Complexing the Marine Sesquiterpene Euplotin C by Means of Cyclodextrin-Based Nanosponges: A Preliminary Investigation.

Euplotin C is a sesquiterpene of marine origin endowed with significant anti-microbial and anti-tumor properties. Despite the promising functional profile, its progress as a novel drug candidate has failed so far, due to its scarce solubility and poor stability in aqueous media, such as biological fluids. Therefore, overcoming these limits is an intriguing challenge for the scientific community. In this work, we synthesized ß-cyclodextrin-based nanosponges and investigated their use as colloidal carriers for stably complex euplotin C. Results obtained proved the ability of the carrier to include the natural compound, showing remarkable values of both loading efficiency and capacity. Moreover, it also allowed us to preserve the chemical structure of the loaded compound, which was recovered unaltered once extracted from the complex. Therefore, the use of ß-cyclodextrin-based nanosponges represents a viable option to vehiculate euplotin C, thus opening up its possible use as pharmacologically active compound.

Synthetic mycomelanin thin films as emergent bio-inspired interfaces controlling the fate of embryonic stem cells.

The fungal pathways of melanin synthesis have so far been little considered as a source of bio-inspiration in the field of functional materials, despite the interesting properties exhibited by Ascomycetes melanins from 1,8-dihydroxynaphthalene (1,8-DHN), including the ability to shield organisms from ionizing radiation. Herein, the processing techniques and characterizations of mycomelanin thin films obtained from the solid state polymerization of 1,8-DHN is reported for the first time. Overall, the results highlighted the role of synthetic mycomelanin thin films as a prototype of next generation bioinspired interfaces featuring high structural regularity and ultrasmooth morphology, high robustness against peroxidative bleaching and adhesion under water conditions, good biocompatibility and unprecedented effects in inducing the spontaneous differentiation of embryonic stem cells prevalently towards the endodermal lineages in the absence of added factors. These data open up new avenues towards the applications of this biomaterial in the fields of tissue engineering and regenerative medicine.

Oxy-imino saccharidic derivatives as a new structural class of aldose reductase inhibitors endowed with anti-oxidant activity.

Aldose reductase is a key enzyme in the development of long term diabetic complications and its inhibition represents a viable therapeutic solution for people affected by these pathologies. Therefore, the search for effective aldose reductase inhibitors is a timely and pressing challenge. Herein we describe the access to a novel class of oxyimino derivatives, obtained by reaction of a 1,5-dicarbonyl substrate with O-(arylmethyl)hydroxylamines. The synthesised compounds proved to be active against the target enzyme. The best performing inhibitor, compound (Z)-8, proved also to reduce both cell death and the apoptotic process when tested in an in vitro model of diabetic retinopathy made of photoreceptor-like 661w cell line exposed to high-glucose medium, counteracting oxidative stress triggered by hyperglycaemic conditions.

Imidazo[1,2-a]pyridine Derivatives as Aldehyde Dehydrogenase Inhibitors: Novel Chemotypes to Target Glioblastoma Stem Cells.

Glioblastoma multiforme (GBM) is the deadliest form of brain tumor. It is known for its ability to escape the therapeutic options available to date thanks to the presence of a subset of cells endowed with stem-like properties and ability to resist to cytotoxic treatments. As the cytosolic enzyme aldehyde dehydrogenase 1A3 turns out to be overexpressed in these kinds of cells, playing a key role for their vitality, treatments targeting this enzyme may represent a successful strategy to fight GBM. In this work, we describe a novel class of imidazo[1,2-a]pyridine derivatives as aldehyde dehydrogenase 1A3 inhibitors, reporting the evidence of their significance as novel drug candidates for the treatment of GBM. Besides showing an interesting functional profile, in terms of activity against the target enzyme and selectivity toward highly homologous isoenzymes, representative examples of the series also showed a nanomolar to picomolar efficacy against patient-derived GBM stem-like cells, thus proving the concept that targeting aldehyde dehydrogenase might represent a novel and promising way to combat GBM by striking its ability to divide immortally.

pH-Responsive Carboxymethylcellulose Nanoparticles for 68Ga-WBC Labeling in PET Imaging.

