Dependence of reactivity of a novel 2,6-diamino pyridine-based enediyne on the extent of salt formation with external acids: A possible implication in pH based drug design

Article abstract of DOI:10.1016/j.bmcl.2005.02.051

A novel pyridine diamine-based cyclic enediyne 1 was synthesized by bis-N-alkylation of 2,6-bis-sulfonamido pyridine 6 followed by deprotection with thiophenol. A variety of salts 1a-c of the parent amine 1 were prepared. Thermal reactivity studies indicated a dependence of the reactivity upon the extent of salt formation. This observation may be useful in pH-based design of enediynes.

Full text of DOI:10.1016/j.bmcl.2005.02.051

Bioorganic & Medicinal Chemistry Letters 15 (2005) 2061–2064
Dependence of reactivity of a novel 2,6-diamino pyridine-based
enediyne on the extent of salt formation with external acids:
a possible implication in pH based drug design
Amit Basak,* Moumita Kar and Subrata Mandal
Department of Chemistry, Indian Institute of Technology, Kharagpur 721 302, India
Received 28 December 2004; revised 16February 2005; accepted 17 February 2005
Available online 22 March 2005
Abstract—A novel pyridine diamine-based cyclic enediyne 1 was synthesized by bis-N-alkylation of 2,6-bis-sulfonamido pyridine 6
followed by deprotection with thiophenol. A variety of salts 1a–c of the parent amine 1 were prepared. Thermal reactivity studies
indicated a dependence of the reactivity upon the extent of salt formation. This observation may be useful in pH-based design of
enediynes.
Ó 2005 Elsevier Ltd. All rights reserved.
Selective cytotoxic activity against tumor cells has
been a major challenge in anticancer chemotherapy.1
Although a large repertoire of anticancer drugs exists,
their side effects arising from their action against normal
cells in addition to the cancer cells puts a serious limita-
tion to their use. Apart from the targeted delivery of
antibody conjugate of an anticancer drug like caliche-
amicin,2 attempts have also been made to exploit the
lower pH of cancer cells as compared to normal cells
by drugs,3 which show higher cytotoxic activity at
higher acidity. In the area of enediynes, the azaenediynes
synthesized by Kerwin and David4 provides a unique
example where the abstraction of H radical from a suit-
able donor has been shown to be pH dependent due to
large singlet to triplet barrier.5 Although enediynes are
activated by electron withdrawal,6 no attempt has so
far been made to prepare an organo base-containing
enediyne and study its properties under salt forming
conditions (to mimic the variation of pH by using a vari-
ety of acid). In this communication we report the syn-
thesis of such a molecule, namely, a novel pyridine
diamine-based cyclic enediyne 1. We have also com-
pared its thermal reactivity with those of its salts pre-
pared from various acids. The aim of our study was to
determine the effect of salt formation, which happens
at acidic pH on the activation barrier to Bergman cycli-
zation (BC).
At the outset, we realized the need to protect the two
primary amines in order to achieve the synthesis of
our target. Our initial plan was to use t-Boc as the pro-
tecting group because of its easy deprotection under acid
conditions. Thus we synthesized N,N0-di-t-Boc-2,6-
diamino pyridine 3 using standard conditions (Boc-
anhydride in EtOAc). Attempted alkylation with the
enediyne dimesylate 4 in the presence of NaH in DMF
or DMSO was not successful; the expected dialkylation
product could never be isolated. Having failed in the ini-
tial strategy, we thought of using 4-nitrobenzene sulfon-
amide as the protecting group.7 The latter offers several
advantages, which include the ease of alkylation and
deprotection under mild conditions. Thus we prepared
the bis-sulfonamide 6 and double alkylation with the
dibromide in the presence of K2CO3 and DMF worked
out nicely under normal dilution to afford the cyclic
enediyne as a brown solid. The free diamino enediyne
was then obtained by deprotection of the sulfonamido
groups with thiophenol and K2CO3 in DMF. It was iso-
lated pure as a brown solid by chromatography over
Si-gel using hexane–ethyl acetate (5:1) as eluent. The
1H NMR and mass spectral data fully corroborated its
structure. The synthesis is shown in Scheme 1.
Having successfully prepared enediyne 1, we first studied
its salt forming abilities with various acids. Three dif-
ferent acids of varying pKa, namely p-toluene sulfonic
acid, pyridine 2,6-dicarboxylic acid and 3,5-dinitrosali-
cylic acid were used. These acids were chosen because
of their reported salt formation with pyridine diamine.8
Keywords: Enediyne; Bergman cyclization; Calorimetry; Drug; Design.
*
Corresponding author. Tel.: +91 322283300; fax: +91 322282252;
e-mail: absk@chem.iitkgp.ernet.in
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.02.051

