Department of Molecular and Medical Pharmacology

Apolipoproteins are the defining functional component of lipoproteins and play critical roles in lipid transport and metabolism. High-density lipoprotein (HDL) and its primary functional constituent, apolipoprotein A-I, are of particular importance because of anti-inflammatory and antioxidant properties. Apolipoprotein mimetic peptides are short-chain amino acids designed to mimic the functions and alpha-helical structure of endogenous apolipoproteins and have demonstrated efficacy in ameliorating animal models of cardiovascular disease (CVD) and cancer. The mechanisms underlying the mimetics are yet to be fully elucidated, but a comprehensive review of the literature suggests that the peptides attack pathways shared in the pathophysiology of both diseases. This review also discusses the many pre-clinical studies on the mimetic peptides, highlighting possible mechanisms at work in each. Proposed mechanisms of protection against CVD and cancer include binding and removal of pro-inflammatory oxidized lipids, reduction in reactive oxygen species, and modulation of immune cell populations. Additionally, nanoparticles (NP) formulations incorporating apolipoprotein mimetic peptides or recombinant apolipoproteins have exhibited anti-atherogenic and anti-cancer activity. To date, clinical trials to assess the effect of reconstituted HDL NPs on CVD outcomes have not shown significant improvement. The large body of successful animal studies on apolipoproteins and apolipoprotein mimetic peptides presents a disconnect between pre-clinical and clinical efficacy, highlighting the need for a more complete understanding of the underlying pathways and mechanisms.

5079 Background: 177 Lu-PSMA-617 (Lu-PSMA) contributes to prolong progression-free survival (PFS) and overall survival (OS) in metastatic castration-resistant prostate cancer (mCRPC) patients who progressed after chemotherapy. Pretherapeutic prostate-specific membrane antigen (PSMA) PET/CT information can be used to predict Lu-PSMA therapy response patterns and outcomes, with various quantitative and visual methods proposed. We aimed to test various proposed PSMA PET/CT-derived outcome predictors in the U.S. expanded-access program (EAP) cohort. Methods: Patients enrolled in the EAP (NCT04825652) for Lu-PSMA at 3 institutions with available pretherapeutic PSMA PET/CT and outcomes were included in this analysis. Quantitative analysis was performed for all tumor lesions on PSMA PET/CT with semi-automatically contouring. Total tumor volume (TV), total tumor SUVmean, total tumor SUVmax, total lesion uptake (TLU = TV * SUVmean), total lesion quotient (TLQ = TV / SUVmean), and quantitative PSMA PET tumor–to–salivary gland ratio (qPSG: high, ≥ 1.5; intermediate, 0.5–1.5; low, ≤ 0.5) were calculated for each patient. For visual analysis, visual PSG (vPSG: high, most of the lesions showed higher uptake than the parotid glands; intermediate, neither low nor high; low, most of the lesions showed lower uptake than the parotid glands) and heterogeneity and intensity of tumors (HIT: 1, SUVmax < 15; 2, 15–79 with heterogeneous intensity; 3, 15–79 with homogeneous intensity; 4, ≥ 80) scores were used for assessment. Outcomes included a prostate-specific antigen (PSA) PFS, and OS. We evaluated the predictive performance of each model using Cox proportional hazards regression analysis and assessed their performance with the concordance index (c-index). Results: In total, 88 patients who received Lu-PSMA within the EAP between May 2021 and March 2022 were eligible and included in this analysis. For the PSA PFS, the total tumor SUVmean achieved the highest c-index of 0.678 (HR 0.91 [95% CI, 0.85–0.97], p = 0.004), followed by the total tumor SUVmax with a c-index of 0.640 (HR 0.99 [95% CI, 0.99–1.00], p = 0.034). For OS, the TLQ achieved the highest c-index of 0.658 (HR 1.01 [95% CI, 1.00–1.01], p < 0.001), followed by the total tumor SUVmean with a c-index of 0.634 (HR 0.89 [95% CI, 0.83–0.96], p = 0.004). The HIT score showed the third highest c-index of 0.632; however, when using score 1 as the reference, the HR did not exhibit a sequential trend across ordinal categories as anticipated. Conclusions: Quantitative analysis outperformed visual analysis in predicting the outcome of mCRPC with Lu-PSMA therapy in the EAP cohort. Total tumor SUVmean was identified as the most robust predictor for PSA PFS, while TLQ showed promise as a predictor for OS. Incorporating these predictors into clinical decision-making for pre-Lu-PSMA therapy could aid in patient selection and treatment planning. Clinical trial information: NCT04825652 .

