Serendipitous Identification of Azine Anticancer Agents Using a Privileged Scaffold Morphing Strategy.

The use of privileged scaffolds as a starting point for the construction of libraries of bioactive compounds is a widely used strategy in drug discovery and development. Scaffold decoration, morphing and hopping are additional techniques that enable the modification of the chosen privileged framework and better explore the chemical space around it. In this study, two series of highly functionalized pyrimidine and pyridine derivatives were synthesized using a scaffold morphing approach consisting of triazine compounds obtained previously as antiviral agents. Newly synthesized azines were evaluated against lymphoma, hepatocarcinoma, and colon epithelial carcinoma cells, showing in five cases acceptable to good anticancer activity associated with low cytotoxicity on healthy fibroblasts. Finally, ADME in vitro studies were conducted on the best derivatives of the two series showing good passive permeability and resistance to metabolic degradation.

Early investigation of a novel SI306 theranostic prodrug for glioblastoma treatment.

Glioblastoma multiforme (GBM) is one of the most aggressive malignancies with a high recurrence rate and poor prognosis. Theranostic, combining therapeutic and diagnostic approaches, arises as a successful strategy to improve patient outcomes through personalized medicine. Src is a non-receptor tyrosine kinase (nRTK) whose involvement in GBM has been extensively demonstrated. Our previous research highlighted the effectiveness of the pyrazolo[3,4-d]pyrimidine SI306 and its more soluble prodrug CMP1 as Src inhibitors both in in vitro and in vivo GBM models. In this scenario, we decided to develop a theranostic prodrug of SI306, ProSI-DOTA(68 Ga) 1, which was designed to target GBM cells after hydrolysis and follow-up on the disease's progression and improve the therapy's outcome. First, the corresponding nonradioactive prodrug 2 was tested to evaluate its ADME profile and biological activity. It showed good metabolic stability, no inhibition of CYP3A4, suboptimal aqueous solubility, and slight gastrointestinal and blood-brain barrier passive permeability. Compound 2 exhibited a drastic reduction of cell vitality after 72 h on two different GBM cell lines (GL261 and U87MG). Then, 2 was subjected to complexation with the radionuclide Gallium-68 to give ProSI-DOTA(68 Ga) 1. The cellular uptake of 1 was evaluated on GBM cells, highlighting a slight but significant time-dependent uptake. The data obtained from our preliminary studies reflect the physiochemical properties of 1. The use of an alternative route of administration, such as the intranasal route, could overcome the physiochemical limitations and enhance the pharmacokinetic properties of 1, paving the way for its future development.

Sweet Cherry Extract as Permeation Enhancer of Tyrosine Kinase Inhibitors: A Promising Prospective for Future Oral Anticancer Therapies.

Although patients would rather oral therapies to injections, the gastrointestinal tract's low permeability makes this method limiting for most compounds, including anticancer drugs. Due to their low bioavailability, oral antitumor therapies suffer from significant variability in pharmacokinetics and efficacy. The improvement of their pharmacokinetic profiles can be achieved by a new approach: the use of natural extracts enriched with polyphenolic compounds that act as intestinal permeability enhancers. Here, we propose a safe sweet cherry extract capable of enhancing oral absorption. The extract was characterized by the HPLC-UV/MS method, evaluated for in vitro antioxidant activity, safety on the Caco-2 cell line, and as a potential permeation enhancer. The sweet cherry extract showed a high antioxidant capacity (ABTS and DPPH assays were 211.74 and 48.65 µmol of Trolox equivalent/g dried extract, respectively), high content of polyphenols (8.44 mg of gallic acid per gram of dry extract), and anthocyanins (1.80 mg of cyanidin-3-glucoside equivalent per g of dry extract), reassuring safety profile (cell viability never lower than 98%), and a significant and fully reversible ability to alter the integrity of the Caco-2 monolayer (+81.5% of Lucifer yellow permeability after 2 h). Furthermore, the ability of the sweet cherry extract to improve the permeability (Papp) and modify the efflux ratio (ER) of reference compounds (atenolol, propranolol, and dasatinib) and selected pyrazolo[3,4-d]pyrimidine derivatives was investigated. The obtained results show a significant increase in apparent permeability across the Caco-2 monolayer (tripled and quadrupled in most cases), and an interesting decrease in efflux ratio when compounds were co-incubated with sweet cherry extract.

