Tài liệu Báo cáo khoa học: Mammalian mitotic centromere-associated kinesin (MCAK) A new molecular

Mammalian mitotic centromere-associated kinesin (MCAK)

A new molecular target of sulfoquinovosylacylglycerols novel

antitumor and immunosuppressive agents

Satoko Aoki1,2, Keisuke Ohta1

, Takayuki Yamazaki1

, Fumio Sugawara1,2 and Kengo Sakaguchi1,2

1 Department of Applied Biological Science, Tokyo University of Science, Noda, Chiba, Japan

2 Genome and Drug Research Center, Tokyo University of Science, Noda, Chiba, Japan

Several kinds of synthetic sulfoquinovosylacylglycerols

(SQAGs) may be potent and clinically promising

agents for cancer chemotherapy and immunosuppres￾sion [1,2]. However, the molecular targets of SQAGs

are ambiguous. The aim of the present study was to

identify molecular targets that could be of significance.

Earlier, effects of SQAGs were found and reported

independently by two laboratories screening directly

for antitumor agents in vivo [1] and for mammalian

DNA polymerase inhibitors in vitro [3–5]. One mole￾cular target of SQAGs is thus DNA polymerases [6]

but there is a strong evidence for other targets [7–9].

SQAGs are found as natural compounds in higher

plants [3], sea algae [4,5] and sea urchins [1]. They

have been reported to have a wide range of bioactivi￾ties: antiviral activity against human immunodeficiency

virus (HIV-1) [7], P-selectin receptor inhibition [8],

antitumor activity [1], tumor cell growth inhibition [6]

and immunosuppressive activity [2]. Sahara et al.

showed that SQAGs effectively inhibit the growth of

implanted human lung adenocarcinoma cells, A549, in

nude mice [1]. Moreover, Ohta et al. reported that a

wide variety of cultured tumor cells were sensitive to

SQAGs [6]. It is very difficult to collect and purify

SQAGs from natural sources but we have succeeded in

the chemical synthesis of a number of forms. We have

found a-anomers that possess potent antitumor activ￾ity but do not have many of the serious side-effects of

Keywords

mammalian DNA polymerases; MT

depolymerization activity of MCAK; SQAGs;

T7 phage display method

Correspondence

K. Sakaguchi, Department of Applied

Biological Science, Tokyo University of

Science, 2641 Yamazaki, Noda, Chiba 278–

8510, Japan

Fax: +81 4 7123 9767

Tel: +81 4 7124 1501 (ext. 3409)

E-mail: kengo@rs.noda.tus.ac.jp

(Received 30 October 2004, revised 20

January 2005, accepted 7 February 2005)

doi:10.1111/j.1742-4658.2005.04600.x

Sulfoquinovosylacylglycerols (SQAGs), in particular compounds with C18

fatty acid(s) on the glycerol moiety, may be clinically promising antitumor

and ⁄ or immunosuppressive agents. They were found originally as inhibitors

of mammalian DNA polymerases. However, SQAGs can arrest cultured

mammalian cells not only at S phase but also at M phase, suggesting they

have several molecular targets. A screen for candidate target molecules

using a T7 phage display method identified an amino acid sequence. An

homology search showed this to be a mammalian mitotic centromere-asso￾ciated kinesin (MCAK), rather than a DNA polymerase. Analyses showed

SQAGs bound to recombinant MCAK with a KD ¼ 3.1 · 10)4 to

6.2 · 10)5 m. An in vivo microtubule depolymerization assay, using EGFP￾full length MCAK fusion constructs, indicated inhibition of the micro￾tubule depolymerization activity of MCAK. From these results, we

conclude that clinically promising SQAGs have at least two different

molecular targets, DNA polymerases and MCAK. It should be stressed

that inhibitors of MCAK have never been reported previously so that there

is a major potential for clinical utility.

Abbreviations

a-SQDG(18:0), saturated 1,2-O-diacyl-3-O-(a-D-sulfoquinovosyl)-glyceride; b-SQDG(18:0), saturated 1,2-O-diacyl-3-O-(b-D-sulfoquinovosyl)-

glyceride; a-SQMG(18:0), saturated 1-O-monoacyl-3-O-(a-D-sulfoquinovosyl)-glyceride; a-SQMG(18:1), unsaturated 1-O-monoacyl-3-O-(a-D￾sulfoquinovosyl)-glyceride; DMSO, dimethylsulfoxide; EGFP, enhanced green fluorescent protein; MCAK, mitotic centromere-associated

kinesin; MTs, microtubules; MCAK184, His6-tagged MCAK truncated form (P184-G593); SQAGs, sulfoquinovosylacylglycerols.

2132 FEBS Journal 272 (2005) 2132–2140 ª 2005 FEBS