Chk1 (Checkpoint kinase 1), an important serine/threonine kinase, is responding to DNA damage and stall DNA replication. Chk1 plays a key role in maintaining the replication fork viability during exposure to DNA antimetabolites. Inhibition of Chk1 will lead to accumulation of double-strand DNA breaks and cell death in human tumor cell lines exposure to antimetabolite. LY2603618 (Rabusertib) is a selective inhibitor of Chk1 with IC50 value of 7nM (measured by protein kinase assays). Treatment with 14-3300nM LY2603618 for 2h can dose-dependently decrease the level of phosphorylated Chk1 serine 296, an autophosphorylation reflecting Chk1 catalytic activity, in HeLa cells incubated with 100nM doxorubicin for 24h. Similar result can also be obtained in Calu6 Cells treated with 60nM gemcitabine. After treatment with LY2603618 1250 or 5000nM for 24h, increased level of p-H2A.X-S139 and p-H3-S10, the markers of DNA damage and mitotic, can be observed in HeLa cells. Meanwhile, a clear decrease in the G1 population and an increase in late S-phase cells, as well as cell arrest in an aberrant prometaphase state by LY2603618 can be observed. These indicated that inhibition of Chk1 by LY2603618 can abrogate the S-phase checkpoint. An increase in staining of the mitotic marker p-H3-Ser10 can be observed in HeLa cells treated for 24h with 100nM doxorubicin to cause a G2 arrest, then for 7h with serially diluted LY2603618. This indicated that LY2603618 can inactivate the G2/M DNA damage checkpoint[1]. Combining 200mg/kg of LY2603618 with 160mg/kg of gemcitabine increased the tumor growth delay compared with gemcitabine alone in Calu-6 and HT-29 xenograft tumor models[2]. Up to now, phase 2 studies of LY2603618 treatment for non small cell lung cancer and pancreatic neoplasms have been completed (see https://www.clinicaltrials.gov/).
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