Pyrrole: A Decisive Scaffold for the Development of Therapeutic Agents and Structure-Activity Relationship.

An overview of pyrroles as distinct scaffolds with therapeutic potential and the significance of pyrrole derivatives for drug development are provided in this article. It lists instances of naturally occurring pyrrole-containing compounds and describes the sources of pyrroles in nature, including plants and microbes. It also explains the many conventional and modern synthetic methods used to produce pyrroles. The key topics are the biological characteristics, pharmacological behavior, and functional alterations displayed by pyrrole derivatives. It also details how pyrroles are used to treat infectious diseases. It describes infectious disorders resistant to standard treatments and discusses the function of compounds containing pyrroles in combating infectious diseases. Furthermore, the review covers the uses of pyrrole derivatives in treating non-infectious diseases and resistance mechanisms in non-infectious illnesses like cancer, diabetes, and Alzheimer's and Parkinson's diseases. The important discoveries and probable avenues for pyrrole research are finally summarized, along with their significance for medicinal chemists and drug development. A reference from the last two decades is included in this review.

Design, Synthesis, and Anti-Breast Cancer Potential of Imidazole-Pyridine Hybrid Molecules In Vitro and Ehrlich Ascites Carcinoma Growth Inhibitory Activity Assessment In Vivo.

Breast cancer remains a challenging medical issue and is a high priority for biomedical research despite significant advancements in cancer research and therapy. The current study aims to determine the anticancer activity of a group of imidazole-pyridine-based scaffolds against a variety of breast cancer cell lines differing in their receptor expression (estrogen receptor (ER), progesterone receptor (PR), and HER-2). A series of 10 molecules (coded 5a-5j) were synthesized through multicomponent and alkylation reactions. FTIR, MS, 1H, and 13C NMR spectral analyses confirmed the structures and purity of the synthesized molecules. Subsequently, these molecules were tested for their ability to inhibit the viability of cell lines representing carcinoma of the breast, viz., MDA-MB-468 (ER-, PR-, and HER-), BT-474 (ER+, PR+, and HER+), T-47D (ER+, PR+, and HER-), and MCF-7 (ER+, PR+, and HER-) in vitro. Among these 10 molecules, 5a, 5c, 5d, and 5e exhibited better potency, as evidenced by IC50 < 50 µM at 24 h of treatment against BT-474 and MDA-MB-468 cell lines. However, except for 5d, the IC50 value is much higher than 50 µM when tested against T47D and MCF-7 cell lines at 24h. Extended treatment for 48 h reduced the effect of these molecules, as an increase in IC50 was observed. In mice, intraperitoneal administration of 5e retarded the Ehrlich ascites carcinoma (EAC) growth without causing any organ toxicity at the doses tested. In summary, we report the synthesis scheme and key structural requirements for a new series of imidazole-pyridine molecules for in vitro inhibition of the feasibility of breast cancer cells and in vivo inhibition of EAC tumors.

Orange Peel Waste as Feedstock for the Production of Glycerol-Free Biodiesel by the Microalgae Nannochloropsis oculata.

The bioconversion of agri-food waste into high-value products is gaining growing interest worldwide. Orange peel waste (OPW) is the main by-product of orange juice production and contains high levels of moisture and carbohydrates. In this study, the orange waste extract (OWE) obtained through acid hydrolysis of OPW was used as a substrate in the cultivation of the marine microalgae Nannochloropsis oculata. Photoheterotrophic (PH) and Photoautotrophic (PA) cultivations were performed in OWE medium and f/2 medium (obtained by supplementing OWE with macro- and micronutrients of f/2 medium), respectively, for 14 days. The biomass yields in PA and PH cultures were 390 mg L-1 and 450 mg L-1, while oil yields were 15% and 28%, respectively. The fatty acid (FA) profiles of PA cultures were mostly represented by saturated (43%) and monounsaturated (46%) FAs, whereas polyunsaturated FAs accounted for about 10% of the FAs. In PH cultures, FA profiles changed remarkably, with a strong increase in monounsaturated FAs (77.49%) and reduced levels of saturated (19.79%) and polyunsaturated (2.72%) FAs. Lipids obtained from PH cultures were simultaneously extracted and converted into glycerol-free biodiesel using an innovative microwave-assisted one-pot tandem protocol. FA methyl esters were then analyzed, and the absence of glycerol was confirmed. The FA profile was highly suitable for biodiesel production and the microwave-assisted one-pot tandem protocol was more effective than traditional extraction techniques. In conclusion, N. oculata used OWE photoheterotrophically, resulting in increased biomass and oil yield. Additionally, a more efficient procedure for simultaneous oil extraction and conversion into glycerol-free biodiesel is proposed.

