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    국제 암학술대회 발표 (Poster, 2007.10)
    CONFERENCE  komipharm 2014.03.18 15:54

KML001 targets telomeres in prostatecancer cells

19thEORTC-NCI-AACR San Francisco, CA  


Abstract

 KML001 (sodium meta arseniteKominox) is an orally bioavailable, trivalent arsenic compound that has entered  phase I/II

clinical trials in prostate cancer. Arsenicals have been described to cause chromosome abnormalities as a result of the

generation of reactive oxygen species (ROS) in cells. Recent studies however, have shown that  trivalent  arsenicals can

specifically alter telomere length, telomerase activity and telomere binding proteins, suggesting that the telomere

/telomerase complex might be a direct target of sodium meta arsenite.  This study was initiated to investigate  whether

KML001 targets telomeres and telomerase in prostate cancer cells.  The prostate cancer cell lines PC3 and  DU145, which

have relatively short telomeres (2.5 and 4.2 kb, respectively)  were chosen for in vitro experiments.  KML001 concentrations

needed to inhibit cell growth to 50% (IC50) and 100% (IC100) were determined by MTT assay.  The IC50 for PC3 was found

to be 0.23 µM and for DU145 2 µM, the IC100s were 9 µM and 5 µM, respectively; the  IC50s and IC100s were used for all

mechanistic analyses. The  TeloTAGGG Quantification Kit was employed to measure mRNA expression of the telomerase

catalytic subunit  hTERThTERT protein expression, induction of  telomere-associated DNA damage by histone H2AX 

phosphorylation, and 8-oxodeoxyguanosine formation were  studied by immunofluorescence microscopy. Direct  binding of

KML001 to 5'-(TTAGGG)x-3' telomericsequences was assessed by high resolution MALDI (Matrix-assisted Laser

Desorption Ionization) mass spectrometry, and by metaphase fluorescence in situhybridization (FISH).  KML001 treatment

of prostate cancer cells did down-regulate  hTERT mRNA expression after 24 hrs of continuous  exposure to drug. hTERT

protein expression in the nucleus was also reduced and a translocation to the cytoplasm  was observed, suggesting the

displacement of hTERT from the  telomeres. Consistent with these observations, we  found that KML001 can bind to

telomeric repeat sequences at a ratio of 1 molecule drug per 3 telomeric repeats and  that these lesions result in a rapid

induction of γ-H2AX, a marker of DNA-damage response and telomere-initiated  senescence. γ-H2AX phosphorylation (γ-H2AX)
occurred earlier (after 2 hrs) and at lower 
KML001 (IC50)  concentrations compared to the ROS-induced formation of
8- 
oxodeoxyguanosine, which was not seen prior to 24 hrs and only at the IC100. Moreover, direct telomere binding by

KML001 resulted in a gradual telomere loss with two telomeres per chromosome missing at 48 hrs and a complete telomere

loss seen at 72 hrs in IC100 treated PC3 cells. Our findings indicate that KML001 can specifically target telomeres and
telomerase. The latter might provide  biomarkers to assess drug response and should be considered in clinical
trials of KML001 in prostate and other cancers