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Growth factors and cytokines released in the lung cancer microenvironment promote an epithelial-to-mesenchymal transition (EMT) that sustains the progression of neoplastic diseases. TGFβ is one of the most powerful inducers of this transition, as it induces overexpression of the fibronectin receptor, αvβ6 integrin, in cancer cells which, in turn, is strongly associated with EMT. Thus, αvβ6 integrin receptors may be exploited as a target for the selective delivery of anti-tumor agents. We introduce three novel synthesized conjugates, in which a selective αvβ6 receptor ligand is linked to nintedanib, a potent kinase inhibitor used to treat advanced adenocarcinoma lung cancer in clinics. The αvβ6 integrin ligand directs nintedanib activity to the target cells of the tumor microenvironment, avoiding the onset of negative side effects in normal cells. We found that the three conjugates inhibit the adhesion of cancer cells to fibronectin in a concentration-dependent manner and that αvβ6-expressing cells internalized the conjugated compounds, thus permitting nintedanib to inhibit 2D and 3D cancer cell growth and suppress the clonogenic ability of the EMT phenotype as well as intervening in other aspects associated with the EMT transition. These results highlight αvβ6 receptors as privileged access points for dual-targeting molecular conjugates engaged in an efficient and precise strategy against non-small cell lung cancer.

Though the bulk of the COVID-19 pandemic is behind, the search for effective and safe anti-SARS-CoV-2 drugs continues to be relevant. A highly pursued approach for antiviral drug development involves targeting the viral spike (S) protein of SARS-CoV-2 to prevent its attachment to the cellular receptor ACE2. Here, we exploited the core structure of polymyxin B, a naturally occurring antibiotic, to design and synthesize unprecedented peptidomimetics (PMs), intended to target contemporarily two defined, non-overlapping regions of the S receptor-binding domain (RBD). Monomers 1, 2, and 8, and heterodimers 7 and 10 bound to the S-RBD with micromolar affinity in cellfree surface plasmon resonance assays (KD ranging from 2.31 µM to 2.78 µM for dimers and 8.56 µM to 10.12 µM for monomers). Although the PMs were not able to fully protect cell cultures from infection with authentic live SARS-CoV-2, dimer 10 exerted a minimal but detectable inhibition of SARS-CoV-2 entry in U87.ACE2+ and A549.ACE2.TMPRSS2+ cells. These results validated a previous modeling study and provided the first proof-of-feasibility of using medium-sized heterodimeric PMs for targeting the S-RBD. Thus, heterodimers 7 and 10 may serve as a lead for the development of optimized compounds, which are structurally related to polymyxin, with improved S-RBD affinity and anti-SARS-CoV-2 potential. ispartof: International Journal Of Molecular Sciences vol:24 issue:10 status: Published online

In recognition of the key role played by integrins in several life-threatening dysfunctions, the search for novel small-molecule probes that selectively recognize these surface receptors is still open and widely pursued. Inspired by previously established aminoproline (Amp)-RGD based cyclopeptidomimetics with attracting αVβ3 integrin affinity and selectivity, the design and straightforward synthesis of 18 new AmpRGD chemotypes bearing additional structural variants were herein implemented, to shift toward peptide-like αVβ6 integrin targeted binders. The ligand competence of the synthesized products toward αVβ6 was evaluated in competitive binding assays on isolated receptors, and αVβ6/αVβ3 selectivity was determined for a subgroup of compounds, resulting in the identification of four very promising candidates. SAR considerations and docking simulations allowed us to appreciate the key structural features responsible for the observed activity.

α

Growth factors and cytokines released in the lung cancer microenvironment promote an epithelial-to-mesenchymal transition (EMT) that sustains the progression of neoplastic diseases. TGFβ is one of the most powerful inducers of this transition, as it induces overexpression of the fibronectin receptor, αvβ6 integrin, in cancer cells which, in turn, is strongly associated with EMT. Thus, αvβ6 integrin receptors may be exploited as a target for the selective delivery of anti-tumor agents. We introduce three novel synthesized conjugates, in which a selective αvβ6 receptor ligand is linked to nintedanib, a potent kinase inhibitor used to treat advanced adenocarcinoma lung cancer in clinics. The αvβ6 integrin ligand directs nintedanib activity to the target cells of the tumor microenvironment, avoiding the onset of negative side effects in normal cells. We found that the three conjugates inhibit the adhesion of cancer cells to fibronectin in a concentration-dependent manner and that αvβ6-expressing cells internalized the conjugated compounds, thus permitting nintedanib to inhibit 2D and 3D cancer cell growth and suppress the clonogenic ability of the EMT phenotype as well as intervening in other aspects associated with the EMT transition. These results highlight αvβ6 receptors as privileged access points for dual-targeting molecular conjugates engaged in an efficient and precise strategy against non-small cell lung cancer.

