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Melanoma is a skin cancer that arises from melanocytes and can spread quickly to the other organs of the body, if not treated early. Generally, melanoma shows an inherent resistance to conventional therapies. In this regard, new potential drugs are being developed as possible treatments for melanoma. In this paper, we report the synthesis of a new decavanadate compound with organic molecules for a potential therapeutic application. The tetra-[methylimidazolium] dihydrogen decavanadate(V) salt (C 4 H 7 N 2 ) 4 [H 2 V 10 O 28 ] is characterized by single-crystal X-ray diffraction, by FT-IR, UV-Vis and 51 V NMR spectroscopy, as well as by thermal analysis (TGA and DSC). The compound crystallizes in the monoclinic centrosymmetric space group P2 1 /c. Its formula unit consists of one dihydrogen decavanadate anion [H 2 V 10 O 28 ] 4- and four organic 4-methylimidazolium cations (C 4 H 7 N 2 ) + . Important intermolecular interactions are N-H···O and O-H···O hydrogen bonds and π-π stacking interactions between the organic cations, revealed by analysis of the Hirshfeld surface and its two-dimensional fingerprint plots. Interestingly, this compound inhibits the viability of IGR39 cells with IC 50 values of 14.65 μM and 4 μM after 24 h and 72 h of treatment, respectively. The analysis of its effect by flow cytometry using an Annexin V-FITC/IP cell labeling, showed that (C 4 H 7 N 2 ) 4 H 2 V 10 O 28 compound induced IGR39 cell apoptosis and necrosis. Molecular docking studies performed against TNFR1 and GPR40, as putative targets, suggest that the (C 4 H 7 N 2 ) 4 [H 2 V 10 O 28 ] compound may act as inhibitor of these proteins, known to be overexpressed in melanoma cells. Therefore, we could consider it as a new potential metallodrug against melanoma., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier Inc.)

Allium roseum is amongst the most important wild medicinal plants. It is known for its diverse biological properties, including antioxidant, antibacterial and antidiabetic activities. In this work, the polysaccharides (PARLs) were ultrasonically extracted from Allium roesum leaves then purified and analyzed by several techniques. Chemical composition and GC-MS analysis showed that the obtained polysaccharides were composed mainly of glucose (40.20 %), mannose (25.30 %), fructose (10.60 %) and galacturonic acid (15.11 %). Moreover, PARLs exhibited a potent antioxidant effect with higher capacities up to 69.61 % and 71.72 % for DPPH and ABTS free radicals, respectively. Furthermore, PARLs significantly modulated inflammatory response by reducing TNF-α, IL-6, and IL-8 pro-inflammatory mediators and promoting the anti-inflammatory IL-10 mediator in LPS stimulated THP-1 derived macrophages. The in-vivo tests proved that the extract was able to decrease carrageenan-induced rat paw swelling by around 68.15 % after 4 h of treatment. PARLs, significantly reduced the growth of U87 (glioblastoma) and IGROV-1 cancer cells with IC 50 values of about 4.27 and 7.89 mg/mL respectively. This research clearly shows that Allium roseum polysaccharides can be used as natural antioxidants with anti-inflammatory and anticancer properties., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Maurotoxin (MTX) is a 34-residue peptide from Scorpio maurus venom. It is reticulated by four disulfide bridges with a unique arrangement compared to other scorpion toxins that target potassium (K + ) channels. Structure-activity relationship studies have not been well performed for this toxin family. The screening of Scorpio maurus venom was performed by different steps of fractionation, followed by the ELISA test, using MTX antibodies, to isolate an MTX-like peptide. In vitro, in vivo and computational studies were performed to study the structure-activity relationship of the new isolated peptide. We isolated a new peptide designated MTX1, structurally related to MTX. It demonstrated toxicity on mice eight times more effectively than MTX. MTX1 blocks the Kv1.2 and Kv1.3 channels, expressed in Xenopus oocytes, with IC 50 values of 0.26 and 180 nM, respectively. Moreover, MTX1 competitively interacts with both 125 I-apamin (IC 50 = 1.7 nM) and 125 I-charybdotoxin (IC 50 = 5 nM) for binding to rat brain synaptosomes. Despite its high sequence similarity (85%) to MTX, MTX1 exhibits a higher binding affinity towards the Kv1.2 and SKCa channels. Computational analysis highlights the significance of specific residues in the β-sheet region, particularly the R27, in enhancing the binding affinity of MTX1 towards the Kv1.2 and SKCa channels.

Several neurodegenerative diseases, such as multiple sclerosis and Parkinson's disease, are linked to alterations in myelin content or structure. Transmembrane receptors such as integrins could be involved in these alterations. In the present study, we investigated the role of αv-integrins in experimental models of neuroinflammation and demyelination with the use of lebecetin (LCT), a C-lectin protein purified from Macrovipera lebetina viper venom, as an αv-integrin modulator. In a model of neuroinflammation, LCT inhibited the upregulation of αv, β3, β5, α5, and β1 integrins, as well as the associated release of pro-inflammatory factor IL-6 and chemokine CXCL-10, and decreased the expression of phosphorylated NfκB. The subsequent "indirect culture" between reactive astrocytes and oligodendrocytes showed a down-regulation of αv and β3 integrins versus upregulation of β1 one, accompanied by a reduced expression of myelin basic protein (MBP). Treatment of oligodendrocytes with LCT rectified the changes in integrin and MBP expression. Through Western blot quantification, LCT was shown to upregulate the expression levels of PI3K and p-mTOR while downregulating expression levels of p-AKT in oligodendrocytes, suggesting the neuroprotective and pro-myelinating effects of LCT may be related to the PI3K/mTor/AKT pathway. Concomitantly, we found that LCT promoted remyelination by tracking the increased expression of MBP in the brains of cuprizone-intoxicated mice. These results point to an involvement of integrins in not only neuroinflammation but demyelination as well. Thus, targeting αv integrins could offer potential therapeutic avenues for the treatment of demyelinating diseases., (© 2024. The Author(s).)

Two bases-decavanadates coordination compounds [(C 6 H 13 N 4 ) 2 ][Mg(H 2 O) 6 ] 2 [O 28 V 10 ].6H 2 O (1) and [(C 7 H 11 N 2 ) 4 ][Mg(H 2 O) 6 ][O 28 V 10 ].4H 2 O (2) have been synthesized and well characterized using vibrational spectroscopy (infrared), UV-Visible analysis and single crystal X-ray diffraction technique. The formula unit, for both compounds, is composed by the decavanadate [V 10 O 28 ] 6- , hydrated magnesium ion, a counter anion and free water molecules. The transition metal adopts octahedral geometries in both compound (1) and (2). The existence of a multitude of hydrogen bonding interactions for both compounds provides a stable three-dimensional supramolecular structure. Optical absorption reveals a band gap energy indicating the semi-conductive nature of the compound. In this study, the cytotoxic and the anti-proliferative activities of compounds (1) and (2) on human cancer cells (U87 and MDA-MB-231) were investigated. Both compounds demonstrated dose-dependent anti-proliferative activity on U87 and MDA-MB-231 with respective IC 50 values of 0.82 and 0.31 μM and 1.4 and 1.75 μM. These data provide evidence on the potential anticancer activity of [(C 6 H 13 N 4 ) 2 ][Mg(H 2 O) 6 ] 2 [O 28 V 10 ].6H 2 O and [(C 7 H 11 N 2 ) 4 ][Mg(H 2 O) 2 ][O 28 V 10 ].4H 2 O. Molecular docking of the compounds was also examined. Molecular docking studies were performed for both compounds against four target receptors and revealed better binding affinity with these targets in comparison to Cisplatin. Moreover, molecular docking investigations suggest that these compounds may function as potential inhibitors of proteins in brain and breast cells, exhibiting greater efficiency compared to Cisplatin., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Plant polyphenols are nutraceutical components with relevant biological effects on human health. They act against development of several diseases including cancer. In this study, the methanolic extracts of four date palm Phoenix dactylifera leaves (Deglet Noor (DN), Barhee (B), Khalas (KS) and Khunezi (KZ)) collected from south Tunisia were preliminary analyzed for their effects against U87 (human glioblastoma) and MDA-MB-231 (human breast cancer) cell line development. Results showed that Barhee extract (30 μg/mL) was the most efficient to reduce the growth of both tumor cells to about 40% (p < 0.05) without inducing cytotoxicity. Significantly, KS, KZ, DN and B extracts (30 μg/mL) decreased MDA-MB-231 and U87 cell adhesion towards fibrinogen and fibronectin. Using integrin blocking antibodies, leaf extracts competitively decreased human glioblastoma cell attachment to immobilized antibodies by interfering to αvβ3 and α5β1 integrin receptors. At the same concentration, extracts decreased MDA-MB-23 and U87 cell migration performed with wound healing assay. Particularly, Barhee and Deglet Noor leaf extracts (30 μg/mL) significantly reduced U87 cell invasion by 52.92% (p < 0.01) and 74.56% (p < 0.01), respectively. Collegially, our findings revealed beneficial proprieties of four varieties of date palm leaf especially those displayed by DN and B extracts that may serve as active candidates against human glioblastoma and breast cancer progression., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Chalcones are polyphenols that belong to the flavonoids family, known for their broad pharmacological properties. They have thus attracted the attention of chemists for their obtention and potential activities. In our study, a library of compounds from 2'-hydroxychalcone's family was first synthesized. A one-step mechanochemical synthesis via Claisen-Schmidt condensation reaction under ball mill conditions was studied, first in a model reaction between a 5'-fluoro-2'-hydroxyacetophenone and 3,4-dimethoxybenzaldehyde. The reaction was optimized in terms of catalysts, ratio of reagents, reaction time, and influence of additives. Among all assays, we retained the best one, which gave the highest yield of 96% when operating in the presence of 1 + 1 eq. of substituted benzaldehyde and 2 eq. of KOH under two grinding cycles of 30 min. Thus, this protocol was adopted for the synthesis of the selected library of 2'-hydroxychalcones derivatives. The biological activities of 17 compounds were then assessed against Plasmodium falciparum , Leishmania donovani parasite development, as well as IGR-39 melanoma cell lines by inhibiting their viability and proliferation. Compounds 6 and 11 are the most potent against L. donovani, exhibiting IC 50 values of 2.33 µM and 2.82 µM, respectively, better than the reference drug Miltefosine (3.66 µM). Compound 15 presented the most interesting antimalarial activity against the 3D7 strain, with IC 50 = 3.21 µM. Finally, chalcone 12 gave the best result against IGR-39 melanoma cell lines, with an IC 50 value of 12 µM better than the reference drug Dacarbazine (IC 50 = 25 µM).

