Resistance-modifying agents. Part 7: 2,6-Disubstituted-4,8-dibenzylaminopyrimido[5,4-d]pyrimidines that inhibit nucleoside transport in the presence of α1-acid glycoprotein (AGP)

Article abstract of DOI:10.1016/S0960-894X(00)00053-6

The synthesis and biological evaluation of potent 4,8-dibenzylaminopyrimidopyrimidine nucleoside transport inhibitors, with reduced binding to α₁-acid glycoprotein, is reported. (C) 2000 Elsevier Science Ltd. All rights reserved.

Full text of DOI:10.1016/S0960-894X(00)00053-6

Bioorganic & Medicinal Chemistry Letters 10 (2000) 585±589
Resistance-Modifying Agents. Part 7: 2,6-Disubstituted-4,8-
dibenzylaminopyrimido[5,4-d]pyrimidines that Inhibit Nucleoside
Transport in the Presence of ꢀ1-Acid Glycoprotein (AGP)y
Hannah C. Barlow, a Karen J. Bowman, b Nicola J. Curtin, b A. Hilary Calvert, b
Bernard T. Golding, a Bing Huang, a Peter J. Loughlin, a David R. Newell, b
Peter G. Smith b and Roger J. Grin a,*
aDepartment of Chemistry, Bedson Building, The University, Newcastle upon Tyne NE1 7RU, UK
bCancer Research Unit, Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, UK
Received 29 November 1999; accepted 24 January 2000
AbstractÐThe synthesis and biological evaluation of potent 4,8-dibenzylaminopyrimidopyrimidine nucleoside transport inhibitors,
with reduced binding to a1-acid glycoprotein, is reported. # 2000 Elsevier Science Ltd. All rights reserved.
Introduction
Antimetabolite drugs which act by inhibition of de novo
purine and pyrimidine biosynthesis play an important
role in the chemotherapy of cancer,1,2 clinically relevant
examples including methotrexate, 5-¯uorouracil (5FU)
and the classical thymidylate synthase (TS) inhibitor
raltitrexed. Tumour resistance to antimetabolites may
arise via a number of mechanisms, one of which
involves the salvage of extracellular nucleosides and
nucleobases via carrier-mediated plasma membrane
transport, resulting in repletion of nucleotide pools.3±5
Dipyridamole binds avidly to a1-acid glycoprotein
(AGP, orosomucoid), an acute phase protein, the
plasma levels of which may be elevated in cancer
patients,13,14 and, as a consequence, AGP reduces the
e€ective concentrations of the drug below those
required to inhibit salvage pathways.15,16 Thus, binding
to AGP may account for the failure of 1 to enhance the
activity of antimetabolite anticancer drugs in clinical
trials. It is likely that a derivative of 1 which exhibited
potent nucleoside transport inhibitory activity, but
which did not bind to AGP, would have potential clin-
ical application as a modulator of antimetabolite cyto-
toxicity in the therapy of cancer.
Potentiation of the in vitro and in vivo cytotoxicity of a
number of antimetabolite drugs has been observed with
the cardiovascular and antithrombotic drug dipyri-
damole 1,6±8 which is an inhibitor of nucleoside and
nucleobase transport.9,10 However, the disappointing
clinical activity of 1 has prevented the development
of this agent as a resistance-modi®er in cancer chemo-
therapy.11,12
*Corresponding author. Tel.: +44-191-222-8591; fax: +44-191-222-
8591; e-mail: r.j.grin@ncl.ac.uk
yFor part 6 see Curtin, N. J.; Bowman, K. J.; Turner, R. N.; Huang,
B.; Loughlin, P. J.; Calvert, A. H.; Golding, B. T.; Grin, R. J.;
Newell, D. R. Br. J. Cancer 1999, 80, 1738.
As part of an ongoing program to develop novel pyr-
imidopyrimidines as resistance-modi®ers in cancer
chemotherapy, we have conducted studies designed to
elucidate structure±activity relationships for binding to
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00053-6

586
H. C. Barlow et al. / Bioorg. Med. Chem. Lett. 10 (2000) 585±589
AGP. In this letter we describe the synthesis and pre-
liminary biological evaluation of a new series of 4,8-
dibenzylaminopyrimidopyrimidines, which are at least
as potent as 1 as inhibitors of thymidine transport, but
which retain activity in the presence of supraphysiolo-
gical concentrations of AGP.
with TBAF, and the 4,8-dmb protecting groups were
®nally removed on treatment with TFA or DDQ at
room temperature, a€ording the target pyrimidopyr-
imidines 5 in good overall yield.20
Results and Discussion
Although a potent inhibitor of nucleoside and nucleo-
base transport, dipyridamole exhibits poor aqueous
solubility, and being hydrophobic and weakly basic
(pKa=6.4), the drug binds avidly to AGP. In order to
address these problems we ®rst sought to identify novel
pyrimidopyrimidines that retain the potency of 1, but
which do not exhibit signi®cant binding to AGP.
