Category: J.Martinez_publications

Site-specific grafting on titanium surfaces with hybrid temporin antibacterial peptides

J. Mater. Chem. B, 2018,6, 1782-1790. doi 10.1039/C8TB00051D

Abstract

Relying on a membrane-disturbing mechanism of action and not on any intracellular target, antimicrobial peptides (AMP) are attractive compounds to be grafted on the surface of implantable materials such as silicone catheters or titanium surgical implants. AMP sequences often display numerous reactive functions (e.g. amine, carboxylic acid) on their side chains and straightforward conjugation chemistries could lead to uncontrolled covalent grafting, random orientation, and non-homogenous density. To achieve an easy and site specific covalent attachment of unprotected peptides on titanium surfaces, we designed hybrid silylated biomolecules based on the temporin-SHa amphipathic helical antimicrobial sequence. With the grafting reaction being chemoselective, we designed five analogues displaying the silane anchoring function at the N-ter, C-ter or at different positions inside the sequence to get an accurate control of the orientation. Grafting density calculations were performed by XPS and the influence of the orientation of the peptide on the surface was clearly demonstrated by the measure of antimicrobial activity. Temporin amphipathic helices are described to permeabilize the bacterial membrane by interacting in a parallel orientation with it. Our results move in the direction of this mechanism as the selective grafting of hybrid temporin 2 through a lysine placed at the center of the peptide sequence, resulted in better biofilm growth inhibition of E. coli and S. epidermis than substrates in which temporins were grafted via their C- or N-terminus.

GHSR-D2R heteromerization modulates dopamine signaling through an effect on G protein conformation

Proc Natl Acad Sci U S A. 2018 Apr 24;115(17):4501-4506. doi: 10.1073/pnas.1712725115. Epub 2018 Apr 9

Damian M, Pons V, Renault P, M’Kadmi C, Delort B, Hartmann L, Kaya AI, Louet M, Gagne D, Ben Haj Salah K, Denoyelle S, Ferry G, Boutin JA, Wagner R, Fehrentz JA, Martinez J, Marie J, Floquet N, Galès C, Mary S, Hamm HE, Banères JL.

Abstract

IThe growth hormone secretagogue receptor (GHSR) and dopamine receptor (D2R) have been shown to oligomerize in hypothalamic neurons with a significant effect on dopamine signaling, but the molecular processes underlying this effect are still obscure. We used here the purified GHSR and D2R to establish that these two receptors assemble in a lipid environment as a tetrameric complex composed of two each of the receptors. This complex further recruits G proteins to give rise to an assembly with only two G protein trimers bound to a receptor tetramer. We further demonstrate that receptor heteromerization directly impacts on dopamine-mediated Gi protein activation by modulating the conformation of its α-subunit. Indeed, association to the purified GHSR:D2R heteromer triggers a different active conformation of Gαi that is linked to a higher rate of GTP binding and a faster dissociation from the heteromeric receptor. This is an additional mechanism to expand the repertoire of GPCR signaling modulation that could have implications for the control of dopamine signaling in normal and physiopathological conditions.

Inorganic polymerization: an attractive route to biocompatible hybrid hydrogels

J. Mater. Chem. B, 2018,6, 3434-3448,  doi 10.1039/C8TB00456K

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Abstract

As an intermediate state between liquid and solid materials, hydrogels display unique properties, opening a wide scope of applications, especially in the biomedical field. Organic hydrogels are composed of an organic network cross-linked via chemical or physical reticulation nodes. In contrast, hybrid hydrogels are defined by the coexistence of organic and inorganic moieties in water. Inorganic polymerization, i.e. the sol–gel process, is one of the main techniques leading to hybrid hydrogels. The chemoselectivity of this method proceeds through hydrolysis and condensation reactions of metal oxide moieties. In addition, the mild reaction conditions make this process very promising for the preparation of water-containing materials and their bio-applications.

Indoloazepinone-Constrained Oligomers as Cell-Penetrating and Blood-Brain Barrier Permeable Compounds

Chembiochem. 2018 Jan 29. doi: 10.1002/cbic.201700678. [Epub ahead of print]

Van der Poorten O, Legrand B, Vezenkov L, Garcia-Pindado J, Bettache N, Knuhtsen A, Sejer Pedersen D, Sanchez-Navarro M, Martinez J, Teixido M, Garcia M, Tourwe D, Amblard M, Ballet S.