Sulfonated Polyether Ketone Membranes Embedded with Nalidixic Acid-An Emerging Controlled Drug Releaser.

The effective administration of medication has advanced over decades, but the medical community still faces significant demand. Burst release and inadequate assimilation are major drawbacks that affect wound healing efficiency, leading to therapy failure. The widespread application of polymers in biomedical research is significant. The polyether ether ketone (PEEK) family is known for its biocompatibility, inertness, and semi-crystalline thermoplastic properties. In our present studies, we have chosen a member of this family, polyether ketone (PEK), to explore its role as a drug carrier. The PEK backbone was subjected to sulfonation to increase its hydrophilicity. The response surface methodology (RSM) was used to optimize the sulfonation process based on the time, degree of sulfonation, and temperature. The PEK polymer was sulfonated using sulfuric acid at 150 °C for 6 h; back titration was performed to quantify the degree of sulfonation, with 69% representing the maximum sulfonation. SPEK and nalidixic sodium salt were dissolved in dichloroacetic acid to create a thin membrane. The physiological and morphological properties were assessed for the SPEK membrane. The studies on drug release in distilled water and a simulated body fluid over the course of 24 h revealed a controlled, gradual increase in the release rate, correlating with a mathematical model and demonstrating the zero-order nature of the drug release. Hemolysis on the SPEK membrane revealed lower toxicity. The SPEK membrane's biocompatibility was established using in vitro cytotoxicity tests on the Vero (IC50: 137.85 g/mL) cell lines. These results confirm that the SPEK membranes are suitable for sustained drug release.

Design of an Antibiotic-Releasing Polymer: Physicochemical Characterization and Drug Release Patterns.

Conventional drug delivery has its share of shortcomings, especially its rapid drug release with a relatively short duration of therapeutic drug concentrations, even in topical applications. Prolonged drug release can be effectively achieved by modifying the carrier in a drug delivery system. Among the several candidates for carriers studied over the years, poly (ether ether ketone), a biocompatible thermoplastic, was chosen as a suitable carrier. Its inherent hydrophobicity was overcome by controlled sulfonation, which introduced polar sulfonate groups onto the polymer backbone. Optimization of the sulfonation process was completed by the variation of the duration, temperature of the sulfonation, and concentration of sulfuric acid. The sulfonation was confirmed by EDS and the degree of sulfonation was determined by an NMR analysis (61.6% and 98.9%). Various physical properties such as morphology, mechanical strength, and thermal stability were studied using scanning electron microscopy, tensile testing, and thermogravimetric analysis. Cytotoxicity tests were performed on the SPEEK samples to study the variation in biocompatibility against a Vero cell line. The drug release kinetics of ciprofloxacin (CP) and nalidixic acid sodium salt (NA)-loaded membranes were studied in deionized water as well as SBF and compared against the absorbance of standardized solutions of the drug. The data were then used to determine the diffusion, distribution, and permeability coefficients. Various mathematical models were used to fit the obtained data to establish the order and mechanism of drug release. Studies revealed that drug release occurs by diffusion and follows zero-order kinetics.

Imidazole-pyridine hybrids as potent anti-cancer agents.

In the current investigation, fifteen novel imidazole-pyridine-based molecules were synthesized and tested against cell lines of the lung (H1299) and colon (HCT116) adenocarcinomas by proliferation assay. The results demonstrated that compounds 5a, 5d, 5e, and 5f were the most active (IC50<30 µM). Based on recent literature and the current results, the glycogen synthase kinase-3ß (GSK-3ß) protein was investigated in-silico as a possible target. The molecular docking and QSAR revealed an excellent binding affinity of the selected imidazole-pyridine compounds to GSK-3ß. Notably, GSK-3ß protein levels were significantly upregulated in hepatocellular liver carcinoma (LIHCs) tissues and negatively affected patient prognosis. Consequently, the compounds were evaluated on liver cancer cell lines (HepG2, HUH-7, and PLC/PRF/5) by the MTT assay, and 5d showed the highest antitumor activity. This study offers new compounds with interesting biological activity on GSK-3ß as a target, exhibiting a potential therapeutic impact for hepatocellular carcinoma patients.

