Design and synthesis of functionalized glycomers as non-peptidic ligands for SH2 binding and as inhibitors of A-431 human epidermoid and HT-29 colon carcinoma cell lines

Article abstract of DOI:10.1016/S0960-894X(00)00022-6

A set of O-substituted aryl β-D-glucopyranosides were prepared and found to have inhibitory activity on the growth of two carcinoma cell lines. (C) 2000 Elsevier Science Ltd. All rights reserved.

Full text of DOI:10.1016/S0960-894X(00)00022-6

Bioorganic & Medicinal Chemistry Letters 10 (2000) 439±442
Design and Synthesis of Functionalized Glycomers as
Non-Peptidic Ligands for SH2 Binding and as Inhibitors of
A-431 Human Epidermoid and HT-29 Colon Carcinoma
Cell Lines
Stephen Hanessian, a,* Oscar M. Saavedra, a Fang Xie, a Nadia Amboldi b
and Carlo Battistini b
a
Â
Â
Â
Department of Chemistry, Universite de Montreal, CP 6128, Succursale Centre-ville, Montreal, PQ, Canada H3C 3J7
bPharmacia & Upjohn Discovery Research Oncology, Nerviano, Italy
Received 23 November 1999; accepted 22 December 1999
AbstractÐA set of O-substituted aryl b-d-glucopyranosides were prepared and found to have inhibitory activity on the growth of
two carcinona cell lines. # 2000 Elsevier Science Ltd. All rights reserved.
Cellular signal transduction is a vital physiological pro-
cess (for recent reviews, see refs 1±4), through which
important recognition events among proteins are regu-
lated. Distinct functional modules composed of inde-
pendently folded amino acids termed Src homology 2
(SH2) domains (for recent reviews, see refs 5±8), and
present in many intracellular proteins, bind to phos-
photyrosine units of peptides. A cascade of sequential
protein±protein interactions follows leading to a com-
plex set of cellular responses. The aberration of some of
these signaling pathways may have important con-
sequences in diverse pathologies such as cancer and
immune disorders. Grb-2 is a key transducer containing
the SH2 domain, and it is particularly important for
EGF (e.g. refs 9±12) and HGF (e.g. refs 13 and 14)
receptors which are expressed in A-431 human epi-
dermoid and HT-29 colon carcinoma cell lines. The
discovery of antagonist molecules to these domains may
have great potential in the development of anticancer
and antiin¯ammatory drugs for example.1±4,15±18
We envisaged the synthesis of a series of aryl b-d-glu-
copyranosides `glycomers' as sca€olds onto which we
would deploy diverse functionality to probe their bind-
ing anities (Fig. 1, Types A, B, and C). Such
glycomers could also be viewed as constrained spacial
glycomimetics of the YVNV tetrapeptide.
The synthesis of the intended glycomers proceeded in a
straightforward fashion and in good yields for indivi-
dual steps (>70%). Scheme 1 depicts the synthesis of
glycomers of Type A, where an 6-O-(m-tri¯uoromethyl
benzenesulfonyl) and a 3-O-allyl groups were present in
each of the 2-ester derivatives (6, R1=acyl). The choice
of a p-iodophenyl aglycone was based on the presence
of the tyrosine unit in YVNV and on the opportunities
o€ered by cross-coupling reactions in subsequent trans-
formations. To that end, the p-iodophenyl, p-bromo-
phenyl and p-hydroxylphenyl b-d-glucopyranosides
were also easily obtained (for glycoside synthesis via
stannyl ethers, see refs 22±25). The protected ethyle-
neglycol glycoside 4 was prepared by the Koenigs±
Knorr method using Hg(CN)2/HgBr2 as activators (for
a review, see ref 26).
Elegant studies on the crystal structures of high anity
phosphotyrosyl peptide ligands such as (YpVNV) to
Grb2-SH2 have been reported (see, for example, refs
19±21) (Fig. 1).
Utilizing a common precursor 2 (R=p-iodophenyl), a
series of 6-carbamates, 6-esters and 6-ethers were also
synthesized by standard methods (Scheme 2).
Scheme 3 shows the elaboration of glycomers in which
a distal carbamate group was introduced at C-3 in
*Corresponding author. Tel.: +1-514-343-6738; fax: +1-514-343-
5728; e-mail: hanessia@ere.umontreal.ca
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00022-6

