Determination of FCN-437c in human plasma for patients with advanced breast cancer by HPLC-MS/MS and its clinical application
Objective:To establish an HPLC-MS/MS method for the determination of FCN-437c in human plasma and its application to the phase Ⅰ clinical study of FCN-437c.Methods:Following protein precipitation,plasma was injected and measured using HPLC-MS/MS method.The analytes were separated on a YMC Triart PFP column(50 mm × 2.1 mm,5 μm)using 0.5％formic acid(containing 5 mmol·L-1 of ammonium acetate,A)and acetonitrile(B)as the mobile phase with gradient elution at the flow rate of 0.5 mL·min-1,the column temperature was set at 35 ℃,the injection amount was 2 µL,and the injector temperature was 10 ℃.MS detec-tion was performed with multiple reaction monitoring(MRM)mode using positive electrospray ionization.The ion transitions were m/z 549.4→449.4 for FCN-437c and m/z 552.3→449.3 for FCN-437-D3,respectively.Other mass spectrometry parameters were TEM,500 ℃,GS1,276 kPa,GS2,207 kPa,DP,100 V,CXP,25 V.Results:The linear range of FCN-437c in human plasma was 5-1 000 ng·mL-1(r=0.999 0).The lower limit of quantification was 5 ng·mL-1.The intra-batch and inter-batch precisions were less than 2.0％and 4.1％,respectively.The average recovery was 104.0％(FCN-437c),78.6％(FCN-437-D3),and the in-ternal standard normalized matrix factor was 100％-102％.The stock solution of FCN-437c was stable at 4 ℃for 202 d,the working solution of FCN-437 c and internal standard were stable at room temperature for 24 h.FCN-437c in human plasma was investigated to be stable at room temperature for 20 h,four cycles of freeze-thaw,-20 ℃ for 134 d,-80 ℃ for 662 d,as well as for 24 h in the autosampler after treatment.The whole blood samples were stable at room temperature for 4 h.This method was applied to the determination FCN-437c in human plasma,and the deviation between the test results and the initial values of 97.0％ISR samples was within±20％.The accuracy was 102.0％-108.0％after 10-fold dilution of plasma samples.The cumulative ratio of RAUC0-24 and RCmax was 1.33 times and 1.59 times when FCN-437c was administered continuously com-pared with single administration.Conclusion:This method is simple,accurate,robust and specific,which can meet the requirements of quantitative analysis of FCN-437c in human plasma and also can be used to determine FCN-473c in human plasma of the phase Ⅰ clinical study.
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