Nanoreporter for Real‐Time Monitoring of Inflammasome Activity and Targeted Therapy

Abstract

Inflammasome activation is associated with a myriad of inflammatory diseases. However, existing methods provides a limited understanding of spatiotemporal kinetics of inflammasome activation, with restricted scope for early detection of associated treatment efficacy. This limitation offers an opportunity for the development of biocompatible in-vivo inflammasome monitoring tools with translational prospects. To achieve this, they report developing a pair of lipid-based nanoparticle systems, a reporter nanoparticle consisting of a caspase-1 activatable probe alone, and a theranostic nanoparticle combining the probe with an inflammasome-inhibiting drug. This biocompatible platform enhances the probe's residence time in circulation by preventing its opsonization and allowing its sustained release over time. Their results demonstrate the specificity of reporter nanoparticles towards caspase-1 activity and provides early-on monitoring of inflammasome activation both in-vitro as well as in-vivo. Additionally, the delivery of disulfiram, an inflammasome-inhibiting drug, along with reporter probe using theranostic nanoparticles enables real-time tracking of treatment efficacy in the gouty-arthritis inflammatory model. In summary, they report an unparalleled pair of the inflammasome-associated reporter and theranostic platforms suited not only for diagnostic applications but can also detect inflammasome-targeted treatment efficiency in real-time. These findings establish two novel, sensitive nanotools for non-invasive evaluation of inflammasome-targeted immunotherapy.

Caspase-1 Responsive Nanoreporter for In Vivo Monitoring of Inflammasome Immunotherapy

“…16 Unlike a few reported nanoparticles such as silica, metal oxides, and DOTAP, DOPC liposomes do not induce inflammasome activation when incubated with the primed macrophages, which makes them suitable to be used during inflamed conditions. 7 Nanoparticle synthesis included physical encapsulation of either just the caspase-1 responsive probe or an additional NLRP3 inhibitor to formulate either the only reporter YVAD Np or the probedrug theranostic YVAD-MCC Np, respectively, named after the components being encapsulated (Figure 2A). The NLRP3 inhibitor that we chose to utilize is MCC-950, which directly inhibits NLRP3 oligomerization, ultimately leading to inhibition in the cleavage of inactive caspase-1 to its respective active subunit.…”

“…In our previous approach, we developed a dual probe-drug theranostic platform delivering a caspase-1 cleavable FLTD peptide probe, mimicking the caspase-1 cleavage site on the pyroptosis-inducing pore-forming protein gasdermin-D, as well as the drug disulfiram (DSR), a gasdermin-D inhibitor (4). Our results elucidated that these liposomes offered potent tissue penetration, sustained release of cargo, and effective monitoring and inhibition of the inflammasome pathway . Our new theranostic platform aims to build off our previous approach by delivering both a caspase-1 responsive YVAD probe (mimicking the cleavage site on IL-1β) and NLRP3-inhibiting MCC950, which inhibits the signal 2 pathway induced by NLRP3 sensing and oligomerization, offering inhibition earlier in the inflammasome pathway.…”

“…Our results elucidated that these liposomes offered potent tissue penetration, sustained release of cargo, and effective monitoring and inhibition of the inflammasome pathway. 7 Our new theranostic platform aims to build off our previous approach by delivering both a caspase-1 responsive YVAD probe (mimicking the cleavage site on IL-1β) and NLRP3-inhibiting MCC950, which inhibits the signal 2 pathway induced by NLRP3 sensing and oligomerization, offering inhibition earlier in the inflammasome pathway. Moreover, this probe allows deeper tissue penetration due to the use of infrared dyes and quencher, which could be utilized in complex inflammatory models, such as colitis.…”

“…Previously, several attempts have been made utilizing caspase-1 enzyme activity to monitor inflammasome activation both in vivo and in vitro . − Some of these platforms applied bioluminescence, FRET (fluorescence resonance energy transfer), and fusion protein approaches and leveraged different caspase-1 specific substrates to develop these tools. Researchers first devised CM-269 by reverse designing caspase-1 inhibitor pralnacasan to include a bioluminescent substrate, allowing for in vitro tracking of its proteolytic activity rather than just physical abundance via usual immunodetection with antibodies or proteomics methods .…”

“…This pertains to its minimal clinical relevance and calls for better translational systems offering greater tissue penetration, bioavailability, and retention capability to generate higher signal-to-noise ratios for a longer period. Our approach is to employ potent in vivo inflammasome probe delivery using liposome nanoparticles, which do not have any effect on inflammasome activation but offer safe, effective delivery of cargo. ,, These liposomes also allow multiplexing capability with different inflammasome inhibiting compounds to develop a theranostic system with the dual capability of delivering anti-inflammatory drugs and monitoring their effectiveness in real-time in both in vitro and in vivo settings. In our previous approach, we developed a dual probe-drug theranostic platform delivering a caspase-1 cleavable FLTD peptide probe, mimicking the caspase-1 cleavage site on the pyroptosis-inducing pore-forming protein gasdermin-D, as well as the drug disulfiram (DSR), a gasdermin-D inhibitor (4).…”