Fluorinated alcohol mediated displacement of the C10 acetoxy group of benzo[a]pyrene-7,8,9,10-tetrahydrotetraol tetraacetates: A new route to diol epoxide-deoxyguanosine adducts

Article abstract of DOI:10.1021/jo701705c

(Chemical Equation Presented) We describe a novel trifluoroethanol (TFE) or hexafluoropropan-2-ol (HFP) mediated substitution reaction of the bay-region C₁₀ acetoxy group in four stereoisomeric 7,8,9,10-tetraacetoxy-7,8,9, 10-tetrahydrobenzo[a]pyrenes (tetraol tetraacetates, two pairs of cis and trans isomers at the 9,10 positions) by the exocyclic N²-amino group of O⁶-allyl-3′,5′-di-O-(tert-butyldimethylsilyl)-2′- deoxyguanosine (3). The tetraacetates are derived from cis and trans hydrolysis of (±)-7β,8α-dihydroxy-9β,10β-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrene (B[a]P DE-1) and of (±)-7β,8α- dihydroxy-9α,10α-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (B[a]P DE-2) at C-10 followed by acetylation. Excellent yields and high regioselectivity were observed. Similar cis/trans product ratios were observed for each set of cis and trans tetraol tetraacetates derived from DE-1 (～75/25) and from DE-2 (～67/33) in HFP. This strongly suggests that the substitution proceeds via an S_N1 mechanism involving a carbocation intermediate that is common to the cis and trans tetraacetates. Since it is likely that the cis and trans products from 3 arise from different conformations of the carbocation, its lifetime must be sufficiently long to permit conformational equilibration before its capture by the purine nucleophile. The corresponding reaction of (±)-9α-bromo-7β,8α,10β- triacetoxy-7,8,9,10-tetrahydrobenzo[a]pyrene with 3 in HFP was highly regio- and stereoselective and gave exclusively trans 10β-adducts. This newly developed substitution reaction provides an attractive alternative synthetic strategy for the preparation of polycyclic hydrocarbon adducted oligonucleotide building blocks.

Full text of DOI:10.1021/jo701705c

Fluorinated Alcohol Mediated Displacement of the C10 Acetoxy
Group of Benzo[a]pyrene-7,8,9,10-tetrahydrotetraol Tetraacetates:
A New Route to Diol Epoxide-Deoxyguanosine Adducts
Haruhiko Yagi* and Donald M. Jerina*
Laboratory of Bioorganic Chemistry, National Institute of Diabetes and DigestiVe and Kidney Diseases,
the National Institutes of Health, DHHS, Bethesda, Maryland 20892
dmjerina@nih.goV
ReceiVed August 3, 2007
We describe a novel trifluoroethanol (TFE) or hexafluoropropan-2-ol (HFP) mediated substitution reaction
of the bay-region C10 acetoxy group in four stereoisomeric 7,8,9,10-tetraacetoxy-7,8,9,10-tetrahydrobenzo-
[a]pyrenes (tetraol tetraacetates, two pairs of cis and trans isomers at the 9,10 positions) by the exocyclic
N2-amino group of O6-allyl-3′,5′-di-O-(tert-butyldimethylsilyl)-2′-deoxyguanosine (3). The tetraacetates
are derived from cis and trans hydrolysis of (()-7â,8R-dihydroxy-9â,10â-epoxy-7,8,9,10-tetrahydrobenzo-
[a]pyrene (B[a]P DE-1) and of (()-7â,8R-dihydroxy-9R,10R-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene
(B[a]P DE-2) at C-10 followed by acetylation. Excellent yields and high regioselectivity were observed.
Similar cis/trans product ratios were observed for each set of cis and trans tetraol tetraacetates derived
from DE-1 (∼75/25) and from DE-2 (∼67/33) in HFP. This strongly suggests that the substitution proceeds
via an SN1 mechanism involving a carbocation intermediate that is common to the cis and trans
tetraacetates. Since it is likely that the cis and trans products from 3 arise from different conformations
of the carbocation, its lifetime must be sufficiently long to permit conformational equilibration before its
capture by the purine nucleophile. The corresponding reaction of (()-9R-bromo-7â,8R,10â-triacetoxy-
7,8,9,10-tetrahydrobenzo[a]pyrene with 3 in HFP was highly regio- and stereoselective and gave
exclusively trans 10â-adducts. This newly developed substitution reaction provides an attractive alternative
synthetic strategy for the preparation of polycyclic hydrocarbon adducted oligonucleotide building blocks.
