M.T. Gabr, M.S. Abdel-Raziq
PhytochemistryLetters27(2018)203–207
3.1.2. General procedure for compounds 5-10
(m, 1 H), 2.06–2.10 (m, 2 H), 2.28–2.35 (m, 2 H), 2.61–2.71 (m, 3 H),
3.52–3.56 (m, 2 H), 7.76–7.79 (m, 1 H), 7.85–7.88 (m, 1 H), 8.04 (d,
J = 4.5 Hz, 1 H), 8.15 (d, J = 8.1 Hz, 1 H), 8.95–9.98 (m, 1 H), 9.23 (d,
J = 4.5 Hz, 1 H). 13C NMR (100 MHz, DMSO-d6) δ 24.1, 24.7, 25.0,
25.1, 26.5, 29.7, 32.7, 34.0, 41.5, 43.5, 55.9, 60.6, 62.4, 64.2, 71.1,
123.2, 124.1, 125.8, 129.0, 129.5, 130.0, 136.3, 148.4, 151.1. HRMS
(ESI): calcd for C24H32N3 [M+H]+, 362.2596; found, 362.2599.
Vaska’s catalyst (1 mol%) and compound 4 (0.3 mmol, 1.0 eq) were
charged into a dry 25 mL flask. Addition of dry DCM (3 mL) was fol-
lowed by TMDS (0.6 mmol, 2.0 eq) while stirring at room temperature.
The reaction mixture was stirred for 15 min, then cooled to −78 °C.
Grignard reagent (2.0 eq) was added dropwise. The solution was stirred
at −78 °C for 10 min, then warmed to room temperature and stirred for
4 h. The reaction mixture was quenched with saturated aqueous NH4Cl
solution and extracted with DCM (2 × 10 mL). The combined organic
layers were dried over sodium sulfate, filtered and the solvent was re-
moved under vacuum. The crude material was further purified by flash
column chromatography using 5% methanol in DCM to furnish com-
pounds 5–10.
3.2. Antiproliferative activity
The in vitro cytotoxicity testing of compounds 5–10 was performed
against four cancer cell lines and mouse embryo fibroblast cells
adopting MTT assay as previously described after 48 h treatment with
3.1.3. 17-Phenylsparteine (5)
Mp 101–103 °C. Yield 61%.1H NMR (400 MHz, DMSO-d6)
δ
3.3. Autophagic protein degradation assay
1.11–1.17 (m, 3 H), 1.37–1.59 (m, 9 H), 1.76–1.82 (m, 3 H), 1.93–1.96
(m, 1 H), 2.09–2.12 (m, 2 H), 2.31–2.33 (m, 2 H), 2.63–2.74 (m, 3 H),
3.32–3.35 (m, 2 H), 7.15–7.21 (m, 3 H), 7.38–7.41 (m, 2 H). 13C NMR
(100 MHz, DMSO-d6) δ 25.9, 26.7, 26.8, 26.9, 28.4, 32.1, 35.1, 35.9,
44.2, 45.9, 58.1, 63.4, 65.1, 66.2, 73.1, 124.3, 125.2, 129.1, 139.8.
HRMS (ESI): calcd for C21H31N2 [M+H]+, 311.2487; found, 311.2492.
The effect of 10 (10 mM) on the degradation of [14C]-valine-labeled
long-lived proteins in Hela cells was performed as previously described
ladenine (10 mM) as positive control.
3.4. Intracellular ATP assay
3.1.4. 17-(4′-Methoxyphenyl)sparteine (6)
Mp 115–116 °C. Yield 59%.1H NMR (400 MHz, DMSO-d6)
δ
The intracellular ATP level in Hela cells was determined using ATP
assay kit purchased from abcam (ab83355). The assay was performed
according to the protocol provided.
1.15–1.23 (m, 3 H), 1.41–1.67 (m, 9 H), 1.78–1.86 (m, 3 H), 1.93–1.96
(m, 1 H), 2.05–2.10 (m, 2 H), 2.27–2.34 (m, 2 H), 2.59–2.69 (m, 3 H),
3.31–3.34 (m, 2 H), 3.68 (s, 3 H), 7.01 (d, J = 7.6 Hz, 2 H), 7.25 (d,
J = 7.6 Hz, 2 H).13C NMR (100 MHz, DMSO-d6) δ 24.9, 25.9, 26.1,
26.6, 27.8, 30.7, 33.5, 36.1, 44.2, 44.9, 56.1, 60.2, 61.8, 63.7, 64.9,
72.1, 111.6, 129.1, 139.6, 156.1. HRMS (ESI): calcd for C22H33N2O [M
+H]+, 341.2592; found, 341.2598.