Accordingly, extensive structure±activity studies have
been conducted with regard to substituents at the 2,6-
and 4,8-positions of this heterocycle, and these will be
reported subsequently in a full paper. Initially, the e€ect
of replacing the 2,6-amino functions of dipyridamole
with alkoxy groups was investigated, in the expectation
that this modi®cation would reduce the basicity of the
molecule and attenuate binding to AGP. This indeed
proved to be the case, as exempli®ed by the 2,6-bis-(2,3-
dihydroxypropoxy) derivative 6, where inhibition of
3H-thymidine transport at an inhibitor concentration of
10 mM was not signi®cantly reduced in the presence
of 5 mg/mL of AGP, an AGP concentration which
ablates the activity of 1 under the same conditions
(Table 1). Unfortunately, this structural modi®cation
also resulted in approximately a 30-fold reduction in
potency (IC50 value=9.3 mM), as compared with 1 (IC50
value=0.37 mM), and the introduction of other alkoxy
substituents at the 2,6-positions did not markedly
enhance potency.
Chemical Synthesis
The required pyrimidopyrimidines were initially pre-
pared analogously to 1, following a two step literature
procedure starting from tetrachloropyrimidopyrimidine
2.17,18 Thus, treatment of 2 with 4 equivalents of the
required benzylamine at room temperature to give 3,
followed by 4 equivalents of the appropriate amine or
alkoxide at elevated temperature (reactivity order:
48>26) a€orded the target compounds. However,
while suitable for the preparation of analogues bearing
N-methylbenzylamino substituents at the 4/8 positions
(4, R1=Me), analogous reactions with benzylamines
a€orded low product yields and necessitated extensive
product puri®cation. This was attributed to side reac-
tions arising from deprotonation of the relatively acidic
4,8-dibenzylamino functionality (3, R1=H) under the
basic conditions and elevated temperatures employed in
the second displacement step.
Protection of the vulnerable benzylamino NH sub-
stituent was achieved with the 3,4-dimethoxybenzyl
(dmb) protecting group. This was introduced by react-
ing the appropriate benzylamine with 3,4-dimethoxy-
benzaldehyde in dry ethanol, and reducing the resultant
imine with sodium borohydride in methanol.19 Treat-
ment of 2 with the requisite N-dmb-protected benzyl-
amine at room temperature, gave the 4,8-disubstituted
derivatives (3, R1=dmb) in excellent yield. Substitution
at the 2,6-positions by the required amine or mono-tri-
isopropylsilyl protected diol was then achieved without
incident to furnish 4. Where necessary, removal of the
triisopropylsilyl groups at the 2,6-positions was e€ected
With a view to increasing potency in the 2,6-dialkoxy-
pyrimidopyrimidine series without restoring AGP
binding, the e€ect of replacing the 4,8-piperidino groups
of 1 with other amines was investigated. The introduc-
tion of amino or simple alkylamino substituents in the
Scheme 1. General synthesis of pyrimidopyrimidines. Reagents and yields: (i) appropriate benzylamine, K2CO3, THF, 25 ꢀC, 75±90%; (ii) where
X=O: R3OH, NaH, THF, re¯ux, where X=NR4: R3NHR4, THF, 120±150 ꢀC, 40±60%; (iii) TBAF, THF, 25 ꢀC, 60±80%; (iv) TFA, 25 ꢀC or DDQ
in CH2Cl2/H2O, 25 ꢀC, 50±70%. dmb=3,4-Dimethoxybenzyl.

H. C. Barlow et al. / Bioorg. Med. Chem. Lett. 10 (2000) 585±589
587
Table 1. Inhibition of 3H-thymidine uptake into L1210 cells by selected pyrimidopyrimidines22
No.