Introduction
epoxide are cis) and DE-2 (2; benzylic C-7 hydroxyl group and
9,10-epoxide are trans).2 These DEs are thought to initiate cancer
by forming stable DNA adducts, primarily at the exocyclic
amino groups of deoxyguanosine (dGuo) and deoxyadenosine
(dAdo) by cis and trans opening of the epoxide ring.3 Subsequent
error-prone replication by human DNA polymerases and inac-
The polycyclic aromatic hydrocarbon benzo[a]pyrene (B[a]P)
is one of the most prevalent environmental carcinogens to which
humans are exposed.1 It is metabolized in mammals by the
combined action of the cytochrome P450 monooxygenase
system and epoxide hydrolase to form two enantiomeric pairs
of diastereomeric bay-region 7,8-diol 9,10-epoxides (DEs),
referred to as DE-1 (1; benzylic C-7 hydroxyl group and 9,10-
(3) (a) Jerina, D. M.; Chadha, A.; Cheh, A. M.; Schurdak, M. E.; Wood,
A. W.; Sayer, J. M. In Biological ReactiVe Intermediates IV; Witmer, C.
M., Snyder, R., Jelow, D. J., Kalf, G. F., Kocsis, J. J., Sipes, I. G., Eds.;
Molecular and cellular effects and their impact on human health; Plenum:
New York, 1991, pp 533-553. (b) Dipple, A. In DNA Adducts: Identifica-
tion and Biological Significance; Hemminiki, K., Dipple, A., Shuker, D.
E. G., Kadlubar, F. F., Segerbaeck, D., Bartsch, H., Eds; IARC Publications
No. 125: Lyon, 1994; pp 107-129.
(4) Slaga, T. J.; Bracken, W. M.; Viaje, A.; Berry, D. L.; Fisher, S. M.;
Miller, D. R.; Levin, W.; Conney, A. H.; Yagi, H.; Jerina, D. M. In
Carcinogenesis Vol. 3: Polynuclear Aromatic Hydrocarbons; Jones, P. W.,
Freudenthal, R. I., Eds.; New York, 1978; pp 371-382.
* Direct correspondence to: Donald M. Jerina, Chief, Section on Oxidation
Mechanisms, National Institutes of Health, NIDDK, Building 8A, Room 1A01,
Bethesda, MD 20892. Telephone: 301 496 4560. Fax: 301 402 0002
(1) Committee on the Biologic Effects of Atmospheric Pollutants.
Particulate Polycyclic Organic Matteri; National Academy of Science:
Washington, DC, 1972.
(2) Thakker, D. R.; Yagi, H.; Levin, W.; Wood, A. W.; Conney, A. H.;
Jerina, D. M. In BioactiVation of Foreign Compounds; Anders, M. W., Ed.;
Academic Press: New York, 1985; pp 177-242.
10.1021/jo701705c CCC: $37.00 © 2007 American Chemical Society
Published on Web 11/30/2007
J. Org. Chem. 2007, 72, 9983-9990
9983

Yagi and Jerina
SCHEME 1 a
a Key: (i) H3+O; (ii) Ac2O/Py; (iii) OsO4/Py.
curate repair lead to mutant cells, some of which result in tumor
formation. The tetrahydrotetraols (Scheme 1) derived from
hydrolysis of B[a]P DE (2) were found to be weaker carcinogens
than their parent DE-2.4 Therefore, the formation of these
tetrahydrotetraols is generally considered to be a detoxification
process.4
Recently, our laboratory developed a convenient method
for the synthesis of modified dGuo adducts by direct reaction
of O6-allyl-3′5′-di-O-(tert-butyldimethylsilyl)-2′-deoxyguano-
sine (3) with B[a]P DEs13-15a as well as with benzo[c]-
phenanthrene DEs.15 Use of the O-allyl protection of the
O6-carbonyl group of dGuo not only prevented adduct formation
at this position but also increased the nucleophilic reactivity
of the N-2 amino group. Choice of the solvent was critical.