Background The high glycolytic activity of multiple myeloma (MM) cells is the rationale for use of Positron Emission Tomography (PET) with 18F-fluorodeoxyglucose ([18F]FDG) to detect both bone marrow (BM) and extramedullary disease. However, FDG-PET has some limitations, since there is a portion of MM patients who are negative, and new tracers are actively searched. Glutamine (Gln) addiction has been recently described as a peculiar metabolic feature of MM cells. Yet, the possible exploitation of Gln as a PET tracer in MM has never been assessed thus far and is investigated in this study. Methods We have firstly synthesized enantiopure (2S,4R)-4-fluoroglutamine (4-FGln) and validated it as a Gln transport analogue in human MM cell lines, comparing its uptake with that of 3H-labelled Gln. We then radiosynthesized of [18F]4-FGln and tested its uptake for MM detection by PET in two different murine models and we checked the effect of Bortezomib treatment. Results Both [18F]4-FGln and [18F]FDG clearly identified the spleen as site of MM cell colonization in C57BL/6 mice challenged with syngeneic Vk12598 cells and assessed by PET. NOD.SCID mice subcutaneously injected with human MM JJN3 cells, showed high values of both [18F]4-FGln and [18F]FDG uptake to T/M: 2.3 ± 0.3 and 7.1 ± 2.6, respectively. With both [18F]4-FGln and [18F]FDG radiotracers, BOR treated animals displayed Standard Uptake Values (SUV) mean values significantly lower than controls at post treatment PET. However, [18F]4-FGln better correlated with the tumour volume in NOD.SCID mice, and a reduction of glutaminolytic, but not of glycolytic, tumour volume was evident in mice showing the highest response to Bortezomib. Conclusion Our data indicate that [18F](2S,4R)-4-FGln is a new PET tracer in pre-clinical MM models, yielding a rationale to design studies in MM patients.

In recognition of the key role played by integrins in several life-threatening dysfunctions, the search for novel small-molecule probes that selectively recognize these surface receptors is still open and widely pursued. Inspired by previously established aminoproline (Amp)-RGD based cyclopeptidomimetics with attracting α

The discovery of chemical methods enabling the construction of carbocycle-fused uracils which embody a three-dimensional and functional-group-rich architecture is a useful tool in medicinal chemistry oriented synthesis. In this work, an unprecedented amine-catalyzed [4+2] cross-cycloaddition is documented; it involves remotely enolizable 6-methyluracil-5-carbaldehydes and β-aryl enals, and chemoselectively produces two novel bicyclic and tricyclic fused uracil chemotypes in good yields with a maximum level of enantiocontrol. In-depth mechanistic investigations and control experiments support an intriguing homo-synergistic organocatalytic approach, where the same amine organocatalyst concomitantly engages both aldehyde partners in a stepwise eliminative [4+2] cycloaddition, whose vinylogous iminium ion intermediate product may diverge-depending upon conditions-to either bicyclic targets by hydrolysis or tricyclic products by a second homo-synergistic trienamine-mediated stepwise [4+2] cycloaddition.

Effective and selective targeting of the αV β3 integrin subtype is of high relevance in cancer research for the development of therapeutic systems with improved efficacy and of diagnostic imaging probes. We report here a new class of highly selective, αV β3 -targeted gold nanoparticles (AuNPs), which carry cyclic 4-aminoproline-RGD semipeptides (cAmpRGD) as the targeting moiety immobilized at low surface density on the poly(ethylene glycol) (PEG)-based nanoparticle coating. We show that these nanoparticles are potent inhibitors of the integrin-mediated melanoma tumor cell adhesion to vitronectin and are selectively internalized via receptor-mediated endocytosis. Furthermore, we have developed bifunctional cAmpRGD-functionalized AuNPs by conjugation of a fluorophore (FAM or TAMRA) to a separate set of reactive groups on the PEG-based coating. These bifunctional AuNPs not only recapitulate the binding properties of cAmpRGD-AuNPs but also can be visualized via confocal laser microscopy, allowing direct observation of nanoparticle internalization. The peculiar molecular design of these nanoparticles and their precisely defined architecture at the molecular level accounts for their selective integrin binding with very low nonspecific background.

