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Abstract: Dihydrochalcones are a class of secondary metabolites, possessing several biological properties such as antitumor, antioxidant, antibacterial, antidiabetic, estrogenic, anti-inflammatory, antithrombotic, antiviral, neuroprotective, and immunomodulatory properties; therefore, they are currently considered promising candidates in the drug discovery process. This review intends to debate their pharmacological actions with particular attention to their antitumor activity against a panel of cancer cell lines and to the description of the inhibition mechanisms of cell proliferation such as the regulation of angiogenesis, apoptosis, etc.

Neuroinflammation is an inflammatory response of the nervous tissue mediated by the production of cytokines, chemokines, and reactive oxygen species. Recent studies have shown that an upregulation of immunoproteasome is highly associated with various diseases and its inhibition attenuates neuroinflammation. In this context, the development of non-covalent immunoproteasome-selective inhibitors could represent a promising strategy for treating inflammatory diseases. Novel amide derivatives, KJ3 and KJ9, inhibit the β5 subunit of immunoproteasome and were used to evaluate their possible anti-inflammatory effects in an in vitro model of TNF-α induced neuroinflammation. Differentiated SH-SY5Y and microglial cells were challenged with 10 ng/mL TNF-α for 24 h and treated with KJ3 (1 µM) and KJ9 (1 µM) for 24 h. The amide derivatives showed a significant reduction of oxidative stress and the inflammatory cascade triggered by TNF-α reducing p-ERK expression in treated cells. Moreover, the key action of these compounds on the immunoproteasome was further confirmed by halting the IkB-α phosphorylation and the consequent inhibition of NF-kB. As downstream targets, IL-1β and IL-6 expression resulted also blunted by either KJ3 and KJ9. These preliminary results suggest that the effects of these two compounds during neuroinflammatory response relies on the reduced expression of pro-inflammatory targets.

Rhodesain is the main cysteine protease of Trypanosoma brucei rhodesiense, the parasite causing the acute lethal form of Human African Trypanosomiasis. Starting from the dipeptide nitrile CD24, the further introduction of a fluorine atom in the meta position of the phenyl ring spanning in the P3 site and the switch of the P2 leucine with a phenylalanine led to CD34, a synthetic inhibitor that shows a nanomolar binding affinity towards rhodesain (Ki = 27 nM) and an improved target selectivity with respect to the parent dipeptide nitrile CD24. In the present work, following the Chou and Talalay method, we carried out a combination study of CD34 with curcumin, a nutraceutical obtained from Curcuma longa L. Starting from an affected fraction (fa) of rhodesain inhibition of 0.5 (i.e., the IC50), we observed an initial moderate synergistic action, which became a synergism for fa values ranging from 0.6 to 0.7 (i.e., 60–70% inhibition of the trypanosomal protease). Interestingly, at 80–90% inhibition of rhodesain proteolytic activity, we observed a strong synergism, resulting in 100% enzyme inhibition. Overall, in addition to the improved target selectivity of CD34 with respect to CD24, the combination of CD34 + curcumin resulted in an increased synergistic action with respect to CD24 + curcumin, thus suggesting that it is desirable to use CD34 and curcumin in combination.

Rhodesain is a cysteine protease crucial for the survival of Trypanosoma brucei rhodesiense, the parasite able to induce the acute lethal form of Human African Trypanosomiasis. PS-1 is a synthetic peptidyl inhibitor of rhodesain, characterised by a picomolar binding affinity (Ki = 1.1 pM). Thus, considering the well-known antiparasitic properties of quercetin, in this study, we decided to carry out drug combination studies of PS-1 and quercetin against rhodesain, according to Chou and Talalay method, which allowed us to obtain for the most relevant fa values a nearly additive effect for the reduction of rhodesain activity from 40% to 90%, thus considering a promising strategy their use in combination.