Dimethylacetamide (DMA)15a and fluorinated alcohols were
found to be effective solvents for this addition reaction. In
particular, fluorinated alcohols such as trifluoroethanol
For the past several years, the synthesis of oligonucleotides
containing site-specific and stereospecific adducts of DEs has
become an active area of research. These modified oligonucle-
otides with a wide variety of sequence contexts have been uti-
lized in structural studies such as NMR solution conformation5
and X-ray crystallographic analysis.6 They have also found ap-
plication in the study of site-specific mutagenicity7a and as mech-
anistic probes for a variety of DNA processing enzymes.7b,c-12
(9) Khan, Q. A.; Kohlhagen, G.; Marshall, R.; Austin, C. A.; Kalena,
G. P.; Heiko, K.; Sayer, J. M.; Jerina, D. M.; Pommier, Y. Proc. Natl.
Acad. Sci. U.S.A. 2003, 100, 12498-12503.
(10) (a) Yakovleva, L.; Tian, L.; Sayer, J. M.; Kalena, G. P.; Kroth, H.;
Jerina, D. M.; Shuman, S. J. Biol. Chem. 2003, 278, 42170-42177. (b)
Yakovleva, L.; Handy, C. J.; Sayer, J. M.; Pirrung, M.; Jerina, D. M.;
Shuman, S. J. Biol. Chem. 2004, 279, 23335-23342. (c) Yakovleva, L.;
Handy, C. J.; Yagi, H.; Sayer, J. M.; Pirrung, M.; Jerina, D. M.; Shuman,
S. Biochemistry 2006, 45, 7644-7653.
(11) (a) Driscoll, H. C.; Matson, S. W.; Sayer, J. M.; Kroth, H.; Jerina,
D. M.; Brosh, R. M., Jr. J. Biol. Chem. 2003, 278, 41126-41135. (b)
Choudhary, S.; Doherty, K. M.; Handy, C. J.; Sayer, J. M.; Yagi, H.; Jerina,
D. M.; Brosh, M. R., Jr. J. Biol. Chem. 2006, 281, 6000-6009.
(12) Johnson, A. A.; Sayer, J. M.; Yagi, H.; Kalena, G. P.; Amin, R.;
Jerina, D. M.; Pommier, Y. J. Biol. Chem. 2004, 279, 7947-7965.
(13) Ramesha, A. R.; Kroth, H.; Jerina, D. M. Tetrahedron Lett. 2001,
42, 1003-1005.
(14) Yagi, H.; Jerina, D. M. J. Org. Chem. 2007, 72, 6037-6045.
(15) (a) Kroth, H.; Yagi, H.; Seidel, A.; Jerina, D. M. J. Org. Chem.
2000, 65, 5558-5564. (b) Yagi, H.; Ramesha, A. R.; Kalena, G.; Sayer, J.
M.; Kumar, S.; Jerina, D. M. J. Org. Chem. 2002, 6, 6678-6689.
(5) For leading references see: (a) Lin, C. H.; Huang, X.; Korbanovskii,
A.; Hingerty, B. E.; Amin, S.; Broyde, S.; Geacintov, N. E.; Patel, D. J. J.
Mol. Biol. 2001, 306, 1059-1080. (b) Pradhan, P.; Tirumala, S.; Liu, X.;
Sayer, J. M.; Jerina, D. M.; Yeh, H. J. C. Biochemistry 2001, 40, 5870-
5881.
(6) Ling, H.; Sayer, J. M.; Polsky, B. S.; Yagi, H.; Boudsocq, F.;
Woodgate, R.; Jerina, D. M.; Yang, W. Proc. Natl. Acad. Sci. U.S.A. 2004,
101, 2265-2269.
(7) (a) Page, J. E.; Liang, C.; Sayer, J. M.; Jerina, D. M.; Dipple, A.
Polycyclic Aromat. Compd. 1999, 16, 99-108. (b) Frank, E. G.; Sayer, J.
M.; Kroth, H.; Ohashi, E.; Ohmori, H.; Jerina, D. M.; Woodgate, R. Nucleic
Acid Res. 2002, 30, 5284-5292. (c) Graziewicz, M. A.; Sayer, J. M.; Jerina,
D. M.; Copeland, W. C. Nucleic Acid Res. 2004, 32, 397-405.
(8) Pommier, Y.; Laco, G. S.; Kohlhagen, G.; Sayer, J. M.; Kroth, H.;
Jerina, D. M. Proc. Natl. Acad. Sci. U.S.A. 2000, 97, 10739-10744.