Glutamine (Gln) addiction has been recently described as a typical metabolic feature of MM by our group. In order to sustain high Gln demand, MM cells upregulate the expression of the Gln transporters ASCT2, SNAT1 and LAT1 and are characterized by fast Gln uptake. Currently, 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) is the standard tracer for the positron emission tomography/computed tomography (PET/CT) scan to detect both medullary and extramedullary disease in MM patients. However, FDG-PET has some limitations, since there is a portion of MM patients who are false-negative. For these reasons, the aim of our study is to characterize Gln metabolism in vivo in different MM mouse models and investigate the possible use of Gln as a PET tracer in MM. To this purpose, we have firstly synthesized enantiopure (2 S,4 R)-4-fluoroglutamine (4-FGln) and validated it in human MM cell lines (RPMI8226 and JJN3) comparing its uptake with that of 3H-labelled Gln. Both Gln and 4-FGln were actively accumulated by MM cells. Inhibition analysis revealed that ASCT2 was the major entry route of both compounds. We then carried out the radiosynthesis of [18F]4-FGln and tested its uptake for MM detection by PET in comparison with [18F]FDG. Firstly, we characterized [18F]4-FGln kinetics using immunodeficient NOD/SCID mice subcutaneously injected with JJN3. In this xenograft model, [18F]FDG- and [18F]4-FGln-PET were performed after plasmacytomas became palpable. In tumor, [18F]4-FGln uptake peaked at 25 min slowly declining thereafter. The Logan plot confirmed linearity starting at 15 min from injection, consistent with largely reversible tracer exchange. Thereafter, [18F]FDG and [18F]4-FGln distribution were assessed in both syngeneic (murine Vk12598 cells) and xenograft ( human JJN3 cells) MM models. C57BL/6 mice were injected intravenously with Vk12598 cells obtained from transgenic Vk*MYC mice. Mice were monitored weekly for M-spike and MM progression by retro-orbital bleeding. At weeks 3, 4 and 5, mice underwent PET with [18F]FDG and [18F]4-FGln on consecutive days. The expression of ASCT2, checked in the femur of Vk12598 MM bearing mice, increased along with MM progression. Uptake of both tracers was measurable in the spleen of MM mice by week 3, reached a peak at week 4 and declined thereafter. In JJN3 model, all the tumors were positive for [18F]FDG and displayed [18F]4-FGln uptake (18F]4-FGln: tumor to muscle ratios (T/M): 1.6 ± 0.1; [18F]FDG: T/M: 3.5 ± 1.1). Tumor volume (896.8 ±349.00 mm3), [18F]4-FGln uptake and [18F]FDG uptake (T/M: 2.3 ± 0.3 and 7.1 ± 2.6, respectively) increased after 1 week ([18F]4-FGln p = 0.0156; [18F]FDG p = 0.0167). Thereafter, the effect of bortezomib (BOR) was investigated to evaluate the potential use of [18F]4-FGln to monitor anti-MM treatment. JJN3-bearing mice were treated with BOR (1mg/kg) by intravenous injections twice weekly. Before and after treatment, animals performed the PET acquisitions with [18F]FDG and [18F]4-FGln on consecutive days and were sacrificed for post-mortem analysis on day 7. As expected, bortezomib treatment reduced tumor size compared with vehicles (172.0 ± 86.6 vs 687.9 ± 286.5 mm3; p = 0.0006). PET analysis performed after BOR treatment showed that BOR significantly reduced the uptake of both radiopharmaceuticals in comparison with vehicles (p< 0.05). BOR-treated mice were classified as responders and non responders according to the adapted RECIST score. Interestingly, responder mice showed a reduction of [18F]4-FGln T/M ratio and [18F]4-FGln-related tumor volume. On the contrary, all mice displayed increased [18F]FDG parameters independently from the response. Lastly, tracer overlap analysis showed that the area of relative exclusive uptake of [18F]4-FGln decreased in responders compared to non responders (9% versus 32%, respectively, p=0.04), while that of [18F]FDG increased in responders compared to non responders (82% versus 23%, respectively, p=0.04). Our data indicate that [18F](2 S,4 R)-4-FGln is a new PET tracer in pre-clinical MM models, providing the rationale to design studies in MM patients. Disclosures Giuliani: Celgene: Membership on an entity's Board of Directors or advisory committees, Other: congress, Research Funding; Millenium Pharmaceutical: Other: clincial studies; Janssen: Membership on an entity's Board of Directors or advisory committees, Other: Clinical studies, congress, Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees; GSK: Other: clinical studies; Bristol Mayers Squibb: Other: congress.