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A. Basak et al. / Bioorg. Med. Chem. Lett. 15 (2005) 2061–2064
NCOOt-Bu
H2N
t-BuOOCHN
b
4
a
N
N
N
NCOOt-Bu
H2N
2
t-BuOOCHN
3
5
c
Not Obtained
a Boc anhydride, Ethyl acetate, 80%
b NaH, DMF, 0%
N
6
NHSO2Ar
c 4-nitrobenzene sulphonyl chloride, Py, 85%
d K2CO3, DMF, rt, 70%
ArO2SHN
e PhSH, K2CO3, DMF
4
d
SO2Ar
H
H
N
HN
N
N
N
N
N
e
HA
A
+
N
H
NSO2Ar
OMs
H
1A-C
1
7
Ar =
NO2
SO3H
COOH
N
For 1a HA =
For 1b HA =
O2N OH
COOH
For 1c HA =
O2N
OMs
COOH
4
Scheme 1. Synthesis of enediyne 1 and its salts.
The salts were prepared by adding a DMSO solution of
the acid to a DMSO solution of 1 equiv of the diamine.
The solution was stirred at rt for 12 h and then lyophi-
lized. Addition of CH2Cl2 precipitated the salts, which
were then dried under vacuum. The extent of salt forma-
tion was checked by 1H NMR. The signal for the meth-
ylene protons as well as for the pyridine 3,5-hydrogens
underwent downfield shifts of varying degrees (Table
1).9
presented in the table, it is clear that the salts were under-
going BC at a lower temperature than the parent ened-
iyne. This is most likely due to the electron withdrawal
from the enediyne network by the positively charged
pyridine nucleus. The lowering of onset temperature is
highest for the salts with toluene sulfonic acid and 3,5-
dinitrosalicylic acid. This can be explained on the basis
of lower pKa of both the acids in comparison to pyridine
dicarboxylic acid. The salt formation being more, there
will be greater withdrawal of electrons from the enediyne
network in the case of the tosylate and the salicylate.12
With negligible salt formation, there is only a marginal
decrease in the onset temperature for cyclization for the
case with pyridine dicarboxylic acid.
With a strong acid like p-toluene sulfonic acid, the
amount of shift was dependent upon the time of interac-
tion between the enediyne and the acid. The NMR taken
immediately after the addition of sulfonic acid showed a
shift of methylene and pyridine 3,5-hydrogens, which
ultimately reached a limiting value after 12 h of stirring.
A similar result was obtained with 3,5-dinitrosalicylic
acid, which is sufficiently strong for complete conversion
into the pyridinium salt. With a weaker acid like pyr-
idine 2,6-dicarboxylic acid, the shift of the above signals
was just marginal indicating little salt formation (Fig. 2).
The shift for the NH hydrogens could not be determined
as the signal was obscured by the aromatic signals
(Fig. 1).
In conclusion we have demonstrated a novel way of acti-
vation of a basic pyridine diamine based enediyne via
salt formation with organic acids. Varying the pKa of
the acid can also regulate the extent of activation. This
may have implication in the design of pH based antican-
cer agents. Current efforts in our laboratory are aimed
towards bringing down the activation further so that
BC takes place under ambient conditions.
The thermal reactivity towards BC for the parent ene-
diyne and its salts were next studied by Differential Scan-
ning Calorimetry, which has now become a useful tool
for studying such a reaction.10 The onset temperature
for BC for the three compounds along with change in
chemical shift values are shown in Table 1. From the data
Selected spectral data
Pyridine diamine bis-sulfonamide 7
dH (200 MHz, d6-DMSO) 4.74 (2H, br s), 7.17–7.24 (4H,
m), 7.61 (2H, dd, J = 2.0, 6.0 Hz), 7.82 (4H, d,
Table 1. NMR shifts and DSC results
Compd no. Dd for CH2 pKa of acids11 Dd for pyridine H-3 and H-5 Onset temp (T2, °C) DT (T1 ꢀ T2) Onset temp for 1 (T1, °C)
1A
1B
1C
0.43
ꢁ0.00
0.39
ꢀ6.5
0.33
ꢁ0.00
0.32
215
232
214
19
4.3
1.53
2
20
234

A. Basak et al. / Bioorg. Med. Chem. Lett. 15 (2005) 2061–2064
2063
Figure 1. DSC curves of parent enediyne 1 and its salts.
Figure 2. NMR spectra of parent enediyne and its salts.
J = 8.8 Hz), 8.09 (4H, d, J = 8.8 Hz), 8.12 (1H, m,
obscured); dC (50 MHz, d6-DMSO) 83.0, 86.5,
121.5, 123.9, 124.3, 128.4, 129.2, 130.9, 141.4,
143.4, 149.6, 150.5; Mass (ES+) 652 (MNa+), 630
(MH+).
Acknowledgements
A.B. expresses thanks to the Council of Scientific and
Industrial Research for funding. M.K. also thanks
the Council of Scientific and Industrial Research for a
junior research fellowship.
Enediyne 1
References and notes
dH (200 MHz, d6-DMSO) 4.18 (4H, d, J = 5.5 Hz),
5.75 (2H, d, J = 9.1 Hz), 6.68 (2H, t, J = 5.5 Hz), 7.08
(1H, t, J = 9.1 Hz), 7.42 (4H, m); Mass (ES+) 260
(MH+).
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