-lysine has been shown to be an effective vector for introducing 2 ' , 5 ' -oligoadenylates and analogues into intact cells. Bayard et . al . , Biochemistry 25: 3730-3736 (1986) Poly(L-lysine) conjugation to trimer molecules is not feasible, owing to the destruction of the 2 '-terminal ribosyl moiety and subsequent inactivation of the molecule. Conjugation to poly (L-lysine) permits efficient intracellular transport of the 2 ' , 5 ' -phosphorothioate/phosphodiester oligoadenylates of the invention, while preserving intact within the conjugate the trimer moiety believed necessary for good biological activity. The conjugates are formed by introducing two aldehyde functions at the 2' end of the tetramer by periodate oxidation of the alpha-glycol group of the ribose residue. The resulting aldehyde groups are then randomly coupled to the epsilon-amino groups of lysine residues of poly (L-lysine) by Schiff base formation, and then reduced with sodium cyanoborohydride at pH 8.0. This procedure converts the 2 ' , 3 ' -terminal ribose ring into a morpholine structure. The poly(L-lysine) peptide preferably contains from about 60 to about 70 lysine residues. From about five to about ten of the lysine residues are coupled in this manner to tetramer moieties. The resulting 2 ' , 5 ' -phosphorothioate/phosphodies- ter/ (poly) L-lysine conjugates may then be isolated by gel filtration chromatography on a Sephadex G-50 column. The 2 ' , 5 ' -phosphorothioate/phosphodiester oligoadenylate poly(L-lysine) conjugates have the for¬ mula: 70 NH, NH, (CH,) (CH 2 ) NH 2 CHCO-R q -NHCHCOOH wherein q is an integer from about 60 to about 70 and each R is independently R' or From about five to about ten of the R groups comprise R' . The R' group has the following formula: - 71 - wherein is zero, 1, 2 or 3 ; and R 3 , R 4 and R ? are independently selected from the group of oxygen and sulfur, provided that all R 3 , R 4 and R 5 may not be oxygen, and further provided that all R 3 , R 4 and R 5 may not be sulfur. The conjugates may be advantageously prepared by the procedure of Bayard et . al . ., Biochemistry 25 : 3730- 3736 (1986) : Example 7 Preparation of Poly(L-lysine) /2 ', 5 ' -Phosphoro- thioate/Phosphodiester Oligoadenylate Conjugates A 4-microliter aliquot of sodium metaperiodate (0.6 micromole in 0.1 M sodium acetate buffer, pH 4.75) is added to an ice-cold solution of 2 ' , 5 ' -phosphoro- thioate/phosphodiester tetramer adenylate in 400 micro¬ liter of distiller water. The reaction mixture is stirred on ice for 30 min; 400 microliter of poly(L- lysine) (0.14 micromole in 0.2 M phosphate buffer, pH 8.0) and 200 microliter of sodium cyanoborohydride (20 micromole in 0.2 M phosphate buffer, pH 8.0) are added. The mixture is incubated for 2 h at room temperature and then loaded on a Sephadex G-50 column equilibrated with 0.1 M sodium acetate buffer, pH 4.75. Each fraction is assayed for its phosphorothioate/phosphodiester oligoadenylate/poly(L-lysine) content by the method described by Lowry e_t al. , J. Biol . Chem. 193 :265-275 (1951) , and by absorbance at 260 nm. Conjugation of the 2 ' , 5 ' -phosphothioate/phospho- diester tetramer to poly(L-lysine) leaves the remaining three 2 ',5' -linked phosphorothioate/phosphodiester adenylic residues intact for optimal RNase L binding and activation. - 72 - LIPOSOME ENCAPSULATION OF 2', 5' PHOSPHOROTHI- OATE/PHOSPHODIESTER OLIGOADENYLATES Encapsulation of the compounds of the present m- vention comprises another attractive non-disruptive technique for introduction into cells. Liposome encap¬ sulation may be advantageously accomplished according to the technique described by Kondorosi et a_l. , FEBS Lett. 120:37-40 (1980) . Example 8 Preparation of Large Unilamellar Vesicles (Lipo- somes) Loaded with 2 ' , 5 ' -Phosphoro- thioate/Phosphodiester