Structure
R1
R2
R3
% Inhibition at 1 mM (no. of determinations in parentheses)
Inhibitor alone
+ 5mg/mL AGPa
Reduction (%)
1
6
7
8
A
A
B
B
B
B
B
B
B
B
B
B
B
B
B
N(CH2CH2OH)2
OCH2CH(OH)CH2OH
N(CH2CH2OH)2
N(CH2CH2OH)2
N(CH2CH2OH)2
N(CH2CH2OH)2
NHCH2CH(OH)CH3
NHCH2CH(OH)CH3
NHCH2CH(OH)CH3
NHCH2CH(OH)CH3
O(CH2)2OH
Ð
Ð
H
H
H
Me
H
H
Me
Me
H
Ð
Ð
H
89‹5b (9)
37‹17 (21)c
38‹1 (3)
56‹10 (7)
71‹5 (9)
75‹10 (12)
57‹11 (15)
85‹7 (9)
39, 33 (2)
73‹9 (9)
40‹19 (11)
57‹ 6 (6)
66‹4 (6)
73‹7 (6)
69‹19 (8)
4‹11 (9)
29‹4 (3)
21‹5 (3)
46‹8 (7)
53‹7 (9)
40‹20 (10)
32‹6 (7)
73‹7 (9)
10,3, 3 (3)
57‹12 (7)
28‹25 (5)
44‹8 (3)
49‹6 (5)
17‹20 (5)
32‹10 (8)
96
22
45
18
26
47
45
14
86
22
30
23
26
77
53
4-OMe
3,4-(OMe)2
4-OMe
4-OMe
3,4-(OMe)2
4-OMe
3,4-(OMe)2
4-OMe
3,4-(OMe)2
4-OMe
4-OMe
4-OMe
9
10
11
12
13
14
15
16
17
18
19
O(CH2)2OH
O(CH2)3OH
O(CH2)2OH
O(CH2)3OH
H
H
Me
Me
aa1-Acid glycoprotein.
bStandard deviation.
cDetermined at 10 mM.
4,8- positions resulted in compounds with very poor
activity as inhibitors of thymidine transport (data not
shown). However, the introduction of benzylamino
groups (7) resulted in only approximately a 2-fold
reduction in potency compared with 1, and electron-
donating substituents on the aryl ring of the benzyl-
amino function enhanced activity, with 8 and 9 (IC50
values=0.25 mM), proving at least equipotent with 1.
Crucially, and in contrast to 1, the activity of 8 was not
signi®cantly reduced in the presence of AGP, and this
was also observed for 12 which bears a 2-hydroxy-
propylamino substituent at the 2,6-positions. Although
the reduced activity of compounds 9 and 11 in the pre-
sence of AGP was signi®cant, this e€ect was modest
compared with that observed for 1. Taken together,
these results suggest that AGP binding is not immutably
associated with the presence of amino functions at the
2,6-positions.
slightly more potent than 15 and 16, all three com-
pounds were less active than the corresponding 2,6-bis-
(diethanolamino) derivative 8, and this is consistent
with the reduced potency of 6 compared with 8. Addi-
tion of AGP had little or no e€ect on the inhibitory
activity of compounds 15, 16 or 17. Finally, a compar-
ison of the inhibitory activities of 4,8-dibenzylamino-
pyrimidopyrimidines with their N-methylbenzylamino
counterparts suggests that the latter compounds may
have a greater binding anity for AGP. Thus, the
activity of 10 is markedly reduced in the presence of
AGP compared with that of compound 8, where no sig-
ni®cant reduction in activity is apparent. A similar rela-
tionship is also evident between the N-methylbenzylamino
compounds 13, 14, 18 and 19, and their respective
benzylamino partners 11, 12, 15 and 17. Although spec-
ulative, these observations suggest that a tertiary-amino
function at pyrimidopyrimidine positions 4- and 8 may
be a determinant of AGP binding, which would be con-
sistent with the high binding anity of dipyridamole (1).