9984 J. Org. Chem., Vol. 72, No. 26, 2007

Route to Diol Epoxide-Deoxyguanosine Adducts
SCHEME 2
(TFE), hexafluoropropan-2-ol (HFP), and perfluoro-tert-butanol
(PFTB) showed an exceptional ability to catalyze this addition
reaction.13-15b These alcohols are acidic because of the electron
withdrawing nature of fluorine and can catalyze the epoxide
opening through their strong hydrogen-bonding capability. Their
high polarity, high ionizing power, and low nucleophilicity make
them ideal solvents for this type of addition reaction by
catalyzing the formation of carbocation intermediates.16 On the
basis of our observations of these fluorinated alcohol-mediated
addition reactions of DEs, we envisaged the possibility of
regioselective substitution reactions using seemingly inert
tetrahydrotetraol derivatives of B[a]P. We thought it might be
possible to generate a stable carbocation regioselectively at the
benzylic C-10 position of the tetrahydrobenzo-ring by introduc-
ing a substituent which has a strong ability to hydrogen bond
with the fluorinated alcohol. The carbocation formed at the bay-
region C-10 benzylic position (∆Edeloc ) 0.794â) is much more
stable and thus more easily formed than the corresponding
carbocation at the C-7 benzylic position (∆Edeloc ) 0.488â).17
Here we report the first example of such a regioselective
substitution reaction between tetrahydrotetraol derivatives of
B[a]P and the dGuo reactant 3 in fluorinated alcohol solvents.
TABLE 1. Fluorinated Alcohol Mediated Synthesis of cis- and
trans-N2-dGuo Adducts (yields in %) from Tetraol Tetraacetates
Derived from B[a]P DE-1 and DE-2 and Bromotriol Triacetatea
trans-1 adducts
tetraol
reaction
cis-1 adducts
tetraacetate solvent time (h)
9a and 9b
10a
10b
cis-1 (4)
trans-1 (5)
HFP
HFP
72
16
76
73
11
12
13
14
cis-2 adducts
trans-2 adducts
11a
11b
12a
12b
cis-2 (6)
HFP
TFE
HFP
42
40
14
37
29
35
31
25
32
20
22
18
13
23
16
trans-2 (7)
trans adducts
14a
14b
bromotriol
triacetate (13) HFP
TFE
24
4
47
48
53
52
a The ratio was determined by HPLC (345 nm). Reaction was carried
out in a sealed glass tube at 70 °C in a 1:5 molar ratio of the acetate and
dGuo reactant (3) in fluorinated alcohol (100 molar equiv).
Results and Discussion
Easily accessible tetraol tetraacetates (4, 5, 6, and 7) derived
from B[a]P DE-1(1) and DE-2 (2) were chosen for the reasons
discussed in the Introduction. The cis-1 tetraol tetraacetate (4)
and the trans-1 tetraol tetraacetate (5) were obtained in a ratio
of 60:4018 by acidic hydrolysis of B[a]P DE-1 (1) followed by
acetylation and HPLC separation. The acidic hydrolysis of B[a]P
DE-2 (2) and successive acetylation afforded the cis-2 tetraol
tetraacetate (6) and trans-2 tetraol tetraacetate (7) in a ratio of
(16) Eberson, L.; Hartshorn, M. P.; Persson, O.; Radner, F. Chem.
Commun. 1996, 2105-2112.
(17) Jerina, D. M.; Lehr, R. E.; Yagi, H.; Hernandez, O.; Dansette, P.
M.; Wislocki, P. G.; Wood, A. W.; Chang, R. L.; Levin, W.; Conney, A.
H. In InVitro Mutagenesis Testing; de Serres, F. J.; Fouts, J. R.; Bend, J.
R.; Philpot, R. M., Eds.; Elsevier/North-Holand Biomedical Press: Am-
sterdam, 1976; pp 179-195.
J. Org. Chem, Vol. 72, No. 26, 2007 9985

Yagi and Jerina
FIGURE 1. HPLC profiles of the products of reaction between O6-allyl dGuo diTBDMS (3) and (()-trans-1 tetraol tetraacetate 5 (A) or (()-
trans-2 tetraol tetraacetate 7 (B) on three coupled Axxiom Sil columns (9.5 mm × 250 mm) eluted with 14% acetone in n-hexane at a flow rate
of 5 mL/min (detected at 346 nm). (A) tR ) 46.4 min: (10S)-trans-1 dGuo adduct (10a); tR ) 48.7 min: (10S)- and (10R)-cis-1 dGuo adducts
(9a,b); tR ) 50.3 min: (10R)-trans-1 dGuo adduct (10b); (B) tR ) 35.2 min: (10S)-cis-2 dGuo adduct (11a); tR ) 36.9 min: (10R)-cis-2 dGuo
adduct (11b); 44.1 min: (10S)-trans-2 dGuo adduct (12a); tR ) 53.8 min: (10R)-trans-2 dGuo adduct (12b).
