Parallel synthesis of novel 3-substituted 1-ethyl-6-methylpyrimido[4,5-c] pyridazine-5,7(1H,6H)-dione Analogs

Article abstract of DOI:10.1080/00397910903013788

A library of novel 1H-pyrimido[4,5-c]pyridazine-5,7-dione analogs was prepared in a parallel fashion via the cyclization of 6-(1-ethylhydrazinyl)-3- methylpyrimidine-2,4(1H,3H)-dione (4) with-bromoketones and was then isolated by a two-step purification method with 90% purity. Both the library synthesis and purification method proved suitable for analogs with a high diversity of substitution at the 3-position including various alkyl, aryl, and heteroaryl groups.

Full text of DOI:10.1080/00397910903013788

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Parallel Synthesis of Novel 3-Substituted
1-Ethyl-6-methylpyrimido[4,5-
c]pyridazine-5,7(1H,6H)-dione Analogs
Yongsheng Chen a , Jack R. Barber a , Shi Chung Ng a & Yuefen Zhou a
a CytRx Corporation , San Diego, California, USA
Published online: 22 Feb 2010.
To cite this article: Yongsheng Chen , Jack R. Barber , Shi Chung Ng & Yuefen Zhou (2010) Parallel
Synthesis of Novel 3-Substituted 1-Ethyl-6-methylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-dione Analogs,
Synthetic Communications: An International Journal for Rapid Communication of Synthetic Organic
Chemistry, 40:6, 821-832, DOI: 10.1080/00397910903013788
To link to this article: http://dx.doi.org/10.1080/00397910903013788
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Synthetic Communications1, 40: 821–832, 2010
Copyright # Taylor & Francis Group, LLC
ISSN: 0039-7911 print=1532-2432 online
DOI: 10.1080/00397910903013788
PARALLEL SYNTHESIS OF NOVEL 3-SUBSTITUTED
1-ETHYL-6-METHYLPYRIMIDO[4,5-c]PYRIDAZINE-
5,7(1H,6H)-DIONE ANALOGS
Yongsheng Chen, Jack R. Barber, Shi Chung Ng, and
Yuefen Zhou
CytRx Corporation, San Diego, California, USA
A library of novel 1H-pyrimido[4,5-c]pyridazine-5,7-dione analogs was prepared in
a
parallel fashion via the cyclization of 6-(1-ethylhydrazinyl)-3-methylpyrimidine-
2,4(1H,3H)-dione (4) with a-bromoketones and was then isolated by a two-step purifi-
cation method with >90% purity. Both the library synthesis and purification method proved
suitable for analogs with a high diversity of substitution at the 3-position including various
alkyl, aryl, and heteroaryl groups.
Keywords: Cyclization; 6-(1-ethylhydrazinyl)-3-methylpyrimidine-2,4(1H,3H)-dione; parallel synthesis;
1H-pyrimido[4,5-c]pyridazine-5,7-dione analogs
1H-Pyrimido[4,5-c]pyridazine-5,7-dione derivatives have been shown to exhibit
interesting biological activities and pharmaceutical potential. For instance, 4-
deazatoxoflavin is a known compound (1) (Fig. 1) that has antibiotic properties.[1]
In the course of our research to develop novel small-molecule chaperone amplifiers
to treat neurodegenerative diseases, we needed to make a small library of novel ana-
logs of 3-substituted 1H-pyrimido[4,5-c]pyridazine-5,7-dione for our primary high-
throughput screening to identify initial hits. Therefore, a versatile method that is
suitable for parallel synthesis of the novel analogs 2 in this class was developed.[2]
There are literature reports of methods for the synthesis of this scaffold
individually where a methyl group is substituted at the 1-position.[3] However, the
limited number of analogs reported usually were prepared in very poor yields with
great difficulty in purification. This turned out to be the major challenge for us in
preparing a library of the novel analogs 2 with high purity in solution-phase parallel
synthesis.
We started our method-development work based on a modified literature
method, as shown in Scheme 1. Commercially available 6-chloro-3-methyl-1H-
pyrimidine-2,4-dione (3) was chosen as a starting material. To prepare the key inter-
mediate 4, existing literature procedures required the use of ethyl hydrazine in its
Received March 25, 2009.
Address correspondence to Yongsheng Chen, CytRx Corporation, 3030 Bunker Hill Street, Suite
101, San Diego, CA 92109, USA. E-mail: ychen@cytrx.com
821