Secondary metabolic profiles and anticancer actions from fruit extracts of immature pomegranates.

Immature fruits from Punica granatum L. thinning are a neglected side product of pomegranate production with cumbersome disposal costs for farmers. To explore value potential of immature fruits from pomegranate 'Wonderful' cultivars, the compositional landscapes and antitumorigenic activities of pomegranate extracts from two different stages of maturation were assessed. Cancer cell proliferation and cytotoxicity was quantified in human lung H1299 and colon HCT116 adenocarcinomas by crystal violet staining, MTS assay and caspase-3 activity. High performance liquid chromatography-diode array detector (HPLC/DAD) and high performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC/ESI-MS) analyses indicate that immature fruits are rich sources of gallotannins and ellagitannins, with the highest amounts contained in immature fruit peels. Biological investigations reveal a robust anticancer activity by those immature P. granatum fruit extracts, which reflected induction of tumor cytotoxicity and cell death mechanisms. Together, present observations suggest P. granatum byproducts from the thinning process may provide unexplored values for virtuous circular economy.

Towards bio-compatible magnetic nanoparticles: Immune-related effects, in-vitro internalization, and in-vivo bio-distribution of zwitterionic ferrite nanoparticles with unexpected renal clearance.

HYPOTHESIS: Iron oxide and other ferrite nanoparticles have not yet found widespread application in the medical field since the translation process faces several big hurdles. The incomplete knowledge of the interactions between nanoparticles and living organisms is an unfavorable factor. This complex subject should be made simpler by synthesizing magnetic nanoparticles with good physical (relaxivity) and chemical (colloidal stability, anti-fouling) properties and no biological activity (no immune-related effects, minimal internalization, fast clearance). Such an innocent scaffold is the main aim of the present paper. We systematically searched for it within the class of small-to-medium size ferrite nanoparticles coated by small (zwitter)ionic ligands. Once established, it can be functionalized to achieve targeting, drug delivery, etc. and the observed biological effects will be traced back to the functional molecules only, as the nanosized scaffold is innocent. EXPERIMENTS: We synthesized nine types of magnetic nanoparticles by systematic variation of core composition, size, coating. We investigated their physico-chemical properties and interaction with serum proteins, phagocytic microglial cells, and a human model of inflammation and studied their biodistribution and clearance in healthy mice. The nanoparticles have good magnetic properties and their surface charge is determined by the preferential adsorption of anions. All nanoparticle types can be considered as immunologically safe, an indispensable pre-requisite for medical applications in humans. All but one type display low internalization by microglial BV2 cells, a process strongly affected by the nanoparticle size. Both small (3 nm) and medium size (11 nm) zwitterionic nanoparticles are in part captured by the mononuclear phagocyte system (liver and spleen) and in part rapidly (≈1 h) excreted through the urinary system of mice. FINDINGS: The latter result questions the universality of the accepted size threshold for the renal clearance of nanoparticles (5.5 nm). We suggest that it depends on the nature of the circulating particles. Renal filterability of medium-size magnetic nanoparticles is appealing because they share with small nanoparticles the decreased accumulation-related toxicity while performing better as magnetic diagnostic/therapeutic agents thanks to their larger magnetic moment. In conclusion, many of our nanoparticle types are a bio-compatible innocent scaffold with unexpectedly favorable clearance.

Zwitterion-Coated Iron Oxide Nanoparticles: Surface Chemistry and Intracellular Uptake by Hepatocarcinoma (HepG2) Cells.