Carboxymethylcellulose (CMC) is a well-known pharmaceutical polymer, recently gaining attention in the field of nanomedicine, especially as a polyelectrolyte agent for the formation of complexes with oppositely charged macromolecules. Here, we report on the application of pH-sensitive pharmaceutical grade CMC-based nanoparticles (NP) for white blood cells (WBC) PET imaging. In this context and as an alternative to 99mTc-HMPAO SPECT labeling, the use of 68Ga3+ as PET radionuclide was investigated since, at early time points, it could provide the greater spatial resolution and patient convenience of PET tomography over SPECT clinical practices. Two operator-friendly kit-type formulations were compared, with the intention of radiolabeling within a short time (10 min), under mild conditions (physiological pH, room temperature) and in agreement with the actual clinically applied guidelines. NP were labeled by directly using 68Ga3+ eluted in HCL 0.05 N, from hospital suited 68Ge/68Ga generator and in absence of chelator. The first kit type approach involved the application of 68Ga3+ as an ionotropic gelation agent for in-situ forming NP. The second kit type approach concerned the re-hydration of a proper freeze-dried injectable NP powder. pH-sensitive NP with 250 nm average diameter and 80% labeling efficacy were obtained. The NP dispersant medium, including a cryoprotective agent, was modulated in order to optimize the Zeta potential value (-18 mV), minimize the NP interaction with serum proteins and guarantee a physiological environment for WBC during NP incubation. Time-dependent WBC radiolabeling was correlated to NP uptake by using both confocal and FT-IR microscopies. The ready to use lyophilized NP formulation approach appears promising as a straightforward 68Ga-WBC labeling tool for PET imaging applications.

Synthesis and investigation of polyhydroxylated pyrrolidine derivatives as novel chemotypes showing dual activity as glucosidase and aldose reductase inhibitors.

Diabetes is a multi-factorial disorder that should be treated with multi-effective compounds. Here we describe the access to polyhydroxylated pyrrolidines, belonging to the d-gluco and d-galacto series, through aminocyclization reactions of two differentially protected d-xylo-hexos-4-ulose derivatives. The prepared compounds proved to inhibit both alpha-glucosidase, responsible for the emergence of hyperglycemic spikes, and aldose reductase, accountable for the development of abnormalities in diabetic tissues. Accordingly, they show the dual inhibitory profile deemed as ideal for diabetes treatment. Significantly, compound 17b reduced the process of cell death and restored the physiological levels of oxidative stress when tested in the photoreceptor-like 661w cell line, thus proving to be effective in an in vitro model of diabetic retinopathy.

Dual Kit/Aur Inhibitors as Chemosensitizing Agents for the Treatment of Melanoma: Design, Synthesis, Docking Studies and Functional Investigation.

Melanoma is the most serious form of skin cancer but its medication is still far from being safe and thoroughly effective. The search of novel therapeutic approaches represents therefore a health emergency to push through eagerly. In this study, we describe a novel class of dual c-Kit/Aur inhibitors, characterized by a 1,2,4-triazole core and developed by a structure-based optimization of a previously developed hit, and report the evidence of their significance as drug candidates for the treatment of melanoma. Compound 6a, merging the best inhibitory profile against the target kinases, showed anti-proliferative efficacy against the human melanoma cell lines A2058, expressing the BRAF V600D mutation, and WM266-4, expressing BRAF V600E. Significantly, it displayed also a highly synergistic profile when tested in combination with vemurafenib, thus proving its efficacy not only per se but even in a combination therapy, which is nowadays acknowledged as the cornerstone approach of the forthcoming tumour management.

Combined Use of Scanning Electron Microscopy-Energy-Dispersive X-ray Spectroscopy and Fourier Transform Infrared Imaging Coupled with Principal Component Analysis in the Study of Ancient Egyptian Papyri.

This paper reports the results of a spectroscopic study on the chemistry of some small papyrus fragments arising from three Egyptian excavation sites. The aim of this investigation was to verify the possibility to identify significant differences in ancient Egyptian papyri using noninvasive analytical methods, that is, ATR (attenuated total reflection)-FTIR (Fourier transform infrared) spectroscopic imaging and scanning electron microscopy-energy-dispersive X-ray spectroscopy. Differences in both lignin and cellulose compositions, which are difficult to detect with traditional FTIR and FTIR imaging spectral analysis, were revealed by the multivariate approach, and the second derivative spectroscopy was applied to enhance the spectrum resolution. Thus, it has been possible to recognize a fragment characterized by the presence of lead and of long chain aliphatic organic compound, which are not present in the other fragments, in the ink region. These data show not only that the combination of these techniques can provide important chemical information, such as to evidence the different compositions and manufacture of each papyrus, but also that metal inks were probably used also in ancient Egyptian papyri.

Acid Derivatives of Pyrazolo[1,5-a]pyrimidine as Aldose Reductase Differential Inhibitors.