Despite several major achievements in the development of vaccines and antivirals, the fight against SARS-CoV-2 and the health problems accompanying COVID-19 are still ongoing. SARS-CoV-2 main protease (Mpro), an essential viral cysteine protease, is a crucial target for the development of antiviral agents. A virtual screening analysis of in-house cysteine protease inhibitors against SARS-CoV-2 Mpro allowed us to identify two hits (i.e., 1 and 2) bearing a methyl vinyl ketone warhead. Starting from these compounds, we herein report the development of Michael acceptors targeting SARS-CoV-2 Mpro, which differ from each other for the warhead and for the amino acids at the P2 site. The most promising vinyl methyl ketone-containing analogs showed sub-micromolar activity against the viral protease. SPR38, SPR39, and SPR41 were fully characterized, and additional inhibitory properties towards hCatL, which plays a key role in the virus entry into host cells, were observed. SPR39 and SPR41 exhibited single-digit micromolar EC50 values in a SARS-CoV-2 infection model in cell culture.

Rhodesain is a cysteine protease that is crucial for the life cycle of

Malaria remains a serious problem in global public health, particularly widespread in South America and in tropical regions of Africa and Asia. Chemotherapy is actually the only way to treat this poverty-related disease, since an effective vaccine is not currently available. However, the onset of resistance to the most common antimalarial drugs sometimes makes the current therapeutic regimen problematic. Therefore, the identification of new targets for a new drug discovery process is an urgent priority. In this context, falcipain-2 and falcipain- 3 of P. falciparum represent the key enzymes in the life-cycle of the parasite. Both falcipain- 2 and falcipain-3 are involved in hemoglobin hydrolysis, an essential pathway to provide free amino acids for the parasite metabolic needs. In addition, falcipain-2 is involved in cleaving ankirin and band 4.1 protein, which are cytoskeletal elements essential for the stability of the red cell membrane. This review article is focused on the most recent and effective inhibitors of falcipain-2 and falcipain-3, with particular attention to peptide, peptidomimetic or nonpeptide inhibitors, which targeted one or both the malarial cysteine proteases, endowed with a consistent activity against P. falciparum.

Human African Trypanosomiasis (HAT) is a neglected tropical disease widespread in sub-Saharan Africa. Rhodesain, a cysteine protease of Trypanosoma brucei rhodesiense, has been identified as a valid target for the development of anti-HAT agents. Herein, we report a series of urea-bond-containing Michael acceptors, which were demonstrated to be potent rhodesain inhibitors with K-i values ranging from 0.15 to 2.51 nM, and five of them showed comparable k(2nd) values to that of K1 1777, a potent antitrypanosomal agent. Moreover, most of the urea derivatives exhibited single-digit micromolar activity against the protozoa, and the presence of substituents at the P3 position appears to be essential for the antitrypanosomal effect. Replacement of Phe with Leu at the P2 site kept unchanged the inhibitory properties. Compound 7 (SPR7) showed the best compromise in terms of rhodesain inhibition, selectivity, and antiparasitic activity, thus representing a new lead compound for future SAR studies.

In this paper, we developed a new series of dipeptide nitriles that were demonstrated to be reversible rhodesain inhibitors at nanomolar level, with EC50 values against cultured T. b. brucei in the micromolar range. We also proved that our dipeptide nitriles directly bind to the active site of rhodesain acting as competitive inhibitors. Within the most interesting compounds, the dipeptide nitrile 2b showed the highest binding affinity towards rhodesain (Ki = 16 nM) coupled with a good antiparasitic activity (EC50 = 14.1 μM). Moreover, for the dipeptide nitrile 3e, which showed a Ki = 122 nM towards the trypanosomal protease, we obtained the highest antiparasitic activity (EC50 = 8.8 μM). Thus, given the obtained results both compounds could certainly represent new lead compounds for the discovery of new drugs to treat Human African Trypanosomiasis.