Integrin α4β1 belongs to the leukocyte integrin family and represents a therapeutic target of relevant interest given its primary role in mediating inflammation, autoimmune pathologies and cancer-related diseases. The focus of the present work is the design, synthesis and characterization of new peptidomimetic compounds that are potentially able to recognize α4β1 integrin and interfere with its function. To this aim, a collection of seven new cyclic peptidomimetics possessing both a 4-aminoproline (Amp) core scaffold grafted onto key α4β1-recognizing sequences and the (2-methylphenyl)ureido-phenylacetyl (MPUPA) appendage, was designed, with the support of molecular modeling studies. The new compounds were synthesized through SPPS procedures followed by in-solution cyclization maneuvers. The biological evaluation of the new cyclic ligands in cell adhesion assays on Jurkat cells revealed promising submicromolar agonist activity in one compound, namely, the c[Amp(MPUPA)Val-Asp-Leu] cyclopeptide. Further investigations will be necessary to complete the characterization of this class of compounds.

The installation of malononitrile into π-extended carbonyl compounds gives rise to vinylogous alkylidene malononitriles (also known as π-extended dicyanovinylidenes), the direct functionalization of which at remote C(sp3) pronucleophilic sites becomes possible and viable. Starting from easily accessible representative polyunsaturated malononitriles, mild conditions were found to directly couple them to complementary enal acceptors. In all cases, the malononitrile handle proved an indispensable (and optionally traceless) activating ingredient for the vinylogous reactions to proceed efficiently and selectively. Merging the vinylogy concept with the malononitrile HOMO-raising activation strategy and complementary organocatalytic activation modalities (i.e. LUMO-lowering iminium ion activation) turned out to be a successful option, as demonstrated by the number of diverse carbocyclic and heterocyclic chiral products that were (stereo)selectively accessed through this chemistry.1 Introduction2 Reactions of Cyclohexenylidene Malononitriles with Enals3 Reactions of Allylidene Malononitriles with Enals4 Reactions of Indolylmethylene Malononitriles with Enals5 Conclusion

Controlled vinylogous carbon–carbon bond-forming reactions are useful options for providing the selective remote functionalization of conjugated carbonyl substrates. Remotely enolizable alkylidene heterocyclic and carbocyclic carbonyl compounds are pro-nucleophilic substrates that may be engaged in highly valuable chemical transformations. This review emphasizes the merits of these recently discovered vinylogous donors in the chemo-, regio- and stereoselective synthesis of many functionality-rich products.1 Introduction2 Alkylidene Oxindoles3 Alkylidene Pyrazolinones4 Alkylidene Furanones5 Alkylidene Azlactones6 Cycloalkylidene Carbonyl Compounds7 Alkylidene Indenones8 Cycloalkylidene Malononitriles9 Conclusion

Ovarian carcinoma, the most lethal gynecological cancer, is characterized by late diagnosis, with drug resistance limiting the efficacy of platinum-based therapy. Since some integrins are upregulated in cancer, including ovarian carcinoma, they represent a potential target for drug delivery. Receptor tyrosine kinases are also deregulated in cancer and their expression has been associated with drug resistance. Here, the antitumor effects of three conjugates possessing a selective binder of the extracellular portion of integrin &alpha, V&beta, 3 covalently linked to the tyrosine kinase inhibitor sunitinib were investigated in cisplatin-sensitive and -resistant ovarian carcinoma cells expressing both tyrosine kinase VEGFR2 and &alpha, 3 at different levels. We found that one of the three compounds was active in inhibiting the growth of both drug-sensitive and -resistant cells in the micromolar range with a slightly increased potency in resistant cells as compared to sunitinib. The same compound markedly impaired cell migratory and invasive abilities and reduced paxillin phosphorylation. Antitumor activity studies in IGROV-1/Pt1 cells xenografted in nude mice revealed a striking activity of this conjugate versus sunitinib. Taken together, our results support the interest of integrin-targeted sunitinib conjugates for the treatment of drug-resistant tumors.

