Characteristics and in vitro imaging study of matrix metalloproteinase-2 targeting activable cell-penetrating peptide

Objective: To study invasively imaging MMP2-positive tumor cell by paramagnetic Gadolinium or fluorescein carried by a activable cell penetrating peptides. Methods: To label Fluorescein-5-isothiocyanate (FITC) and gadopentetate with the activable cell-penetrating peptides by a solid-phase synthesis method. Identification by TOF mass spectrum (TOF-MS). Isoelectric point of the activable cell penetrating peptides is determined by disc electrophoresis. T1 relaxion of gadopentetate labeled with the activable cell-penetrating peptides (B) in water were determined on 400 MHZ NMR. Human lung cancer cell lines: A549 were respectively stained by FITC labeled with ACPPs (A) or FITC alone. MMP2 expression and activity were determined by zymography. T1 WI signal of A549 incubated with 120 nmol/ ml B or Diethylenetriaminepentaacetic Acid-Gadolinium (Gd-DTPA) for different times were obtained by 1.5T MRI. The location of B in A549 was detected by Transmission Electron Microscopy. Visualization analysis and half-quantitative analysis were used to determine the signal characteristics. The ANOVA analysis and the paired samples t test were performed by SPSS 13.0. Results: MALDI TOF-MS molecular weigh of A and B respectively is 3789.74 and 3911.93. Isoelectric point is 11.005T1 Relaxion of 0.5 mmo/L Gd-DTPA and B at 17°C respectively is (0.052 ± 0.01) sec and (0.050 ± 0.001) sec. Fluorescein uptake assays showed that A translocated into A549 but would be inhibited by MMP2 antibody. Zymography showed both actived-MMP2 (67000) and pro-MMP2 (72000) expressed byA549. MR imaging showed A549 incubating with B had a high T₁ signal, and the signal of A549 incubating with Gd-DTPA is similar with that of the control group. Furthermore, ANOVA suggested that the T₁ signal intensity of A549 incubating with B was effected by incubating time(F = 267.569, P < 0.001) and increasing in a time-dependent fashion at the observed time. There is no defference between the T₁ signal intensity of A549 incubating with Gd-DTPA and the control group (P > 0.05). TEM showed A in cytoplasm and nucleus. Conclusion: The study in vitro suggests that the MMP-2 activable cell-penetrating peptides bearing contrast media can detect the MMP2-positive tumor cell.