822
Y. CHEN ET AL.
Figure 1. 1H-Pyrimido[4,5-c]pyridazine-5,7-dione derivatives.
basic form, which is not readily available commercially and required synthesis from
ethyl bromide and hydrazine followed by distillation.[4] We chose to use the readily
available oxalate salt form of ethyl hydrazine with the addition of a base to the reac-
tion mixture to generate ethyl hydrazine in situ. A variety of bases have been tried
including cesium carbonate, potassium carbonate, sodium acetate, pyridine, and dii-
sopropylethylamine (DIPEA), in combination with different solvents including
dimethylformamide (DMF), dimethylsulfoxide (DMSO), dioxane, and ethanol.
The majority of the tested reaction conditions resulted in quite complex reaction
mixtures or difficulty in the isolation of product 4. It turned out that heating the
reaction at 100 ꢀC overnight in EtOH in the presence of diisopropylethyl amine
(DIPEA) in a sealed tube gave the best result, and intermediate 4 was easily
separated by simple filtration with moderate isolated yield.
Once the key intermediate 4 was made, converting it to the desired product 2
could be achieved by either cyclizing 4 with an a-bromoketone at 120 ꢀC in a single
step or stepwise reacting 4 with an aldehyde followed by triethyl orthoformate at a
higher temperature. As fewer steps of synthesis are generally beneficial for a library
synthesis, we chose the former method, and the reactions were usually successful
judging by the presence of the desired mass peak in liquid chromatographic-mass
spectrometry (LC-MS) spectra. The challenge came with the purification of the final
products, as most of them had poor solubility in organic solvents commonly used for
normal-phase chromatographic purification such as methanol, ethyl acetate, and
hexane. In addition, an unknown side product (as the major impurity with mass unit
twice the desired mass), which could not be completely removed by preparative
reverse-phase high-performance liquid chromatography (HPLC) alone, often
occurred and appeared very closely to the desired product. Eventually, a two-step
purification method was developed using preparative TLC normal-phase
Scheme 1. Synthesis of 3-substituted 1-ethyl-6-methylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-dione
derivatives.

NOVEL PYRIMIDO[4,5-c]PYRIDAZINE-5,7-DIONE ANALOGS
823
Table 1. 3-Substituted 1-ethyl-6-methylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-
diones 2
Compound
2{1}
R
2{2}
2{3}
2{4}
2{5}
2{6}
2{7}
2{8}
2{9}
2{10}
2{11}
(Continued )

824
Y. CHEN ET AL.
Table 1. Continued
Compound
2{12}
R
2{13}
2{14}
2{15}
2{16}
2{17}
2{18}
2{19}
2{20}
2{21}
2{22}
2{23}
2{24}
(Continued )

NOVEL PYRIMIDO[4,5-c]PYRIDAZINE-5,7-DIONE ANALOGS
825
Table 1. Continued
Compound
R
2{25}
2{26}
2{27}
2{28}
2{29}
2{30}
2{31}
2{32}
2{33}
2{34}
2{35}
2{36}
2{37}
purification (MeOH=DCM ¼ 1=40) followed by preparative reverse-phase HPLC
purification with a suitable gradient of CH3CN and H2O. However, the isolated
yields were usually poor after multiple purifications.
Once the reaction conditions and purification methods were optimized and
developed, the key intermediate 4 was prepared on a large scale with high purity
as a building block. A library of 37 compounds (with variation of R at the 3-
position) was synthesized in one step in a parallel-batch fashion (Table 1). All of