This article described the synthesis and biological investigation of a series of symmetric diarylpentanoids, characterized by a dienone moiety and by a different pattern of substitution on the two phenyl rings. The series of compounds 1a-p were tested against drug-sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 cells to evaluate their cytotoxic profile, and all the diarypentanoids revealed to be active against both the leukemia cell lines, with the best activity shown by compound 1o that showed a submicromolar activity against both CCRF-CEM and CEM/ADR5000 cell lines (EC

Multiple myeloma (MM) is a currently incurable hematologic cancer. This disease is characterized by immunological alterations of myeloid cells and lymphocytes. The first-line therapy involves the use of classic chemotherapy; however, many patients have a relapsed form that could evolve into a refractory MM. The new therapeutic frontiers involve the use of new monoclonal antibodies (Mab) such as daratumumab, isatuximab, and elotuzumab. In addition to monoclonal antibodies, new immunotherapies based on modern bispecific antibodies and chimeric antigen receptor (CAR) T cell therapy have been investigated. For this reason, immunotherapy represents the greatest hope for the treatment of MM. This review intends to focus the attention on the new approved antibody targets. The most important are: CD38 (daratumumab and isatuximab), SLAM7 (elotuzumab), and BCMA (belantamab mafodotin) for the treatment of MM currently used in clinical practice. Although the disease is still incurable, the future perspective is to find the best therapeutic combination among all available drugs.

Rhodesain is a cysteine protease crucial for the survival of

Rhodesain is an enzyme essential for the life of Trypanosoma brucei rhodesiense, a parasite causing a rapid-onset form of Human African Trypanosomiasis. RK-52 is a synthetic inhibitor of rhodesain,...

The development of immunoproteasome-selective inhibitors is a promising strategy for treating hematologic malignancies, autoimmune and inflammatory diseases. In this context, we report the design, synthesis, and biological evaluation of a new series of amide derivatives as immunoproteasome inhibitors. Notably, the designed compounds act as noncovalent inhibitors, which might be a promising therapeutic option because of the lack of drawbacks and side effects associated with irreversible inhibition. Among the synthesized compounds, we identified a panel of active inhibitors with Ki values in the low micromolar or sub-micromolar ranges toward the b5i and/or b1i subunits of immunoproteasomes. One of the active compounds was shown to be the most potent and selective inhibi-tor with a Ki value of 21 nm against the single b1i subunit. Docking studies allowed us to determine the mode of binding of the molecules in the catalytic site of immunoproteasome subunits.

During almost all 2020, coronavirus disease 2019 (COVID-19) pandemic has constituted the major risk for the worldwide health and economy, propelling unprecedented efforts to discover drugs for its prevention and cure. At the end of the year, these efforts have culminated with the approval of vaccines by the American Food and Drug Administration (FDA) and the European Medicines Agency (EMA) giving new hope for the future. On the other hand, clinical data underscore the urgent need for effective drugs to treat COVID-19 patients. In this work, we embarked on a virtual screening campaign against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Mpro chymotrypsin-like cysteine protease employing our in-house database of peptide and non-peptide ligands characterized by different types of warheads acting as Michael acceptors. To this end, we employed the AutoDock4 docking software customized to predict the formation of a covalent adduct with the target protein. In vitro verification of the inhibition properties of the most promising candidates allowed us to identify two new lead inhibitors that will deserve further optimization. From the computational point of view, this work demonstrates the predictive power of AutoDock4 and suggests its application for the in silico screening of large chemical libraries of potential covalent binders against the SARS-CoV-2 Mpro enzyme.

To date, SARS-CoV-2 infectious disease, named COVID-19 by the World Health Organization (WHO) in February 2020, has caused millions of infections and hundreds of thousands of deaths. Despite the scientific community efforts, there are currently no approved therapies for treating this coronavirus infection. The process of new drug development is expensive and time-consuming, so that drug repurposing may be the ideal solution to fight the pandemic. In this paper, we selected the proteins encoded by SARS-CoV-2 and using homology modeling we identified the high-quality model of proteins. A structure-based pharmacophore modeling study was performed to identify the pharmacophore features for each target. The pharmacophore models were then used to perform a virtual screening against the DrugBank library (investigational, approved and experimental drugs). Potential inhibitors were identified for each target using XP docking and induced fit docking. MM-GBSA was also performed to better prioritize potential inhibitors. This study will provide new important comprehension of the crucial binding hot spots usable for further studies on COVID-19. Our results can be used to guide supervised virtual screening of large commercially available libraries.