Targeting GLI1 and GLI2 with small molecule inhibitors to suppress GLI-dependent transcription and tumor growth.

Aberrant activation of Hedgehog (HH) signaling in cancer is the result of genetic alterations of upstream pathway components (canonical) or other oncogenic mechanisms (noncanonical), that ultimately concur to activate the zinc-finger transcription factors GLI1 and GLI2. Therefore, inhibition of GLI activity is a good therapeutic option to suppress both canonical and noncanonical activation of the HH pathway. However, only a few GLI inhibitors are available, and none of them have the profile required for clinical development due to poor metabolic stability and aqueous solubility, and high hydrophobicity. Two promising quinoline inhibitors of GLI were selected by virtual screening and subjected to hit-to-lead optimization, thus leading to the identification of the 4-methoxy-8-hydroxyquinoline derivative JC19. This molecule impaired GLI1 and GLI2 activities in several cellular models interfering with the binding of GLI1 and GLI2 to DNA. JC19 suppressed cancer cell proliferation by enhancing apoptosis, inducing a strong anti-tumor response in several cancer cell lines in vitro. Specificity towards GLI1 and GLI2 was demonstrated by lower activity of JC19 in GLI1- or GLI2-depleted cancer cells. JC19 showed excellent metabolic stability and high passive permeability. Notably, JC19 inhibited GLI1-dependent melanoma xenograft growth in vivo, with no evidence of toxic effects in mice. These results highlight the potential of JC19 as a novel anti-cancer agent targeting GLI1 and GLI2.

Progress towards Adjuvant Development: Focus on Antiviral Therapy.

In recent decades, vaccines have been extraordinary resources to prevent pathogen diffusion and cancer. Even if they can be formed by a single antigen, the addition of one or more adjuvants represents the key to enhance the response of the immune signal to the antigen, thus accelerating and increasing the duration and the potency of the protective effect. Their use is of particular importance for vulnerable populations, such as the elderly or immunocompromised people. Despite their importance, only in the last forty years has the search for novel adjuvants increased, with the discovery of novel classes of immune potentiators and immunomodulators. Due to the complexity of the cascades involved in immune signal activation, their mechanism of action remains poorly understood, even if significant discovery has been recently made thanks to recombinant technology and metabolomics. This review focuses on the classes of adjuvants under research, recent mechanism of action studies, as well as nanodelivery systems and novel classes of adjuvants that can be chemically manipulated to create novel small molecule adjuvants.

Biological Evaluation and In Vitro Characterization of ADME Profile of In-House Pyrazolo[3,4-d]pyrimidines as Dual Tyrosine Kinase Inhibitors Active against Glioblastoma Multiforme.

The therapeutic use of tyrosine kinase inhibitors (TKIs) represents one of the successful strategies for the treatment of glioblastoma (GBM). Pyrazolo[3,4-d]pyrimidines have already been reported as promising small molecules active as c-Src/Abl dual inhibitors. Herein, we present a series of pyrazolo[3,4-d]pyrimidine derivatives, selected from our in-house library, to identify a promising candidate active against GBM. The inhibitory activity against c-Src and Abl was investigated, and the antiproliferative profile against four GBM cell lines was studied. For the most active compounds endowed with antiproliferative efficacy in the low-micromolar range, the effects toward nontumoral, healthy cell lines (fibroblasts FIBRO 2-93 and keratinocytes HaCaT) was investigated. Lastly, the in silico and in vitro ADME properties of all compounds were also assessed. Among the tested compounds, the promising inhibitory activity against c-Src and Abl (Ki 3.14 µM and 0.44 µM, respectively), the irreversible, apoptotic-mediated death toward U-87, LN18, LN229, and DBTRG GBM cell lines (IC50 6.8 µM, 10.8 µM, 6.9 µM, and 8.5 µM, respectively), the significant reduction in GBM cell migration, the safe profile toward FIBRO 2-93 and HaCaT healthy cell lines (CC50 91.7 µM and 126.5 µM, respectively), the high metabolic stability, and the excellent passive permeability across gastrointestinal and blood-brain barriers led us to select compound 5 for further in vivo assays.

