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ether, and the layers were separated. The ether layer was washed
with saturated NaHCO3 (3 ꢁ 25 mL). Combined aqueous layers
were extracted with CH2Cl2 (1 ꢁ 25 mL) and the combined organic
layers dried over anhydrous Na2SO4, filtered, and concentrated to
give the crude aldehyde as a yellow oil. The aldehyde was purified
by flash chromatography (silica gel, 80% hexane, 15% EtOAc, 5%
CH2Cl2) to give aldehyde 5l as a pale-yellow oil which was used
immediately in the next step: 1H NMR (300 MHz, CDCl3) d 1.47
(d, 3H, lateral isoxazole C-50 Me) 2.29 (s, 3H, isoxazole C-30 Me)
3.39–3.42 (m, 2H, CH2) 3.76–3.82 (m, 1H, CH) 7.01 (d, 1H, Ar-H)
7.20–7.29 (t, 1H, Ar-H) 7.42–7.47 (m, 2H, Ar-H) 7.67 (d, 1H, Ar-
H) 7.78 (d, 1H, Ar-H) 7.86 (d, 1H, Ar-H) 9.17 (s, 1H, aldehyde H).
The aldehyde 5l 0.37 g (1.33 mmol) was dissolved in ethanol
(10 mL) and transferred to an aerosol dispersion tube to which ethyl
acetoacetate (357 mg, 2.75 mmol) and aqueous ammonia (2 mL,
29.6%) were added. The mixture was heated to 100–110 °C for
48 h, the pressure rising to 30–40 psi. After cooling, the solvents
were removed under vacuum to give a brown oil. The crude product
was chromatographed on silica gel (CHCl3) and purified by radial
chromatography (40% hexane, 20% EtOAc, 10% CH2Cl2), giving
100 mg of 1l as white crystals (15%): mp 144–146 °C; 1H NMR
(500 MHz, CDCl3) d 1.11–1.14 (t, 3H, CH3CH2O–) 1.17–1.19 (t, 3H,
CH3CH2O–) 1.20 (d, 3H, lateral isoxazole C-50 Me) 1.59 (s, 3H, DHP-
Me) 2.18 (s, 3H, DHP-Me) 2.35 (s, 3H, isoxazole C-30 Me) 3.30–3.35
(m, 2H, lateral isoxazole C-50 CH and diastereotopic CH) 3.53–3.56
(m, 1H, other diastereotopic CH) 3.95–4.00 (m, 2H, CH3CH2O–)
4.05–4.13 (m, 2H, CH3CH2O–) 4.76 (s, 1H, DHP-CH) 5.25 (br s, 1H,
NH) 7.15 (d, 1H, Ar-HG) 7.27–7.30 (t, 1H, Ar-HF) 7.43–7.52 (m, 2H,
Ar-HE, HD) 7.68 (d, 1H, Ar-HC) 7.85 (d, 1H, Ar-HB) 8.03 (d, 1H, Ar-
HA); 13C NMR (125 MHz, CDCl3) d 10.1 , 14.32, 14.38, 19.1, 19.5,
19.9, 28.9, 33.0, 37.0, 59.7, 59.8, 102.1, 102.4, 120.7, 123.5, 125.2,
125.4, 125.8, 126.8, 126.9, 128.9, 131.9, 134.0, 136.0, 142.3, 142.8,
159.8, 166.8, 167.3, 172.7; MS (EI) 502. Anal. Calcd for C30H34N2O5:
C, 71.69; H, 6.82; N, 5.57. Found: C, 71.54; H, 6.63; N, 5.30.
drous Na2SO4, filtered, and concentrated to give an orange oil.
The crude product was hydrolyzed with 20 mL of 1 M aqueous
HCl in 27 mL of 4:1 THF/H2O. The reaction mixture was poured into
50 mL of ether, and the layers were separated. The ether layer was
washed with saturated NaHCO3 (3 ꢁ 25 mL). Combined aqueous
layers were extracted with CH2Cl2 (1 x 25 mL) and the combined
organic layers dried over anhydrous Na2SO4, filtered, and concen-
trated to give the crude aldehyde 5k as a yellow oil. The aldehyde
5k was purified by flash chromatography (silica gel, 80% hexane,
15% ETOAc, 5% CH2Cl2) to give 1.96 g as a pale-yellow oil (58 %):
1H NMR (200 MHz, CDCl3) d 1.40 (d, 3H, lateral isoxazole C-50
Me) 2.41 (s, 3H, isoxazole C-30 Me) 2.85–3.07 (m, 2H, CH2)
3.61–3.72 (q, 1H, CH) 6.97 (d, 1H, Ar-H) 7.06–7.13 (t, 1H, Ar-H)
7.22–7.33 (m, 1H, Ar-H) 9.66 (s, 1H, aldehyde H).