Dear Editor,We would like to reply to the letter to the Editor of Steverding (2018) on our research article “Drug combination studies of curcumin and genistein against rhodesain of Trypanosoma bruc...

Proteasome inhibition is a promising strategy for the treatment of multiple myeloma; unfortunately, this disease is often associated with an increasing chemoresistance. One novel approach may be to target the immunoproteasome, a proteasomal isoform mainly present in cells of hematopoietic origin. We investigated the activity of a panel of amides against immunoproteasome core particles as potential agents for the treatment of multiple myeloma (MM). Amide 6 showed an ideal profile since it was able to inhibit both the chymotrypsin-like activities of the immunoproteasome with Ki values of 4.90 µM and 4.39 µM for β1i and β5i, respectively, coupled with an EC50 =17.8 µM against MM.1R cells. Compound 6 inhibited also ubiquitinated protein degradation and was able to act on different phases of MM cell cycle reducing the levels of cyclin A/CDK1, cyclin B/CDK1 and cyclin D/CDK4/6 complexes, which turns in cell cycle arrest.

Starting from the reversible rhodesain inhibitors 1 a-c, which have Ki values towards the target protease in the low-micromolar range, we have designed a series of peptidomimetics, 2 a-g, that contain a benzodiazepine scaffold as a β-turn mimetic; they are characterized by a specific peptide sequence for the inhibition of rhodesain. Considering that irreversible inhibition is strongly desirable in the case of a parasitic target, a vinyl ester moiety acting as Michael-acceptor was introduced as the warhead; this portion was functionalized in order to evaluate the size of corresponding enzyme pocket that could accommodate this substituent. With this investigation, we identified an irreversible rhodesain inhibitor (i. e., 2 g) with a k2nd value of 90 000 M-1 min-1 that showed antitrypanosomal activity in the low-micromolar range (EC50 =1.25 μM), this may be considered a promising lead compound in the drug-discovery process for treating human African trypanosomiasis (HAT).

Curcumin and genistein are two natural products obtained from Curcuma longa L. and soybeans, endowed with many biological properties. Within the last years they were shown to possess also a promising antitrypanosomal activity. In the present paper, we investigated the activity of both curcumin and genistein against rhodesain, the main cysteine protease of Trypanosoma brucei rhodesiense; drug combination studies, according to Chou and Talalay method, allowed us to demonstrate a potent synergistic effect for the combination curcumin-genistein. As a matter of fact, with our experiments we observed that the combination index of curcumin-genistein is

The inhibition of immunoproteasome is considered nowadays a promising strategy for the treatment of hematologic malignancies. In this paper we report the design, synthesis, and biological evaluation as immunoproteasome inhibitors of a new series of isoquinolinone derivatives characterized by a (E)-prop-1-ene fragment that connects the heterocycle to a distal amide functionality. Among all the synthesized compounds, we identified an inhibitor with Ki values in the low micromolar or submicromolar range towards the chymotrypsin-like activities of both proteasome and immunoproteasome (β5c, β5i and β1i subunits). Molecular modeling studies suggest that the most potent compound of the series may act a single-site binder. In particular, through its isopentyl group, it might dock into P1 site in the case of the β1i catalytic subunit, while in the case of β5c and β5i subunits, the P3 site might be the preferred binding site.