Having identi®ed substituted-benzylamino as an alter-
native to piperidino at positions 4- and 8, the e€ect of
combining this functionality with simple hydroxyalkoxy
substitution at the 2,6-positions was investigated. Thus,
derivatives with a 2-hydroxyethoxy- (15 and 16) or 3-
hydroxypropoxy- (17) substituent at the 2,6-positions,
and a 4-methoxybenzylamino- (15 and 17) or 3,4-di-
methoxybenzylamino- (16) group at the 4,8-positions,
were prepared and evaluated. Although 17 was perhaps
Several of the more promising dibenzylaminopyrimido-
pyrimidines have been selected for in vitro and in vivo
biological evaluation. Indeed, we have recently demon-
strated that compound 8 (NU3076) potentiates the in
vitro cytotoxicity of 5-FU, and the antifolate TS
inhibitors CB3717 and nolatrexed in L1210 murine leu-
kaemia cells.21

588
H. C. Barlow et al. / Bioorg. Med. Chem. Lett. 10 (2000) 585±589
Acknowledgements
2ÂOCH3), 3.82 (s, 6H, 2ÂOCH3), 6.78 (m, 6H, 6ÂAr-H); m/z
(EI) 317 (M+, 7%), 166 ([MeO]2C7H5NH+), 151 ([MeO]2
C7H+5 , 100%).
The authors thank the Cancer Research Campaign for
the award of studentships to H.C.B., B.H. and P.L., and
Eli Lilly Co for P.S.
20. A representative synthesis (compound 16) is as follows: (A)
a mixture of 2,4,6,8-tetrachloropyrimidopyrimidine (0.26 g,
0.98 mmol), N,N-bis-(3,4-dimethoxybenzyl)amine (1.24 g, 3.91
mmol) and potassium carbonate (0.59 g, 5.87 mmol) in THF
(10 mL) was stirred at 25 ꢀC for 1 h. Water (75 mL) was added
and the resulting solid was collected, washed with water and
dried in vacuo to give 2,6-dichloro-4,8-bis-(N,N-di-[30,40-di-
methoxybenzyl]amino)pyrimidopyrimidine (0.55 g, 68%) as a
yellow solid, mp 197±199 ꢀC. Found C, 59.22; H, 4.91; N,
References and Notes
1. Rustum, Y. M.; Harstrick, A.; Cao, S.; Vanhoefer, U.; Yin,
M.-B.; Wilke, H.; Seeber, S. J. Clin. Oncol. 1997, 15, 389.
2. Jackman, A. L.; Calvert, A. H. Annals of Oncology 1995, 6,
871.
.
9.81%. C42H44Cl2N6O8 1.0 H2O requires C, 59.36; H, 5.22; N,
3. Jackman, A. L.; Taylor, G. A.; Calvert, A. H.; Harrap, K.
R. Biochem. Pharmacol. 1984, 33, 3269.
4. Fox, M.; Boyle, J. M.; Kinsella, A. R. Br. J. Cancer 1991,
64, 428.
5. Belt, J. A.; Marina, N. M.; Phelps, D. A.; Crawford, C. R.
Advan. Enzyme Regul. 1993, 33, 235.
6. Goel, R.; Howell, S.B. In New Drugs, Concepts and Results
in Cancer Chemotherapy; Muggia, F. M., Ed.; Kluwer Aca-
demic Press: Boston, MA,1992; pp 19.
7. Turner, R. N.; Aherne, G. W.; Curtin, N. J. Br. J. Cancer
1997, 76, 1300.
8. Smith, P. G.; Marshman, E.; Calvert, A. H.; Newell, D. R.;
Curtin, N. J. Semin. Oncol. 1999, 26, 63.
9. Plagemann, P. G. W.; Wohlhueter, R. M.; Wo€endin, C.
Biochim. Biophys. Acta. 1988, 947, 405.
10. Buolamwini, J. K. Current Med. Chem. 1997, 4, 35.
11. Wadler, S.; Subar, M.; Green, M. D.; Wiernik, P. H.;
Muggia, F. M. Cancer Treatment Reports 1987, 71, 821.
12. Schmoll, H.-J.; Harstrick, A.; Kohne-Wompner, C.-H.;
Schober, C.; Wilke, H.; Poliwoda, H. Cancer Treatment
Reviews 1990, 17, 57.
13. Turner, G. A.; Skillen, A. W.; Buamah, P.; Guthrie, D.;
Welsh, J.; Harrison, J.; Kowalski, A. J. Clin. Pathol. 1985, 38,
588.
14. Kremer, J. M. H.; Wilting, J.; Janssen, L. H. M. Pharma-
col. Revs. 1988, 40, 1.
15. Curtin, N. J.; Newell, D. R.; Harris, A. L. Biochem. Phar-
macol. 1989, 38, 3281.