826
Y. CHEN ET AL.
the products were purified by normal-phase preparative TLC followed by prepara-
tive reverse-phase HPLC with >90% purity and confirmed by LC-MS and=or
1H NMR spectra.
As shown in Table 1, a broad range of R groups were used in the library,
including alkyl (acyclic or cyclic), phenyl, mono- and di-substituted phenyl, and dif-
ferent types of heterocyclic rings. Despite the big variation of electronic properties
and steric hindrance of the R groups, all of the reactions worked fairly well on a
small scale (mg) to provide an adequate amount of pure products for biological
screening.
In summary, we have successfully optimized the reaction conditions based on
literature methods for the parallel synthesis of novel 3-substituted 1-ethyl-6-
methylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-dione derivatives and developed a
two-step purification method to provide a library of 37 analogs in this class with high
purity. Our library synthesis and purification method proved suitable for diversified
substituents at the 3-position regardless of the variation of electronic and steric
properties.
EXPERIMENTAL
General
Anhydrous solvents purchased from EMD were used directly without
additional treatment. All other reagents and solvents were reagent grade and used
as received from commercial sources. TLC was performed using EM Science pre-
coated silica gel 60F 254 plates. LC-MS data was recorded on a LCQ Advantage
MAX mass spectrometer with electrospray ionization (ESIþ). NMR spectra were
obtained on a Bruker AMX-500 II or DMX-500 spectrometer, and chemical shifts
are reported in ppm (d) using internal sovent signals as references.
Synthesis of 6-(1-Ethylhydrazinyl)-3-methylpyrimidine-2,4(1H,3H)-
dione (4)
A mixture of 6-chloro-(1-ethylhydrazinyl)-3-methylpyrimidine-2,4(1H,3H)-
dione (3; 7.12 g, 0.044 mol), ethylhydrazine oxalate (10 g, 0.066 mol), and N,N-
di-isopropyl-ethylamine (17.2 g, 0.133 mol) in dry EtOH (200 mL) was heated in a
sealed bottle at 100 ꢀC for 16 h. After cooling to ambient temperature, the desired
product was filtered off from the mixture under reduced pressure to give the key
intermediate 4 (4.6 g, 56%). 1H NMR (DMSO): d 4.71 (1H, s), 3.38 (2H, J ¼ 6.9 Hz,
q), 3.03 (3H, s), 1.11 (3H, J ¼ 6.9 Hz, t); LC-MS (ESI) m=z ¼ 185 [M þ H]þ.
General Synthetic Procedure for Preparation of Library
Compounds 2{1–37}
The mixtures of 6-(1-ethylhydrazinyl)-3-methylpyrimidine-2,4(1H,3H)-dione
(4; 100 mg, 0.543 mmol) and a-bromoketones (0.543 mmol) in 2-methoxyethanol
(3 mL) were heated at 120 ꢀC in sealed vials for 16 h in a parallel-batch fashion. Each
vial contained a single reaction mixture with the appropriate a-bromoketone.

NOVEL PYRIMIDO[4,5-c]PYRIDAZINE-5,7-DIONE ANALOGS
827
The reactions were concentrated in vacuo to give crude products, which were first
purified by normal-phase preparative TLC (20 ꢁ 20 cm2) (DCM=MeOH ¼ 40:1),
then further purified by preparative reverse-phase HPLC to give the desired pro-
ducts. Preparative reverse-phase HPLC purification was carried out on a Gilson
215 HPLC (XBP 250 ꢁ 20 mm ꢁ 10 mm C18, flow rate: 15 mL=min) with dual wave-
length UV detection (214, 254 nm) using a specified gradient of acetonitrile in water
with 0.1% of trifluoroaceticacid (TFA) over 15 min.
1-Ethyl-6-methyl-3-phenylpyridazino[3,4-d]pyrimidne-5,7(1H,6H)-
dione (2{1})
Prep-HPLC gradient: 25–95%; yield: 10.7 mg (7.6% from 92 mg of 4). 1H NMR
(CDCl3) d 8.66 (1H, s), 7.93 (2H, m), 7.54 (3H, m), 4.77 (2H, q, J ¼ 7.2 Hz), 3.48 (3H,
s), 1.58 (3H, t, J ¼ 7.2 Hz); 13C NMR (CDCl3) d 160.9, 156.3, 154.6, 148.6, 133.4,
131.0, 129.6, 128.8, 126.4, 123.6, 51.1, 28.7, 13.9; LC-MS (ESI) m=z ¼ 283 [M þ H]þ.
3-(2-Chlorophenyl)-1-ethyl-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{2})
Prep-HPLC gradient: 25–90%; yield: 36 mg (20.9%). LC-MS (ESI) m=z ¼ 317
[M þ H]þ.
1-Ethyl-3-(2-hydroxyphenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{3})
Prep-HPLC gradient: 25–90%; yield: 15 mg (9.3%). 1H NMR (DMSO) d 10.47
(1H, s), 8.74 (1H, s), 7.71 (1H, d, J ¼ 7.8 Hz), 7.40 (1H, m), 7.06 (2H, m), 4.61 (2H, d,
J ¼ 6.9 Hz), 3.24 (3H, s), 1.49 (3H, m); LC-MS (ESI) m=z ¼ 299 [M þ H]þ.
1-Ethyl-3-(2-methoxyphenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{4})
1
Prep-HPLC gradient: 10–90%; yield: 27 mg (15.9%). H NMR (CDCl3) d 8.72
(1H, s), 7.64 (1H, d, J ¼ 7.5 Hz), 7.49 (1H, m), 7.11 (2H, m), 4.71 (2H, m), 3.89 (3H,
s), 3.45 (3H, s), 1.56 (3H, m); LC-MS (ESI) m=z ¼ 313 [M þ H]þ.
3-(3-Chlorophenyl)-1-ethyl-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{5})
Prep-HPLC gradient: 50–90%; yield: 20 mg (11.6%). LC-MS (ESI) m=z ¼
317 [M þ H]þ.
1-Ethyl-3-(3-fluorophenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{6})
Prep-HPLC gradient: 25–95%; yield: 20 mg (12.2%). LC-MS (ESI) m=z ¼
301 [M þ H]þ.

