4-(Isoxazol-3-yl)pyrimidines from Pyrimidinyl Nitrile Oxides

Abstract

The 1,3-dipolar cycloaddition reaction of pyrimidinyl aldoxime derived nitrile oxides and alkynes delivers 4-(isoxazol-3-yl)pyrimidines. The procedures reported accommodate three points of diversification around this bisheterocyclic scaffold and the resulting library of compounds has been added to the National Institutes of Health repository (ca. 10 mg of each with >90% purity) for pilot-scale biomedical studies with bioassay data available at the National Center for Biotechnology Information PubChem database.

Key words

nitrile oxide - 2-aminopyrimidine - isoxazoles - isoxazolines - ipso substitution - cycloaddition

References and Notes

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• General Procedure for Bioassays
• 19a

  The primary screen involved two high-throughput insect assays, both incorporating 96-well microtiter plates. The first assay was a 96-well high-throughput bioassay for beet armyworm (Spodoptera exigua: Lepidoptera) larvae, based on modifications of Lewer et al.(2006). Eggs of S. exigua were placed on top of artificial diet (100 mL) in each well of 96-well microtiter plate. The diet had been pretreated with test compounds (12 mg dissolved in 30 mL of DMSO-acetone-H₂O mixture) layered on top of the diet using a Sagian (Bechman-Coulter, Fullerton, CA) liquid handling system and then allowed to dry for several hours. Infested plates were then covered with a layer sterile cotton batting and the plate lid, and then held in the dark at 29 ˚C (Lewer et al. 2006). Mortality was recoded at 6 d post-treatment. Each plate had six replicates. The second assay used the yellow fever mosquito (Aedes aegypti: Diptera). One-day old larvae of A. aegypti were pippetted into each well of a 96-well microtiter plate that had been pretreated with 6 mg (dissolved in 15 mL of DMSO-acetone-H₂O mixture, which had been allowed to dry in the well) of the test compound. The microtiter plates, covered with a lid of prevent evaporation, were held 3 d at r.t., and then graded for mortality. There were six replicates per treatment.
  Compounds found to be active were further evaluated a larger format diet-based bioassay using 128-well diet trays (Bio-Serv, Frenchtown, NJ). Three-to-five-second instar larvae of either S. exigua or Helicoverpa zea (corn earworm: Lepidoptera) were placed in each well (3 mL) of the diet tray that had been previously filled with 1 mL of artificial diet to which 25 mg the test compound [dissolved in 50 mL of an acetone-H₂O mixture (90:10)] had been applied (to each of eight wells) and then allowed to dry. Trays were covered with a clear self-adhesive cover, and held at 25 ˚C, 14:10 light/dark for 6 d. Percent mortality was recorded for the larvae in each well; activity in the eight wells was then averaged.

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• 20 Krieger RL. Feeny PP. Wilkinson CF. Science  1971,  172:  579 

15

Demeter, D., log Kow model developed at Dow AgroSciences, unpublished work

17

Representative Examples of 5
4-Methyl-6-(5-phenylisoxazol-3-yl)- N -propylpyrimidin-2-amine
^¹H NMR (300 MHz, CDCl₃): δ = 7.80 (m, 2 H), 7.42 (m,
3 H), 7.10 (s, 1 H), 7.00 (s, 1 H), 3.42 (m, 2 H), 2.31 (s, 3 H), 2.18 (s, 1 H), 1.62 (m, 2 H), 0.93 (m, 3 H). ^¹³C NMR (75 MHz, CDCl₃): δ = 170.9, 162.9, 162.7, 156.3, 130.6, 129.3, 127.5, 126.1, 106.8, 105.8, 98.7, 43.5, 23.3, 23.1, 11.8.
N -(Cyclopropylmethyl)-4-isopropyl-6-(5-phenylisoxazol-3-yl)pyrimidin-2-amine
^¹H NMR (300 MHz, CDCl₃): δ = 7.60 (m, 2 H), 7.24 (m,
3 H), 6.91 (s, 1 H), 6.84 (s, 1 H), 5.22 (br s, 1 H), 3.11 (m,
2 H), 2.64 (m, 1 H), 1.27 (d, J = 7.0 Hz, 6 H), 0.82 (m, 1 H), 0.30 (m, 2 H), 0.05 (m, 2 H). ^¹³C NMR (75 MHz, CDCl₃):
δ = 178.2, 170.9, 163.6, 162.8, 156.2, 130.6, 129.3, 127.5, 126.1, 104.4, 98.7, 46.8, 36.3, 21.9, 11.9, 3.7.
4-Isopropyl-6-(5-phenylisoxazol-3-yl)- N -propyl-pyrimidin-2-amine
^¹H NMR (300 MHz, CDCl₃): δ = 7.87 (m, 2 H), 7.50 (m,
3 H), 7.19 (s, 1 H), 7.11 (s, 1 H), 3.50 (m, 2 H), 2.91 (m, 1 H), 1.70 (m, 2 H), 1.30 (d, J = 7.2 Hz, 6 H), 1.02 (t, J = 6.2 Hz, 3 H); note: NH proton not detected. ^¹³C NMR (75 MHz, CDCl₃): δ = 170.9, 163.8, 162.6, 156.4, 130.6, 130.2, 129.3, 127.5, 126.1, 104.1, 98.7, 43.6, 36.1, 23.1, 21.9, 11.9.
4-[5-(4-Chlorophenyl)isoxazol-3-yl]-6-isopropyl- N -propyl-pyrimidin-2-amine
^¹H NMR (300 MHz, CDCl₃): δ = 7.80 (d, J = 8.7 Hz, 2 H), 7.48 (d, J = 8.7 Hz, 2 H), 7.16 (s, 1 H), 7.09 (s, 1 H), 5.90 (br s, 1 H), 3.48 (br s, 2 H), 2.90 (m, 1 H), 1.70 (m, 2 H), 1.32 (d, J = 7.2 Hz, 6 H), 1.03 (t, J = 7.5 Hz, 3 H). ^¹³C NMR (75 MHz, CDCl₃): δ = 178.2, 170.9, 163.6, 162.8, 156.2, 130.6, 129.3, 127.5, 126.1, 104.4, 98.7, 46.8, 36.3, 21.9, 11.9, 3.7.