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Caspases are a family of highly homologous cysteine proteases that play critical roles in inflammation and apoptosis. Small molecule inhibitors are useful tools for studying caspase biology, complementary to genetic approaches. However, achieving inhibitor selectivity for individual members of this highly homologous enzyme family remains a major challenge in developing such tool compounds. Prior studies have revealed that one strategy to tackle this selectivity gap is to target the precursor or zymogen forms of individual caspases, which share reduced structural homology when compared to active proteases. To establish a screening assay that favors the discovery of zymogen-directed caspase-10 selective inhibitors, we engineered a low-background and high-activity tobacco etch virus (TEV)-activated caspase-10 protein. We then subjected this turn-on protease to a high-throughput screen of approximately 100 000 compounds, with an average Z' value of 0.58 across all plates analyzed. Counter screening, including against TEV protease, delineated bona fide procaspase-10 inhibitors. Confirmatory studies identified a class of thiadiazine-containing compounds that undergo isomerization and oxidation to generate cysteine-reactive compounds with caspase-10 inhibitory activity. In parallel, mode-of-action studies revealed that pifithrin-μ (PFTμ), a reported TP53 inhibitor, also functions as a promiscuous caspase inhibitor. Both inhibitor classes showed preferential zymogen inhibition. Given the generalized utility of activation assays, we expect our screening platform to have widespread applications in identifying state-specific protease inhibitors.

SARS-CoV-2 can infect cells through endocytic uptake, a process that is targeted by inhibition of lysosomal proteases. However, clinically this approach to treat viral infections has afforded mixed results, with some studies detailing an oral regimen of hydroxychloroquine accompanied by significant off-target toxicities. We rationalized that an organelle-targeted approach will avoid toxicity while increasing the concentration of the drug at the target. Here, we describe a lysosome-targeted, mefloquine-loaded poly(glycerol monostearate-co-ε-caprolactone) nanoparticle (MFQ-NP) for pulmonary delivery via inhalation. Mefloquine is a more effective inhibitor of viral endocytosis than hydroxychloroquine in cellular models of COVID-19. MFQ-NPs are less toxic than molecular mefloquine, are 100-150 nm in diameter, and possess a negative surface charge, which facilitates uptake via endocytosis allowing inhibition of lysosomal proteases. MFQ-NPs inhibit coronavirus infection in mouse MHV-A59 and human OC43 coronavirus model systems and inhibit SARS-CoV-2 WA1 and its Omicron variant in a human lung epithelium model. Organelle-targeted delivery is an effective means to inhibit viral infection.

To identify genes that regulate the response to the potentially addictive drug cocaine, we performed a control experiment using genome-wide association studies (GWASs) and RNA-Seq of a panel of inbred and recombinant inbred mice undergoing intravenous self-administration of saline. A linear mixed model increased statistical power for analysis of the longitudinal behavioral data, which was acquired over 10 days. A total of 145 loci were identified for saline compared to 17 for the corresponding cocaine GWAS. Only one locus overlapped. Transcriptome-wide association studies (TWASs) using RNA-Seq data from the nucleus accumbens and medial frontal cortex identified 5031434O11Rik and Zfp60 as significant for saline self-administration. Two other genes, Myh4 and Npc1, were nominated based on proximity to loci for multiple endpoints or a cis locus regulating expression. All four genes have previously been implicated in locomotor activity, despite the absence of a strong relationship between saline taking and distance traveled in the open field. Our results indicate a distinct genetic basis for saline and cocaine self-administration, and suggest some common genes for saline self-administration and locomotor activity.

BACKGROUND: The Editorial Board of EJNMMI Radiopharmacy and Chemistry releases a biannual highlight commentary to update the readership on trends in the field of radiopharmaceutical development and application of radiopharmaceuticals. MAIN BODY: This selection of highlights provides commentary on 24 different topics selected by each co-authoring Editorial Board member addressing a variety of aspects ranging from novel radiochemistry to first-in-human application of novel radiopharmaceuticals. CONCLUSION: Trends in radiochemistry and radiopharmacy are highlighted. Hot topics cover the entire scope of EJNMMI Radiopharmacy and Chemistry, demonstrating the progress in the research field in many aspects.

Pancreatic ductal adenocarcinoma (PDAC) is a leading cause of cancer-related mortality, largely due to late-stage diagnosis. Reliable early detection methods are critically needed. PDAC-derived extracellular vesicles (EVs) carry molecules that reflect their parental tumor cells and are detectable in early disease stages, offering a promising noninvasive diagnostic approach. Here, a streamlined PDAC EV Surface Protein Assay for quantifying PDAC EV subpopulations in 300-µL plasma through a two-step workflow is presented: i) click chemistry-mediated EV enrichment using EV Click Beads and trans-cyclooctene-grafted antibodies targeting three PDAC EV-specific surface proteins (MUC1, EGFR, and TROP2), and ii) quantification of enriched PDAC EVs through reverse transcription-quantitative polymerase chain reaction. The three PDAC EV-specific surface proteins are identified using a bioinformatics framework and validated on PDAC cell lines and tissue microarrays. The resultant PDAC EV Score, derived from signals of the three PDAC EV subpopulations, demonstrates robust differentiation of PDAC patients from noncancer controls, with area under the receiver operating characteristic curves of 0.94 in the training (n = 124) and 0.93 in the validation (n = 136) cohorts. This EV-based diagnostic approach successfully exploits PDAC EV subpopulations as novel biomarkers for PDAC early detection, translating PDAC surface proteins into an EV-based liquid biopsy platform.