4. Conclusion
In a summary, arylsparteine derivatives were synthesized and
evaluated for their antiproliferative activity. Incorporation of electron
withdrawing groups (EWGs) to the aryl substituents enhanced the an-
tiproliferative activity of the arylsparteine derivatives. Compound 10
bearing 4-quinolinyl moiety as the aryl substituent demonstrated pro-
mising antiproliferative activity against A549 and Hela cancer cell
lines. The antiproliferative activity of 10 is directly related to the
blockade of autophagic flux in cancer cells as revealed by monitoring
autophagic protein degradation and intracellular ATP level in Hela
cells.
3.1.5. 17-(3′,4′,5′-Trimethoxyphenyl)sparteine (7)
Mp 97–99 °C. Yield 51%.1H NMR (400 MHz, DMSO-d6) δ 1.10–1.18
(m, 3 H), 1.38–1.62 (m, 9 H), 1.75–1.83 (m, 3 H), 1.92–1.95 (m, 1 H),
2.08–2.13 (m, 2 H), 2.30–2.37 (m, 2 H), 2.63–2.75 (m, 3 H), 3.45–3.49
(m, 2 H), 3.75 (s, 3 H), 3.85 (s, 6 H), 7.25 (s, 2 H). 13C NMR (100 MHz,
DMSO-d6) δ 25.7, 26.2, 26.4, 26.7, 28.1, 31.3, 34.3, 35.6, 43.1, 45.0,
57.5, 57.6, 61.6, 62.2, 64.0, 65.8, 70.2, 106.2, 133.2, 144.3, 154.8.
HRMS (ESI): calcd for
401.2801.
C , 401.2804; found,
24H37N2O3 [M+H]+
Appendix A. Supplementary data
3.1.6. 17-(4′-Chlorophenyl)sparteine (8)
Mp 123–125 °C. Yield 72%.1H NMR (400 MHz, DMSO-d6)
Supplementary material related to this article can be found, in the
δ
1.08–1.14 (m, 3 H), 1.30–1.51 (m, 9 H), 1.71–1.76 (m, 3 H), 1.88–1.91
(m, 1 H), 2.01–2.06 (m, 2 H), 2.27–2.30 (m, 2 H), 2.59–2.65 (m, 3 H),
3.57–3.60 (m, 2 H), 7.89 (d, J = 6.9 Hz, 2 H), 7.97 (d, J = 6.9 Hz, 2 H).
13C NMR (100 MHz, DMSO-d6) δ 24.1, 25.8, 25.9, 26.0, 27.5, 30.6,
33.8, 35.0, 44.2, 45.5, 56.8, 63.1, 65.1, 65.9, 71.2, 127.9, 128.8, 139.1,
141.2. HRMS (ESI): calcd for C21H30ClN2 [M+H]+, 345.2097; found,
345.2092.
References
Amaravadi, R.K., Yu, D., Lum, J.J., Bui, T., Christophorou, M.A., Evan, G.I., et al., 2007.
Bao, Y., Kong, X., Yang, L., Liu, R., Shi, Z., Li, W., Hua, B., Hou, W., 2014. Complementary
Bi, C., Zhang, C., Li, Y., Tang, S., Deng, H., Zhao, W., Wang, Z., Shao, R., Song, D., 2014.
Bi, C., Ye, C., Li, Y., Zhao, W., Shao, R., Song, D., 2016. Synthesis and biological eva-
Bi, C., Zhang, N., Yang, P., Ye, C., Wang, Y., Fan, T., Shao, R., Deng, H., Song, D., 2017.
3.1.7. 17-(4′-Nitrophenyl)sparteine (9)
Mp 108–110 °C. Yield 55%.1H NMR (400 MHz, DMSO-d6)
δ
1.11–1.17 (m, 3 H), 1.37–1.59 (m, 9 H), 1.76–1.82 (m, 3 H), 1.93–1.96
(m, 1 H), 2.09–2.12 (m, 2 H), 2.31–2.33 (m, 2 H), 2.63–2.74 (m, 3 H),
3.53–3.56 (m, 2 H), 8.16 (d, J = 7.4 Hz, 2 H), 8.40 (d, J = 7.4 Hz, 2 H).
13C NMR (100 MHz, DMSO-d6) δ 25.7, 26.2, 26.5, 26.7, 28.1, 31.3,
34.3, 35.6, 43.1, 45.0, 57.5, 62.2, 64.0, 65.8, 70.4, 125.7, 132.1, 141.5,
144.3. HRMS (ESI): calcd for C21H30N3O2 [M+H]+, 356.2338; found,
356.2343.
3.1.8. 17-(4′-Quinolinyl)sparteine (10)
Mp 137–139 °C. Yield 68%.1H NMR (400 MHz, DMSO-d6)
δ
1.07–1.14 (m, 3 H), 1.34–1.60 (m, 9 H), 1.74–1.79 (m, 3 H), 1.91–1.94
206