This paper describes the development of a class of peptide-based inhibitors as novel antitrypanosomal and antimalarial agents. The inhibitors are based on a characteristic peptide sequence for the inhibition of the cysteine proteases rhodesain of Trypanosoma brucei rhodesiense and falcipain-2 of Plasmodium falciparum. We exploited the reactivity of novel unsaturated electrophilic functions such as vinyl-sulfones, -ketones, -esters, and -nitriles. The Michael acceptors inhibited both rhodesain and falcipain-2, at nanomolar and micromolar levels, respectively. In particular, the vinyl ketone 3b has emerged as a potent rhodesain inhibitor (k2nd = 67 × 106 M-1 min-1), endowed with a picomolar binding affinity (Ki = 38 pM), coupled with a single-digit micromolar activity against Trypanosoma brucei brucei (EC50 = 2.97 μM), thus being considered as a novel lead compound for the discovery of novel effective antitrypanosomal agents.

[Image: see text] Rhodesain is an enzyme essential for the life of Trypanosoma brucei rhodesiense, a parasite causing a rapid-onset form of Human African Trypanosomiasis. RK-52 is a synthetic inhibitor of rhodesain, characterized by an impressive k(second) value (k(second) = 67000 × 10(3) M(–1) min(–1)) and by a picomolar affinity toward the trypanosomal protease (K(i) = 38 pM). Differently, curcumin, the golden multitarget nutraceutical obtained from Curcuma longa L., was proven to inhibit rhodesain noncompetitively with an IC(50) of 7.75 μM. In the present study, we carried out studies of a combination of RK-52 and curcumin toward rhodesain, by applying the Chou and Talalay approach, which led us to obtain a combination index

The identification of effective and selective immunoproteasome inhibitors could greatly promote smart treatment of many autoimmune, inflammatory and tumor diseases. This paper explores the activity of some pyridone derivatives which are taking advantage of peptide mimicking moieties and other functional groups whose related effects are taken into consideration. The target compounds were designed and synthesized starting from a lead, next their in vitro biological activity was tested; the following structural and conformational analysis sheds light over the different activity of the tested compounds. Compound 3 bearing a phenylalanine at the P2 site and an allyloxycarbonyl group spanning the P3 region, was found to be a selective inhibitor of β5 subunit of immunoproteasome. It has been demonstrated by Nuclear Magnetic Resonance (NMR) that, in solution, this substrate displays a more rigid conformation respect to the others, especially in apolar media, this feature is likely crucial for the enhancement of the activity towards immunoproteasome. The specific conformational analysis enables insights of the biological activity of new chemicals driving possible advancements based on the suitable structural features to identify novel selective β5 immunoproteasome inhibitor.

Human African Trypanosomiasis (HAT) is an endemic parasitic disease of sub-Saharan Africa, caused by two subspecies of protozoa belonging to Trypanosoma genus: T. brucei gambiense and T. brucei rhodesiense. In this context the inhibition of the papain-family cysteine proteases rhodesain and TbCatB has to be considered a promising strategy for HAT treatment. Rhodesain, the major cathepsin L-like cysteine protease of T. brucei rhodesiense, is a lysosomal protease essential for parasite survival. It is involved in parasite invasivity, allowing it to cross the blood-brain barrier (BBB) of the human host, causing the second lethal stage of the disease. Moreover, it plays an important role in immunoevasion, being involved in the turnover of variant surface glycoproteins of the T. brucei coat and in the degradation of immunoglobulins, avoiding a specific immune response by the host cells. On the other hand TbCatB, a cathepsin B-like cysteine protease, present in minor abundance in T. brucei, showed a key role in the degradation of host transferrin, which is necessary for iron acquisition by the parasite. In this review article we now discuss the most active peptide, peptidomimetic and non-peptide rhodesain and TbCatB inhibitors as valuable strategy to treat HAT, due also to the complementary role of the two T. brucei proteases.

Novel 1,4-benzodiazepines, endowed with a Michael acceptor moiety, were designed taking advantage of a computational prediction of their pharmacokinetic parameters. Among all the synthesized derivatives, we identified a new lead compound (i.e., 4a), bearing a vinyl ketone warhead and endowed with a promising antitrypanosomal activity against Trypanosoma brucei brucei (IC50 = 5.29 µM), coupled with a lack of cytotoxicity towards mammalian cells (TC50>100 µM).