3:97.18 The minor cis-2 tetraol tetraacetate (6) could also be
obtained as a major product (86% yield) together with a minor
product, the cis-1 tetraol tetraacetate (4; 8.4% yield) from the
osmium tetroxide oxidation of B[a]P 7,8-dihydrodiol (8) fol-
lowed by acetylation (Scheme 1).
As predicted, fluorinated alcohols have a dramatic catalytic
effect on the regioselective substitution reaction at the C-10
acetoxy group for all of the above tetraol tetraacetates (4, 5, 6,
and 7) with the N2-amino group of 3. Reaction of a 5-fold molar
excess of 3 with the trans-1 (5) and trans-2 tetraol tetraacetate
(7) was complete within 16 and 14 h, respectively, at 70 °C in
the presence of 5 molar equiv of HFP (cf. Table 1). The
corresponding reaction for the cis-1 (4) and cis-2 tetraol
tetraacetate (6) required more prolonged heating (72 and ∼40
h, respectively). The yields for the substitution in all cases were
nearly quantitatiVe (>95%). Unlike the addition reaction of 3
with DEs, no solvent adducts13,14,15b were formed. The reactions
of the cis-1 (4) and trans-1 tetraol tetraacetate (5) each afforded
a mixture of cis-1 dGuo adducts (9a and 9b) and trans-1 dGuo
adducts (10a and 10b) in similar ratios [the total cis-1 dGuo
adducts: the total trans-1 dGuo adducts from 4 (76:24) and from
5 (73:27)]. The corresponding ratios of the total cis-2 dGuo
adducts (11a and 11b) to the total trans-2 dGuo adducts (12a
and 12b) from 6 (68:32) and from 7 (66:34) were also similar
to each other (Scheme 2 and Table 1). The corresponding
reaction of these tetraacetates in the less acidic TFE required
more prolonged heating. Even the trans-2 tetraol tetraacetate
(7), which was the most reactive isomer under these conditions,
required heating at 70 °C for 40 h (∼95% completion).
Interestingly, a somewhat different cis/trans ratio (55:45) was
observed for the reaction of 7 in TFE compared with those
obtained in HFP as shown in Table 1. The reaction rates for
the less reactive tetraacetates (4, 5, and 6) in TFE were too
slow to be practical for synthetic purposes. Unlike the addition
reaction of 3 with DEs,13-15 these adduct ratios were insensitive
to the molar ratio of solvent to reactants. As shown in Table 1,
a small but significant stereoselectivity was observed for the
set of diastereomeric adducts formed by the reaction of 6 as
well as 7 (e.g., 11a and 11b vs 12a and 12b). The product dGuo
(18) Yagi, H.; Thakker, D. R.; Hernandez, O.; Koreeda, M.; Jerina, D.
M. J. Am. Chem. Soc. 1977, 99, 1604-1611.
9986 J. Org. Chem., Vol. 72, No. 26, 2007

Route to Diol Epoxide-Deoxyguanosine Adducts
SCHEME 3
adducts were separated by HPLC (Figure 1) and were character-
the common carbocation formed from 5(e,e) and from 4(e,a) is
B-1 (with three quasi-equatorial acetoxy groups) which is
expected to be the preferred one at equilibrium with A-1 (with
three quasi-axial acetoxy groups). Carbocation formation from
4 most likely proceeds via axial departure (the microscopic
reverse of axial attack) of the acetoxy group from (4e,a) to give
initially carbocation B-1. Similarly, axial loss of the acetoxy
group from (5a,a) would give carbocation A-1. Energetically
favorable axial attack20 on B-1 will lead to cis-1 dGuo adducts
9a and 9b, whereas conformation A-1 should give trans-1 dGuo
adducts 10a and 10b. Thus if A-1 and B-1 are trapped by 3
faster than they equilibrate with each other, 5 would be predicted
to give predominantly trans adducts whereas 4 would give
predominantly cis adducts. The fact that a similar cis/trans
adduct ratio was obtained from both the cis-1 (4) and trans-1
tetraol tetraacetate (5) regardless of their conformational prefer-
ence strongly suggests the carbocations A-1 and B-1 undergo
conformational equilibration at a faster rate than their capture
by 3 (Scheme 3).