Background: Integrins, important extracellular matrix (ECM) receptor proteins, are affected by inflammation and can participate in the maintenance of many painful conditions. Although they are ubiquitous and changeable across all cell types, the roles of these cell adhesion molecules in pathological pain have not been fully explored., Objective: We evaluated the effects of the subcutaneous injection of lebecetin, a C-type lectin isolated from Macrovipera lebetina snake venom, previously reported to inhibit α5β1 and αv integrin activity, on different components of inflammation induced by the formalin administration in the hind paw of mice., Methods: The formalin-induced nocifensive behavior, edema, and histopathological changes in the hind paw associated with cytokine, iNOS, and COX2 expression, nociceptive-specific neuron activity, and microglial activation analysis in the spinal cord were evaluated in mice receiving vehicle or lebecetin pretreatment., Results: Lebecetin inhibited the nocifensive responses in the formalin test, related edema, and cell infiltration in the injected paw in a biphasic, hormetic-like, and dose-dependent way. According to that hormetic trend, a reduction in pro-inflammatory cytokines IL-6, IL-8, and TNF-alpha and upregulation of the anti-inflammatory cytokine IL-10 in the spinal cord were found with the lowest doses of lebecetin. Moreover, COX2 and iNOS expression in serum and spinal cord followed the same biphasic pattern of cytokines. Finally, nociceptive neurons sensitization and activated microglia were normalized in the dorsal horn of the spinal cord by lebecetin., Conclusion: These findings implicate specific roles of integrins in inflammation and tonic pain, as well as in the related central nervous system sequelae., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Inflammation is associated with many pathology disorders and the malignant progression of most cancers. Therefore, targeting inflammatory pathways could provide a promising strategy for disease prevention and treatment. In this study, we experimentally investigated the anti-inflammatory effect of CC5 and CC8, two disintegrin isoforms isolated from Cerastes cerastes snake venom, on LPS-stimulated macrophages, both on human THP-1 and mouse RAW264.7 cell adherence and their underlying mechanisms by measuring cytokine release levels and Western blot assay. Equally, both molecules were evaluated on a carrageenan-induced edema rat model. Our findings suggest that CC5 and CC8 were able to reduce adhesion of LPS-stimulated macrophages both on human THP-1 and mouse RAW264.7 cells to fibrinogen and vitronectin through the interaction with the αvβ3 integrin receptor. Moreover, CC5 and CC8 reduced the levels of reactive oxygen species (ROS) mediated by the NF-κB, MAPK and AKT signaling pathways that lead to decreased production of the pro-inflammatory cytokines TNF-α, IL-6 and IL-8 and increased secretion of IL-10 in LPS-stimulated THP-1 and RAW264.7 cells. Interestingly, both molecules potently exhibited an anti-inflammatory effect in vivo by reducing paw swelling in rats. In light of these results, we can propose the CC5 and CC8 disintegrins as interesting tools to design potential candidates against inflammatory-related diseases.

The emerging concept of small conductance Ca 2+ -activated potassium channels (SK Ca ) as pharmacological target for cancer treatment has significantly increased in recent years. In this study, we isolated the P01 toxin from Androctonus australis (Aa) scorpion venom and investigated its effect on biological properties of glioblastoma U87, breast MDA-MB231 and colon adenocarcinoma LS174 cancer cell lines. Our results showed that P01 was active only on U87 glioblastoma cells. It inhibited their proliferation, adhesion and migration with IC 50 values in the micromolar range. We have also shown that P01 reduced the amplitude of the currents recorded in HEK293 cells expressing SK2 channels with an IC 50 value of 3 pM, while it had no effect on those expressing SK3 channels. The investigation of the SK Ca channels expression pattern showed that SK2 transcripts were expressed differently in the three cancer cell lines. Particularly, we highlighted the presence of SK2 isoforms in U87 cells, which could explain and rely on the specific activity of P01 on this cell line. These experimental data highlighted the usefulness of scorpion peptides to decipher the role of SK Ca channels in the tumorigenesis process, and develop potential therapeutic molecules targeting glioblastoma with high selectivity., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Mlayah-Bellalouna, Aissaoui-Zid, Chantome, Jebali, Souid, Ayedi, Mejdoub, Belghazi, Marrakchi, Essafi-Benkhadir, Vandier and Srairi-Abid.)

Newcastle disease virus (NDV) is one of the most serious contagions affecting domestic poultry and other avian species. It causes high morbidity and mortality, resulting in huge economic losses to the poultry industry worldwide. Despite vaccination, NDV outbreaks increase the need for alternative prevention and control means. In this study, we have screened fractions of Buthus occitanus tunetanus ( Bot ) scorpion venom and isolated the first scorpion peptide inhibiting the NDV multiplication. It showed a dose dependent effect on NDV growth in vitro, with an IC 50 of 0.69 µM, and a low cytotoxicity on cultured Vero cells (CC50 > 55 µM). Furthermore, tests carried out in specific pathogen-free embryonated chicken eggs demonstrated that the isolated peptide has a protective effect on chicken embryos against NDV, and reduced by 73% the virus titer in allantoic fluid. The N-terminal sequence, as well as the number of cysteine residues of the isolated peptide, showed that it belongs to the scorpion venom Chlorotoxin-like peptides family, which led us to designate it "BotCl". Interestingly, at 10 µg/mL, BotCl showed an inhibiting effect three times higher than its analogue AaCtx, from Androctonus australis (Aa) scorpion venom, on NDV development. Altogether, our results highlight the chlorotoxin-like peptides as a new scorpion venom AMPs family.

