{"doi":"10.2147/jir.s333725","title":"CASPorter: A Novel Inducible Human CASP1/NALP3/ASC Inflammasome Biosensor","abstract":"Background: Following our 2015 elucidation of the CASP1/NALP3 inflammasome mechanism of glucocorticoid (GC)-resistance in pediatric acute lymphoblastic leukemia (ALL) patients, we engineered a cell-based CASP1/NALP3 reporter system suitable for high-throughput screening (HTS) of small molecule libraries, with the purpose of identifying compounds capable of inhibiting the CASP1/NALP3 inflammasome and synergizing with GC drugs for the treatment of GC-resistant ALL patients and various autoinflammatory diseases. Methods: A Dox-controlled system was utilized to induce the expression of the ASC transgene in HEK293 cells while simultaneously overexpressing NLRP3 and CASP1 . ASC/CASP1/NALP3 inflammasome complex formation was confirmed by co-immunoprecipitation (co-IP) experiments. Next, a LV fluorescence-based biosensor ( CASP orter) was transduced in the HEK293-iASC-NLRP3/CASP1 cell line to monitor the real-time activation of CASP1/NALP3 inflammasome in live cells. The applicability and effectiveness of the CASP orter cell line were tested by co-treatment with Dox and four known CASP1/NLRP3 inhibitors (MCC950, Glyburide, VX-765 and VRT-043198). Inflammasome activation and inhibitions were assessed by Western blotting, fluorescence microscopy and flow cytometry (FC) methods. Results: Dox treatment significantly induced ASC expression and increased levels of cleaved and catalytically active CASP1, co-IPs further demonstrated that CASP1 was pulled-down with NLRP3 in HEK293-iASC-NLRP3/CASP1 cells after induction of ASC by Dox treatment. In HEK293-iASC-NLRP3/CASP1- CASP orter cell system, cleavage of the CASP1 consensus site (YVAD) in the CASP orter protein after Dox treatment causing excitation/emission of green fluorescence and the 71% GFP+ cell population increase quantified by FC (78.1% vs 6.90%). Dox-induced activation of the NLRP3 inflammasome was dose-dependently inhibited by Dox co-treatment with four known CASP1/NLRP3 inhibitors. Conclusion: We have established a cell-based CASP1/NLRP3 inflammasome model, utilizing a fluorescence biosensor as readout for qualitatively observing and quantitatively determining the activation of caspase 1 and NLRP3 inflammasomes in living cells and easily define the inhibitory effect of inhibitors with high efficacy. Keywords: cell-based biosensor, NALP3 inflammasome, CASP1, ASC","journal":"Journal of Inflammation Research","year":2022,"id":268627,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":2,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9562,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2022-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":931412,"name":"Jordan A Beard","orcid":null,"position":1,"is_corresponding":false},{"id":930819,"name":"Guoping Yang","orcid":"0000-0001-5930-586X","position":2,"is_corresponding":false},{"id":931413,"name":"William E Evans","orcid":null,"position":3,"is_corresponding":false},{"id":931414,"name":"Erik J Bonten","orcid":null,"position":4,"is_corresponding":false},{"id":930818,"name":"Chan Zou","orcid":"0000-0001-5228-8593","position":0,"is_corresponding":true}],"reference_count":33,"raw_metadata":null,"created_at":"2026-07-19T00:27:14.088663Z","pmid":"35221708","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}