G. Chi, B. I. Seo, and V. Nair
483
TABLE 1 Anti-HIV-1 Integrase Data for Dinucleotides
Compounds
3’-Processing IC50 (mM)
Strand transfer IC50 (mM)
1
2
3
6
19
>1000
3[6]
25[7]
>1000
chose to utilize the phosphotriester method in this synthesis (Scheme 1). Thus, the
fully protected dinucleotide 6 was synthesized from 4[9,12] and 5[12] in 92% yield by
a condensation reaction in the presence of 1-mesitylenesulfonyl-3-nitro-1,2,4-triazole
(MSNT) in pyridine. Selective removal of the cyanoethyl protecting group of 6 with
triethylamine and pyridine and subsequent deprotection of the trityl group with 2%
dichloroacetic acid in CH2Cl2 afforded partially deblocked dimer 7. Intramolec-
ular cyclization of 7 under conditions of high dilution with MSNT in pyridine gave
the fully protected cyclic dimer 8 (44% yield for 2 steps). Protecting group removal
using syn-2-pyridinealdoximine and tetramethyl-guanidine followed by treatment
with ammonium hydroxide[13] gave 3 in 46% yield. The complete structure of 3 was
*
established by multinuclear NMR spectral data, HRMS, and quantitative UV data.
Consistent with the absence of base stacking, no observed hypochromicity could be
discerned from the UV data. Support of the cyclic nature of 3 also came from the
NMR data, through observation of the downfield shift of both H-3’ hydrogens
compared to the uncyclized dinucleotide and from the splitting of the carbon
resonances for both C-5’ carbons to doublets.
Integrase inhibition assays were conducted with purified recombinant HIV-1
integrase using a 21-mer oligonucleotide substrate.[6,7] The data (Table 1) clearly
showed that the cyclic dinucleotide 3 was not an inhibitor of HIV-1 integrase, in
contrast to its non-cyclic counterparts 1 and 2.
REFERENCES
1. Frankel, A.D.; Young, J.A.T. HIV-1: fifteen proteins and an RNA. Annu. Rev. Biochem. 1998, 67, 1–25.
2. Asante-Appiah, E.; Skalka, A.M. HIV-1 integrase: structural organization, conformational changes, and
catalysis. Adv. Virus Res. 1999, 52, 351–369.
3. Esposito, D.; Craigie, R. HIV integrase structure and function. Adv. Virus Res. 1999, 52, 319–333.
4. Nair, V. HIV integrase as a target for antiviral chemotherapy. Rev. Med. Virol. 2002, 12, 179–193.
5. Engelman, A.; Mizuuchi, K.; Craigie, R. HIV-1 DNA integration: mechanism of viral DNA cleavage and
DNA strand transfer. Cell 1991, 67, 1211–1221.
6. Mazumder, A.; Uchida, H.; Neamati, N.; Sunder, S.; Jaworska-Maslanka, M.; Wickstrom, E.; Zeng, F.; Jones,
R.A.; Mandes, R.F.; Chenault, H.K.; Pommier, Y. Probing interactions between viral DNA and human
immunodeficiency virus type 1 integrase using dinucleotides. Mol. Pharmacol. 1997, 51, 567–575.
*
HNMR (D2O): 7.91 (s, 1H), 7.54 (s, 1H), 6.13 (m, 1H), 6.07 (m, 1H), 4.88 (m, 1H), 4.75 (m, 1H), 4.00–4.09
(m, 4H), 3.89–3.93 (m, 2H), 2.80 (m, 1H), 2.44–2.61 (m, 3H), 1.68 (s, 3H). 13CNMR (D2O): 166.5, 159.0, 153.9,
151.6, 151.0, 137.4 (two carbons, T-6, G-8), 116.4, 111.4, 84.4, 83.1, 82.5, 82.2, 71.5, 70.5, 62.4, 62.1, 38.3, 38.1, 11.6.
31PNMR (D2O): À0.075, À0.29. FAB-HRMS: [M + Na]+ calcd. for C20H25N7NaO13P2 656.0883, found 656.0905.
UV (H2O): lmax 256 (e 18,900).