440
S. Hanessian et al. / Bioorg. Med. Chem. Lett. 10 (2000) 439±442
Figure 1.
Scheme 1. (a) i. Acetone, H2SO4, ii. NaH, DMF, iii. allyl bromide; (b) IRa-1 20 (H+), H2O; (c) Ac2O, pyridine; (d) HBr, AcOH; (e) for 3: tribu-
tylstannyl p-iodophenoxide, SnCl4, CH2Cl2, 85%; for 3a: tributyistannyl p-bromophenoxide, SnCl4, CH2Cl2, 88%; for 3b: p-resorcinol, CdCO3,
toluene, 70%; for 4: b-diphenyl tert-butylsilyloxyethanol, Hg(CN)2, HgBr2, benzene, 75%; (f) MeONa/MeOH; (g) benzaldehyde dimethylacetal,
DMF, HBF4; (h) RCOCl, Et3N, CH2Cl2; (i) TFA, H2O; (j) m-tri¯uorobenzenesulfonyl chloride, pyridine; (k) R1=acetyl 6a, R1=propionyl 6b,
R1=butyryl 6b,A1=benzoyl 6d, R1=phenylacetyl 6e, R1=phenylpropionyl 6f (yields 80±87%).
Scheme 2.
Scheme 3. (a) O3, MeOH; (b) NaBH4; (c) for 9: i. p-nitrophenyl chloroformate, ii. ammonium formate; for 10, PhNCO.
conjunction with
Related carbamates in the m-tri¯uoromethyl benzene-
a
6-O-(2-naphthylsulfonyl) ester.
and C-3, respectively, was synthesized from precursor 5f
as shown in Scheme 4.
sulfonate series were also prepared.
The results for the inhibition of Grb2-SH2 were dis-
appointing, with at best 25±30% inhibition at 100 mM
for compounds in entries 5, 9 and 15 (Table 1). Much
A representative glycomer of the Type C family in
which ester and carbamate groups were deployed at C-2

S. Hanessian et al. / Bioorg. Med. Chem. Lett. 10 (2000) 439±442
441
Scheme 4. (a) O3, MeOH; (b) NaBH4; (c) phenylisocyanate, CH2Cl2; (d) TFA, H2O; (e) m-tri¯uorobenzenesulphonyl chloride, pyridine.
Table 1. Cytotoxicity of glycomers against A-431 and HT-29 cell linesa
Entry
Structure
A-431 (IC50 mM)
HT-29 (IC50 mM)
1
2
3
4
5
6
7
8
R=mTFMB
R=mTFMB
R=mTFMB
R=mTFMB
R=mTFMB
R=mTFMB
R=mTFMB
R=mTFMB
R=TP
R1=H
R1=MeCO
R1=EtCO
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
R2=allyl
36.80
6.10
2.17
2.60
3.30
7.00
5.15
2.00
12.75
9.77
8.81
5.20
35.45
23.07
4.65
4.00
4.20
7.50
4.95
3.00
Ð
R1=PrCO
R1=iPrCO
R1=PhCO
R1=PhCH2CO
R1=Ph(CH2)2CO
R1=MeCO
R1=MeCO
R1=H
9
10
11
12
R=pClBn
R=2-naphthyl
R=2-naphthyl
Ð
Ð
11.55
R1=MeCO
13
14
15
R=2-naphthyl
R=2-naphthyl
R1=H
R1=H
R2=
R2=
9.10
5.10
4.52
23.10
3.67
7.15
R=mTFMB
16
17
R=mTFMB
R=2-naphthyl
38.80
>5 0
35.45
>5 0
18
19
20
21
22
R=phenyl
>5 0
20.5
24.6
>5 0
10.6
>5 0
R=4-chlorophenyl
R=4-methoxyphenyl
R=2-naphthyl
25.16
55.10
>5 0
19.72
aCytotoxicity was determined as described in refs 27 and 28.

