Abstract
BackgroundThe high expression of BLM (Bloom syndrome) helicase in tumors involves its strong association with cell expansion. Bisbenzylisoquinoline alkaloids own an antitumor property and have developed as candidates for anticancer drugs. This paper aimed to screen potential antiproliferative small molecules from 12 small molecules (the derivatives of bisbenzylisoquinoline alkaloids tetrandrine and fangchinoline) by targeting BLM642–1290 helicase. Then we explore the inhibitory mechanism of those small molecules on proliferation of MDA-MB-435 breast cancer cells.MethodsFluorescence polarization technique was used to screen small molecules which inhibited the DNA binding and unwinding of BLM642–1290 helicase. The effects of positive small molecules on the ATPase and conformation of BLM642–1290 helicase were studied by the malachite green-phosphate ammonium molybdate colorimetry and ultraviolet spectral scanning, respectively. The effects of positive small molecules on growth of MDA-MB-435 cells were studied by MTT method, colony formation and cell counting method. The mRNA and protein levels of BLM helicase in the MDA-MB-435 cells after positive small molecule treatments were examined by RT-PCR and ELISA, respectively.ResultsThe compound HJNO (a tetrandrine derivative) was screened out which inhibited the DNA binding, unwinding and ATPase of BLM642–1290 helicase. That HJNO could bind BLM642–1290helicase to change its conformationcontribute to inhibiting the DNA binding, ATPase and DNA unwinding of BLM642–1290 helicase. In addition, HJNO showed its inhibiting the growth of MDA-MB-435 cells. The values of IC50 after drug treatments for 24 h, 48 h and 72 h were 19.9 μmol/L, 4.1 μmol/L and 10.9 μmol/L, respectively. The mRNA and protein levels of BLM helicase in MDA-MB-435 cells increased after HJNO treatment. Those showed a significant difference (P < 0.05) compared with negative control when the concentrations of HJNO were 5 μmol/L and 10 μmol/L, which might contribute to HJNO inhibiting the DNA binding, ATPase and DNA unwinding of BLM helicase.ConclusionThe small molecule HJNO was screened out by targeting BLM642–1290 helicase. And it showed an inhibition on MDA-MB-435 breast cancer cells expansion.
Highlights
The high expression of Bloom syndrome helicase (BLM) (Bloom syndrome) helicase in tumors involves its strong association with cell expansion
Screening out small molecules with inhibiting BLM642–1290 helicase from 12 derivatives of tetrandrine and fangchinoline When concentration of small molecules was 6.67 μmol/ L, among 12 derivatives of tetrandrine and fangchinoline, the inhibiting values of HL-22, A tetrandrine derivative (HJNO), HL-6, HL-27 and HY-2 on BLM642–1290 helicase binding to dsDNA were 14, 19, 30, 47 and 65, respectively (Fig. 1)
MMC had no significant effect on BLM helicase binding to dsDNA and a weak inhibiting effect on BLM helicase binding to ssDNA (21 nt) with the Ki value of 3.62 ± 0.84 μmol/L (Fig. 2d)
Summary
The high expression of BLM (Bloom syndrome) helicase in tumors involves its strong association with cell expansion. As one of the biggest public health problems around the world, malignant tumors do great harm to human health and will become the first killer of human in the new century [1] Conventional cancer treatments such as radiotherapy and chemotherapy cause great damage to normal cells as well as human themselves. The lack of three coding genes BLM, WRN and RecQ4 leads to occur related diseases, which are Bloom syndrome (BS), Werner syndrome (WS) and Rothmund-Thomson syndrome (RTS) [3,4,5], respectively. The patients of these diseases are commonly susceptible to cancer [6]
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