828
Y. CHEN ET AL.
1-Ethyl-3-(3-hydroxyphenyl)-6-methylpyridazino[3,4-d]pyrimidne-
5,7(1H,6H)-dione (2{7})
Prep-HPLC gradient: 20–50%; yield: 25 mg (15.4%). 1H NMR (DMSO) d 9.76
(1H, s), 8.56 (1H, s), 7.49 (3H, m), 6.92 (1H, d, J ¼ 7.5 Hz), 4.60 (2H, m), 3.24 (3H, s),
1.46 (3H, m); LC-MS (ESI) m=z ¼ 299 [M þ H]þ.
1-Ethyl-3-(3-methoxyphenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{8})
1
Prep-HPLC gradient: 40–80%; yield: 20 mg (11.7%). H NMR (CDCl3) d 8.64
(1H, s), 7.48 (3H, s), 7.08 (1H, s), 4.78 (2H, d, J ¼ 6 Hz), 3.90 (3H, s), 3.48 (3H, s),
1.58 (3H, s); LC-MS (ESI) m=z ¼ 313 [M þ H]þ.
3-(4-Chlorophenyl)-1-ethyl-6-methylpyrimido[4,5-c]pyridazine-
5,7(1H,6H)-dione (2{9})
1
Prep-HPLC gradient: 25–95%; yield: 20 mg (11.6%). H NMR (CDCl3) d 8.62
(1H, s), 7.90 (2H, d, J ¼ 8.1 Hz), 7.50 (2H, d, J ¼ 8.1 Hz), 4.80 (2H, m), 3.48 (3H, s),
1.60 (3H, m); LC-MS (ESI) m=z ¼ 317 [M þ H]þ.
4-(1-Ethyl-6-methyl-5,7-dioxo-1,5,6,7-tetrahydropyridazino[3,4-d]
pyrimidin-3-yl)benzonitrile (2{10})
1
Prep-HPLC gradient: 40–80%; yield: 15 mg (8.9%). H NMR (CDCl3) d 8.65
(1H, s), 8.08 (2H, d, J ¼ 7.8 Hz), 7.86 (2H, d, J ¼ 7.5 Hz), 4.80 (2H, m), 3.49 (3H, s),
1.62 (3H, m); LC-MS (ESI) m=z ¼ 308 [M þ H]þ.
1-Ethyl-3-(4-fluorophenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{11})
1
Prep-HPLC gradient: 25–90%; yield: 32 mg (19.6%). H NMR (CDCl3) d 8.63
(1H, s), 7.97 (2H, m), 7.25 (2H, d, J ¼ 8.4 Hz), 4.79 (2H, m), 3.49 (3H, s), 1.62
(3H, m); LC-MS (ESI) m=z ¼ 301 [M þ H]þ.
1-Ethyl-3-(4-methoxyphenyl)-6-methylpyridazino[3,4-d]pyrimidine-
5,7(1H,6H)-dione (2{12})
1
Prep-HPLC gradient: 40–80%; yield: 38 mg (22.4%). H NMR (CDCl3) d 8.65
(1H, s), 7.93 (2H, d, J ¼ 7.2 Hz), 7.09 (2H, d, J ¼ 7.5 Hz), 4.80 (2H, d, J ¼ 6.9 Hz),
3.93 (3H, s), 3.52 (3H, s), 1.63 (3H, d, J ¼ 6.9 Hz); LC-MS (ESI) m=z ¼ 313 [M þ H]þ.
1-Ethyl-6-methyl-3-(4-(Trifluoromethoxy)phenyl)pyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{13})
Prep-HPLC gradient: 25–90%; yield: 20 mg (10.0%). LC-MS (ESI) m=z ¼
367 [M þ H]þ.