Aldose reductase (AKR1B1), the key enzyme of the polyol pathway, plays a crucial role in the development of long-term complications affecting diabetic patients. Nevertheless, the expedience of inhibiting this enzyme to treat diabetic complications has failed, due to the emergence of side effects from compounds under development. Actually AKR1B1 is a Janus-faced enzyme which, besides ruling the polyol pathway, takes part in the antioxidant defense mechanism of the body. In this work we report the evidence that a class of compounds, characterized by a pyrazolo[1,5-a]pyrimidine core and an ionizable fragment, modulates differently the catalytic activity of the enzyme, depending on the presence of specific substrates such as sugar, toxic aldehydes, and glutathione conjugates of toxic aldehydes. The study stands out as a systematic attempt to generate aldose reductase differential inhibitors (ARDIs) intended to target long-term diabetic complications while leaving unaltered the detoxifying role of the enzyme.

Challenging clinically unresponsive medullary thyroid cancer: Discovery and pharmacological activity of novel RET inhibitors.

It is now known that "gain of function" mutations of RET (REarranged during Transfection) kinase are specific and key oncogenic events in the onset of thyroid gland cancers such as the Medullary Thyroid Carcinoma (MTC). Although a number of RET inhibitors exist and are capable of inhibiting RET variants, in which mutations are outside the enzyme active site, the majority becomes dramatically ineffective when mutations are within the protein active site (V804L and V804M). Pursuing a receptor-based virtual screening against the kinase domain of RET, we found that compound 5 is able to inhibit efficiently both wild type and V804L mutant RET. Compound 5 was able to significantly reduce proliferation of both commercially available TT cell lines and surgical thyroid tissues obtained from patients with MTC and displayed a suitable drug-like profile, thus standing out as a promising candidate for further development towards the treatment of clinically unresponsive MTC.

Chiral ionic liquids supported on natural sporopollenin microcapsules.

Supported chiral ionic liquids (SILs) were prepared choosing the starting material for the ionic liquid part from the enantiopure stock of the chiral pool (monoterpenoids and an amino acid) and the sporopollenin as an environmentally friendly support. Sporopollenins are microcapsules with naturally well-defined sizes and shapes that can be obtained from pollen grains after removal of the internal cytoplasm and the second shell layer. As thermally stable organic biocompatible structures, sporopollenins have attracted increasing interest in recent years for several applications. Herein, bio-based ILs were anchored onto the surface of sporopollenins obtained from the pollen of Populus deltoides, selected as a model pollen grain. These new structures, which present an external positively charged shell, were characterized by physico-chemical techniques (ATR-FTIR, TGA, SEM, EDX, and solid-state 13C NMR). A metathesis reaction was also performed on selected bio-based IL modified sporopollenins, demonstrating the possibility to switch the surface properties by exploiting well-known IL chemistry.

Synthesis and Functional Evaluation of Novel Aldose Reductase Inhibitors Bearing a Spirobenzopyran Scaffold.

BACKGROUND: Aldose reductase, the first enzyme of the polyol pathway, is the key determinant for the pathogenesis of long term diabetic complications. Accordingly, its inhibition represents the major therapeutic strategy to treat this kind of pathologies. OBJECTIVES: In this work we describe the synthesis and the functional evaluation of a number of spiro-oxazolidinone and spiro-morpholinone acetic acid derivatives, and their benzyloxy analogs, developed as aldose reductase inhibitors. RESULTS: Most of them proved to inhibit the target enzyme, showing IC50 values in the micromolar/low micromolar range. SARs observed among the three different series allowed to highlight their key pharmacophoric elements, thus creating sound basis for the design of novel and more effective inhibitors. CONCLUSION: Although further substitution patterns are needed, the novel compounds here proposed represent a good starting point for the development of novel and effective ARIs.

Cyclodextrin-based nanosponges for the targeted delivery of the anti-restenotic agent DB103: A novel opportunity for the local therapy of vessels wall subjected to percutaneous intervention.

Nano-sized colloidal carriers represent innovative drug delivery systems, as they allow a targeted and prolonged release of poorly water-soluble drugs, improving their bioavailability and modifying their pharmacokinetic parameters. In this work we describe cyclodextrin-based nanosponges, obtained through polimerization of ß-cyclodextrin with diphenyl carbonate as the cross-linking agent, loaded with a novel multi-effective heterocyclic compound, DB103, able to regulate key cellular events involved in the remodelling of vessels wall. Fabrication and drug-loading procedures, as well as physical-chemical characterization and drug-release profile of the novel colloidal system are reported. Results achieved demonstrate the ability of nanosponges to enclose efficiently the target drug and release it slowly and continuously, thus suggesting the exploitability of the novel system for the local therapy of vessels wall subjected to percutaneous intervention.