The aldehyde 5k 1.96 g (6.36 mmol) was dissolved in ethanol
(17 mL) and transferred to an aerosol dispersion tube to which ethyl
acetoacetate (1.74 g, 13.3 mmol) and aqueous ammonia (1.5 mL,
29.6%) were added. The mixture was heated to 100–110 °C for
48 h, the pressure rising to 30–40 psi. After cooling, the solvents
were removed under vacuum to give a brown oil. The crude product
was chromatographed on silica gel (70% hexane, 30% ETOAc) and
purified by radial chromatography (40% hexane, 20% EtOAc, 10%
CH2Cl2), giving 600 mg of 1k as white crystals (18%): mp 134–
136 °C; 1H NMR (500 MHz, CDCl3)
d
1.13–1.21 (m, 9H, 2
CH3CH2O–, lateral isoxazole C-50 Me) 2.19 (s, 3H, DHP-Me) 2.23 (s,
3H, DHP-Me) 2.33 (s, 3H isoxazole C-30 Me) 2.67–2.72 (q, 1H, lateral
isoxazole C-50 CH) 3.03–3.16 (m, 2H, CH2) 4.00–4.14 (m, 4H, 2
CH3CH2O–) 4.79 (s, 1H, DHP-CH) 5.48 (br s, 1H, NH) 6.92 (d, 1H,
Ar-H) 7.04–7.07 (t, 1H, Ar-H) 7.24 (t, 1H, Ar-H) 7.27–7.30 (m, 1H,
Ar-H); 13C NMR (125 MHz, CDCl3) d 10.12, 14.37, 14.39, 19.0, 19.6,
19.7, 28.9, 33.8, 40.5, 59.8, 59.9, 102.3, 102.6, 120.7, 122.2, 127.5,
129.1, 129.8, 131.6, 142.2, 142.6, 142.7, 159.7, 166.8, 167.2, 172.1;
MS (EI) 528, 530. Anal. Calcd for C26H31N2O5Br: C, 58.76; H, 5.88;
N, 5.27. Found: C, 58.79; H, 5.96; N, 5.31.
6.3. Diethyl 2,6-dimethyl-4-[5-(RS-10-m-bromophenyl-prop-20-
yl)-3-methylisoxazol-4-yl]-1,4-dihydropyridine-3,5-dicarboxyl
ate (1k)
6.4. Pharmacophore computational modeling
Ligand structures were drawn and energy minimized (Powell
method, 0.01 kcal/mol⁄A gradient termination, MMFF94s force
field, MMFF94 charges, 1000 maximum iterations) using the Sybyl
modeling program (Tripos, St. Louis, MO, USA). Construction of the
overlaying pharmacophore was achieved by assembling the energy
minimized structures and merging the collection of structures into
the same field. This was followed by energy minimization, molec-
ular dynamics, and an energy minimization simulation. Aggregates
for molecular dynamics and minimization simulations were de-
fined as Carbons 2,3,4,5 and 6 of the 1,4-dihydropyridine ring. All
ligands were then energy minimized to allow for the lowest energy
confirmation of each ligand.
To a stirred solution of ethyl isoxazolyl-oxazoline (3) from
above (2.69 g, 12.9 mmol) in 100 mL dry THF cooled to ꢀ78 °C un-
der N2 was added 1.1 equiv of 2.45 M n-BuLi (5.8 mL) dropwise.
The yellow solution was stirred at ꢀ78 °C for 2 h. Then m-Br-ben-
zylbromide (3.41 g, 13.7 mmol) dissolved in 10 mL dry THF was
added dropwise and the reaction allowed to come to room temper-
ature overnight. The mixture was concentrated under vacuum to
give a brown oil which was purified by flash chromatography (sil-
ica gel, 80% hexane, 20% EtOAc) to give 4.30 g of branched isoxaz-
olyl-oxazoline 4k as a yellow oil (88%): 1H NMR (200 MHz, CDCl3) d
1.27–1.31 (m, 9H, oxazoline Me’s, lateral isoxazole C-50 Me); 2.38
(s, 3H, isoxazole C-30 Me) 2.69–2.80 (m, 1H, diastereotopic CH)
and 2.96–3.06 (q, 1H, other diastereoptopic CH) 3.84–3.95 (m,
3H, oxazoline CH2 and lateral isoxazole C-50 CH) 6.97–7.11 (m,
2H, Ar-H) 7.26–7.34 (m, 1H, Ar-H); MS (EI) 376, 378.
To a stirred solution of the product 4k (4.52 g, 12.0 mmol) in
100 mL of dry CH2Cl2 was added 2.0 mL (2.90 g, 17.7 mmol) of dis-
tilled CF3SO3CH3, and the mixture stirred under N2 until TLC (silica,
80% hexane, 20% EtOAc) showed only baseline material. The mix-
ture was cooled to 0ꢀC, and a solution of 0.82 g (21.7 mmol) of
NaBH4 in 30 mL of 4:1 THF/MeOH was added in one portion. This
mixture was stirred at 0 °C for 30 min. Then 7 mL of saturated
NH4Cl was added and the mixture allowed to warm to room tem-
perature. Ether, 50 mL, was added, and the layers were separated.
The ether layer was washed with saturated NaCl (1 ꢁ 25 mL). The
combined aqueous layers were extracted with CH2Cl2
(1 ꢁ 25 mL). The combined organic layers were dried over anhy-
Acknowledgment
The authors thank the NIH for grants GM42029, NS038444, and
P20RR015583.
MDR1 data was generously provided by the National Institute
of Mental Health’s Psychoactive Drug Screening Program (NIMH
PDSP), Contract # HHSN-271-2008-00025-C (NIMH PDSP). The
NIMH PDSP is Directed by Bryan L. Roth MD, PhD at the University
of North Carolina at Chapel Hill and Project Officer Jamie Driscoll at
NIMH, Bethesda MD, USA.
Supplementary data
Supplementary data associated with this article can be found, in