This paper describes an optimization strategy of the highly active vinyl ketone 3 which was recognized as a strong inhibitor of rhodesain of Trypanosoma brucei rhodesiense, endowed with a ksecond v...

Novel rhodesain inhibitors were developed by combining an enantiomerically pure 3-bromoisoxazoline warhead with a 1,4-benzodiazepine scaffold as specific recognition moiety. All compounds were proven to inhibit rhodesain with Ki values in the low-micromolar range. Their activity towards rhodesain was found to be coupled to an in vitro antitrypanosomal activity, with IC50 values ranging from the mid-micromolar to a low-micromolar value for the most active rhodesain inhibitor (R,S,S)-3. All compounds showed a good selectivity against the target enzyme since all of them were proven to be poor inhibitors of human cathepsin L. Novel rhodesain inhibitors were developed by combining an enantiomerically pure 3-bromoisoxazoline warhead with a 1,4-benzodiazepine scaffold as specific recognition moiety. All compounds were proven to inhibit rhodesain with K-i values in the low-micromolar range. Their activity towards rhodesain was found to be coupled to an in vitro antitrypanosomal activity, with IC50 values ranging from the mid-micromolar to a low-micromolar value for the most active rhodesain inhibitor (R,S,S)-3. All compounds showed a good selectivity against the target enzyme since all of them were proven to be poor inhibitors of human cathepsin L.

Conformational analysis of a potent class of cysteine protease inhibitors is thoroughly studied by NMR, in both, polar and apolar solvents to get a better insight over the known biological activity and migration through biological media. These molecules are composed by a benzodiazepine (BDZ) scaffold connected to a bromo-isoxazoline (IOX) ring through an alkyl spacer (AS) with up to four-carbon atoms. Data, supported by theoretical calculations at DFT level, reveal that both BDZ and IOX keep a pretty rigid and asymmetric conformation, so that four diastereo-atropisomers (two mirror-image couples) are generated. The relative stiffness of these substrates, maintained also in different solvents, is confirmed by: (a) remarkable separation of diastereotopic protons; (b) specific “through the space contacts” (NOESY); and (c) very good fitting of the coupling constants evaluations. The prototypic compound with the longer AS shows two main conformations and a certain dynamic freedom around the AS torsional angles close to IOX; according to our data, the AS length is not fundamental for the functional BDZ and IOX fitting into the macromolecular complex; however, it does play a crucial role to cross the parasite cell membranes.

This paper describes the design, synthesis, and biological evaluation of 2-thioxoimidazolidin-4-one derivatives as inhibitors of proteasome and immunoproteasome, potential targets for the treatment of hematological malignancies. In particular, we focused our efforts on the design of noncovalent inhibitors, which might be a promising therapeutic option potentially devoid of drawbacks and side-effects related to irreversible inhibition. Among all the synthesized compounds, we identified a panel of active inhibitors with Ki values towards one or two chymotrypsin-like activities of proteasome (β5c) and immunoproteasome (β5i and β1i subunits) in the low micromolar range. Docking studies suggested a unique binding mode of the molecules in the catalytic site of immunoproteasome proteolytic subunits.