Touched for the very first time! It is herein highlighted how acetaldehyde silyl enol ethers undergo enantioselective Mukaiyama aldol reaction with aliphatic and aromatic aldehydes. The chemistry relies on the use of the highly efficient and substrate-selective imidodiphosphorimidate catalyst, which displays some of the features of enzymatic catalysis.

On the basis of a previously discovered anti-?V?3 integrin peptidomimetic (c(AmpRGD)) and the clinically approved antiangiogenic kinase inhibitor sunitinib, three novel dual conjugates were synthesized (compounds 1-3), featuring the covalent and robust linkage between these two active modules. In all conjugates, the ligand binding competence toward ?V?3 (using both isolated receptors and ?V?3-overexpressing endothelial progenitor EP cells) and the kinase inhibitory activity (toward both isolated kinases and EPCs) remained almost untouched and comparable to the activity of the single active units. Compounds 1-3 showed interesting antiangiogenesis properties in an in vitro tubulogenic assay; furthermore, dimeric-RGD conjugate 3 strongly inhibited in vivo angiogenesis in Matrigel plug assays in FVB mice. These results offer proof-of-concept of how the covalent conjugation of two angiogenesis-related small modules may result in novel and stable molecules, which impair tumor-related angiogenesis with equal or even superior ability as compared to the single modules or their simple combinations.

Drug resistance and off-organ toxicity remain unsolved issues in chemotherapy of advanced-stage melanoma patients. Thus, the creation of new molecular conjugates able to combine a selective accumulation, high ability of internalization and signaling pathway inhibition, are highly requested. Recently, we reported a new class of molecular conjugates, compounds 1-3, where the anti-αVβ3 integrin peptidomimetic c(AmpRGD), which is a selective ligand for αVβ3 integrin, was covalently bound to the tyrosine kinase inhibitor sunitinib. Here, we report that these c(AmpRGD)-sunitinib conjugates and, in particular, compound 3, are selectively internalized by human melanoma cells through αVβ3 receptor-mediated endocytosis. Compound 3 is more effective than sunitinib in reducing in vitro melanoma cells proliferation, cloning efficiency, migration, and invasion. More interestingly, compound 3 is able to significantly reduce the growth of xenografted melanoma tumor developed in immune-compromised mice, more efficiently than an equimolar dose of sunitinib. Indeed, its targeting ability was demonstrated by the selective localization at the tumor level with respect to healthy tissues. Thus, c(AmpRGD)-sunitinib conjugates such as compound 3 could serve as intriguing multiple-target agents to selectively reach melanoma cells and interfere with the progression of the disease.

We report here the preparation, physico-chemical characterization, and biological evaluation of a new liposome formulation as a tool for tumor angiogenesis inhibition. Liposomes are loaded with sunitinib, a tyrosine kinase inhibitor, and decorated with cyclo-aminoprolineRGD units (cAmpRGD), efficient and selective ligands for integrin αVβ3. The RGD units play multiple roles since they target the nanovehicles at the integrin αVβ3-overexpressing cells (e.g. activated endothelial cells), favor their active cell internalization, providing drug accumulation in the cytoplasm, and likely take part in the angiogenesis inhibition by interfering in the αVβ3-VEGFR2 cross-talk. Both in vitro and in vivo studies show a better efficacy of this integrated antiangiogenic tool with respect to the free sunitinib and untargeted sunitinib-loaded liposomes. This system could allow a lower administration of the drug and, by increasing the vector specificity, reduce side-effects in a prolonged antiangiogenic therapy.