9.89%; dH (200 MHz, CDCl3) 3.78 (s, 12H, 4ÂOCH3), 3.81 (s,
12H, 4ÂOCH3), 4.77 (br s , 4H, 2ÂArCH2), 5.33 (br s, 4H,
2ÂArCH2), 6.78 (m, 12H, 4Â[MeO]2Ar-H); m/z (EI) 830, 832,
834 (M+, 9:6:1, 16%), 679 (M+-[MeO]2C6H3CH2), 529 (M+-
2Â[MeO]2C6H3CH2), 151 ([MeO]2C6H3CH+2 , 100%). (B)
Sodium hydride (0.07 g, 2.76 mmol) was added to a solution
of 2-triisopropylsilyloxyethan-1-ol (0.55 g, 2.51 mmol) in dry
THF (15 mL), and the mixture was stirred at 25 ꢀC for 1 h. A
solution of 2,6-dichloro-4,8-bis-(N,N-di-[30,40-dimethoxybenzyl]-
amino)pyrimidopyrimidine (1.00 g, 0.84 mmol) in dry THF
(15 mL) was added and the mixture was heated under re¯ux for
12 h. Water (30 mL) was added and the mixture was extracted
with ethyl acetate (4Â20 mL) and dried (Na2SO4). The yellow
oil remaining after solvent removal was puri®ed by chroma-
tography on silica, employing petroleum ether (boiling range
40±60 ꢀC):ethyl acetate (3:2) as eluent, to give the required 2,6-
di-(20-triisopropylsilyloxy)ethoxy-4,8-bis-(N,N-di-[30,40-dimeth-
oxybenzyl]amino)pyrimidopyrimidine (0.40 g, 40%) as a white
solid, mp 81±83 ꢀC. Found C, 64.58; H, 8.26; N, 6.99%.
C64H94N6O12Si2 requires C, 64.29; H, 7.92; N, 7.03%; dH (200
MHz, CDCl3) 0.90±0.95 (m, 42H, 2ÂSi[CH(CH3)2]3) 3.72 (m,
16H, 4ÂOCH3 and 2ÂSiOCH2CH2O), 3.80 (s, 12H,
4ÂOCH3), 3.97 (t, 4H, 2ÂSiOCH2CH2O, J=5.4 Hz), 5.16 (br
s, 8H, 4ÂArCH2), 6.76 (m, 12H, 12ÂAr-H); m/z (EI) 1194
(M+, 0.5%), 1151 (M+-C3H7), 1043 (M+-[MeO]2C7H5), 151
([MeO]2C7H+5 , 100%). (C) To a solution of 2,6-di-(20-triiso-
propylsilyloxy)ethoxy-4,8-bis-(N,N-di-[30,40-dimethoxybenzyl]-
amino)pyrimidopyrimidine (0.40 g, 0.34 mmol) in THF (15
mL) was added tetrabutylammonium ¯uoride (1 M soln in
dry THF, 1.36 mL, 1.36 mmol). The mixture was stirred for
12 h at room temperature, and solvents were removed in vacuo.
The yellow residue was suspended in water (30 mL), stirred for
12 h, and the product was collected to give 2,6-di-(20-hydroxy-
ethoxy)-4,8-bis-(N,N-di-[30,40-dimethoxybenzyl]amino)pyrimido-
pyrimidine (0.20 g, 68%) as a white solid, mp 196±198 ꢀC.
Found C, 61.93; H, 6.40; N, 9.20%. C46H54N6O12.0.2H2O
requires C, 62.32; H, 6.14; N, 9.48%; dH (200 MHz, d6-
DMSO) 3.54 (m, 4H, 2ÂHOCH2CH2O), 3.76 (s, 12H,
4ÂOCH3), 3.81 (s, 12H, 4ÂOCH3), 3.94 (t, 4H, 2ÂHOCH2
CH2O), 4.84 (t, 2H, 2ÂOH), 5.30 (br s, 8H, 4ÂArCH2), 6.96
(m, 12H, 12ÂAr-H); m/z (EI) 882 (M+, 0.2%), 731 (M+-
[MeO]2C7H5), 151 ([MeO]2C7H+5 , 100%). (D) A solution of
2,6-di-(20 -hydroxyethoxy)-4,8-bis-(N,N-di-[30,40 -dimethoxy-
benzyl]amino)pyrimidopyrimidine (0.13 g, 0.15 mmol) in tri-
¯uoroacetic acid (2.5 mL) was stirred for 3 h at 25 ꢀC. Excess
tri¯uoroacetic acid was removed in vacuo and the residue was
suspended in sat. NaHCO3 solution (20 mL), and stirred for
12 h. The precipitated solid was collected by ®ltration and
puri®ed by chromatography on silica, with dichloro-
methane:methanol (98:2) as eluent, to furnish 2,6-di-(20-
hydroxyethoxy)-4,8-N,N-di-(30,40-dimethoxybenzyl)aminopyr-
imidopyrimidine (0.03 g, 36%) as a pale yellow solid, mp 216±
218 ꢀC; dH (200 MHz, d6-DMSO) 3.80 (m, 6H, 2ÂOCH3), 3.82
(s, 6H, 2ÂOCH3), 4.41 (m, 4H, 2ÂHOCH2CH2O), 4.69 (m,
8H, 2ÂHOCH2CH2O and 2ÂArCH2), 4.92 (t, 2H, 2ÂOH,
J=5.3 Hz), 7.05 (m, 6H, 6ÂAr-H), 8.51 (t, 2H, 2ÂNH); m/z
16. Budd, G. T.; Jayaraj, A.; Grabowski, D.; Adelstein, D.;
Bauer, L.; Boyett, J.; Bukowski, R.; Murthy, S.; Weick, J.