Immunotherapy by blocking immune checkpoint regulators has emerged as a new targeted therapy for some cancers. Among them V-domain Ig suppressor of Tcell activation (VISTA) which is identified as a novel checkpoint regulator in ovarian cancer. This study aimed to investigate the VISTA role in Epithelial ovarian cancer (EOC), and its relationship with tumor-infiltrating lymphocytes (TILs) markers and its prognostic value. The expression of VISTA, CD3, CD8, CD4, FOXP3, and CD56 was assessed in 168 EOC tissue microarrays (TMA) by immunohistochemistry (IHC). In addition, associations between VISTA, TILs, clinicopathological variables, and overall survival (OS) were analyzed. VISTA expression in IGRov1 cells, as well as in PBMC of EOC patient, was evaluated by western blot. VISTA expression was detected in 64,28% of tissues, among which 42.3% were positive for tumor cells (TCs), and 47,9% were positive for immune cells (ICs). In univariate analysis, VISTA expression was significantly associated with a high density of TILs:CD3+ (p = 0,001), CD4+ (p = 0,002) and CD8+ (p≤0,001), in ICs but not in TCs. In terms of OS, multivariate analysis showed a significant association between the high density of CD8+ TILs and VISTA positive staining in ICs (p = 0,044), but not in TCs (p = 0,108). Kaplan-Meier curves demonstrated no correlation between VISTA expression and prolonged OS in both ICs (p = 0,841) and TCs (p = 0,090). Classification of EOC tumor microenvironment based on VISTA and CD8+TILs expression, demonstrated four immune subtypes: VISTA+/CD8+, VISTA+/CD8-, VISTA-/CD8+ and VISTA-/CD8-. The dual positive VISTA+/CD8+ subtype was significantly associated with prolonged OS in both TCs and ICs (p = 0,012 and p≤0,01, respectively), whereas patients with VISTA+/CD8- had the worst OS. Our results showed that VISTA is highly expressed in the IGRov1 cell line and LT-CD8 from a patient with EOC. Our results highlighted the association of VISTA expression and CD8+ TILs in EOC, with prolonged OS in patients with VISTA+/CD8+ and proposed VISTA as a potential immunotherapeutic target in EOC., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Jlassi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: The saliva of sand flies, vectors of Leishmania parasites, contains several components that exert pharmacological activity facilitating the acquisition of blood by the insect and contributing to the establishment of infection. Previously, we demonstrated that PpSP32 is the immunodominant salivary antigen in humans exposed to Phlebotomus papatasi bites and validated its usefulness as a predictive biomarker of disease. PpSP32, whose functions are little known to date, is an intriguing protein due to its involvement in the etiopathogenesis of pemphigus, an auto-immune disease. Herein, we aimed to better decipher its role through the screening of several immunomodulatory activity either on lymphocytes or on monocytes/macrophages., Methods: Peripheral mononuclear cells from healthy volunteers were stimulated with anti-CD3/anti-CD28 antibodies, phytohemagglutinin, phorbol 12-myristate 13-acetate/ionomycin, or lipopolysaccharide in the presence of increasing doses of PpSP32. Cell proliferation was measured after the addition of tritiated thymidine. Monocyte activation was tested by analyzing the expression of CD86 and HLA-DR molecules by flow cytometry. Cytokine production was analyzed in culture supernatants by ELISA. THP-1-derived macrophages were stimulated with LPS in the presence of increasing doses of PpSP32, and cytokine production was analyzed in culture supernatants by ELISA and multiplex technique. The effect of PpSP32 on NF-kB signaling was tested by Western blot. The anti-inflammatory activity of PpSP32 was assessed in vivo in an experimental inflammatory model of carrageenan-induced paw edema in rats., Results: Our data showed that PpSP32 down-modulated the expression of activation markers in LPS-stimulated monocytes and THP1-derived macrophages. This protein negatively modulated the secretion of Th1 and Th2 cytokines by human lymphocytes as well as pro-inflammatory cytokines by monocytes, and THP1-derived macrophages. PpSP32 treatment led to a dose-dependent reduction of IκB phosphorylation. When PpSP32 was injected into the paw of carrageenan-injected rats, edema was significantly reduced., Conclusions: Our data indicates that PpSP32 induces a potent immunomodulatory effect on monocytes and THP-1-derived macrophages. This inhibition could be mediated, among others, by the modulation of the NF-kB signaling pathway. The anti-inflammatory activity of PpSP32 was confirmed in vivo in the carrageenan-induced paw edema model in rats., (© 2023. The Author(s).)

Scorpion venom is a rich source of promising therapeutic compounds, such as highly selective ion channel ligands with potent pharmacological effects. Bot33 is a new short polypeptide of 38 amino acid residues with six cysteines purified from the venom of the Buthus occitanus tunetanus scorpion. Bot33 has revealed less than 40% identity with other known alpha-KTx families. This peptide displayed a neutral amino acid (Leucine), in the position equivalent to lysine 27, described as essential for the interaction with Kv channels. Bot33 did not show any toxicity following i.c.v. injection until 2 µg/kg mouse body weight. Due to its very low venom concentration (0.24%), Bot33 was chemically synthesized. Unexpectedly, this peptide has been subjected to a screening on ion channels expressed in Xenopus laevis oocytes, and it was found that Bot33 has no effect on seven Kv channel subtypes. Interestingly, an in silico molecular docking study shows that the Leu27 prevents the interaction of Bot33 with the Kv1.3 channel. All our results indicate that Bot33 may have a different mode of action from other scorpion toxins, which will be interesting to elucidate.

Potassium ion channels are expressed on the cell membranes, implicated in wide variety of cell functions and intimately linked to cancer cell behaviors. This work reports the first bioplatform described to date allowing simple and rapid detection of ion channel activity and the effect of their inhibitors in cancer cells. The methodology involves interrogation of the channel of interest from cells specifically captured on magnetic immunoconjugates using specific detection antibodies that are labeled with horseradish peroxidase enzyme. The channel activity is reflected by an amperometric signal transduction of the resulting magnetic bioconjugates onto screen-printed carbon electrodes. The bioplatform feasibility was proven for the detection of the Kv channels in U87 human glioblastoma cells and their blocking by scorpion venom KAaH1 and KAaH2 peptides. The obtained results confirm the high sensitivity (detection of 5 U87 cells⋅mL -1 and 0.06 μg mL -1 of KAaH2) of the proposed bioplatform and their versatility to detect both potassium channel activity and their potential inhibitors, in a given cancer cell line, with high sensitivity in a simple and fast way. This bioplatform presents potential applications in cancer and theranostic of channelopathies., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Noureddine Raouafi reports financial support was provided by Tunisian Ministry of Higher Education and Scientific Research., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Inflammatory breast cancer (IBC) is the most pro-metastatic form of breast cancer (BC). We previously demonstrated that protein overexpression of Myristoylated Alanine-Rich C Kinase Substrate (MARCKS) protein was associated with shorter survival in IBC patients. MARCKS has been associated with the PI3K/AKT pathway. MARCKS inhibitors are in development. Our objective was to investigate MARCKS, expressed preferentially in IBC that non-IBC (nIBC), as a novel potential therapeutic target for IBC. The biologic activity of MPS, a MARCKS peptide inhibitor, on cell proliferation, migration, invasion, and mammosphere formation was evaluated in IBC (SUM149 and SUM190) and nIBC (MDA-MB-231 and MCF7) cell lines, as well as its effects on protein expression in the PTEN/AKT and MAPK pathways. The prognostic relevance of MARCKS and phosphatase and tensin homolog (PTEN) protein expression as a surrogate marker of metastasis-free survival (MFS) was evaluated by immunohistochemistry (IHC) in a retrospective series of archival tumor samples derived from 180 IBC patients and 355 nIBC patients. In vitro MPS impaired cell proliferation, migration and invasion, and mammosphere formation in IBC cells. MARCKS inhibition upregulated PTEN and downregulated pAKT and pMAPK expression in IBC cells, but not in nIBC cells. By IHC, MARCKS expression and PTEN expression were negatively correlated in IBC samples and were associated with shorter MFS and longer MFS, respectively, in multivariate analysis. The combination of MARCKS-/PTEN+ protein status was associated with longer MFS in IBC patient only ( p = 8.7 × 10 -3 ), and mirrored the molecular profile (MARCKS-downregulated/PTEN-upregulated) of MPS-treated IBC cell lines. In conclusion, our results uncover a functional role of MARCKS implicated in IBC aggressiveness. Associated with the good-prognosis value of the MARCKS-/PTEN+ protein status that mirrors the molecular profile of MPS-treated IBC cell lines, our results suggest that MARCKS could be a potential therapeutic target in patients with MARCKS-positive IBC. Future preclinical studies using a larger panel of IBC cell lines, animal models and analysis of a larger series of clinical samples are warranted in order to validate our results.

In the present study, we assess tyrosol derivatives bearing 3,5-disubstituted isoxazoles and 1,4-disubstituted triazoles for their ability to inhibit the proliferation of K562 cells derived from leukemia as well as primary chronic myeloid leukemia (CML) cells obtained from the peripheral blood of 15 CML patients including 10 patients with untreated chronic phase and 5 patients with resistance against imatinib or multiple TKI. Our results showed that most derivatives displayed significant anti-proliferative activity against K562 cells in a dose-dependent manner. Among them, compounds 3d and 4a exhibited greater potent anticancer activity with respective IC 50 values of 16 and 18 µg/mL (45 µM and 61 µM). Interestingly, compound 3d inhibited CML cell proliferation not only in newly diagnosed but also in imatinib-resistant patients. We demonstrated that the anti-proliferative effect of this compound is mediated by a pro-apoptotic activity by promoting oxidative stress and modulating the activity of the Akt, p38 MAPK and Erk 1/2 pathways. In conclusion, our data highlight the potential of this class of derivative as a novel promising therapeutic agent for CML therapy.