Nanoparticles (NPs) have received much attention in recent years for their diverse potential biomedical applications. However, the synthesis of NPs with desired biodistribution and pharmacokinetics is still a major challenge, with NP size and surface chemistry being the main factors determining the behavior of NPs in vivo. Here we report on the surface chemistry and in vitro cellular uptake of magnetic iron oxide NPs coated with zwitterionic dopamine sulfonate (ZDS). ZDS-coated NPs were compared to similar iron oxide NPs coated with PEG-like 2-[2-(2-methoxyethoxy)ethoxy]acetic acid (MEEA) to investigate how surface chemistry affects their in vitro behavior. ZDS-coated NPs had a very dense coating, guaranteeing high colloidal stability in several aqueous media and negligible interaction with proteins. Treatment of HepG2 cells with increasing doses (2.5-100 µg Fe/mL) of ZDS-coated iron oxide NPs had no effect on cell viability and resulted in a low, dose-dependent NP uptake, inferior than most reported data for the internalization of iron oxide NPs by HepG2 cells. MEEA-coated NPs were scarcely stable and formed micrometer-sized aggregates in aqueous media. They decreased cell viability for dose ≥50 µg Fe/mL, and were more efficiently internalized than ZDS-coated NPs. In conclusion, our data indicate that the ZDS layer prevented both aggregation and sedimentation of iron oxide NPs and formed a biocompatible coating that did not display any biocorona effect. The very low cellular uptake of ZDS-coated iron NPs can be useful to achieve highly selective targeting upon specific functionalization.

Single-agent Smac-mimetic compounds induce apoptosis in B chronic lymphocytic leukaemia (B-CLL).

Defective apoptosis is a hallmark of the progression of B chronic lymphocytic leukaemia (B-CLL). Smac-mimetics have been shown to induce apoptosis in several tumours. We describe the in vitro pro-apoptotic activity and regulation of the molecular pathway induced by new Smac-mimetics in B-CLL. The cytotoxic effect was significantly higher in B-CLL samples than in healthy controls. No significant synergistic effect was observed in combined treatment. In conclusion one of our compounds (Smac66), used as monotherapy and not in combination, is highly active against B-CLL cells thus suggesting a promising therapeutic potential as a new class of antileukemic drugs in haematology.

Colloidal stability of iron oxide nanocrystals coated with a PEG-based tetra-catechol surfactant.

Long-term colloidal stability of magnetic iron oxide nanoparticles (NPs) is an important goal that has not yet been fully achieved. To make an advance in our understanding of the colloidal stability of iron oxide NPs in aqueous media, we prepared NPs comprising a monodisperse (13 nm) iron oxide core coated with a PEG-based (PEG: polyethyleneglycol) surfactant. This consists of a methoxy-terminated PEG chain (MW = 5000 Da) bearing four catechol groups via a diethylenetriamine linker. The surfactant was grafted onto the nanocrystals by ligand exchange monitored by infrared spectroscopy. The colloidal stability of these nanoparticles was probed by monitoring the time evolution of the Z-average intensity-weighted radius R(h) and volume-weighted size distribution P(v) obtained from analysis of dynamic light scattering data. The nanoparticles showed no sign of aggregation for four months in deionized water at room temperature and also when subjected to thermal cycling between 25 and 75 °C. In 0.01 M PBS (phosphate buffered saline), aggregation (if any) is slow and partial; after 66 h, about 50% of NPs have not aggregated. Aggregation is more effective in 0.15 M NH(4)AcO buffer, where isolated particles are not observed after 66 h, and especially in acidic NH(4)AcO/AcOH buffer, where aggregation is complete within 1 h and precipitation is observed. The differing stability of the NPs in the above aqueous media is closely related to their ζ potential.

Structural insight into inhibitor of apoptosis proteins recognition by a potent divalent smac-mimetic.