We introduce 3-(alkoxycarbonyl-2-alkyliden)-2-oxindoles as pronucleophilic donors in the direct, vinylogous Michael addition to nitroolefins orchestrated by a chiral, bifunctional cinchona-thiourea organocatalyst. This reaction displays excellent levels of γ-site-, diastereo- and enantioselectivity delivering valuable enantioenriched functionalized oxindoles. Of note, the C-γ enolization of these pronucleophiles by the organocatalyst generates a multidentate, captodative dienolate that delivers vinylogous adducts with an unprecedented Z-selectivity through a peculiar interaction with the catalyst and the nitroolefin. The optimized procedure is operatively simple: the reaction is conducted in air, at room temperature, with low catalyst loading (up to 1 mol%). The synthetic versatility of these Michael adducts is demonstrated by several transformations leading to a valuable quaternary oxindolyl proline analogue and a chiral spirocyclic furoindolone structure. Finally, a mechanistic rationale and a suitable transition state accounting for the observed selectivities are proposed, which are supported by DFT calculations. (Figure presented.).

The asymmetric synthesis of a set of hydroxyphenyl γ-valerolactones was achieved starting from 2-silyloxyfuran and alkoxy-substituted benzaldehydes as common precursors. Key synthesis steps included an enantioselective vinylogous Mukaiyama aldol reaction and a Barton–McCombie deoxygenation. Five enantioenriched γ-valerolactone targets were obtained in 5–6 steps, 18–63% overall yields and 82–98 % ee, paving the way for the straightforward entry to this class of biologically effective and poorly available flavan-3-ol metabolites. In parallel, an unprecedented one-pot reductive ring expansion process was fortuitously discovered, yielding racemic δ-lactone analogues from phenolic butanolide precursors.

A direct aminocatalytic synthesis has been developed for the chemo-, regio-, diastereo-, and enantioselective construction of densely substituted polycyclic carbaldehydes containing fused cyclohexadiene rings. The chemistry utilizes, for the first time, remotely enolizable π-extended allylidenemalononitriles as electron-rich 1,3-diene precursors in a direct eliminative [4+2] cycloaddition with both aromatic and aliphatic α,β-unsaturated aldehydes. The generality of the process is demonstrated by approaching 6,6-, 5,6-, 7,6-, 6,6,6-, and 6,5,6-fused ring systems, as well as biorelevant steroid-like 6,6,6,6,5- and 6,6,6,5,6-rings. A stepwise reaction mechanism for the key [4+2] addition is proposed as a domino bis-vinylogous Michael/Michael/retro-Michael reaction cascade. The utility of the malononitrile moiety as traceless activating group of the dicyano nucleophilic substrates is demonstrated.

The first example of catalytic, enantioselective hy- pervinylogous Mukaiyama aldol reaction (HVMAR) involving multiply unsaturated 2-silyloxyindoles is reported. The reac- tion utilizes a chiral Lewis base-catalyzed Lewis acid-mediat- ed technology to deliver homoallylic 3-polyenylidene 2-oxin- doles with extraordinary levels of regio-, enantio-, and geo- metrical selectivity. This work highlights a subtle yet decisive influence of the indole N-substituents on the propagation of the vinylogous reactivity space of the donor substrates up to ten bonds away from the origin of the vinylogy effect. Analysis of the 13 C NMR chemical shifts of the C-w remote site within homologous silyloxyindole donors enabled ra- tionalization of the results and easy qualitative prediction of the HVMAR reactivity/inertia toward a given aldehyde acceptor.

In recent years, many efforts have been addressed to the development of new imaging techniques enabling early diagnosis and non-invasive monitoring of primary tumors and metastases. Among the integrin family, αVβ3 and α5β1 receptors have been characterized as prototypic markers of angiogenic tumor-associated endothelial cells and their overexpression in tumor cells has been correlated with the progression of various tumor types such as melanoma. Herein, we report the synthesis, characterization and preclinical evaluation of an 111In-labelled DOTA conjugate embodying a cyclic aminoproline-RGD-peptide motif as a competent αVβ3 integrin ligand, to be used as a radiotracer in preclinical models of human melanoma. Practical and efficient chemical and radiochemical synthetic procedures were set up, the in vitro stability and hydrophilicity of a cold c(AmpRGD)-DOTA conjugate were demonstrated, the binding affinities toward isolated αVβ3/α5β1 receptors were assayed and inhibition of cell adhesion to vitronectin and fibronectin was tested in human melanoma and endothelial progenitor cell lines. The anti-angiogenic activity of the peptide conjugates was also tested and assessed in vitro by tubulogenesis assays. The in vivo biodistribution SPECT/CT studies in healthy mice revealed high renal uptake at earlier observation times (30 min to 4 h p.i.) and complete clearance from the kidney at 48 h p.i.; the displacement experiments in human melanoma xenografts confirmed the αVβ3/α5β1 integrin specificity of tumor uptake, suggesting the 111In-labelled c(AmpRGD)-DOTA bioconjugate as a promising starting point in the search for new SPECT-imaging small-molecular probes for non-invasive visualization of tumor angiogenesis, human melanoma and other αVβ3/α5β1-positive tumors.