[1,2,4]Triazolo[3,4-b]benzothiazole Scaffold as Versatile Nicotinamide Mimic Allowing Nanomolar Inhibition of Different PARP Enzymes.

We report [1,2,4]triazolo[3,4-b]benzothiazole (TBT) as a new inhibitor scaffold, which competes with nicotinamide in the binding pocket of human poly- and mono-ADP-ribosylating enzymes. The binding mode was studied through analogues and cocrystal structures with TNKS2, PARP2, PARP14, and PARP15. Based on the substitution pattern, we were able to identify 3-amino derivatives 21 (OUL243) and 27 (OUL232) as inhibitors of mono-ARTs PARP7, PARP10, PARP11, PARP12, PARP14, and PARP15 at nM potencies, with 27 being the most potent PARP10 inhibitor described to date (IC50 of 7.8 nM) and the first PARP12 inhibitor ever reported. On the contrary, hydroxy derivative 16 (OUL245) inhibits poly-ARTs with a selectivity toward PARP2. The scaffold does not possess inherent cell toxicity, and the inhibitors can enter cells and engage with the target protein. This, together with favorable ADME properties, demonstrates the potential of TBT scaffold for future drug development efforts toward selective inhibitors against specific enzymes.

Proteins from Tenebrio molitor: An interesting functional ingredient and a source of ACE inhibitory peptides.

The angiotensin-converting enzyme (ACE) inhibitory potential of the main protein fractions from Tenebrio molitor larvae (TML) was examined to evaluate their use as a novel antihypertensive functional food. Both fractions contained YAN tripeptide, previously reported as responsible for ACE inhibition. Although YAN has been synthesized and was used as a standard for LC-MS/MS quantification and IC50 against ACE was determined, low yields of YAN from TML did not explain adequately the activity of the whole protein fraction. LC-HRMS/MS investigation led to the identification of other three peptides, which were evaluated in silico, synthesized and tested against ACE. Among them, tetrapeptide NIKY showed the most promising activity (52 µM), highlighting once more the potential of TML and paving the way for exploitation in novel foods.

Reversible Monoacylglycerol Lipase Inhibitors: Discovery of a New Class of Benzylpiperidine Derivatives.

Monoacylglycerol lipase (MAGL) is the enzyme responsible for the metabolism of 2-arachidonoylglycerol in the brain and the hydrolysis of peripheral monoacylglycerols. Many studies demonstrated beneficial effects deriving from MAGL inhibition for neurodegenerative diseases, inflammatory pathologies, and cancer. MAGL expression is increased in invasive tumors, furnishing free fatty acids as pro-tumorigenic signals and for tumor cell growth. Here, a new class of benzylpiperidine-based MAGL inhibitors was synthesized, leading to the identification of 13, which showed potent reversible and selective MAGL inhibition. Associated with MAGL overexpression and the prognostic role in pancreatic cancer, derivative 13 showed antiproliferative activity and apoptosis induction, as well as the ability to reduce cell migration in primary pancreatic cancer cultures, and displayed a synergistic interaction with the chemotherapeutic drug gemcitabine. These results suggest that the class of benzylpiperidine-based MAGL inhibitors have potential as a new class of therapeutic agents and MAGL could play a role in pancreatic cancer.