Genetic alterations enhancing cell survival and suppressing apoptosis are hallmarks of cancer that significantly reduce the efficacy of chemotherapy or radiotherapy. The Inhibitor of Apoptosis Protein (IAP) family hosts conserved proteins in the apoptotic pathway whose over-expression, frequently found in tumours, potentiates survival and resistance to anticancer agents. In humans, IAPs comprise eight members hosting one or more structural Baculoviral IAP Repeat (BIR) domains. Cellular IAPs (cIAP1 and 2) indirectly inhibit caspase-8 activation, and regulate both the canonical and the non-canonical NF-κB signaling pathways. In contrast to cIAPs, XIAP (X chromosome-linked Inhibitor of Apoptosis Protein) inhibits directly the effector caspases-3 and -7 through its BIR2 domain, and initiator caspase-9 through its BIR3 domain; molecular docking studies suggested that Smac/DIABLO antagonizes XIAP by simultaneously targeting both BIR2 and BIR3 domains. Here we report analytical gel filtration, crystallographic and SAXS experiments on cIAP1-BIR3, XIAP-BIR3 and XIAP-BIR2BIR3 domains, alone and in the presence of compound 9a, a divalent homodimeric Smac mimetic. 9a is shown to bind two BIR domains inter- (in the case of two BIR3) and intra-molecularly (in the case of XIAP-BIR2BIR3), with higher affinity for cIAP1-BIR3, relative to XIAP-BIR3. Despite the different crystal lattice packing, 9a maintains a right handed helical conformation in both cIAP1-BIR3 and XIAP-BIR3 crystals, that is likely conserved in solution as shown by SAXS data. Our structural results demonstrate that the 9a linker length, its conformational degrees of freedom and its hydrophobicity, warrant an overall compact structure with optimal solvent exposure of its two active moieties for IAPs binding. Our results show that 9a is a good candidate for pre-clinical and clinical studies, worth of further investigations in the field of cancer therapy.

Homo- and heterodimeric Smac mimetics/IAP inhibitors as in vivo-active pro-apoptotic agents. Part I: Synthesis.

Novel pro-apoptotic, homo- and heterodimeric Smac mimetics/IAPs inhibitors based on the N-AVPI-like 4-substituted 1-aza-2-oxobicyclo[5.3.0]decane scaffold were prepared from monomeric structures connected through a head-head (8), tail-tail (9) or head-tail (10) linker. The selection of appropriate decorating functions for the scaffolds, and of rigid and flexible linkers connecting them, is described. The synthesis, purification and analytical characterization of each prepared dimer 8-10 is thoroughly described.

Dimeric Smac mimetics/IAP inhibitors as in vivo-active pro-apoptotic agents. Part II: Structural and biological characterization.

Novel pro-apoptotic, homodimeric and heterodimeric Smac mimetics/IAPs inhibitors connected through head-head (8), tail-tail (9) or head-tail linkers (10), were biologically and structurally characterized. In vitro characterization (binding to BIR3 and linker-BIR2-BIR3 domains from XIAP and cIAP1, cytotoxicity assays) identified early leads from each dimer family. Computational models and structural studies (crystallography, NMR, gel filtration) partially rationalized the observed properties for each dimer class. Tail-tail dimer 9a was shown to be active in a breast and in an ovary tumor model, highlighting the potential of dimeric Smac mimetics/IAP inhibitors based on the N-AVPI-like 4-substituted 1-aza-2-oxobicyclo[5.3.0]decane scaffold as potential antineoplastic agents.

Novel second mitochondria-derived activator of caspases (Smac) mimetic compounds sensitize human leukemic cell lines to conventional chemotherapeutic drug-induced and death receptor-mediated apoptosis.

The Inhibitor of Apoptosis Proteins (IAPs) are important regulators of programmed cell death. XIAP is the most potent among them and is over-expressed in several hematological malignancies. Its activity is endogenously antagonized by SMAC/DIABLO, and also by small molecules mimicking Smac that can induce apoptosis in tumor cells. Here we describe the activity of 56 newly synthesized Smac-mimetics in human leukemic cell lines and normal CD34(+) progenitor cells. Our compounds bind to XIAP with high affinity, reduce the levels of cIAP1 and are cytotoxic at nanomolar or low micromolar concentrations. Furthermore, the Smac-mimetics synergize with Cytarabine, Etoposide and especially with TRAIL in combination treatments. Apoptosis activation was clearly detectable by the occurrence of sub G(1) apoptotic peak and the accumulation of cleaved PARP, caspase 8 and caspase 3. Interestingly, the down-regulation of XIAP sensitized Jurkat cells to drugs too, confirming the role of this protein in drug-resistance. In conclusion, while being very active in leukemic cells, our Smac-mimetics have modest effects on normal hematopoietic progenitors, suggesting their promising therapeutic potential as a new class of anticancer drugs in onco-hematology, particularly when combined with TRAIL, to overcome the resistance of cancer cells.