Background: Multiple sclerosis (MS) is characterized by a combination of inflammatory and demyelination processes in the spinal cord and brain. Conventional drugs generally target the autoimmune response, without any curative effect. For that reason, there is a great interest in identifying novel agents with anti-inflammatory and myelinating effects, to counter the inflammation and cell death distinctive of the disease., Methods and Results: An in vitro assay showed that curcumin (Cur) at 10 µM enhanced the proliferation of C8-D1A cells and modulated the production of Th1/Th2/Th17 cytokines in the cells stimulated by LPS. Furthermore, two in vivo pathophysiological experimental models were used to assess the effect of curcumin (100 mg/kg). The cuprizone model mimics the de/re-myelination aspect in MS, and the experimental autoimmune encephalomyelitis model (EAE) reflects immune-mediated events. We found that Cur alleviated the neurological symptomatology in EAE and modulated the expression of lymphocytes CD3 and CD4 in the spinal cord. Interestingly, Cur restored motor and behavioral deficiencies, as well as myelination, in demyelinated mice, as indicated by the higher index of luxol fast blue (LFB) and the myelin basic protein (MBP) intensity in the corpus callosum., Conclusions: Curcumin is a potential therapeutic agent that can diminish the MS neuroimmune imbalance and demyelination through its anti-inflammatory and antioxidant effects.

Natural polyphenols are widely reported to have a large range of pharmacological properties, especially antioxidant activities and free radical scavenging capacities. In this study, we investigate the effects of naringin, chlorogenic acid, and quercetin mixtures (NCQ) on renal fibrosis in streptozotocin (STZ)-induced diabetic aged rats and its underlying mechanisms for ten consecutive weeks. The oxidative defense system in the kidneys of treated rats was found to be improved. Several biomarkers were investigated including the blood urea nitrogen, creatinine, and uric acid. Moreover, antioxidant parameters were evaluated and we found that superoxide dismutase, catalase, glutathione peroxidase, Na + -K + -ATPase activities, the nitric oxide production, the protein carbonyl, the advanced oxidation protein products, lipid peroxidation, and reduced glutathione levels were all significantly balanced and close to control values. In addition, NCQ restored renal injuries and fibrosis as assessed by histological method and molecular biology investigation of the matrix metalloproteinase, the transforming growth factor-beta TGF-β, the tumor necrosis factor TNFα, and p53 expression. Our study proposes the NCQ combination as potential plant-derived bioactive compounds to prevent diabetic nephropathy.

PIVL is a Kunitz-type serine protease inhibitor that was previously characterized from Tunisian snake venom, Macrovipera lebetina transmediterranea . It reduced glioblastoma cells' development and significantly blocked angiogenesis in in-vitro and ex-vivo models. PIVL exerted these effects by interfering with αvβ3 integrin. In order to produce a biological active recombinant, the cDNA cloning and expression of PIVL was performed in Escherichia coli (BL21)-DE3 cells using pET-22b (+) vector. The recombinant PIVL protein (rPIVL) was purified by nickel affinity chromatography and has recognized monoclonal anti-His antibody. Functionally, rPIVL exhibited potent anti-tumor cell effects as well as anti-angiogenesis properties. Interestingly, we found that both native PIVL (nPIVL) and rPIVL modulated PI3/AKT and MAPK signaling pathways. In all, our results showed that we have successfully expressed the first active anti-oncogenic snake venom Kunitz-type protease inhibitor that can be a potential therapeutic drug against glioblastoma, in its native or recombinant form.

The Receptor Binding Domain (RBD) of SARS-CoV-2 virus harbors a sequence of Arg-Gly-Asp tripeptide named RGD motif, which has also been identified in extracellular matrix proteins that bind integrins as well as other disintegrins and viruses. Accordingly, integrins have been proposed as host receptors for SARS-CoV-2. However, given that the microenvironment of the RGD motif imposes a structural hindrance to the protein-protein association, the validity of this hypothesis is still uncertain. Here, we used normal mode analysis, accelerated molecular dynamics microscale simulation, and protein-protein docking to investigate the putative role of RGD motif of SARS-CoV-2 RBD for interacting with integrins. We found, that neither RGD motif nor its microenvironment showed any significant conformational shift in the RBD structure. Highly populated clusters of RBD showed no capability to interact with the RGD binding site in integrins. The free energy landscape revealed that the RGD conformation within RBD could not acquire an optimal geometry to allow the interaction with integrins. In light of these results, and in the event where integrins are confirmed to be host receptors for SARS-CoV-2, we suggest a possible involvement of other residues to stabilize the interaction., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Othman, Messaoud, Khamessi, Ben-Mabrouk, Ghedira, Bharuthram, Treurnicht, Achilonu, Sayed and Srairi-Abid.)

Glioblastoma is the most aggressive and invasive form of central nervous system tumors due to the complexity of the intracellular mechanisms and molecular alterations involved in its progression. Unfortunately, current therapies are unable to stop its neoplastic development. In this context, we previously identified and characterized AaTs-1, a tetrapeptide (IWKS) from Androctonus autralis scorpion venom, which displayed an anti-proliferative effect against U87 cells with an IC 50 value of 0.57 mM. This peptide affects the MAPK pathway, enhancing the expression of p53 and altering the cytosolic calcium concentration balance, likely via FPRL-1 receptor modulation. In this work, we designed and synthesized new dendrimers multi-branched molecules based on the sequence of AaTs-1 and showed that the di-branched (AaTs-1-2B), tetra-branched (AaTs-1-4B) and octo-branched (AaTs-1-8B) dendrimers displayed 10- to 25-fold higher effects on the proliferation of U87 cells than AaTs-1. We also found that the effects of the newly designed molecules are mediated by the enhancement of the ERK1/2 and AKT phosphorylated forms and by the increase in p53 expression. Unlike AaTs-1, AaTs-1-8B and especially AaTs-1-4B affected the migration of the U87 cells. Thus, the multi-branched peptide synthesis strategy allowed us to make molecules more active than the linear peptide against the proliferation of U87 glioblastoma cells.

Mint species ( Lamiaceae family) have been used as traditional remedies for the treatment of several diseases. In this work, we aimed to characterize the biological activities of the total phenolic and flavonoid contents of Mentha pulegium L. extracts collected from two different regions of Tunisia. The highest amounts of total phenols (74.45 ± 0.01 mg GAE/g DW), flavonoids (28.87 ± 0.02 mg RE/g DW), and condensed tannins (4.35 ± 0.02 mg CE/g DW) were found in the Bizerte locality. Methanolic leaf extracts were subjected to HPLC-UV analysis in order to identify and quantify the phenolic composition. This technique allowed us to identify seven phenolic compounds: two phenolic acids and five flavonoid compounds, such as eriocitrin, hesperidin, narirutin, luteolin, and isorhoifolin, which were found in both extracts with significant differences between samples collected from the different regions ( p < 0.05). Furthermore, our results showed that the methanolic extract from leaves collected from Bizerte had the highest antioxidant activities (DPPH IC 50 value of 16.31 μg/mL and 570.08 μmol Fe 2+ /g, respectively). Both extracts showed high radical-scavenging activity as well as significant antimicrobial activity against eight tested bacteria. The highest antimicrobial activities were observed against Gram-positive bacteria with inhibition zone diameters and MIC values ranging between 19 and 32 mm and 40 and 160 µg/mL, respectively. Interestingly, at 10 μg/mL, the extract had a significant effect on cell proliferation of U87 human glioblastoma cells. These findings open perspectives for the use of Mentha pulegium L. extract in green pharmacy, alternative/complementary medicine, and natural preventive therapies for the development of effective antioxidant, antibacterial, and/or antitumoral drugs.