442
S. Hanessian et al. / Bioorg. Med. Chem. Lett. 10 (2000) 439±442
more encouraging were the results of inhibitions of the
growth of A-431 and HT-29 human cell lines (Table 1).
In particular m-tri¯uoromethyl benzenesulfonate esters
of Type A bearing a carboxylate moiety at C-2 (entries
4±8), 2-naphthylsulfonate esters of Type B (entry 14),
and the m-tri¯uoromethyl benzenesulfonate esters with
carboxylate and carbamate esters at C-2 and C-3,
respectively (Type C, entry 15), exhibited IC50 values
ranging from 2.0 to 7.5 mM for both cell lines. Ester and
ether groups at C-6 were much less active (Table 1,
entries 18±22). In an e€ort to mimic the phosphotyro-
sine group, we prepared the glycomers shown in entries
16 and 17, but they were found to be inactive, perhaps
because of limited cell penetration due to the charged
phosphate.
3. Battistini, C.; Penco, S.; Comoglio, P. M. In Biochemistry of
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Although the mechanism of action of these compounds
is still unknown, it should be noted that the A-431 cells
overexpress the EGF receptor whose signal transduc-
tion also involves a Grb2±SH2 domain interaction (e.g.
refs 9±12). Perhaps of greater interest is the ®nding that
the active glycomers shown in Table 1 are neutral
molecules.
13. di Renzo, M. F.; Olivero, M.; Giacomini, A.; Porte, H.;
Chastre, E.; Mirossay, L.; Nordlinger, B.; Bretti, S.; Bottardi,
S.; Giordano, S.; Plebani, M.; Gespach, G.; Comoglio, D. M.
Clin. Cancer Res. 1995, 1, 147.
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glio, D. M. Cell 1994, 77, 261.
15. Beattie, J. Cell Signal 1996, 2, 75.
16. Smithgall, T. E. J. Pharmacol. Toxicol. Methods 1995, 34,
125.
The observed activity against the A-431 and H-29 cell
lines is exciting since it implies that such glycomers are
capable of penetrating the cells. Thus, the prospects of
improving antitumor activity by functional group tun-
ing and modi®cation of the glycomer skeleton may
prove to be highly bene®cial, particularly for the much
dreaded human epidermoid carcinomas.
17. Gishiwzky, M. L. Annu. Rep. Med. Chem. 1995, 30, 247.
18. Brugge, J. S. Science 1993, 260, 918.
19. Rahuel, J.; Gay, B.; Erdmann, D.; Strauss, A.; Garcia-
Echeverria, G.; Furet, P.; Caravatti, G.; Fretz, H.; Schoepfer,
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cia, P.; Sgarella, L.; Bolis, G.; Battistini, C. The Crystal
Structure of the Complex between the Grb2-SH2 Domain and
the Tetrapeptide pYVNV. Abstracts No. 231, Keystone Sym-
posium, Keystone, CO, 31 March 1997.
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hedron 1981, 37, 2779.
In conclusion, we have devised a new carbohydrate-
based sca€old which is amenable to systematic functio-
nalizations using the hydroxyl groups as anchors to
provide a series of glycomers. Exploration of a limited
subset of hydrophobic functional groups, and allowing
for some latitude for H-bonding or polar interactions at
some sites, has led to compounds that inhibit A-431 and
A-29 human tumor cell lines at levels as low as 2 mM.
These encouraging results should pave the way to the
design and synthesis of related non-peptidic neutral
compounds,29 with improved antitumor activities, espe-
cially utilizing parallel or combinatorial synthesis pro-
tocols.30
25. David, S.; Hanessian, S. Tetrahedron 1985, 41, 643.
26. For a review, see Igarashi, K. Advan. Carbohydr. Chem.
Biochem. 1977, 34, 243.
Acknowledgement
27. Skehan, P.; Storeng, R.; Suidiero, D.; Monks, A.; McMa-
hon, J.; Vistica, D.; Warren, J. T.; Bokesch, H.; Kenney, S.;
Boyd, M. R. J. Natl. Cancer Inst. 1990, 82, 1107.
28. Rubinstein, L. V.; Shoemaker, R. H.; Paull, K. D.; Simon,
R. M.; Tosini, S.; Skehan, P.; Surdiero, D. A.; Monks, A.;
Boyo, M. R. J. Natl. Cancer Inst. 1990, 82, 1113.
29. See for example, Plummer. M.; Holland, D.; Shahripour,
A.; Lunney. E. A.; Fergus, J. H.; Marks, J. S.; McConnell, P.;
Mueller, W. T.; Sawyer. T. K. J. Med. Chem. 1997, 40, 3719.
30. See for example, Lee, T. R.; Lawrence, D. S. J. Med.
Chem. 1999, 42, 784.
We thank NSERCC for generous ®nancial assistance
through the Medicinal Chemistry Chair Program.
References and Notes
1. Smittberger, T.; Waldmann, H. Synlett 1998, 574.
2. Burke, Jr., T. R.; Yao, Z. J.; Smyth, M. S.; Ye, B. Curr.
Pharm. Des. 1997, 3, 291.