Oligoadenylates Briefly, a phospholipid mixture from bovine brain (Sigma Chemical Co., Folch fraction III composed of 80- 85% phosphatidylserine with the remaining 15% composed of other brain lipids; 35 mg) is suspended in 5 ml of buffer A [0.1 M NaCl, 2 mM histidine, 2 mM N- tris (hydroxymethyl)methyl-2-aminoethane sulfonic acit ("TES") , 0.4 mM EDTA (pH 7.4) by vortexing. The sus¬ pension is sonicated under nitrogen for 10 minutes at 0°C. The suspension is further incubated for 1 hr at 37°C after adjusting the final concentration of Ca++ to 20 mM by the addition of 125 microliters of 800 M CaCl- . The resulting precipitate is sedimented by centrifugation (2500 x g, 10 min) , vortexing and mixing with 100 microliters of 1 x 10 "4 M 2 ' , 5 ' -phosphorothioate/phospho- diester oligoadenylate, which is dissovled in phosphate- buffered saline. The final concentration of EDTA is then adjusted to 120 mM by the addition of 400 microliters of buffer B [150 mM EDTA, pH 7.4, 0.1 M NaCl, 2 mM histidine, 2 mM TES] . Liposomes are formed after incubation of this mixture for 30 minutes at 37°C. The excess of EDTA and non-encapsulated components N are removed by passing the liposomes through a Sephadex G-25 column which is equilibrated with phosphate-buffered saline . About 10% of the 2 ' , 5 ' -phosphorothioate/phospho- - 73 - diester oligoadenylate is encapsulated into liposomes by this procedure. The liposome suspension is stable at 4°C for one week following preparation. PREPARATION OF RECONSTITUTED SENDAI VIRUS ENVELOPES CONTAINING 2 ' , 5 ' -PHOSPHOROTHIOATE/PHOSPHODIESTER OLIGOADENYLATES Reconstituted Sendai virus envelopes may be used as efficient vehicles for the introduction of poly- nucleotides into cells. Arad et . al . ., Biochimica et Biophysica Acta 859 : 88-94 (1986) , discloses introduction of poly(I) • poly(C) into cultured cells by the use of reconstituted Sendai virus envelopes. Fusion of the aforesaid reconstituted Sendai virus envelopes leads to introduction of the enclosed macromolecules into the recipient cell cytoplasm. Reconstituted Sendai virus envelopes may be obtained by detergent solubilization of intact Sendai virus particles. The reconstituted envelopes are fusogenic vesicles consisting of the viral envelope phospholids and their glycoproteins, devoid of the viral genomic RNA. Incorporation of the compounds of the present in¬ vention into reconstituted Sendai virus envelopes for fusion-mediated micro-injection may be accomplished by following the procedure of Arad et . a_l . , Biochimica et Biophysica Acta 859 : 88-94 (1986) . Briefly, a pellet of Sendai virus particles (1.5 mg protein) is dissolved in 30 microliters of a solution containing 10% Triton X-100, 100 mM NaCl, 50 mM Tris-HCl (pH 7.4) and 0.1 mM phenylmethylsulfonyl fluoride (Triton X-100 :protein ratio, 2:1, w/w) . To the clear supernatant obtained after centrifugation, 2 ' , 5 ' -phosphorothioate/phos- phodiester oligoadenylate dissolved in a solution A (160 mM NaCl, 20 mM Tris-HCl, (pH 7.4)) is added to give a final concentration of active ingredient of 5-20 mg/ml and a final volume of 150 microliters. Triton X-100 is removed from the supernatant by direct addition of 40 - 74 - mg of SM-2 Bio-Beads. The turbid suspension obtained (containing reconstituted Sendai virus envelopes) is centrifuged at 100,000 x g for 1 h. The pellet, containing about 10% of the original viral protein, is then suspended in solution A to give a final protein concentration of 25 micrograms/ml. All references cited with respect to synthetic, preparative and analytical procedures are incorporated by reference. The present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof and, accordingly, reference should be made to the appended claims, rather than to the foregoing specification, as indicating the scope of the invention.