Snake venom proteins, which are responsible for deadly snakebite envenomation, induce severe injuries including neurotoxicity, myotoxicity, cardiotoxicity, hemorrhage, and the disruption of blood homeostasis. Yet, many snake-venom proteins have been developed as potential drugs for treating human diseases due to their pharmacological effects. In this study, we evaluated the use of, an L-amino acid oxidase isolated from Cerastes cerastes snake venom CC-LAAO, as a potential anti-glioblastoma drug, by investigating its in vivo and in vitro pharmacological effects. Our results showed that acute exposure to CC-LAAO at 1 and 2.5 µg/mL does not induce significant toxicity on vital organs, as indicated by the murine blood parameters including aspartate transaminase (AST), alanine transaminase (ALT), lactate dehydrogenase (LDH) activities, and creatinine levels. The histopathological examination demonstrated that only at high concentrations did CC-LAAO induce inflammation and necrosis in several organs of the test subjects. Interestingly, when tested on human glioblastoma U87 cells, CC-LAAO induced a dose-dependent apoptotic effect through the H 2 O 2 generated during the enzymatic reaction. Taken altogether, our data indicated that low concentration of CC-LAAO may be safe and may have potential in the development of anti-glioblastoma agents.

Glioblastoma is an aggressive cancer, against which medical professionals are still quite helpless, due to its resistance to current treatments. Scorpion toxins have been proposed as a promising alternative for the development of effective targeted glioblastoma therapy and diagnostic. However, the exploitation of the long peptides could present disadvantages. In this work, we identified and synthetized AaTs-1, the first tetrapeptide from Androctonus australis scorpion venom (Aa), which exhibited an antiproliferative effect specifically against human glioblastoma cells. Both the native and synthetic AaTs-1 were endowed with the same inhibiting effect on the proliferation of U87 cells with an IC 50 of 0.56 mM. Interestingly, AaTs-1 was about two times more active than the anti-glioblastoma conventional chemotherapeutic drug, temozolomide (TMZ), and enhanced its efficacy on U87 cells. AaTs-1 showed a significant similarity with the synthetic peptide WKYMVm, an agonist of a G-coupled formyl-peptide receptor, FPRL-1, known to be involved in the proliferation of glioma cells. Interestingly, the tetrapeptide triggered the dephosphorylation of ERK, p38, and JNK kinases. It also enhanced the expression of p53 and FPRL-1, likely leading to the inhibition of the store operated calcium entry. Overall, our work uncovered AaTs-1 as a first natural potential FPRL-1 antagonist, which could be proposed as a promising target to develop new generation of innovative molecules used alone or in combination with TMZ to improve glioblastoma treatment response. Its chemical synthesis in non-limiting quantity represents a valuable advantage to design and develop low-cost active analogues to treat glioblastoma cancer.

Hepatocellular carcinoma (HCC) is the most common primary liver cancer in adults, the fifth most common malignancy worldwide and the third leading cause of cancer related death. An alternative to the surgical treatments and drugs, such as sorafenib, commonly used in medicine is necessary to overcome this public health problem. In this study, we determine the anticancer effect on HCC of Moroccan cobra Naja haje venom and its fraction obtained by gel filtration chromatography against Huh7.5 cancer cell line. Cells were grown together with WI38 human fibroblast cells, LX2 human hepatic stellate cell line, and human endothelial cells (HUVEC) in MCTS (multi-cellular tumor spheroids) models. The hepatotoxicity of venom and its fractions were also evaluated using the normal hepatocytes cell line (Fa2N-4 cells). Our results showed that an anti HCC activity of Moroccan cobra Naja haje venom and, more specifically, the F7 fraction of gel filtration chromatography exhibited the greatest anti-hepatocellular carcinoma effect by decreasing the size of MCTS. This effect is associated with a low toxicity against normal hepatocytes. These results strongly suggest that the F7 fraction of Moroccan cobra Naja haje venom obtained by gel filtration chromatography possesses the ability to inhibit cancer cells proliferation. More research is needed to identify the specific molecule(s) responsible for the anticancer effect and investigate their mechanism of action.

Alzheimer's disease (AD) is one of the most common neurodegenerative diseases leading to dementia. Despite research efforts, currently there are no effective pharmacotherapeutic options for the prevention and treatment of AD. Recently, numerous studies highlighted the beneficial effects of curcumin (CUR), a natural polyphenol, in the neuroprotection. Especially, its dual antioxidant and anti-inflammatory properties attracted the interest of researchers. In fact, besides its antioxidant and anti-inflammatory properties, this biomolecule is not degraded in the intestinal tract. Additionally, CUR is able to cross the blood-brain barrier and could therefore to be used to treat neurodegenerative pathologies associated with oxidative stress, inflammation and apoptosis. The present study aimed to assess the ability of CUR to induce neuronal protective and/or recovery effects on a rat model of neurotoxicity induced by aluminum chloride (AlCl 3 ), which mimics the sporadic form of Alzheimer's disease. Our results showed that treatment with CUR enhances pro-oxidant levels, antioxidant enzymes activities and anti-inflammatory cytokine production and decreases apoptotic cells in AlCl 3 -exposed hippocampus rats. Additionally, histopathological analysis of hippocampus revealed the potential of CUR in decreasing the hallmarks in the AlCl 3 -induced AD. We also showed that CUR post-treatment significantly improved the behavioral, oxidative stress and inflammation in AlCl 3 -exposed rats. Taken together, our data presented CUR as a nutraceutical potential through its protective effects that are more interesting than recovery ones in sporadic model of AD.

The excessive production of inflammatory mediators results in an overactive immune response leading to the worsening of various human diseases. Thus, there is a still need to identify molecules able to regulate the inflammatory response. Lebecetin, a C-type lectin protein isolated from Macrovipera lebetina snake venom, was previously characterized as a platelet aggregation inhibitor and antitumor active biomolecule. In the present work, we investigated its effect on the production of some cytokines linked to inflammatory response and the underlying mechanisms in lipopolysaccharide (LPS)-induced THP1 macrophages. Interestingly, we found that lebecetin reduced the levels of the pro-inflammatory cytokines TNF-α, IL-6, and IL-8 while it partially increased LPS-induced secretion of the immunomodulatory cytokine IL-10. Furthermore, this modulatory effect was accompanied by decreased activation of ERK1/2, p38, AKT kinases and NF-κB along with reduced expression of αvβ3 integrin. Thus, this study highlights the promising role of lebecetin as a natural biomolecule that could manage the inflammatory response involved in the development and progression of inflammatory diseases., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The spread of COVID-19 caused by the SARS-CoV-2 outbreak has been growing since its first identification in December 2019. The publishing of the first SARS-CoV-2 genome made a valuable source of data to study the details about its phylogeny, evolution, and interaction with the host. Protein-protein binding assays have confirmed that Angiotensin-converting enzyme 2 (ACE2) is more likely to be the cell receptor through which the virus invades the host cell. In the present work, we provide an insight into the interaction of the viral spike Receptor Binding Domain (RBD) from different coronavirus isolates with host ACE2 protein. By calculating the binding energy score between RBD and ACE2, we highlighted the putative jump in the affinity from a progenitor form of SARS-CoV-2 to the current virus responsible for COVID-19 outbreak. Our result was consistent with previously reported phylogenetic analysis and corroborates the opinion that the interface segment of the spike protein RBD might be acquired by SARS-CoV-2 via a complex evolutionary process rather than a progressive accumulation of mutations. We also highlighted the relevance of Q493 and P499 amino acid residues of SARS-CoV-2 RBD for binding to human ACE2 and maintaining the stability of the interface. Moreover, we show from the structural analysis that it is unlikely for the interface residues to be the result of genetic engineering. Finally, we studied the impact of eight different variants located at the interaction surface of ACE2, on the complex formation with SARS-CoV-2 RBD. We found that none of them is likely to disrupt the interaction with the viral RBD of SARS-CoV-2., Competing Interests: Declaration of competing interest No conflicts of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Prostate cancer is the most highly diagnosed cancer in men worldwide. It is characterized by high proliferation, great invasion and metastatic potential. Sodium channel subtypes have been identified as highly expressed in different prostate cancer cell lines. In this study, we have screened the negatively charged fractions of Androctonus australis (Aa) scorpion venom to identify active peptides on DU145 prostate cancer cells proliferation. The most active compound was identified to be the sodium channel peptide AaHIV with an IC 50 value of 15 μM. At this concentration, AaHIV had low effect on the adhesion of DU145 cells to fibronectin. When compared to other Na + channel Aa toxins, AaHIV was found to be 2 times more active than AaHI and AaHII on DU145 cells proliferation and slightly less active than AaHII on their adhesion. The three peptides are inactive on DU145 cells migration. AaHIV was found to be 16 times more active than veratridine, asteroidal alkaloid from plants of the lily family widely used as a sodium channel activator. Electrophysiological experiments showed that the AaHIV toxin activates Nav1.6 channel, suggesting that this sodium channel subtype is implicated in the proliferation of DU145 prostate cancer cells., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The use of predictive biomarkers provides potential individualized cancer therapeutic options to prevent therapy failure as well as serious toxicities. Several recent studies showed that predictive and prognostic biomarkers are a notable personalized strategy to improve patients' care in several cancers. Trabectedin (Yondelis®) is a cytotoxic agent, derived from a marine organism, harbouring a significant antitumor activity against several cancers such as soft tissue sarcoma, ovarian, and breast cancers. Recently and with the advent of molecular genetic testing, BRCA mutational status was found as an important predictor of response to this anticancer drug, especially in gynecological cancers. The aim of this updated review is to discuss the mechanisms of action of trabectedin against the wellknown cancer hallmarks described until today. The current advances were also examined related to genomic biomarkers that can be used in the future to predict the efficacy of this potent anticancer natural molecule in various gynecological cancers., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