cope The contribution of the gut microbiota to the metabolism of catechins and proanthocyanidins remains unclear. Phenyl-γ-valerolactones have been identified as the most representative metabolites of these dietary flavan-3-ols, but their accurate quantification has posed problems because of a lack of appropriate bioanalytical standards. This work aimed at synthesizing a novel set of sulphate- and glucuronide-conjugated phenyl-γ-valerolactones and at developing an analytical platform using UHPLC-ESI-MS/MS for their quantification in urine. Methods and results Eight glucuronide and sulphate conjugates of hydroxyphenyl-γ-valerolactones were synthesized and used as analytical standards, together with five phenyl-γ-valerolactone aglycones, for the development of a high-throughput, validated analytical method. Chromatographic and MS conditions were optimized. The method validation showed acceptable linearity, intra-day and inter-day repeatability, and accuracy, with the analytical range, limit of detection (LOD), and lower limit of quantification (LLOQ) varying notably among compounds. The method was used to calculate the excretion of phenyl-γ-valerolactones in healthy subject consuming green tea, providing novel information on the real concentrations of phenyl-γ-valerolactones in urine. Conclusion This work opens the door to better studying the bioavailability of flavan-3-ols and the real exposition to flavan-3-ol sources, as well as to define the bioactivity of these colonic metabolites in cell assays.

Polycyclic indole architectures, including carbazole and hydrocarbazole skeletons, represent privileged structural motifs found in a number of natural alkaloids, many of which displaying interesting activities against a diverse set of biological targets.[1] Accordingly, the search for efficient methods for the assembly of these structures has drawn a great deal of attention in the synthetic organic chemistry community.[2,3] An unprecedented technique for the in situ generation of indolyl ortho-quinodimethanes from 2-methylindole-based methylenemalononitriles 1 by amine-mediated remote C(sp3)-H deprotonation was developed. These intermediates were efficiently trapped by diverse enals to provide a rapid entry to 2,9-dihydro-1H-carbazole-3-carboxyaldehyde structures 3 through a formal asymmetric [4+2] eliminative cycloaddition governed by a ?,?-diphenylprolinol trimethylsilyl ether catalyst.[4] A stepwise reaction mechanism of the formal catalytic asymmetric [4+2] cycloaddition of methylenemalononitrile 1a with enal 2a catalyzed by (S)-C1 is proposed. Parole chiave: asymmetric synthesis, cycloaddition heterocycles, organocatalysis, vinylogy Riferimenti: 1) A. G?uszynska, Eur. J. Med. Chem. 2015, 94, 405. 2) N. H. Krishna, A. P. Saraswati, M. Sathish, N. Shankaraiah, A. Kamal, Chem. Commun. 2016, 52, 4581. 3) Y. Li, F. Tur, R. P. Nielsen, H. Jiang, F. Jensen, K. A. Jørgensen, Angew. Chem. Int. Ed. 2016, 55, 1020; Angew. Chem. 2016, 128, 1032. 4) G. Rassu, C. Curti, V. Zambrano, L. Pinna, N. Brindani, G. Pelosi, and F. Zanardi, Chem. Eur. J. 2016, 22, 12637 - 12640 Ringraziamenti: Financial support provided by the Università degli Studi di Parma.