N-(Aroyl)-N-(arylmethyloxy)-α-alanines: Selective inhibitors of aldose reductase.

Aldose reductase (ALR2), a NADPH-dependent reductase, is the first and rate-limiting enzyme of the polyol pathway of glucose metabolism and is implicated in the pathogenesis of secondary diabetic complications. In the last decades, this enzyme has been targeted for inhibition but despite the numerous efforts made to identify potent and safe ALR2 inhibitors, many clinical candidates have been a failure. For this reason the research of new ALR2 inhibitors highly effective, selective and with suitable pharmacokinetic properties is still of great interest. In this paper some new N-(aroyl)-N-(arylmethyloxy)alanines have been synthesized and tested for their ability to inhibit ALR2. Some of the synthesized compounds exhibit IC50 in the low micromolar range and all have proved to be highly selective towards ALR2. The N-(aroyl)-N-(arylmethyloxy)-α-alanines are a promising starting point for the development of new ALR2 selective drugs with the aim of delaying the onset of diabetic complications.

FOXD1-ALDH1A3 Signaling Is a Determinant for the Self-Renewal and Tumorigenicity of Mesenchymal Glioma Stem Cells.

Glioma stem-like cells (GSC) with tumor-initiating activity orchestrate the cellular hierarchy in glioblastoma and engender therapeutic resistance. Recent work has divided GSC into two subtypes with a mesenchymal (MES) GSC population as the more malignant subtype. In this study, we identify the FOXD1-ALDH1A3 signaling axis as a determinant of the MES GSC phenotype. The transcription factor FOXD1 is expressed predominantly in patient-derived cultures enriched with MES, but not with the proneural GSC subtype. shRNA-mediated attenuation of FOXD1 in MES GSC ablates their clonogenicity in vitro and in vivo Mechanistically, FOXD1 regulates the transcriptional activity of ALDH1A3, an established functional marker for MES GSC. Indeed, the functional roles of FOXD1 and ALDH1A3 are likely evolutionally conserved, insofar as RNAi-mediated attenuation of their orthologous genes in Drosophila blocks formation of brain tumors engineered in that species. In clinical specimens of high-grade glioma, the levels of expression of both FOXD1 and ALDH1A3 are inversely correlated with patient prognosis. Finally, a novel small-molecule inhibitor of ALDH we developed, termed GA11, displays potent in vivo efficacy when administered systemically in a murine GSC-derived xenograft model of glioblastoma. Collectively, our findings define a FOXD1-ALDH1A3 pathway in controling the clonogenic and tumorigenic potential of MES GSC in glioblastoma tumors. Cancer Res; 76(24); 7219-30. ©2016 AACR.

1,2-Benzisothiazole Derivatives Bearing 4-, 5-, or 6-Alkyl/arylcarboxamide Moieties Inhibit Carbonic Anhydrase Isoform IX (CAIX) and Cell Proliferation under Hypoxic Conditions.

Three novel series of 1,2-benzisothiazole derivatives have been developed as inhibitors of carbonic anhydrase isoform IX. Compounds 5c and 5j, tested in vitro on the human colon cell line HT-29, blocked the growth of cells cultured under chemically induced hypoxic conditions, displaying a specific activity against cancer cells characterized by CAIX up-regulation. Moreover, a synergistic activity of 5c with SN-38 (the active metabolite of irinotecan) and 5-fluorouracil on cell proliferation under hypoxic conditions was demonstrated.

A Series of COX-2 Inhibitors Endowed with NO-Releasing Properties: Synthesis, Biological Evaluation, and Docking Analysis.

Herein we report the synthesis, biological evaluation, and docking analysis of a class of cyclooxygenase-2 (COX-2) inhibitors with nitric oxide (NO)-releasing properties. In an earlier study, a number of selective COX-2 inhibitors/NO donors were developed by conjugating a diarylpyrrole scaffold endowed with selective COX-2 inhibitory properties with various nitrooxyalkyl side chains such as esters, α-amino esters, amides, α-amino amides, ethers, ß-amino ethers, inverse esters, and amides. These candidates were found to have high in vitro potencies (COX-2 inhibition at 10 µm: ≥96 %), great efficacy in determining NO-vasorelaxing responses, and good antinociceptive activity in an abdominal writhing test. Among the compounds synthesized in the present work, derivative 2 b [2-(2-(1-(3-fluorophenyl)-2-methyl-5-(4-sulfamoylphenyl)-1H-pyrrol-3-yl)acetamido)ethyl nitrate] showed particularly outstanding activity, with efficacy similar to that of celecoxib even at very low concentrations.