18

Representative Examples of 8
N -Cyclopentyl-3-[2-(2-methoxyethylamino)-6-methyl-pyrimidin-4-yl]isoxazole-5-carboxamide
^¹H NMR (300 MHz, CDCl₃): d = 7.42 (s, 1 H), 7.10 (s, 1 H), 6.55 (br s, 1 H), 5.53 (br s, 1 H), 4.41 (m, 1 H), 3.69-3.55 (m, 4 H), 3.38 (s, 3 H), 2.39 (s, 3 H), 2.13-2.04 (m, 2 H), 1.85-1.52 (m, 6 H). ^¹³C NMR (75 MHz, CDCl₃): d = 169.7, 164.3, 163.9, 162.8, 155.5, 154.9, 107.1, 106.2, 71.5, 59.0, 51.9, 41.4, 33.3, 24.4, 24.0.
N -Benzyl-3-[2-(cyclopentylamino)-6-methylpyrimidin-4-yl]isoxazole-5-carboxamide
^¹H NMR (300 MHz, CDCl₃): δ = 7.49 (s, 1 H), 7.35 (m, 5 H), 7.06 (br s, 2 H), 5.22 (d, J = 6.9 Hz, 1 H), 4.64 (d, J = 6.9 Hz, 2 H), 4.33 (m, 1 H), 2.37 (s, 3 H), 2.04 (m, 2 H), 1.72-1.24 (m, 6 H). ^¹³C NMR (75 MHz, CDCl₃): δ = 169.5, 163.9, 162.6, 162.4, 155.9, 154.8, 137.2, 129.1, 129.0, 128.2, 106.7, 106.5, 53.2, 43.8, 33.5, 24.5, 23.9.
N -(4-Methoxybenzyl)-3-[2-(2-methoxyethylamino)-6-methyl-pyrimidin-4-yl]isoxazole-5-carboxamide
^¹H NMR (300 MHz, CDCl₃): δ = 8.46 (br s, 1 H), 7.46 (s,
1 H), 7.30 (d, J = 8.7 Hz, 2 H), 7.09 (s, 1 H), 6.90 (d, J = 8.7 Hz, 2 H), 5.59 (br s, 1 H), 4.59 (d, J = 5.4 Hz, 2 H), 3.80 (s, 3 H), 3.69-3.56 (m, 4 H), 3.38 (s, 3 H), 2.39 (s, 3 H). ^¹³C NMR (75 MHz, CDCl₃): δ = 169.6, 164.0, 163.8, 162.7, 159.6, 155.7, 154.9, 129.7, 129.2, 114.5, 107.1, 106.5, 70.4, 59.0, 55.6, 43.4, 41.4, 24.4.
3-[2-(Cyclopentylamino)-6-isopropylpyrimidin-4-yl]- N -(4-methoxybenzyl)isoxazole-5-carboxamide
^¹H NMR (300 MHz, CDCl₃): δ = 7.48 (s, 1 H), 7.29 (d, J = 7.5 Hz, 2 H), 7.08 (s, 1 H), 6.97 (br s, 1 H), 6.88 (d, J = 8.7 Hz, 2 H), 5.23 (br s, 1 H), 4.58 (d, J = 5.1 Hz, 2 H), 4.30 (m, 1 H), 3.79 (s, 3 H), 2.84 (m, 1 H), 2.06 (m, 2 H), 1.73-1.61 (m, 4 H), 1.45 (m, 2 H), 1.25 (d, J = 5.1 Hz, 6 H). ^¹³C NMR (75 MHz, CDCl₃): δ = 178.2, 164.1, 163.9, 162.5, 159.6, 155.8, 155.0, 129.7, 129.3, 114.5, 106.6, 104.2, 55.6, 53.3, 43.4, 36.3, 33.5, 24.0, 21.8.