Fig. 3: TKD shows a typical kinetics of protein drug metabolism and little side effects.

a Protein stability analysis performed with 1 μg/μL TKD using Uncle (Unchained Labs, USA). Tm, denaturation temperature; BCM, centroid wavelengths of protein endogenous fluorescence emission spectra. b 0.1 μg purified TKD was mixed with 27 μL of human sera and incubated at 37 °C for different durations as indicated, the degradation of TKD was evaluated by western blotting assays, and the degradation curve was drawn using nonlinear regression based on the gray value. Input, purified TKD; F, sera derived from the female; M, sera derived from the male. c Main organs from mice treated with 60 μM Cy5 control or TKD-Cy5 through tail vein injection (2 h) were collected for imaging. n = 3; black arrow, gallbladder. d Mice treated with different doses of TKD, and the percent survival of each group was analyzed. n = 10. e Routine blood test of mice treated with 100 mg/kg TKD. n = 10; WBC white blood cell, Neu neutrophil, Lym lymphocyte, Mon monocyte, Eos eosinophil, RBC red blood cell, HGB hemoglobin, HCT hematocrit, MCV (fL) mean corpuscular volume, MCH mean corpuscular hemoglobin, MCHC mean corpuscular hemoglobin concentration, RDW-CV coefficient variation of red blood cell volume distribution width, RDW-SD (fL) standard deviation in red cell distribution width, PLT platelet, MPV (fL) mean platelet volume, PDW platelet distribution width, PCT thrombocytocrit. f Young (4–6 weeks of age, n = 10) and elder mice (12–13 weeks of age, n = 10) were treated with vehicle (0 mg/kg) or TKD at 50 mg/kg or 100 mg/kg every 3 days and their body weights were measured for statistics. g Pathological analysis of lungs, livers, spleens, and kidneys derived from the vehicle (TKD 0 mg/kg) or TKD-treated mice by using hematoxylin-eosin (H&E) staining. n = 5. Scale bars = 100 μm.