1
ized by comparison of H NMR, mass, and CD spectra with
authentic samples.13-15
The similar cis/trans product ratios obtained from each set
of the cis- and trans- tetraol tetraacetates strongly suggest the
involvement of a common carbocation intermediate. Conforma-
tions of the starting tetraol tetraacetates and their intermediate
carbocations (Schemes 3 and 4) provide a plausible explanation
for the cis/trans product ratios observed. The conformational
preferences for these tetraol tetraacetates and the C-10 carboca-
tions derived from them are determined by the following two
factors: (i) the acetoxy group at the hindered C-10 position
prefers a quasi-axial conformation due to steric hindrance in
the bay region,18 and (ii) the acetoxy groups at the other
positions prefer quasi-equatorial conformations19,20 in the car-
1
bocation. On the basis of H NMR spectra, 5 has an equal
preference for conformation 5(a,a) and 5(e,e), whereas 4 is
expected to prefer conformation 4(e,a). The conformation of
The preferred conformation of 7 is 7(a,a), whereas 6 exists
as a mixture of 6(e,a) and 6(a,e). Preferred axial departure of
the acetoxy group should initially give cation A-2 from 6 and
B-2 from 7. Again, the cis/trans ratios from 6 and 7 are very
similar, suggesting that the cation intermediate undergoes
conformational equilibration faster than trapping by 3. The
(19) (a) Doan, L.; Yagi, H.; Jerina, D. M.; Whalen, D. L. J. Am. Chem.
Soc. 2002, 124, 14382-14367. (b) Doan, L.; Yagi, H.; Jerina, D. M.;
Whalen, D. L. J. Org. Chem. 2004, 69, 8012-8017.
(20) (a) Sayer, J. M.; Yagi, H.; Silverton, J. V.; Friedman, S. L.; Whalen,
D. L.; Jerina, D. M. J. Am. Chem. Soc. 1982, 104, 1972-1978. (b) Goering,
H. L.; Josephson, R. R. J. Am. Chem. Soc. 1962, 84, 2779-2785. (c)
Goering, H. L.; Vlazny, J. C. J. Am. Chem. Soc. 1982, 104, 1972-1978.
J. Org. Chem, Vol. 72, No. 26, 2007 9987

Yagi and Jerina
SCHEME 4
energy difference between A-2 (with two quasi-axial acetoxy
groups) and B-2 (with one quasi-axial acetoxy) is expected to
be slightly smaller than that between A-1 (with all quasi-equa-
torial acetoxy groups) and B-1 (with three quasi-axial acetoxy
groups).20 Accordingly, the preference for cis adduct formation
from 6 or 7 in the DE-2 series is slightly less pronounced
(∼67%) than from 4 or 5 in the DE-1 series (∼75%).
Interestingly, the corresponding reaction of the bromotriol
triacetate (13) with 3 in TFE or in HFP was highly regio- and
stereoselective affording exclusively the trans-dGuo adduct
diastereomers (14a and 14b) (Scheme 5 and Table 1). In this
case, due to the steric hindrance of the bulky bromine substituent
at the C-9 position, 13 is almost locked in the conformation
13(a,a). Furthermore, the rate of capture of carbocation A-3
derived from 13(e,e) by the dGuo reactant 3 is expected to be
much slower than that for the capture of carbocation B-3 due
to steric hindrance as well as electronic repulsion between the
adjacent cis equatorial bromine in A-3 at the C-9 position and
the incoming dGuo reactant (3). The 1,3 diaxial interaction
between the C-8 acetoxy group and the incoming 3 should also
retard the rate of the cis adduct formation.
FIGURE 2. CD spectra (normalized to 1 absorbance unit at 8max in
methanol) of the trans-N2-dG adducts (14a and 14b).
were close to those of the trans-2 dGuo adducts (12a and
12b).15a Absolute configurations of 14a and 14b were assigned
on the basis of comparison of their CD spectra (cf. Figure 2,
Experimental Section) with those of the trans-2 dGuo adducts
Assignment of trans stereochemistry for 14a and 14b was
on the basis of the observed coupling constants of the tetrahydro
benzo-ring methine protons (J7,8 ) 8.7; J8,9 ) 2.9; J9,10 ) 2.9
Hz) and the chemical shift of H-10 (6.50 and 6.44 ppm), which
(21) See: Johnson, F.; Bonala, R.; Tawde, D.; Torres, M. C.; Iden, C.