We previously reported that immunoreactivity of recombinant CFP32 (Rv0577), a virulence factor of Mycobacterium tuberculosis, was higher when produced in transformed Pichia pastoris as compared to transformed E. coli. In this study, we show that this difference is partly due to the N-glycosylation of the recombinant CFP32 (rCFP32) by the yeast Pichia pastoris. In addition, SDS-PAGE and western blotting analysis of Mycobacterium bovis BCG and yeast-produced rCFP32 showed the presence of a band corresponding to a homodimeric state of the protein, unlike that of rCFP32 produced in E. coli. Computational modeling indicates that a single cysteine residue at position 193 of each monomer might bond to stabilize the homodimeric state of CFP32. Computational study showed that this residue is buried inside the protein core of E. coli-produced rCFP32 suggesting that rCFP32 may adopt a different folding in P. pastoris and BCG, in which C193 is solvent exposed. Surprisingly, an enzyme-linked immunosorbent assay using a generated monoclonal antibody (14D4) reveals the presence of a differential epitope that appears to be the consequence of the protein dimerization of the yeast- and BCG-, but not E.coli- produced, CFP32 recombinant form. We conclude that, in addition to N-glycosylation, homodimeric folding significantly enhances the immunoreactivity of rCFP32 and may these post-translational modifications may factor into the structure and function of native M. tuberculosis CFP32., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Scorpion toxins have been the subject of many studies exploring their pharmacological potential. The high affinity and the overall selectivity to various types of ionic channels endowed scorpion toxins with a potential therapeutic effect against many channelopathies. These are diseases in which ionic channels play an important role in their development. Cancer is considered as a channelopathy since overexpression of some ionic channels was highlighted in many tumor cells and was linked to the pathology progression. Interestingly, an increasing number of studies have shown that scorpion venoms and toxins can decrease cancer growth in vitro and in vivo. Furthermore through their ability to penetrate the cell plasma membrane, certain scorpion toxins are able to enhance the efficiency of some clinical chemotherapies. These observations back-up the applicability of scorpion toxins as potential cancer therapeutics. In this review, we focused on the anti-cancer activity of scorpion toxins and their effect on the multiple hallmarks of cancer. We also shed light on effectors and receptors involved in signaling pathways in response to scorpion toxins effect. Until now, the anticancer mechanisms described for scorpion peptides consist on targeting ion channels to (i) inhibit cell proliferation and metastasis; and (ii) induce cell cycle arrest and/or apoptosis through membrane depolarization leading to hemostasis deregulation and caspase activation. Putative targets such as metalloproteinases, integrins and/or growth factor receptors, beside ion channels, have been unveiled to be affected by scorpion peptides., (Copyright © 2019. Published by Elsevier Ltd.)

Resistance to 5-Fluorouracil chemotherapy is a major cause of therapeutic failure in colon cancer cure. Development of combined therapies constitutes an effective strategy to inhibit cancer cells and prevent the emergence of drug resistance. For this purpose, we investigated the anti-tumoral effect of thirteen phenolic compounds, from the Tunisian quince Cydonia oblonga Miller, alone or combined to 5-FU, on the human 5-FU-resistant LS174-R colon cancer cells in comparison to parental cells. Our results showed that only Kaempferol was able to chemo-sensitize 5-FU-resistant LS174-R cells. This phenolic compound combined with 5-FU exerted synergistic inhibitory effect on cell viability. This combination enhanced the apoptosis and induced cell cycle arrest of both chemo-resistant and sensitive cells through impacting the expression levels of different cellular effectors. Kaempferol also blocked the production of reactive oxygen species (ROS) and modulated the expression of JAK/STAT3, MAPK, PI3K/AKT and NF-κB. In silico docking analysis suggested that the potent anti-tumoral effect of Kaempferol, compared to its two analogs (Kaempferol 3-O-glucoside and Kampferol 3-O-rutinoside), can be explained by the absence of glucosyl groups. Overall, our data propose Kaempferol as a potential chemotherapeutic agent to be used alone or in combination with 5-FU to overcome colon cancer drug resistance.