NOVEL PYRIMIDO[4,5-c]PYRIDAZINE-5,7-DIONE ANALOGS
829
1-Ethyl-6-methyl-3-p-tolylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-
dione (2{14})
1
Prep-HPLC gradient: 40–70%; yield: 14 mg (8.7%). H NMR (CDCl3) d 8.63
(1H, s), 7.83 (2H, d, J ¼ 8.1 Hz), 7.35 (2H, d, J ¼ 8.1 Hz), 4.77 (2H, d, J ¼ 6.6 Hz),
3.48 (3H, s), 2.43 (3H, s), 1.59 (3H, d, J ¼ 6.9 Hz); LC-MS (ESI) m=z ¼ 297 [M þ H]þ.
1-Ethyl-6-methyl-3-(4-(trifluoromethyl)phenyl)pyridazino
[3,4-d]pyrimidine-5,7(1H,6H)-dione (2{15})
1
Prep-HPLC gradient: 25–95%; yield: 25 mg (13.1%). H NMR (CDCl3) d 8.66
(1H, s), 8.07 (2H, d, J ¼ 8.1 Hz), 7.81 (2H, d, J ¼ 8.1 Hz), 4.81 (2H, m), 3.47 (3H, s),
1.61 (3H, m); LC-MS (ESI) m=z ¼ 351 [M þ H]þ.
3-(2,4-Dichlorophenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{16})
1
Prep-HPLC gradient: 25–90%; yield: 30 mg (15.7%). H NMR (CDCl3) d 8.53
(1H, s), 7.56 (3H, m), 4.77 (2H, m), 3.46 (3H, s), 1.57 (3H, m); LC-MS (ESI) m=z ¼
352 [M þ H]þ.
3-(2,4-Difluorophenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{17})
Prep-HPLC gradient: 40–85%; yield: 40 mg (23.1%). LC-MS (ESI) m=z ¼
319 [M þ H]þ.
3-(2,4-Dihydroxyphenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{18})
Prep-HPLC gradient: 20–80%; yield: 15 mg (8.8%). 1H NMR (DMSO) d 11.73
(1H, s), 11.10 (1H, s), 9.98 (1H, s), 7.34 (1H, d, J ¼ 8.7 Hz), 6.41 (2H, m), 3.90 (2H, d,
J ¼ 6.6 Hz), 3.19 (3H, m), 1.21 (3H, m); LC-MS (ESI) m=z ¼ 315 [M þ H]þ.
3-(2,4-Dimethoxyphenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{19})
1
Prep-HPLC gradient: 25–90%; yield: 19 mg (10.2%). H NMR (CDCl3) d 8.76
(1H, s), 6.58 (2H, s), 4.78 (2H, s), 3.91 (6H, d, J ¼ 3.3 Hz), 3.49 (3H, s), 1.58 (3H, s);
LC-MS (ESI) m=z ¼ 343 [M þ H]þ.
3-(3-Chloro-4-fluorophenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{20})
1
Prep-HPLC gradient: 25–95%; yield: 15 mg (8.2%). H NMR (CDCl3) d 8.58
(1H, s), 8.03 (2H, d, J ¼ 6.6 Hz), 7.81 (1H, s), 7.35 (1H, m), 4.80 (2H, m), 3.48
(3H, s), 1.58 (3H, m); LC-MS (ESI) m=z ¼ 335 [M þ H]þ.