R. Chem. Res. Toxicol. 2002, 15, 1489-1494. Lakshman, M. K.; Gunda,
P. Org. Lett. 2003, 5, 39-42. Champeil, E; Pradhan, P.; Lakshman, M. K.
J. Org. Chem. 2007, 72, 5035-5045 and references theirin.
9988 J. Org. Chem., Vol. 72, No. 26, 2007

Route to Diol Epoxide-Deoxyguanosine Adducts
SCHEME 5
reaction time). After completion of the reaction, the solvent was
evaporated, and the residue was subjected to HPLC purification
(Figure 1). The combined yields of the adducts in all cases were
nearly quantitative (>95%). The structures of these adducts (9a,
9b, 10a, 10b, 11a, 11b, 12a, and 12b) were confirmed by
comparison of their 1H NMR, CD, and mass spectra with those of
authentic samples.15a
(()-9r-Bromo-7â,8r,10â-triacetoxy-7,8,9,10-tetrahydroben-
zo[a]pyrene (13). Acetylation of (()-9R-bromo-7â,8R,10â,-
trihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene18 was carried out
by standard procedures in 93% yield; mp 166-168 °C (EtOAc-
n-hexane); 1HNMR (CDCl3) δ: 2.17, 2.23, and 2.32 (each s,
each 3H, CH3CO), 4.89 (t, 1H, H9, J ) 3.1 Hz), 5.75 (dd,
1H, H8, J ) 8.7, 3.4 Hz), 6.92 (d, 1H, H7, J ) 8.7 Hz), 7.20 (d,
1H, H10, J ) 3.1 Hz), 8.00-8.30 (m, 8 pyrene aromatic
protons). C26H21BrO6: C, 61.13; H, 4.16. Found: C, 61.22; H,
4.06.
Reaction of Bromotriol Triacetate (13) with O6-Allyl-dGuo
Di-TBDMS Ether (3). To a solution of 3 (268 mg; 5 molar equiv)
in TFE or HFP (100 molar equiv) was added the bromotriol
triacetate (51 mg, 0.1 mmol; 1 molar equiv), and the mixture was
stirred at 70 °C (bath temperature) in a sealed glass tube (see Table
1 for the reaction time). After completion of the reaction, the solvent
was evaporated, and the residue was subjected to HPLC purification
on a silica column using 20% EtOAc in n-hexane at a flow rate of
10 mL/min (detected at 280 and 346 nm). Evaporation of the
fraction corresponding to the peak at tR ) 16.9 min afforded the
unused 3 (200 mg, 75% recovery).
(12a and 12b) whose absolute configurations were previously
determined.15a
In summary, a novel and highly efficient, regioselective
substitution reaction occurs between the O6-allyl protected dGuo
derivative (3) and the tetraol tetraacetates derived from B[a]P
DEs. The present method compares very favorably with earlier
approaches (reviewed in ref 15a) as well as to more recent
palladium catalyzed C-N bond forming reactions.21 For practi-
cal synthetic purposes, it is unnecessary to separate the cis/
trans mixture of tetraol tetraacetates from either DE-1 or DE-2
in the present approach because the cis and trans tetraol
tetraacetates yield the same mixture of dGuo adducts (see Table
1). Compared to the direct solvent assisted reaction of the DEs
with 3, the protected dGuo adducts obtained in this study provide
an attractive alternate route for the synthesis of the O6-allyl-
protected N2-dGuo phosphoramidites which can be incorporated
into oligonucleotides with selectable sequence contexts.
Experimental Section
Caution: Benzo[a]pyrene 7,8-dihydrodiol and diol epoxides
DE-1 and DE-2 are mutagenic and carcinogenic and must be
handled carefully in accordance with NIH guidelines.22
General Procedures for the Reaction of B[a]P Tetraol
Tetraacetates (4, 5, 6, or 7) with O6-Allyl-dGuo Di-TBDMS
Ether (3). To a solution of 3 (548 mg, 1.02 mmol, 5 molar equiv)
in TFE (1.5 mL, 20.4 mmol, 100 molar equiv) or HFP (2.1 mL,
20.4 mmol, 100 molar equiv) was added the tetraol tetraacetate
(100 mg, 0.204 mmol, 1 molar equiv), and the mixture was stirred
at 70 °C (bath temperature) in a sealed glass tube (see Table 1 for
Evaporation of the fraction corresponding to the peak at tR
)
18.4 min afforded N2-[10S-(9R-bromo-7R,8S-dihydroxy-7,8,9,-
10-tetrahydrobenzo[a]pyrenyl)]-O6-allyl-3′,5′-di-O-(tert-butyldi-
methylsilyl)-2′-deoxyguanosine (14a) as a colorless solid (36 mg,
39% yield for the reaction in HFP); 1H NMR (CDCl3-CD3OD) δ:
0.01-0.12 (br. m, 6H, Si bonded methyls), 0.80-0.94 (br. m, 18H,
tert-butyl methyls), 2.20 and 2.29 (each s, each 3H, CH3CO), 2.40
(22) NIH Guidelines for the Laboratory Use of Chemical Carcinogens;
NIH publication No. 81-2385; U.S. Government Printing Office: Wash-
ington, DC, 1981.