Cancer Res. 1990, 50, 7206.
17. Thomae, K.G. 1959, Brit. Pat. 807826. (Chem. Abstr.
1959, 53, 12317e).
18. Thomae, K.G. 1963, Ger. Pat. 1151806. (Chem. Abstr.
1964, 60, 2974a).
19. In a typical preparation, a mixture of 3,4-dimethoxy-
benzylamine (1.80 mL, 12.0 mmol) and 3,4-dimethoxy-
benzaldehyde (2.00 g, 12.0 mmol) was re¯uxed in dry ethanol
(20 mL) under N2 for 1 h. Solvents were removed in vacuo,
and the residual oil was triturated with methanol to give N-
(3,4-dimethoxybenzyl)-N-3,4-dimethoxybenzylimine (3.69 g,
97%) as a white solid, mp 77±79 ꢀC. Found C, 68. 31; H, 6.53;
H, 4.18%. C18H21NO4 requires C, 68.55; H, 6.71; N, 4.44%;
dH (200 MHz, CDCl3) 3.80 (s, 3H, OCH3), 3.82 (s, 3H,
OCH3), 3.85 (s, 3H, OCH3), 3.86 (s, 3H, OCH3), 4.67 (s, 2H,
ArCH2), 7.05 (m, 6H, 6ÂAr-H), 8.21 (s, 1H, ArCH=N); m/z
(EI) 316 (MH+), 315 (M+, 18%), 151 ([MeO]2C7H+7 , 100%).
To a stirred solution of N-(3,4-dimethoxybenzyl)-N-3,4-di-
methoxybenzylimine (3.50 g, 11.1 mmol) in dry methanol
(20 mL) at 40 ꢀC, was added sodium borohydride (0.42 g,
11.1 mmol) over a 20 min period. The mixture was re¯uxed for
1 h and the solvents were removed to a€ord N,N-bis-(3,4-di-
methoxybenzyl)amine (3.10 g, 88%) as a white solid, mp 69±
71 oC. Found C, 67.71; H, 7.13; N, 4.35%. C18H23NO4
requires C, 68.12; H, 7.30; N, 4.41%; dH (200 MHz, CDCl3)
1.51 (br s, 1H, NH), 3.68 (s, 4H, 2ÂArCH2), 3.81 (s, 6H,

H. C. Barlow et al. / Bioorg. Med. Chem. Lett. 10 (2000) 585±589
589
(EI) 582 (M+, 67%), 166 ([MeO]2C7H5NH+), 151 ([MeO]2
C7H+5 , 100%).
21. Curtin, N. J.; Bowman, K. J.; Turner, R. N.; Huang, B.;
Loughlin, P. J.; Calvert, A. H.; Golding, B. T.; Grin, R. J.;
Newell, D. R. Br. J. Cancer 1999, 80, 1738.
assayed as described in ref 21. Brie¯y, the uptake of 100 mM
thymidine by 106 cells was followed at 2 s intervals over a 12 s
time course in the presence or absence of inhibitor in 1 or 5%
(v/v) DMSO. In experiments to determine the e€ect of AGP,
the uptake of thymidine was also measured in the presence of
5 mg/mL human AGP.
22. Thymidine uptake in murine L1210 leukaemia cells was