830
Y. CHEN ET AL.
3-(5-Chloro-2-hydroxyphenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{21})
Prep-HPLC gradient: 50–90%; yield: 15 mg (8.3%). 1H NMR (DMSO) d 10.82
(1H, s), 8.73 (1H, s), 7.70 (1H, s), 7.43 (1H, d, J ¼ 8.4 Hz), 7.08 (1H, d, J ¼ 8.4 Hz),
4.62 (1H, d, J ¼ 6.9 Hz), 3.34 (1H, s), 1.49 (3H, m); LC-MS (ESI) m=z ¼ 333 [M þ H]þ.
1-Ethyl-3-(5-fluoro-2-hydroxyphenyl)-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{22})
Prep-HPLC gradient: 10–90%; yield: 10 mg (5.8%). LC-MS (ESI) m=z ¼
317 [M þ H]þ.
3-(2,5-Dimethoxyphenyl)-1-ethyl-6-methylpyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{23})
1
Prep-HPLC gradient: 25–95%; yield: 30 mg (16.1%). H NMR (CDCl3) d 8.76
(1H, s), 7.22 (1H, s), 7.01 (2H, m), 4.79 (2H, m), 3.86 (6H, d, J ¼ 8.4 Hz), 3.47 (3H, s),
1.58 (3H, m); LC-MS (ESI) m=z ¼ 343 [M þ H]þ.
1-Ethyl-6-methyl-3-(naphthalen-2-yl)pyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{24})
1
Prep-HPLC gradient: 70–100%; yield: 12 mg (6.6%). H NMR (CDCl3) d 8.85
(1H, s), 8.41 (1H, s), 8.14 (4H, m), 7.64 (2H, d, J ¼ 3 Hz), 4.86 (2H, d, J ¼ 6.9 Hz),
3.54 (3H, s), 2.04 (1H, s), 1.69 (3H, m); LC-MS (ESI) m=z ¼ 333 [M þ H]þ.
1-Ethyl-6-methyl-3-(2-oxo-2H-chromen-3-yl)pyridazino[3,4-d]
pyrimidine-5,7(1H,6H)-dione (2{25})
1
Prep-HPLC gradient: 20–50%; yield: 10 mg (5.2%). H NMR (CDCl3) d 9.09
(1H, s), 8.42 (1H, s), 7.70 (2H, m), 7.46 (2H, m), 4.79 (2H, d, J ¼ 7.2 Hz), 3.48
(3H, s), 1.61 (3H, m); LC-MS (ESI) m=z ¼ 351 [M þ H]þ.
1-Ethyl-3-(furan-2-yl)-6-methylpyridazino[3,4-d]pyrimidne-
5,7(1H,6H)-dione (2{26})
Prep-HPLC gradient: 25–95%; yield: 9.3 mg (6.8% from 92 mg of 4). 1H NMR
(CDCl3) d 8.57 (1H, s), 7.63 (1H, m), 7.08 (1H, m), 6.61 (1H, m), 4.72 (2H, q,
J ¼ 7.2 Hz), 3.47 (3H, s), 1.55 (3H, t, J ¼ 7.2 Hz); LC-MS (ESI) m=z ¼ 273 [M þ H]þ.
1-Ethyl-6-methyl-3-(5-methylfuran-2-yl)pyridazino[3,4-d]
pyrimidne-5,7(1H,6H)-dione (2{27})
1
Prep-HPLC gradient: 25–95%; yield: 11 mg (7.7% from 92 mg of 4). H NMR
(CDCl3) d 8.53 (1H, s), 6.97 (1H, m), 6.20 (1H, m), 4.70 (2H, q, J ¼ 7.2 Hz), 3.47 (3H,
s), 2.43 (3H, s), 1.54 (3H, t, J ¼ 7.2 Hz); LC-MS (ESI) m=z ¼ 287 [M þ H]þ.

