A novel theranostic activity-based probe targeting kallikrein 7 for the diagnosis and treatment of skin diseases

Abstract

We applied a new in silico approach for fishing protease-substrate motifs to design a kallirein 7 (KLK7)-specific phosphonate activity-based probe (ABP) to quantify the active KLK7 in situ. Epidermal application...

Generation of a quenched phosphonate activity-based probe for labelling the active KLK7 protease

“…Regarding the carbon stereocenter, we have previously showed that the R state is prevalent. 18 In respect to the P atom stereochemistry, the R diastereomer of compound 13 showed much higher binding scores towards both chymotrypsin A (Fig. 6A) and KLK7 (Fig.…”

“…Overall, these dockings are consistent in terms of general docking orientation in respect to similar compounds described in our previous work. 18 The Cy5 group interacts with the same region of KLK7 as well as chymotrypsin A, namely S1′. Notable differences are the benzyl – F218 and tert -butyl – W215 and L175 interactions with KLK7 (Fig.…”

“…Using an in silico approach to screen the MEROPS database (), we identified the dipeptide sequence Phe–Phe as a specific recognition sequence for cleavage by KLK7. 18 This sequence was modified here to yield a quenched ABP (qABP) that emits fluorescence upon binding to the active (mature) KLK7 (Fig. 1 and 2).…”

“…To investigate whether addition of the bulky organic groups (Cy5 and QSY21) could alter the kinetics of qABP binding onto active KLK7 as compared to the Boc-FFP 18 and the Cy5-FFP ABP (compound 13 ), the IC 50 values were determined. As shown in Table 1, the IC 50 for qABP 15 was approximately 80-fold higher than the IC 50 for the Cy5-FFP that lacks the quencher.…”

“…The qABP 15 was designed based on a mixed aryl alkyl phosphonate reactive warhead carrying a Phe-Phe recognition sequence. 18 It is shown that the KLK7-qABP binds both the recombinant and endogenously expressed human KLK7 but also the mouse KLK7, which makes it a useful tool for preclinical studies employing animal models, as well. The advantage of the generated qABP for future applications in molecular imaging is the use of a quencher that zeroes background fluorescence for the unbound qABP, thus, optimizing the quantification of the active enzyme based on fluorescence emission by the active enzyme-bound qABP.…”