Cell cycle perturbation induced by gemcitabine in human tumor cells in cell culture, xenografts and bladder cancer patients: implications for clinical trial designs combining gemcitabine with a Chk1 inhibitor
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Ryan Montano1, Nadeem Khan2,4, Huagang Hou2,4, John Seigne3,4, Marc S. Ernstoff3,4,5, Lionel D. Lewis3,4 and Alan Eastman1,4
1Department of Molecular and Systems Biology, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA
2Department of Radiology, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA
3Department of Medicine, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA
4Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Lebanon, NH, USA
5Department of Medicine, Roswell Park Cancer Institute, Buffalo, NY, USA
Alan Eastman, email: Alan.R.Eastman@Dartmouth.edu
Keywords: gemcitabine, S phase arrest, cell cycle checkpoint, Chk1, bladder cancer
Received: May 07, 2017 Accepted: June 03, 2017 Published: June 28, 2017
Gemcitabine irreversibly inhibits ribonucleotide reductase and induces S phase arrest but whether this occurs in tumors in mice or patients has not been established. Tumor cells in culture were incubated with gemcitabine for 6 h to approximate the administration schedule in a patient. Concentrations that induced persistent S phase arrest thereafter correlated with cell killing. Administration of gemcitabine to mice also demonstrated a persistent S phase arrest in their tumor. The minimum dose that induced almost complete S phase arrest after 24 h (40 mg/kg) was well below the maximum tolerated dose in mice. S phase arrest was also observed in tumors of bladder cancer patients receiving gemcitabine. The Chk1 inhibitor MK-8776 sensitized cells to gemcitabine with the greatest cell killing when added 18 h after gemcitabine. In mice, the administration of MK-8776 18 h after gemcitabine elicited positivity for the DNA damage marker γH2AX; this also occurred at relatively low dose (40 mg/kg) gemcitabine. Hence, in both cell culture and xenografts, MK-8776 can markedly enhance cell killing of cells reversibly arrested in S phase by gemcitabine. Some cell lines are hypersensitive to MK-8776 as monotherapy, but this was not observed in xenograft models. Effective monotherapy requires a higher dose of Chk1 inhibitor, and target inhibition over a longer time period as compared to its use in combination. These results have important implications for combining Chk1 inhibitors with gemcitabine and suggest that Chk1 inhibitors with increased bioavailability may have improved efficacy both in combination and as monotherapy.
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