Abstract

Non-cationic and amphipathic indoloazepinone-constrained (Aia) oligomers have been synthesized as new vectors for intracellular delivery. The conformational preferences of the [L-Aia-Xxx]n oligomers were investigated using circular dichroism and NMR spectroscopy. While Boc-[L-Aia-Gly]2,4-OBn 12-13 and Boc-[L-Aia-β3-h-L-Ala]2,4-OBn 16-17 oligomers were totally or partially disordered, Boc-[L-Aia-L-Ala]2-OBn 14 induced a typical turn stabilized by C5- and C7-membered H-bond pseudo-cycles and aromatic interactions. Boc-[L-Aia-L-Ala]4-OBn 15 exhibited a unique structure with remarkable T-shaped pie-stacking interactions involving the indole rings of the four L-Aia residues forming a dense hydrophobic cluster. All the proposed FITC-6-Ahx-[L-Aia-Xxx]4-NH2 oligomers 19-23, with exception of FITC-6-Ahx-[L-Aia-Gly]4-NH2 oligomer 18, were internalized by MDA-MB-231 cells with higher efficiency than the positive references penetratin and Arg8. In parallel, this series of compounds was successfully explored on an in vitro blood-brain barrier (BBB) permeation assay. While no passive diffusion permeability was observed for any of the tested Ac-[L-Aia-Xxx]4-NH2 oligomers in the PAMPA model, Ac-[L-Aia-L-Arg]4-NH2 26 showed significant permeation in the in vitro cell-based human model of the BBB, suggesting an active mechanism of cell-penetration.

Heteromultivalent targeting of integrin αvβ3 and neuropilin 1 promotes cell survival via the activation of the IGF-1/insulin receptors

Biomaterials 2018 Feb;155:64-79. doi: 10.1016/j.biomaterials.2017.10.042. Epub 2017 Oct 29.

Jia T, Choi J, Ciccione J, Henry M, Mehdi A, Martinez J, Eymin B, Subra G, Coll JL.

Abstract

Angiogenesis strongly depends on the activation of integrins, especially integrin αvβ3, and of neuropilin-1 (NRP-1), a co-receptor of VEGFR2. Dual-targeted molecules that simultaneously block both of them are expected have increased anti-angiogenic and antitumor activity. Toward this goal, we generated bifunctional 40 nm-sized silica nanoparticles (NPs) coated with controlled amounts of cRGD and ATWLPPR peptides and studied their affinity, selectivity and biological activity in HUVECs. Sub-nanomolar concentrations of NPs grafted either with ATWLPPR alone or in combination with cRGD exhibit potent and specific antagonist activity against VEGFR2/AKT signaling. However, a 1 nM concentration of the cRGD/ATWLPPR-heteromultivalent particles (RGD/ATW-NPs) also blocks the phosphorylation of VEGFR2 while co-inducing an unexpected long-lasting activation of AKT via IGF-1R/IR-AKT/GSK3β/eNOS signaling that stimulates cell survival and abrogates the intrinsic toxicity of silica-NPs to serum-starved HUVECs. We also showed that their repeated intravenous administration was associated with the proliferation of human U87MG tumor cells engrafted in nude mice and a dilatation of the tumor blood vessels. We present biochemical evidence for the complex cross-talk generated by the binding of the heteromultivalent NPs with αvβ3-integrin and with NRP1. In particular, we show for the first time that such heteromultivalent NPs can trans-activate IGF-1/insulin receptors and exert dose-dependent pro-survival activity. This study demonstrates the difficulties in designing targeted silica-based NPs for antiangiogenic therapies and the possible risks posed by undesirable side effects.

The GHR-R antagonist JMV 2959 neither induces malaise nor alters the malaise property of LiCl in the adult male rat

Physiol Behav. 2018 Jan 1;183:46-48. doi: 10.1016/j.physbeh.2017.10.017. Epub 2017 Oct 19.

Rodriguez JA, Fehrentz JA, Martinez J, Ben Haj Salah K, Wellman PJ.

Abstract

The orexigenic peptide ghrelin (GHR) interacts with ghrelin receptors (GHR-Rs) to modulate brain reinforcement and feeding circuits. Pharmacological inactivation of GHR-Rs via administration of the drug JMV 2959 attenuates the rewarding/reinforcing effects of several drugs of abuse including alcohol, morphine, amphetamine and nicotine. One view of these results is that inactivation of GHR-Rs taps into brain reinforcement/feeding circuits acted upon by drugs of abuse. An alternate explanation is that JMV 2959 may induce malaise, which in turn may limit reinforcement as well as food ingestion. This is a variable of interest given that nicotine alone can induce malaise which may be enhanced by JMV 2959. In the present study, we assessed the capacity of JMV 2959 to produce malaise using a conditioned taste aversion (CTA) task. Adult male rats were allowed to consume a 0.1% sodium saccharin solution and then injected IP with either vehicle, 0.4mg/kg nicotine, 3mg/kg JMV 2959, a combination of 0.4mg/kg nicotine and 3mg/kg JMV 2959, or 32mg/kg lithium chloride (a positive control known to support induction of CTA). Lithium chloride produced a robust avoidance of the saccharin solution in subsequent 2 bottle (water and saccharin) tests, whereas JMV 2959 alone did not induce CTA. The combination of JMV 2959 and nicotine induced a moderate degree of CTA that was similar to that produced by nicotine alone. These results suggest that JMV 2959 is unlikely to limit either reinforcement or food ingestion via induction of malaise.