Scorpion peptides are well known for their pharmaceutical potential on different targets. These include mainly the ion channels which were found to be highly expressed in many diseases, including cancer, auto-immune pathologies and Alzheimer. So far, however, the disintegrin activity had only been characterized for snake venom molecules. Herein, we present the first short peptide, purified from the venom of Buthus occitanus tunetanus, (termed RK) able to inhibit the cell adhesion of Glioblastoma, Melanoma and Rat pheochromocytoma to different extracellular matrix (ECM) receptors. Anti-integrin antibody assay suggests that RK interacts with both α 1 β 1 and α v β 3 with a more pronounced effect for the former. The examination of the primary structure of RK suggests the involvement of two motifs: KSS, analogue to KTS which was characterized for α 1 β 1 Snake venom disintegrins, and ECD, analogue to RGD which was found to be active on α v β 3 . To assess their roles in the disintegrin activity of RK, we conducted a computational analysis. The molecular docking study shows that RK involves mainly two segments to interact with the α 1 β 1 integrin, but the peptide does not implicate the KSS motif in the interaction. The molecular modeling study, suggests the key contribution of the ECD segment in the interaction with α v β 3 integrin., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Snail secretion is currently revolutionizing the world of cosmetics and human skin care. The efficacy of snail secretion in wounds healing has been proven both in vitro and by clinical studies. However, the potential anti-tumor effect of snail secretion was poorly investigated. In this report, our in vitro study showed that Helix aspersa maxima species snail slime (SS) could not only treat melanogenesis but also endowed with anti-tumoral activity against human melanoma cells. Indeed, SS reduced melanin content and tyrosinase activity on B16F10 cells with IC 50 values of 288 μg/mL and 286 μg/mL, respectively, without altering cell viability. This effect was also observed, at a lesser extent, on human melanoma IGR-39 and SK-MEL-28 cell lines. On another hand, SS specifically inhibited the viability of IGR-39 and SK-MEL-28 cells associated to an apoptotic effect highlighted by PARP cleavage. It is worth to note that SS did not affect the viability of B16F10 cells and non tumorigenic HaCaT cells. Interestingly, this extract was found to inhibit migration and invasion of both human melanoma cells through reducing the expression of Matrix metalloproteinase MMP2. Snail slime also exerted a high inhibitory effect on IGR-39 cell adhesion through blocking the function of α 2 β 1 (45%), α v β 3 (38%) integrins and by reducing the expression levels of α v and β 1 integrins. The presented results shed light on the potential anti-melanoma effect of SS and support its use against skin diseases., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Voltage-gated potassium (Kv) channels are known to play a pivotal role in the progression of various cancer types and considered as new targets for designing anti-cancer therapy. However, the fact that many Kv channels are expressed in different cell lines makes it difficult to ascribe a functional role for a given Kv channel on a specific aspect of the tumorogenesis. In this work, we showed that although both Kv1.1 and Kv1.3 channels are expressed in U87 (glioblastoma), MDA-MB-231 (breast cancer) and LS174 (colon adenocarcinoma) cells, these respond differently to KAaH1 or KAaH2, two homologous Kv1 blockers from scorpion venom. KAaH1 is active on Kv1.1 and Kv1.3 and was found to inhibit migration and adhesion of U87 cells whereas KAaH2 which is slightly active only on Kv1.1 channel, inhibits their proliferation via the EGFR signaling pathway. The correlation between the electro-physiological activity of the scorpion peptides and their anti-migratory effects suggests the involvement of the Kv1.1 and Kv1.3 channels in the mobility of the three cancer cell lines. Our results showed that besides they can elucidate the implication of Kv1.1 and Kv1.3 channels in molecular mechanisms of neoplastic progression, KAaH1 and KAaH2 may be used as therapeutic tools against glioblastoma., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Here we report the cDNA cloning of a phospholipase A 2 (PLA 2 ) from five Sparidae species. The deduced amino acid sequences show high similarity with pancreatic PLA 2 . In addition, a phylogenetic tree derived from alignment of various available sequences revealed that Sparidae PLA 2 are closer to avian PLA 2 group IB than to mammals' ones. In order to understand the structure-function relationships of these enzymes, we report here the recombinant expression in E.coli, the refolding and characterization of His-tagged annular seabream PLA 2 (AsPLA 2 ). A single Ni-affinity chromatography step was used to obtain a highly purified recombinant AsPLA 2 with a molecular mass of 15kDa as attested by gel electrophoresis and MALDI-TOF mass spectrometry data. The enzyme has a specific activity of 400U.mg -1 measured on phosphatidylcholine at pH 8.5 and 50°C. The enzyme high thermo-activity and thermo-stability make it a potential candidate in various biological applications. The 3D structure models of these enzymes were compared with structures of phylogenetically related pancreatic PLA 2 . By following these models and utilizing molecular dynamics simulations, the resistance of the AsPLA 2 at high temperatures was explained. Using the monomolecular film technique, AsPLA 2 was found to be active on various phospholipids spread at the air/water interface at a surface pressure between 12 and 25dyncm -1 . Interestingly, this enzyme was shown to be mostly active on dilauroyl-phosphatidylglycerol monolayers and this behavior was confirmed by molecular docking and dynamics simulations analysis. The discovery of a thermo-active new member of Sparidae PLA 2 , provides new insights on structure-activity relationships of fish PLA 2 ., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: The resistance of melanoma cells to cisplatin restricts its clinical use. Therefore, the search for novel tumor inhibitors and effective combination treatments that sensitize tumor cells to this drug are still needed. We purified macrovipecetin, a novel heterodimeric C-type lectin, from Macrovipera lebetina snake venom and investigated its anti-tumoral effect on its own or combined with cisplatin, in human melanoma cells., Methods: Biochemical characterization, in vitro cells assays such as viability, apoptosis, adhesion, migration, invasion, Western blotting and in silico analysis were used in this study., Results: Macrovipecetin decreased melanoma cell viability 100 times more than cisplatin. Interestingly, when combined with the drug, macrovipecetin enhanced the sensitivity of SK-MEL-28 cells by augmenting their apoptosis through increased expression of the apoptosis inducing factor (AIF) and activation of ERK 1/2 , p38, AKT and NF-κB. Moreover, macrovipecetin alone or combined with cisplatin induced the expression of TRADD, p53, Bax, Bim and Bad and down-regulated the Bcl-2 expression and ROS levels in SK-MEL-28 cells. Interestingly, these treatments impaired SK-MEL-28 cell adhesion, migration and invasion through modulating the function and expression of αvβ3 integrin along with regulating E-cadherin, vimentin, β-catenin, c-Src and RhoA expression. In silico study suggested that only the α chain of macrovipecetin interacts with a region overlapping the RGD motif binding site on this integrin., Conclusions: We validated the antitumor effect of macrovipecetin when combined, or not, with cisplatin on SK-MEL-28 cells., General Significance: The presented work proposes the potential use of macrovipecetin and cisplatin in combination as an effective anti-melanoma treatment., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Glioblastoma is the deadliest type of brain cancer. Treatment could target the Matrix metalloproteinase-2 (MMP-2), which is known to be involved in the invasion process of glioblastoma cells. But current available inhibitors are not selective to MMP-2 due to their interaction with the catalytic binding site, which is highly conserved in all MMPs structures. Interestingly, members of the chloride channel blocker scorpion toxins, such as chlorotoxin (ClTx) and AaCTx, inhibit glioblastoma cell invasion and show a promising therapeutic potential. Indeed, it has been shown that CITx inhibits selectively MMP-2 and was also able to cross the blood brain and tissue barriers. Although ClTx and AaCTx show high sequence similarity, AaCTx is ten times less active than ClTx. By using molecular modeling, molecular dynamics and MM-PB(GB)SA free energy estimation, we present the first computational study reporting the interaction mode of ClTx/AaCTx with MMP-2. We found that the two peptides probably act on an exosite of MMP-2 comprising mainly residues from the collagen binding domain, a feature that could be exploited to enhance the selectivity toward MMP-2. van der Waals and hydrophobic forces are the primary mediators of this interaction. The N- and C-termini of the two peptides harbor the key residues of the interaction spread across a conserved amino acid patch. In particular, F6 contributes mostly to the binding free energy in ClTx. We also suggest that the lack of the C-terminal arginine and the residues P10 and R24, might be responsible for altering the activity of AaCTx toward glioblastoma cells compared to ClTx.

Novel phospholipase (PLA 2 ) genes from the Sparidae family were cloned. The sequenced PLA 2 revealed an identity with pancreatic PLA 2 group IB. To better understand the structure/function relationships of these enzymes and their evolution, the Diplodus annularis PLA 2 (DaPLA 2 ) was overexpressed in E. coli. The refolded enzyme was purified by Ni-affinity chromatography and has a molecular mass of 15 kDa as determined by MALDI-TOF spectrometry. Interestingly, unlike the pancreatic type, the DaPLA 2 was active and stable at higher temperatures, which suggests its great potential in biotechnological applications. The 3D structure of DaPLA 2 was constructed to gain insights into the functional properties of sparidae PLA 2 . Molecular docking and dynamic simulations were performed to explain the higher thermal stability and the substrate specificity of DaPLA 2 . Using the monolayer technique, the purified DaPLA 2 was found to be active on various phospholipids ranging from 10 to 20 mN·m -1 , which explained the absence of the hemolytic activity for DaPLA 2 .

Lebein, is an heterodimeric disintegrin isolated from Macrovipera lebetina snake venom that was previously characterized as an inhibitor of ADP-induced platelet aggregation. In this study, we investigated the effect of Lebein on the p53-dependent growth of human colon adenocarcinoma cell lines. We found that Lebein significantly inhibited LS174 (p53wt), HCT116 (p53wt), and HT29 (p53mut) colon cancer cell viability by inducing cell cycle arrest through the modulation of expression levels of the tumor suppression factor p53, cell cycle regulating proteins cyclin D1, CDK2, CDK4, retinoblastoma (Rb), CDK1, and cyclin-dependent kinase inhibitors p21 and p27. Interestingly, Lebein-induced apoptosis of colon cancer cells was dependent on their p53 status. Thus, in LS174 cells, cell death was associated with PARP cleavage and the activation of caspases 3 and 8 while in HCT116 cells, Lebein induced caspase-independent apoptosis through increased expression of apoptosis inducing factor (AIF). In LS174 cells, Lebein triggers the activation of the MAPK ERK1/2 pathway through induction of reactive oxygen species (ROS). It also decreased cell adhesion and migration to fibronectin through down regulation of α5β1 integrin. Moreover, Lebein significantly reduced the expression of two angiogenesis stimulators, Vascular Endothelial Growth Factor (VEGF) and Neuropilin 1 (NRP1). It inhibited the VEGF-induced neovascularization process in the quail embryonic CAM system and blocked the development of human colon adenocarcinoma in nude mice. Overall, our work indicates that Lebein may be useful to design a new therapy against colon cancer. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