The ubiquitin-proteasome system (UPS) is the major non-lysosomal proteolytic system for the degradation of abnormal or damaged proteins no longer required. The proteasome is involved in degradation of numerous proteins which regulate the cell cycle, indicating a role in controlling cell proliferation and maintaining cell survival. Defects in the UPS can lead to anarchic cell proliferation and to tumor development. For these reasons UPS inhibition has become a significant new strategy for drug development in cancer treatment. In addition to the constitutive proteasome, which is expressed in all cells and tissues, higher organisms such as vertebrates possess two immune-type proteasomes, the thymoproteasome and the immunoproteasome. The thymoproteasome is specifically expressed by thymic cortical epithelial cells and has a role in positive selection of CD8+ T cells, whereas the immunoproteasome is predominantly expressed in monocytes and lymphocytes and is responsible for the generation of antigenic peptides for cell-mediated immunity. Recent studies demonstrated that the immunoproteasome has a preservative role during oxidative stress and is up-regulated in a number of pathological disorders including cancer, inflammatory and autoimmune diseases. As a consequence, immunoproteasome-selective inhibitors are currently the focus of anticancer drug design. At present, the commercially available proteasome inhibitors bortezomib and carfilzomib which have been validated in multiple myeloma and other model systems, appear to target both the constitutive and immunoproteasomes, indiscriminately. This lack of specificity may, in part, explain some of the side effects of these agents, such as peripheral neuropathy and gastrointestinal effects, which may be due to targeting of the constitutive proteasome in these tissues. In contrast, by selectively inhibiting the immunoproteasome, it may be possible to maintain the antimyeloma and antilymphoma efficacy while reducing these toxicities, thereby increasing the therapeutic index. This review article will be focused on the discussion of the most promising immunoproteasome specific inhibitors which have been developed in recent years. Particular attention will be devoted to the description of their mechanism of action, their structure-activity relationship, and their potential application in therapy.

We recently reported the synthesis, of a potent papain-family cathepsin L-like cysteine protease inhibitor, as new lead compound for the development of new drugs that can be used as antiprotozoal agents. The investigation of its conformational profile is crucial for the in-depth understanding of its biological behaviour. Our careful NMR analysis has been based on the complete and total assignment of 1H, 13C, 15N and 19F signals of the molecule in both CDCl3 and CD3OH, which could reproduce in some way a scenario of polar and not polar phases into the biological environment. In this way it has been unveiled a different behaviour of the molecule in polar and apolar media. In CDCl3 it is possible to define stable conformational arrangements on the basis of the detected through space contacts, whereas, in CD3OH a greater conformational freedom is envisaged: (a) by the overlap of any of the CH2 diastereotopic resonances (unable to distinguish asymmetric molecular sides because of the free rotation about the single bonded chains), (b) by the less definite measured vicinities not consistent with just one conformation and (c) by the evident loss or switching of key intramolecular hydrogen interactions.

A new series of pseudopeptide boronate proteasome inhibitors (2–3) was developed, through optimization of our previously described analogs of bortezomib, bearing a bicyclic 1,6-naphthyridin-5(6H)-one scaffold as P3 fragment (1). The biological evaluation on human 20S proteasome displayed a promising inhibition profile, especially for compounds bearing a P2 ethylene fragment, which exhibited Ki values in the nanomolar range for the ChT-L activity (e.g. 2a, Ki = 0.057 μM) and considerable selectivity for proteasome over bovine pancreatic α-chymotrypsin. Docking experiments into the yeast 20S proteasome revealed that the ligands are accommodated predominantly into the ChT-L site and that they covalently bind to the active site threonine residue via boron atom. Within the cellular assays performed against a 60 cancer cell line panel, compounds 3e and 3f demonstrated also good antiproliferative activity and compound 3f emerged as promising lead compound for the development of anticancer agents targeting melanoma and non-small cell lung cancer.

The identification of the key role of the eukaryotic 26S proteasome in regulated intracellular proteolysis and its importance as a target in many pathological conditions wherein the proteasomal activity is defective (e.g., malignancies, autoimmune diseases, neurodegenerative diseases, etc.) prompted several research groups to the development of specific inhibitors of this multicatalytic complex with the aim of obtaining valid drug candidates. In regard to the anticancer therapy, the peptide boronate bortezomib (Velcade®) represents the first molecule approved by FDA for the treatment of multiple myeloma in 2003 and mantle cell lymphoma in 2006. Since then, a plethora of molecules targeting the proteasome have been identified as potential anticancer agents and a few of them reached clinical trials or are already in the market (i.e., carfilzomib; Kyprolis®). In most cases, the design of new proteasome inhibitors (PIs) takes into account a proven peptide or pseudopeptide motif as a base structure and places other chemical entities throughout the peptide skeleton in such a way to create an efficacious network of interactions within the catalytic sites. The purpose of this review is to provide an in-depth look at the current state of the research in the field of peptide-based PIs, specifically those ones that might find an application as anticancer agents.

