In order to discover new compounds with good fungicidal activities, 32 pyrazole derivatives were designed and synthesized. The structures of the target compounds were confirmed by ¹H-NMR, ¹³C-NMR, and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), and their fungicidal activities against Botrytis cinerea, Rhizoctonia solani Kuhn, Valsa mali Miyabe et Yamada, Thanatephorus cucumeris (Frank) Donk, Fusarium oxysporum (S-chl) f.sp. cucumerinum Owen, and Fusarium graminearum Schw were tested. The bioassay results indicated that most of the derivatives exhibited considerable antifungal activities, especially compound 26 containing a p-trifluoromethyl- phenyl moiety showed the highest activity, with EC₅₀ values of 2.432, 2.182, 1.787, 1.638, 6.986, and 6.043 μg/mL against B. cinerea, R. solani, V. mali, T. cucumeris, F. oxysporum, and F. graminearum, respectively. Moreover, the activities of compounds such as compounds 27–32 were enhanced by introducing isothiocyanate and carboxamide moieties to the 5-position of the pyrazole ring.

Cell-based, phenotypic screening of small molecules often identifies compounds with provocative biological properties. However, determining the cellular target(s) and/or mechanism of action (MoA) of lead compounds remains an extremely challenging and time-consuming exercise. To provide insights into a compound's cellular action and greatly reduce the time required for MoA determination, we have developed a screening platform consisting of an extensive series of reporter gene assays (RGAs). A collection of > 11,000 compounds of known MoA (e.g., World Drug Index entries) were screened against the entire panel. The output provided evidence that an RGA signature could be ascribed to numerous, biologically diverse MoAs. The reference database generated suggested novel biological activity for particular compounds. For example, the profiling data led to the prediction that the cellular target of the natural product terprenin was dihydroorotate dehydrogenase (DHODH), which was confirmed experimentally. The screening methodology developed for this endeavor renders it amenable to the future examination of compounds with unknown MoA, in an automated, inexpensive, and time-efficient manner.

Cherry tomatoes (Solanum lycopersicum) are becoming increasingly popular due to their nutrition and delicious flavor. However, cherry tomatoes are highly perishable and susceptible to various pathogenic microorganisms after harvest, such as Botrytis cinerea. In the pretest experiment, we screened out three kinds of plant essential oils (EOs) (Torreya grandis oil, Eriobotrya japonica oil, and Citrus medica oil) that have strong fungicidal activity on B. cinerea from cherry tomatoes. To further evaluate the postharvest preservation application prospect of these three oils for cherry tomatoes, the oils were extracted from different parts of three plants by hydrodistillation, and their chemical constituents were analyzed by gas chromatography-mass spectrometry. The main representative components of T. grandis oil, E. japonica oil, and C. medica oil were δ-cadinene (11.76%), transnerolidol (9.70%), and 5,7-dimethoxycoumarin (23.22%), respectively. These three EOs effectively inhibited the mycelial growth of B. cinerea in vitro, with EC₅₀ values of 81.672, 144.046, and 221.500 μl/liter, respectively. Compared with the blank control and other oil treatments, the T. grandis oil (at a concentration of 200 µl/liter) fumigation treatment was more effective at inhibiting the growth rate of the pathogen. In addition, the phenolic content and phenylalanine ammonia lyase, β-1,3-glucanase, chitinase, and peroxidase activities of tomatoes significantly increased on the seventh day due to the T. grandis oil treatment. The present study shows that these three oils with high extraction rates have preservation potential for cherry tomatoes. Among these three EOs, T. grandis oil can be used to further develop preservative products as a fumigant.

Background Metastatic castration-resistant prostate cancer (mCRPC) remains an incurable disease. Bispecific T cell engagers (TCEs) targeting prostate-specific membrane antigen (PSMA) and CD3 on T cells showed great clinical potential for the treatment of mCRPC. However, cytokine release syndrome (CRS) and poor pharmacokinetic (PK) profile hinder their further development. To overcome these challenges, Janux has developed JANX007, a tumor-activated T cell engager (TRACTr) with enhanced safety and PK properties. JANX007 is a humanized trispecific protein that contains PSMA- and CD3-binding domains, an albumin binding domain to extend circulating half-life, and a CD3 inhibitory peptide mask fused to the molecule through tumor protease cleavable linker. Only when tumor-resident proteases cleave the TRACTr and enable mask separation can the resulting active molecule bind CD3. This cleavage-dependent CD3 agonism can potentially limit systemic toxicity associated with broad T cell activation. Methods Peptide masks against the CD3 binding domain were identified via phage display. Mask efficiency was evaluated using human CD3 ELISAs. Masking and cleavable linker stability was characterized in human (healthy and mCRPC donor) and cynomolgus monkey serum. JANX007-induced cleavage-dependent activation of T cells was evaluated in human PBMC/prostate tumor cell in vitro co-culture assays. The pharmacokinetic and safety profile of JANX007 was evaluated in non-human primate (NHP) studies. Results Engagement of CD3 target by JANX007 was shown to be cleavage dependent where masking reduced CD3 binding by >600x. In vitro, JANX007 exhibited a ~500x decrease in potency to activate T cells and induce T cell-mediated tumor cell killing relative to non-masked TCE. JANX007 was highly stable in healthy and mCRPC human donor serum, with ≤1% cleavage per day. While proteolytic cleavage of JANX007 in the tumor microenvironment is expected to drive anti-tumor activity, the maintenance of masking in the blood compartment is expected to mitigate the safety risks associated with potential off-tumor toxicity and CRS. JANX007 was found to be highly stable in NHPs with minimally detectable cleavage. The lack of TCE accumulation in NHPs mitigated on-target healthy tissue toxicities and minimized CRS. Clinical chemistry, hematology, and pathology data package support No-Observed-Adverse-Effect-Level (NOAEL) ≥1.5 mg/kg/dose. Finally, the cleavable albumin-binding domain extended the circulating half-life of JANX007 to ~120h in NHPs, relative to the 2h half-life of non-masked TCE, supporting its projected once-weekly clinical dosing. Conclusions Cleavage-dependent activity, half-life extended PK, the potential for superior safety and manufacturability properties of JANX007 support its further development as an attractive mCRPC therapeutic. Acknowledgements We acknowledge Marque Todd for providing insightful comments and help with the interpretation of NHP safety studies. Ethics Approval All animal experiments were approved by the Institutional Animal Use and Care Committee of the institutions conducting the studies and in compliance with the Animal Welfare Act, the Guide for the Care and Use of Laboratory Animals, and the Office of Laboratory Animal Welfare.