Rational design, synthesis and characterization of potent, non-peptidic Smac mimics/XIAP inhibitors as proapoptotic agents for cancer therapy.

Novel proapoptotic Smac mimics/IAPs inhibitors have been designed, synthesized and characterized. Computational models and structural studies (crystallography, NMR) have elucidated the SAR of this class of inhibitors, and have permitted further optimization of their properties. In vitro characterization (XIAP BIR3 and linker-BIR2-BIR3 binding, cytotox assays, early ADMET profiling) of the compounds has been performed, identifying one lead for further in vitro and in vivo evaluation.

Structural basis for bivalent Smac-mimetics recognition in the IAP protein family.

XIAP is an apoptotic regulator protein that binds to the effector caspases -3 and -7 through its BIR2 domain, and to initiator caspase-9 through its BIR3 domain. Molecular docking studies suggested that Smac-DIABLO may antagonize XIAP by concurrently targeting both BIR2 and BIR3 domains; on this basis bivalent Smac-mimetic compounds have been proposed and characterized. Here, we report the X-ray crystal structure of XIAP-BIR3 domain in complex with a two-headed compound (compound 3) with improved efficacy relative to its monomeric form. A small-angle X-ray scattering study of XIAP-BIR2BIR3, together with fluorescence polarization binding assays and compound 3 cytotoxicity tests on HL60 leukemia cell line are also reported. The crystal structure analysis reveals a network of interactions supporting XIAP-BIR3/compound 3 recognition; moreover, analytical gel-filtration chromatography shows that compound 3 forms a 1:1 stoichiometric complex with a XIAP protein construct containing both BIR2 and BIR3 domains. On the basis of the crystal structure and small-angle X-ray scattering, a model of the same BIR2-BIR3 construct bound to compound 3 is proposed, shedding light on the ability of compound 3 to relieve XIAP inhibitory effects on caspase-9 as well as caspases -3 and -7. A molecular modeling/docking analysis of compound 3 bound to cIAP1-BIR3 domain is presented, considering that Smac-mimetics have been shown to kill tumor cells by inducing cIAP1 and cIAP2 ubiquitination and degradation. Taken together, the results reported here provide a rationale for further development of compound 3 as a lead in the design of dimeric Smac mimetics for cancer treatment.

Targeting the X-linked inhibitor of apoptosis protein through 4-substituted azabicyclo[5.3.0]alkane smac mimetics. Structure, activity, and recognition principles.

The X-linked inhibitor of apoptosis protein (XIAP) is overexpressed in several malignant cells where it prevents apoptosis by binding to, and blocking, the activation of caspase-3, -7, and -9. Human XIAP (479 residues) is composed of three tandem-repeated baculoviral IAP repeat (BIR) domains (BIR1-3), and by a C-terminal RING domain. Smac-DIABLO [second mitochondria-derived activator of caspases (Smac)-direct IAP binding protein with low pI (DIABLO)], the natural antagonist of XIAP, binds through its N-terminal sequence AVPI to the same surface groove, in the BIR domains, that binds caspases. Synthetic compounds mimicking such tetrapeptide motif effectively block the interaction between IAP and active caspases, thus triggering apoptosis. Peptidomimetics based on an azabicyclo[x.y.0]alkane scaffolds, have been shown to bind the BIR3 domain of XIAP with micromolar to nanomolar affinities, thus presenting attractive features for drug lead optimization. Here we report a study on three newly synthesized Smac mimetics, which have been characterized in their complexes with XIAP BIR3 domain through X-ray crystallography and molecular modelling/docking simulations. Based on analysis of the crystal structures, we show that specific substitutions at the 4-position of the azabicyclo[5.3.0]alkane scaffold results in sizeable effects on the peptidomimetic-BIR3 domain affinity. By means of functional, biophysical and simulative approaches we also propose that the same Smac mimetics can bind XIAP BIR2 domain at a location structurally related to the BIR3 domain AVPI binding groove. Details of the XIAP-Smac mimetic recognition principles highlighted by this study are discussed in light of the drug-like profile of the three (potentially proapoptotic) compounds developed that show improved performance in ADMET (adsorption, distribution, metabolism, excretion and toxicity) tests.