High glycolitic activity of multiple myeloma (MM) cells is the rationale for the use of Positron Emission Tomography (PET) with 18F-fluorodeoxyglucose ([18F]FDG) to detect both medullary and extramedullary disease. However, FDG-PET has some limitations, since there is a good portion of MM patients who are false-negative. Besides enhanced glycolysis, glutamine (Gln) addiction has been recently described as a metabolic feature of MM by our group. To sustain high Gln demand, MM cells increase the expression of several Gln transporters (ASCT2, SNAT1, LAT1) and are endowed with fast Gln uptake. Yet, at variance with other Gln-addicted cancers, the possible exploitation of Gln as a PET tracer in MM has never been assessed and was investigated in this study. To this purpose, we have firstly synthesized enantiopure (2S,4R)-4-Fluoroglutamine (4-FGln) and validated it as a Gln analogue in human MM cell lines (RPMI8226 and JJN3) comparing its uptake with that of 3H-labelled Gln. The intracellular levels of 4-FGln were determined by HPLC-MS/MS employing a HILIC gradient separation and multiple reaction monitoring (MRM) detection. Both Gln and 4-FGln were actively accumulated by MM cells and exhibited a strong reciprocal competition, pointing to shared transporters. Inhibition analysis revealed that ASCT2 was the major entry route of both compounds, with minor contributions from the other transporters. However, compared with Gln, 4-FGln exhibited higher affinity for both ASCT2 and LAT1 transporters. On the basis of these results, we then tested [18F]4-FGln uptake for MM detection by Positron Emission Tomography (PET) in two different in vivo murine models. Firstly, to investigate sensitivity of human MM to [18F]4-FGln in vivo, JJN3 cells were subcutaneously injected in immunodeficient NSG mice In this xenograft model, [18F]4-FGln- and[18F]FDG-PET scans were performed after plasmacytomas became palpable and repeated after one week. All the tumours were positive for [18F]FDG and displayed [18F]4-FGln uptake with Standard Uptake Values (SUV) of 1.21±1.9 and 0.99±0.07 after 2 weeks, respectively. Thereafter, the effect of bortezomib (BOR) was investigated to evaluate the potential use of [18F]4-FGln to monitor anti-MM treatment. Ten NGS mice were injected with JJN3 cells and, after 14 days, treated twice weekly with BOR, 1mg/kg, or vehicle for two weeks. PET scans were performed before and after 5 and 12 days of BOR treatment. As expected, BOR reduced tumour size as compared to vehicle. At the first post-BOR PET scan, [18F]4-FGln (SUV mean: pre 0.85±0.31; post 0.45±0.10, P Thereafter, to mimic BOR-resistant MM in a syngeneic mouse model, C57BL/6 mice were injected intravenously with Vk12598 cells obtained from transgenic Vk*MYC mice repeatedly treated with sub-optimal doses of BOR. Upon injection into C57BL/6 mice, Vk12598 cells colonize the BM without lytic lesions and extensively colonize the spleen generating an aggressive MM that brings animals to death within five weeks. PET scans were performed with [18F]4-FGln and [18F]FDG before Vk*MYC MM cells injection and after three, four and five weeks. Blood samples for M-spike evaluation were obtained in parallel. Four weeks after MM cells injection a significant increase of both [18F]4-FGln and [18F]FDG uptake was detected in spleens (SUV mean: 1.14±0.23, P=0.018; 0.94±0.24, P= 0.005). In both MM models, the volume of distribution of [18F]4-F-Gln did not overlap that of [18F]FDG. In conclusion, our data indicate that [18F]-(2S,4R)-4-Fluoroglutamine is a new potential PET tracer in pre-clinical MM models especially of extramedullary disease, either in a BOR-sensitive or in a BOR-resistant context, supporting the exploitation of Gln addiction for diagnostic purposes in MM patients. Disclosures Giuliani: Janssen: Research Funding.

On the basis of a previously discovered anti-α

Previously elusive 2,9-dihydro-1H-carbazoles become accessible by a novel enantioselective catalytic [4 + 2] eliminative cycloaddition of indolyl methylenemalononitriles with various cinnamaldehydes.

The asymmetric synthesis of a set of hydroxyphenyl γ-valerolactones was achieved starting from 2-silyloxyfuran and alkoxy-substituted benzaldehydes as common precursors. Key synthesis steps included an enantioselective vinylogous Mukaiyama aldol reaction and a Barton–McCombie deoxygenation. Five enantioenriched γ-valerolactone targets were obtained in 5–6 steps, 18–63% overall yields and 82–98 % ee, paving the way for the straightforward entry to this class of biologically effective and poorly available flavan-3-ol metabolites. In parallel, an unprecedented one-pot reductive ring expansion process was fortuitously discovered, yielding racemic δ-lactone analogues from phenolic butanolide precursors.

A two-step, one-pot sequence under chiral amine—NHC catalysis allows the approach to spiro[4.5]decanone structures in a diastereoselective manner with high e.e.

The reactions proceed with high to excellent Z-selectivity and can be scaled-up without loss of stereoselectivity.