Research Article Zangarini, Rajan, Danilenko, Berry, Traversa & Veal
Table 6. Interday precision and accuracy of the method for the analysis of pegcantratinib in human tumor homogenate.
Measured concentration Day 1 Day 2 Day 3
Mean (n = 11) SD
Accuracy (%) Precision (%)
8.00 8.47 7.52 7.18 7.27 8.14 9.02 8.49 8.85 7.88 7.21 7.47 7.95 0.68 99.4 8.50
Linearity & range Examination of linearity over six concentrations of pegcantratinib (range: 2.5–250 ng/ml) yielded a lin- ear correlation of ≥0.995 from three separate experi- ments (Figure 4). The range of the method was exam- ined between 1.0 and 500 ng/ml over one working day. Results are shown in Table 4.
Intra-/interassay precision & accuracy Intra-assay experiments showed precision rang- ing from 3.6 to 7.3% and accuracy of 95.4–99.0% (n = 5). Interassay experiments carried out over three days showed precision ranging from 4.67 to 8.50% and accuracy of 96.1–99.4% (n = 11) (Tables 5 & 6).
Carryover & stability No carryover was detected for pegcantratinib or IS in chromatograms of blank samples injected imme- diately following ULOQ samples. The stability of pegcantratinib in homogenate tumor was assessed by analyzing QC samples (low and high) in triplicate. Pegcantratinib was stable in tumor for at least 4 h
Patient number
1 2
Top layer 321 279
Nominal concentration (ng/ml) 80.0 72.7 69.6 82.1 81.9 75.4 78.2 77.5 76.7 78.2 76.5 77.0 76.9 3.59 96.1 4.67
160 154 151 160 163 164 138 150 154 162 171 177 159
10.74 99.1 6.77
at room temperature and for 10 days at 4°C before and after extraction. Control tumor homogenate, stored at -20°C was shown to be stable for prepara- tion of standards and QCs. Pegcantratinib was stable in tumor homogenate at -20°C over 3 freeze–thaw cycles.
Clinical trial tumor distribution The present method was applied to the quantita- tive analysis of pegcantratinib in human skin tumor biopsies obtained following 4 weeks of daily topical pegcantratinib treatment, from two CYLD carrier patients enrolled in a Phase lb open label clinical trial. The punch biopsies obtained were sliced in three dif- ferent layers to determine drug penetration. Results presented in Table 7 show a clear trend toward higher drug concentration in the top layers, with drug also able to penetrate to the inner layers.
Discussion The TRK inhibitor, pegcantratinib, currently being investigated in early phase clinical trials for the
Table 7. Pegcantratinib tumor concentrations obtained in three tumor layer levels from two CYLD mutation carrier patients treated in a Phase Ib trial.
Tumor concentration (ng/ml) Middle layer 41.1 19.5
Bottom layer 11.0 16.2
286 23
Bioanalysis (2017) 9(3)
future science group
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