This manuscript describes the synthesis of a new series of 2-amino-6-(trifluoromethoxy)benzoxazole derivatives (1–5) and a benzothiazine derivative (6) structurally related to riluzole, the only neuroprotective drug currently approved for the treatment of the amyotrophic lateral sclerosis. Preliminary results indicate that the synthesized compounds, in particular benzamide derivatives 3 and 4, are able to antagonize voltage-dependent Na+ channel currents, and therefore they could be exploited as new inhibitors of these channels. Moreover, all compounds possess low binding affinity for GABA and NMDA receptors.

In this work, we designed and synthesized novel N-3-bromoisoxazolin-5-yl substituted 2,3-benzodiazepines as noncompetitive AMPAR antagonists, with the aim that this heterocycle could establish favourable interactions with a putative binding pocket of the receptor, like the thiadiazole nucleus of GYKI 47409 does. Within this investigation, we identified some active molecules and, among these 2,3-benzodiazepines, 4c showed a much improved inhibitory potency as compared with unsubstituted 2,3-benzodiazepines.

Herein we report the synthesis of a set of constrained peptidomimetics endowed with an electrophilic vinyl ester warhead and structurally related to a previously identified lead compound, a potent and irreversible inhibitor of falcipain-2 (FP-2). FP-2 is the main hemoglobinase of the malaria parasite P. falciparum. The new compounds were evaluated for their inhibition against FP-2, and the results were rationalized on the basis of docking experiments. These studies underscore the pivotal role of both the ester function at the P1' site and the trifluoromethyl group of the P3 side chain in determining the correct orientation of the Michael acceptor warhead in the catalytic site, and as a consequence, the potency of the inhibitors as well as their reversible or irreversible mode of inhibition.

This article describes the synthesis and the biological evaluation of two sets of benzothiazole derivatives bearing at C-2 an arylamide (1a–e, 2a–e) or an arylurea (3a–d, 4a–d) moiety. Five compounds (3d and 4a–d) were selected and screened by the National Cancer Institute for the in vitro primary anticancer assay against a panel of 60 human tumor cell lines. Compounds 4a and 4c showed interesting anticancer activities, more marked for compound 4c. All compounds were also submitted to a preliminary in vitro assay as potential inhibitors of the ubiquitin-activating enzyme (E1), but they lacked significant activity.

Herein we report the synthesis of a series of novel constrained peptidomimetics 2-10 endowed with a dipeptide backbone (D-Ser-Gly) and a vinyl ester warhead, structurally related to a previously identified lead compound 1, an irreversible inhibitor of falcipain-2, the main haemoglobinase of lethal malaria parasite Plasmodium falciparum. The new compounds were evaluated for their inhibition against falcipain-2, as well as against cultured P. falciparum. The inhibitory activity of the synthesized compounds was also evaluated against another protozoal cysteine protease, namely rhodesain of Trypanosoma brucei rhodesiense.

This paper describes the synthesis of novel peptidomimetics bearing a protected aspartyl aldeyde warhead leading to the thioacylals 2a,b and the acylals 3a,b. Compounds 2a and 3a proved to possess an increased antiplasmodial activity with respect to the parent molecule 1. Furthermore thioacylal 2a can be considered as a promising trypanocidal agent.

A series of 1-aryl-6,7-disubstituted-2H-isoquinolin-3-ones (2-10) was synthesized and evaluated for their inhibition against Plasmodium falciparum cysteine protease falcipain-2, as well as against cultured P. falciparum strain FCBR parasites. All compounds displayed inhibitory activity against recombinant falcipain-2 and against in vitro cultured intraerythrocytic P. falciparum, with the exception of 9. The new compounds exhibited no selectivity against human cysteine proteases such as cathepsins B and L. The inhibitory activity of the synthesized compounds was also evaluated against another protozoal cysteine protease, namely rhodesain of Trypanosoma brucei rhodesiense.