The present study investigated the kinetic and interfacial properties of two secreted phospholipases isolated from Tunisian vipers'venoms: Cerastes cerastes (CC-PLA2) and Macrovipera lebetina transmediterranea (MVL-PLA2). Results show that these enzymes have great different abilities to bind and hydrolyse phospholipids. Using egg-yolk emulsions as substrate at pH 8, we found that MVL-PLA2 has a specific activity of 1473U/mg at 37°C in presence of 1mM CaCl2. Furthermore the interfacial kinetic and binding data indicate that MVL-PLA2 has a preference to the zwitterionic phosphatidylcholine monolayers (PC). Conversely, CC-PLA2 was found to be able to hydrolyse preferentially negatively charged head group phospholipids (PG and PS) and exhibits a specific activity 9 times more important (13333U/mg at 60°C in presence of 3mM CaCl2). Molecular models of both CC-PLA2 and MVL-PLA2 3D structures have been built and their electrostatic potentials surfaces have been calculated. A marked anisotropy of the overall electrostatic charge distribution leads to a significantly difference in the dipole moment intensity between the two enzymes explaining the great differences in catalytic and binding properties, which seems to be governed by the electrostatic and hydrophobic forces operative at the surface of the two phospholipases., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Kbot55 is a 39 amino acid peptide isolated from the venom of the Tunisian scorpion Buthus occitanus tunetanus. This peptide is cross-linked by 3 disulfide bridges and has a molecular mass of 4128.65Da. Kbot55 is very low represented in the venom and thus represents a challenge for biochemical characterization. In this study, Kbot55 has been subjected to a screening on ion channels expressed in Xenopus laevis oocytes. It was found that Kbot55 targets voltage-gated potassium channels with high affinity. Kbot55 shows very low amino acid identity with other scorpion potassium toxins and therefore was considered a bona fide novel type of scorpion toxin. Sequence alignment analysis indicated that Kbot55 is the first representative of the new α-Ktx31 subfamily and therefore was classified as α-Ktx31.1., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Angiogenesis constitutes a fundamental step in tumor progression. Thus, targeting tumour angiogenesis has been identified to be promising in cancer treatment. In this work, CC5 and CC8, two highly homologous disintegrins isolated from the venom Cerastes cerastes viper from the south of Tunisia, were assessed for their anti-angiogenic effect by testing their ability to interfere with viability, adhesion, migration and angiogenesis of Human Microvascular Endothelial Cells, HMEC-1 and HBMEC. We found that CC5 and CC8 displayed pro-apoptotic potential in HMEC-1 cells. Anoïkis like induced by these two disintegrins was evidenced by cell detachment, down regulation of FAK/AKT/PI3K axis and caspase activation. In addition, both CC5 and CC8 exhibited in vitro anti-adhesive, anti-migratory and anti-proliferative effects on endothelial cells HBMEC. These effects appeared to require RGD and/or WGD loops disintegrin. CC5 and CC8 also inhibited tube-formation on matrigel and displayed potent anti-angiogenic activities as assessed ex vivo, using both the embryo chick chorioallantoic membrane model (CAM) and rat aortic ring assay. Altogether our results demonstrate that CC5 and CC8, are potent inhibitors of angiogenesis, by disrupting αvβ3 and α5β1 binding. The use of CC5 and/or CC8 could provide a beneficial tool to inhibit abnormal angiogenesis and to induce cancer regression., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Snake venom l-amino acid oxidases are multifunctional enzymes that exhibited a wide range of pharmacological activities. Although it has been established that these activities are primarily caused by the H2O2 generated in the enzymatic reaction, the molecular mechanism, however, has not been fully investigated. In this work, LAAO interaction with cytoplasmic membranes using different cell types and Langmuir interfacial monolayers was evaluated. The Cerastes cerastes venom LAAO (CC-LAAO) did not exhibit cytotoxic activities against erythrocytes and peripheral blood mononuclear cells (PBMC). However, CC-LAAO caused cytotoxicity on several cancer cell lines and induced platelet aggregation in dose-dependent manner. Furthermore, the enzyme showed remarkable effect against Gram-positive and Gram-negative bacteria. These activities were inhibited on the addition of catalase or substrate analogs, suggesting that H2O2 liberation× is required for these effects. Binding studies revealed that CC-LAAO binds to the cell surface and enables the production of highly localized concentration of H2O2 in or near the binding interfaces. On another hand, the interaction of CC-LAAO with a mimetic phospholipid film was evaluated, for the first time, using a monomolecular film technique. Results indicated that phospholipid/CC-LAAO interactions are not involved in their binding to membrane and in their pharmacological activities., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Early secreted antigenic target 6 kDa (ESAT-6) and culture filtrate protein 10 kDa (CFP-10) are complex proteins secreted by Mycobacterium tuberculosis that play a major role in the pathogenesis of tuberculosis. However, studies focusing on the biological functions of ESAT-6 led to discordant results and the role of ESAT-6 remains controversial. In the present study, we aim to address a potential explanation for this discrepancy and to highlight the physiological impact of two conformational states of ESAT-6. Analysis of a recombinant form of ESAT-6 by native gel electrophoresis, size exclusion chromatography and CD spectroscopy revealed that ESAT-6 forms dimers/multimers with higher molecular weight, which disappeared under the action of the detergent amidosulfobetaine-14 (ASB), giving rise to another conformational state of the protein. NMR has further indicated that ASB-treated versus nontreated ESAT-6 adopted distinct structural forms but with no well defined tertiary structure. However, protein-protein docking analysis favored a dimeric state of ESAT-6. Interestingly, the two preparations presented opposing effects on mycobacterial infectivity, as well as macrophage survival, interferon-γ secretion and membrane pore formation. Thereafter, we generated a recombinant form of the physiological heterodimer ESAT-6/CFP-10 that ASB was also able to dissociate and which showed functions similar to those of ESAT-6 dimers/multimers. Our data suggest that, in the absence of CFP-10, the hydrophobic regions of the ESAT-6 can form dimers/multimers, mimicking the ESAT-6/CFP-10 heterodimer, whereas their dissociation generates a protein presenting entirely different activities. Overall, the present study clarifies the intriguing divergences between reports that could be attributed to the ESAT-6 oligomeric state and sheds light on its importance for a better comprehension of the physiopathology of tuberculosis., (© 2015 FEBS.)

Scorpion toxins are important pharmacological tools for probing the physiological roles of ion channels which are involved in many physiological processes and as such have significant therapeutic potential. The discovery of new scorpion toxins with different specificities and affinities is needed to further characterize the physiology of ion channels. In this regard, a new short polypeptide called Kbot21 has been purified to homogeneity from the venom of Buthus occitanus tunetanus scorpion. Kbot21 is structurally related to BmBKTx1 from the venom of the Asian scorpion Buthus martensii Karsch. These two toxins differ by only two residues at position 13 (R /V) and 24 (D/N).Despite their very similar sequences, Kbot21 and BmBKTx1 differ in their electrophysiological activities. Kbot21 targets KV channel subtypes whereas BmBKTx1 is active on both big conductance (BK) and small conductance (SK) Ca2+-activated K+ channel subtypes, but has no effects on Kv channel subtypes. The docking model of Kbot21 with the Kv1.2 channel shows that the D24 and R13 side-chain of Kbot21 are critical for its interaction with KV channels.

The purified (phospho)lipase of Fusarium solani (FSL), was known to be active on both triglycerides and phospholipids. This study aimed at assessing the potential of this enzyme in hydrolyzing galactolipids. FSL was found to hydrolyze at high rates of synthetic medium chains monogalactosyldiacylglycerol (4658±146U/mg on DiC8-MGDG) and digalactosyldiacylglycerol (3785±83U/mg on DiC8-DGDG) and natural long chain monogalactosyldiacylglycerol extracted from leek leaves (991±85U/mg). It is the microbial enzyme with the highest activity on galactolipids identified so far with a level of activity comparable to that of pancreatic lipase-related protein 2. FSL maximum activity on galactolipids was measured at pH8. The analysis of the hydrolysis product of natural MGDG from leek showed that FSL hydrolyzes preferentially the ester bond at the sn-1 position of galactolipids. To investigate the structure-activity relationships of FSL, a 3D model of this enzyme was built. In silico docking of medium chains MGDG and DGDG and phospholipid in the active site of FSL reveals structural solutions which are in concordance with in vitro tests., (Copyright © 2015 Elsevier B.V. All rights reserved.)

We have purified the AaTX1 peptide from the Androctonus australis (Aa) scorpion venom, previously cloned and sequenced by Legros and collaborators in a venom gland cDNA library from Aa scorpion. AaTX1 belongs to the α-Ktx15 scorpion toxins family (αKTx15-4). Characterized members of this family share high sequence similarity and were found to block preferentially IA-type voltage-dependent K(+) currents in rat cerebellum granular cells in an irreversible way. In the current work, we studied the effects of native AaTX1 (nAaTX1) using whole-cell patch-clamp recordings of IA current in substantia nigra pars compacta dopaminergic neurons. At 250 nM, AaTX1 induces 90% decrease in IA current amplitude. Its activity was found to be comparable to that of rAmmTX3 (αKTx15-3), which differs by only one conserved (R/K) amino acid in the 19th position suggesting that the difference between R19 and K19 in AaTX1 and AmmTX3, respectively, may not be critical for the toxins' effects. Molecular docking of both toxins with Kv4.3 channel is in agreement with experimental data and suggests the implication of the functional dyade K27-Y36 in toxin-channel interactions. Since AaTX1 is not highly abundant in Aa venom, it was synthesized as well as AmmTX3. Synthetic peptides, native AaTX1 and rAmmTX3 peptides showed qualitatively the same pharmacological activity. Overall, these data identify a new biologically active toxin that belongs to a family of peptides active on Kv4.3 channel., (Copyright © 2014 Elsevier Ltd. All rights reserved.)