Nanostructured ultra-thin patches for ultrasound-modulated delivery of anti-restenotic drug.

This work aims to demonstrate the possibility to fabricate ultra-thin polymeric films loaded with an anti-restenotic drug and capable of tunable drug release kinetics for the local treatment of restenosis. Vascular nanopatches are composed of a poly(lactic acid) supporting membrane (thickness: ~250 nm) on which 20 polyelectrolyte bilayers (overall thickness: ~70 nm) are alternatively deposited. The anti-restenotic drug is embedded in the middle of the polyelectrolyte structure, and released by diffusion mechanisms. Nanofilm fabrication procedure and detailed morphological characterization are reported here. Barium titanate nanoparticles (showing piezoelectric properties) are included in the polymeric support and their role is investigated in terms of influence on nanofilm morphology, drug release kinetics, and cell response. Results show an efficient drug release from the polyelectrolyte structure in phosphate-buffered saline, and a clear antiproliferative effect on human smooth muscle cells, which are responsible for restenosis. In addition, preliminary evidences of ultrasound-mediated modulation of drug release kinetics are reported, thus evaluating the influence of barium titanate nanoparticles on the release mechanism. Such data were integrated with quantitative piezoelectric and thermal measurements. These results open new avenues for a fine control of local therapies based on smart responsive materials.

Pyrazolopyrimidine Derivatives as Antineoplastic Agents: with a Special Focus on Thyroid Cancer.

Tyrosine kinase inhibitors (TKIs) are molecules that compete with ATP on tyrosine kinase receptors (TKRs), blocking tyrosine kinase (TK) activation and then oncogenic pathways; they have been studied, and some of them are right approved for the treatment of many types of cancer. Among TKIs, one of the most explored chemical template is the pyrazolo[3,4-d]pyrimidine (PP) heterocyclic core, which proved to be a useful scaffold for the obtainment of effective compounds. Actually, derivatives belonging to this structural class show a large spectrum of activity, thus standing out as multi-target agents. Different PP compounds have been shown to act as: a) ABL inhibitors and antiproliferative agents against human leukemia cell lines; b) Src kinase inhibitors in neuroblastoma, medulloblastoma and osteosarcoma; c) Phospholipase D inhibitors in different neoplasias; d) Urokinase plasminogen activator inhibitors, in breast cancer. In thyroid cancer (TC), PP1 and PP2 (inhibitors of RET, Hck, lck, and fynT kinases, and a good inhibitor of c-Src and platelet-derived growth factor receptor) showed antineoplastic actvity in human papillary TC cell lines that carry spontaneous RET/PTC1 rearrangements. More recently, new derivatives, (R)-1-phenethyl-N-(1-phenylethyl)-1H-pyrazolo[3,4-d]pyrimidin-4- amine, namely, CLM3 and CLM29, have been demonstrated to exert a multiple signal transduction inhibition (including the RET-TK, BRAF, EGFR, and with antiangiogenic activity), showing antineoplastic activity, in vitro and in vivo, in papillary dedifferentiated, medullary and anaplastic TC. These data have shown the antineoplastic activity of PP in different neoplasias, opening the way to a future clinical evaluation in human cancers.

Investigation of new 2-aryl substituted Benzothiopyrano[4,3-d]pyrimidines as kinase inhibitors targeting vascular endothelial growth factor receptor 2.

Vascular Endothelial Growth Factor (VEGF) pathway has emerged as one of the most important positive modulators of Angiogenesis, a central process implicated in tumour growth and metastatic dissemination. This led to the design and development of anti-VEGF monoclonal antibodies and small-molecule ATP-competitive VEGFR-inhibitors. In this study, we describe the synthesis and the biological evaluation of novel 2-aryl substituted benzothiopyrano-fused pyrimidines 1a-i, 2a-i and 3a-i. The ability of the compounds to target the VEGF pathway was determined in vitro exploiting the compounds' antiproliferative efficacy against HUVEC cells. The VEGFR-2 inhibition was confirmed by enzymatic assays on recombinant human kinase insert domain receptor (KDR), by cell-based phospho-VEGFR-2 inhibition assays, and by ex vivo rat aortic ring tests. The selectivity profile of the best performing derivatives belonging to series 2 was further explored combining modeling studies and additional assays in a panel of human cell lines and other kinases.