NOVEL PYRIMIDO[4,5-c]PYRIDAZINE-5,7-DIONE ANALOGS
831
1-Ethyl-6-methyl-3-(thiophen-2-yl)pyridazino[3,4-d]pyrimidne-5,7
(1H,6H)-dione (2{28})
Prep-HPLC gradient: 25–95%; yield: 4.7 mg (6.0% from 50 mg of 4). LC-MS
(ESI) m=z ¼ 289 [M þ H]þ.
1-Ethyl-6-methyl-3-(thiazol-2-yl)pyridazino[3,4-d]pyrimidne-5,7
(1H,6H)-dione (2{29})
Prep-HPLC gradient: 25–95%; yield: 18.4 mg (12.7% from 92 mg of 4).
1H NMR (CDCl3) d 9.05 (1H, s), 7.99 (1H, m), 7.54 (1H, m), 4.74 (2H, q,
J ¼ 7.2 Hz), 3.48 (3H, s), 1.58 (3H, t, J ¼ 7.2 Hz); LC-MS (ESI) m=z ¼ 290 [M þ H]þ.
1-Ethyl-6-methyl-3-(1-methyl-1H-Benzo[d]imidazol-2-yl)pyrimido
[4,5-c]pyridazine-5,7(1H,6H)-dione (2{30})
Prep-HPLC gradient: 25–95%; yield: 5.9 mg (3.5% from 92 mg of 4). LC-MS
(ESI) m=z ¼ 337 [M þ H]þ.
3-(Benzofuran-2-yl)-1-ethyl-6-methylpyrimido[4,5-c]pyridazine-5,7
(1H,6H)-dione (2{31})
Prep-HPLC gradient: 25–95%; yield: 18.9mg (11.7% from 92mg of 4). 1H NMR
(CDCl3) d 8.75 (1H, s), 7.69 (1H, m), 7.61 (1H, m), 7.46 (1H, s), 7.43 (1H, m), 7.33
(1H, m), 4.77 (2H, q, J ¼ 7.2 Hz), 3.49 (3H, s), 1.59 (3H, t, J ¼ 7.2 Hz); LC-MS
(ESI) m=z ¼ 323 [M þ H]þ.
3-(Benzo[b]thiophen-2-yl)-1-ethyl-6-methylpyrimido[4,5-c]
pyridazine-5,7(1H,6H)-dione (2{32})
Prep-HPLC gradient: 25–95%; yield: 11.6 mg (6.9% from 92 mg of 4). LC-MS
(ESI) m=z ¼ 339 [M þ H]þ.
3-(Benzo[b]thiazol-2-yl)-1-ethyl-6-methylpyrimido[4,5-c]pyridazine-
5,7(1H,6H)-dione (2{33})
Prep-HPLC gradient: 25–95%; yield: 13.4 mg (7.9% from 92 mg of 4). 1H NMR
(CDCl3) d 9.18 (1H, s), 8.14 (1H, m), 7.96 (1H, m), 7.57 (1H, s), 7.49 (1H, m), 7.33
(1H, m), 4.77 (2H, q, J ¼ 7.2 Hz), 3.49 (3H, s), 1.62 (3H, t, J ¼ 7.2 Hz); LC-MS (ESI)
m=z ¼ 340 [M þ H]þ.
1-Ethyl-6-methyl-3-(trifluoromethyl)pyridazino[3,4-d]pyrimidine-5,7
(1H,6H)-dione (2{34})
Prep-HPLC gradient: 70–100%; yield: 16 mg (10.7%). 1H NMR (CDCl3) d 8.41
(1H, s), 4.77 (2H, m), 3.47 (3H, s), 1.60 (3H, m); LC-MS (ESI) m=z ¼ 275 [M þ H]þ.

832
Y. CHEN ET AL.
1,3-Diethyl-6-methylpyridazino[3,4-d]pyrimidine-5,7(1H,6H)-dione
(2{35})
1
Prep-HPLC gradient: 10–90%; yield: 11 mg (8.7%). H NMR (CDCl3) d 8.12
(1H, s), 4.69 (2H, m), 3.44 (3H, s), 2.89 (2H, m), 1.52 (3H, m), 1.36 (3H, m); LC-MS
(ESI) m=z ¼ 235 [M þ H]þ.
3-tert-Butyl-1-ethyl-6-methylpyridazino[3,4-d]pyrimidine-5,7
(1H,6H)-dione (2{36})
1
Prep-HPLC gradient: 25–95%; yield: 23 mg (16.2%). H NMR (CDCl3) d 8.32
(1H, s), 4.69 (2H, m), 3.45 (3H, s), 1.53 (3H, m), 1.38 (9H, s); LC-MS (ESI) m=z ¼
263 [M þ H]þ.
1-Ethyl-3-adamantyl-6-methylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-
dione (2{37})
1
Prep-HPLC gradient: 40–80%; yield: 12 mg (6.5%). H NMR (CDCl3) d 8.32
(1H, s), 4.71 (2H, m), 3.46 (3H, s), 2.17 (3H, s), 1.97 (6H, s), 1.87 (7H, m), 1.54
(8H, m), 1.27 (9H, s), 0.89 (4H, d, J ¼ 9 Hz); LC-MS (ESI) m=z ¼ 341 [M þ H]þ.
ACKNOWLEDGMENT
We thank Dr. Linyi Wei for his helpful suggestion on intermediate synthesis.
REFERENCES
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