J. Org. Chem, Vol. 72, No. 26, 2007 9989

Yagi and Jerina
(m, 1H, H2′′), 2.62 (m, 1H, H2′), 3.78 (m, 2H, H5′,5′′), 4.0 (m, 1H,
H4′), 4.56 (m, 1H, H3′), 5.06 (m, 2H, allyl CH2), 5.16 (t, 1H, H9, J
) 2.9 Hz), 5.2-5.58 (m, 2H, Hc and Ht), 5.95 (dd, 1H, H8, J )
8.7, 2.9 Hz), 6.14 (m, 1H, Hv), 6.50 (d, 1H, H10, J ) 2.9 Hz), 6.80
(d, 1H, H7, J ) 8.7 Hz), 8.00-8.25 (m, 9H, 8 pyrene aromatic
protons and H8G). HRMS (FAB+) calcd for C49H62O8N5BrCs:
1116.2375 (79Br); 1118.2354 (81Br). Found: 1116.2352 (79Br);
1118.2384 (81Br). The CD spectrum (Figure 2) was similar to that
of the trans-(10S)-12a.
aromatic protons and H8G). HRMS (FAB+) calcd for C49H62O8N5
BrCs: 1116.2375 (79Br); 1118.2354 (81Br). Found: 1116.2368 (79-
Br); 1118.2324 (81Br). The CD spectrum (Figure 2) was similar to
that of the trans-(10R)-12b. Yields of 14a and 14b for the reaction
in TFE were similar to those in HFP.
Acknowledgment. The authors are greatly indebted to Dr.
Jane M. Sayer at this Institution for helpful discussions. This
research was supported by the Intramural Research Program of
the NIH, National Institute of Diabetes and Digestive and
Kidney Diseases.
Evaporation of the fraction corresponding to the peak at tR
)
20.1 min afforded N2-[10R-(9S-bromo-7S,8R-dihydroxy-7,8,9,-
10-tetrahydrobenzo[a]pyrenyl)]-O6-allyl-3′,5′-di-O-(tert-butyldi-
methylsilyl)-2′-deoxyguanosine (14b) as a colorless solid (40 mg,
43.4% yield for the reaction in HFP); 1H NMR (CDCl3-CD3OD)
δ: 0.01-0.12 (br. m, 6H, Si bonded methyls), 0.80-0.94 (br. m,
18H, tert-butyl methyls), 2.21 and 2.29 (each s, each 3H, CH3-
CO), 2.42 (m, 1H, H2′′), 2.62 (m, 1H, H2′), 3.80 (m, 2H, H5′,5′′),
3.99 (m, 1H, H4′), 4.58 (m, 1H, H3′), 5.10 (m, 2H, allyl CH2), 5.19
(t, 1H, H9, J ) 2.9 Hz), 5.2-5.58 (m, 2H, Hc and Ht), 5.94 (dd,
1H, H8, J ) 8.7, 2.9 Hz), 6.14 (m, 1H, Hv), 6.44 (d, 1H, H10, J )
2.9 Hz), 6.81 (d, 1H, H7, J ) 8.7 Hz), 8.00-8.25 (m, 9H, 8 pyrene
Supporting Information Available: Synthetic procedures for
the reactions of the 7,8-dihydrodiol (8) with osmium tetroxide and
the hydrolysis of B[a]P DE-1 and DE-2 to tetraols under acidic
conditions are presented as well as proton NMR spectra of the new
compounds 13, 14a, and 14b are provided. This material is available
free of charge via the Internet at http://pubs.acs.org.
JO701705C
9990 J. Org. Chem., Vol. 72, No. 26, 2007