Claims: - 75 - CLAIMS 1. A compound of the formula wherein m is zero, 1, 2, or 3; n and q are selected from the group of zero and 1, provided that n and q may not both be zero; and where R, R x and R 2 , are independently selected from the group of oxygen and sulfur, provided that all R, R and R 2 may not be oxygen, and further provided that all R, R x and R 2 may not be sulfur; or water-soluble salt thereof. - 76 - A compound according to claim 1 wherein m is 1. 3. A compound according to claim 1 wherein m is 0 4. A compound according to any of claims 1-3 wherein n is 1 and q is zero. 5. A compound according to any of claims 1-3 wherein n is 1 and q is 1. 6. A compound according to claim 1 selected from the group consisting of adenylyl- (2 ' , 5 ' ) -P-thioadenylyl- (2 ', 5 ') -adenosine, the 5 '-mono-, di-, and triphosphates thereof, and water-soluble salts of any of them. 7. A compound according to claim 1 selected from the group consisting of adenylyl- (2 ' , 5 ' ) -P-thioadenylyl- (2 ', 5 ') -adenylyl- (2 ', 5 ') -adenosine, the 5 '-mono-, di-, and triphosphates thereof, and water-soluble salts of any of them. 8. A compound according to claim 1 selected from the group consisting of adenylyl- (2 ', 5 ') -adenylyl- (2 ' , 5 ' ) -P-thioadenylyl- (2 ' ,5 ' ) -adenosine, the 5 ' -mono- , di-, and triphosphates thereof, and water-soluble salts of any of them. 9. A compound according to claim 1 selected from the group consisting of adenylyl- (2 ' , 5 ' ) -P-thioadenylyl- (2 ' , 5 ' ) -P-thioadenylyl- (2 ' , 5 ' ) -adenosine, the 5 ' -mono- , di-, and triphosphates thereof, and water-soluble salts of any of them. - 77 - 10. A pharmaceutical composition comprising a pharmaceutical carrier and a compound according to any of claims 1-9. 11. A pharmaceutical composition according to claim 10 wherein m is 1. 12. A pharmaceutical composition according to claim 10 wherein the carrier is selected from the group consisting of reconstituted Sendai virus envelope and liposome . 13. A method of treating viral infection in a plant or mammal comprising administering thereto an antiviral effective amount of a compound according to any of claims 1-9. 14. A method according to claim 13 wherein m is 1. 15. An isolated optical isomer according to any of claims 1-9, or water-soluble salt of such isolated isomer. 16. An isomer according to claim 15 having one or two internucleotide phosphorothioate groups at least one of which is of the PR configuration. 17. An isomer according to claim 16 selected from the group consisting of adenylyl- (2 ' , 5 ' ) - (PR) -P- thioadenylyl- (2 ', 5 ') -adenosine, the 5' mono-, di-, and - 78 - triphosphates thereof, and water-soluble salts of any of them. 18. An isomer according to claim 16 selected from the group consisting of adenylyl- (2 ' , 5 ' ) - (PS) -P- thioadenylyl- (2 ' , 5 ' ) adenosine, the 5' mono-, di-, and triphosphates thereof, and water-soluble salts of any of them. 19. An isomer according to claim 16 selected from the group consisting of adenylyl- (2 ' , 5 ' ) - (PR) -P- thioadenylyl- (2 ' , 5 ' ) -adenylyl- (2 ' , 5 ,' ) -adenosine, the 5 ' mono-, di-, and triphosphates thereof, and water-soluble salts of any of them. 20. An isomer according to claim 16 selected from the group consisting of adenylyl- (2 ' , 5 ' ) - (PS) -P- thioadenylyl- (2 ' , 5 ' ) -adenylyl- (2 ' , 5 ' ) -adenosine, the 5 ' mono-, di-, triphosphates thereof, and water-soluble salts of any of them. 21. An isomer according to claim 16 selected from the group consisting of adenylyl- (2 ', 5 ') -adenylyl- (2 ', 5 ') - (PR) -P-thioadenylyl- (2 ' ,5 ') -adenosine, the 5' mono-, di-, and triphosphates thereof, and water-soluble salts of any of them. 22. An isomer compound according to claim 16 selected from the group consisting of adenylyl- (2 ', 5 ') - adenylyl- (2 ' , " 5 ' ) - (PS) -P-thioadenylyl- (2 ' , 5' ) -adenosine, the 5' mono-, di-, and triphosphates thereof, and water- soluble salts of any of them. 23. A method of treating viral infection n a plant or mammal comprising administering thereto an - 79 - antiviral effective amount of an isolated optical isomer according to claim 15. 24. A pharmaceutical composition comprising a pharmaceutical carrier and an isolated optical isomer according to claim 15. 25. A conjugate of poly(L-lysine) and a 2 '-5' phosphorothioate/phosphodiester oligoadenylate, said conjugate having the formula wherein q is an integer from about 60 to about 70 and each R is independently R' or provided from about five to about ten of the R groups are R', which R' has the following formula - 80 wherein m is zero, 1, 2, or 3; and where R j , R 4 and R 5 , are independently selected from the group of oxygen and sulfur, provided that all R 3 , R 4 and R 5 may not be oxygen, and further provided that all R 3 , R 4 and R,, may not be sulfur.

Inventors: PFLEIDERER WOLFGANG, SUHADOLNIK ROBERT F, PFLEIDERER, WOLFGANG, SUHADOLNIK, ROBERT, F.
Applicants: UNIV TEMPLE, TEMPLE UNIVERSITY - OF THE COMMONWEALTH SYSTEM OF
Application year: 1995
Main classification: A61K31
IPC codes: A01N57, A61K9, A61K31, A61K38, A61K47, A61P17, A61P31, A61P35, C07F9, C07H21, C07K14