Growth hormone secretagogues hexarelin and JMV2894 protect skeletal muscle from mitochondrial damages in a rat model of cisplatin-induced cachexia

Sci Rep. 2017 Oct 12;7(1):13017. doi: 10.1038/s41598-017-13504-y.

Sirago G, Conte E, Fracasso F, Cormio A, Fehrentz JA, Martinez J, Musicco C, Camerino GM, Fonzino A, Rizzi L, Torsello A, Lezza AMS, Liantonio A, Cantatore P, Pesce V.

Abstract

Chemotherapy can cause cachexia, which consists of weight loss associated with muscle atrophy. The exact mechanisms underlying this skeletal muscle toxicity are largely unknown and co-therapies to attenuate chemotherapy-induced side effects are lacking. By using a rat model of cisplatin-induced cachexia, we here characterized the mitochondrial homeostasis in tibialis anterior cachectic muscle and evaluated the potential beneficial effects of the growth hormone secretagogues (GHS) hexarelin and JMV2894 in this setting. We found that cisplatin treatment caused a decrease in mitochondrial biogenesis (PGC-1α, NRF-1, TFAM, mtDNA, ND1), mitochondrial mass (Porin and Citrate synthase activity) and fusion index (MFN2, Drp1), together with changes in the expression of autophagy-related genes (AKT/FoxO pathway, Atg1, Beclin1, LC3AII, p62) and enhanced ROS production (PRX III, MnSOD). Importantly, JMV2894 and hexarelin are capable to antagonize this chemotherapy-induced mitochondrial dysfunction. Thus, our findings reveal a key-role played by mitochondria in the mechanism responsible for GHS beneficial effects in skeletal muscle, strongly indicating that targeting mitochondrial dysfunction might be a promising area of research in developing therapeutic strategies to prevent or limit muscle wasting in cachexia.

Involvement of PPARγ in the Anticonvulsant Activity of EP-80317, a Ghrelin Receptor Antagonist

Front Pharmacol. 2017 Sep 22;8:676. doi: 10.3389/fphar.2017.00676. eCollection 2017.

Lucchi C, Costa AM, Giordano C, Curia G, Piat M, Leo G, Vinet J, Brunel L, Fehrentz JA, Martinez J, Torsello A, Biagini G.

Abstract

Ghrelin, des-acyl ghrelin and other related peptides possess anticonvulsant activities. Although ghrelin and cognate peptides were shown to physiologically regulate only the ghrelin receptor, some of them were pharmacologically proved to activate the peroxisome proliferator-activated receptor gamma (PPARγ) through stimulation of the scavenger receptor CD36 in macrophages. In our study, we challenged the hypothesis that PPARγ could be involved in the anticonvulsant effects of EP-80317, a ghrelin receptor antagonist. For this purpose, we used the PPARγ antagonist GW9662 to evaluate the modulation of EP-80317 anticonvulsant properties in two different models. Firstly, the anticonvulsant effects of EP-80317 were studied in rats treated with pilocarpine to induce status epilepticus (SE). Secondly, the anticonvulsant activity of EP-80317 was ascertained in the repeated 6-Hz corneal stimulation model in mice. Behavioral and video electrocorticographic (ECoG) analyses were performed in both models. We also characterized levels of immunoreactivity for PPARγ in the hippocampus of 6-Hz corneally stimulated mice. EP-80317 predictably antagonized seizures in both models. Pretreatment with GW9662 counteracted almost all EP-80317 effects both in mice and rats. Only the effects of EP-80317 on power spectra of ECoGs recorded during repeated 6-Hz corneal stimulation were practically unaffected by GW9662 administration. Moreover, GW9662 alone produced a decrease in the latency of tonic-clonic seizures and accelerated the onset of SE in rats. Finally, in the hippocampus of mice treated with EP-80317 we found increased levels of PPARγ immunoreactivity. Overall, these results support the hypothesis that PPARγ is able to modulate seizures and mediates the anticonvulsant effects of EP-80317.

C9/12 Ribbon-Like Structures in Hybrid Peptides Alternating α- and Thiazole-Based γ-Amino Acids

Chemistry. 2017 Dec 11;23(69):17584-17591. doi: 10.1002/chem.201704001. Epub 2017 Nov 15.

Bonnel C, Legrand B, Simon M, Martinez J, Bantignies JL, Kang YK, Wenger E, Hoh F, Masurier N, Maillard LT.

Abstract

According to their restricted conformational freedom, heterocyclic γ-amino acids are usually considered to be related to Z-vinylogous γ-amino acids. In this context, oligomers alternating α-amino acids and thiazole-based γ-amino acids (ATCs) were expected to fold into a canonical 12-helical shape as described for α/γ-hybrid peptides composed of cis-α/β-unsaturated γ-amino acids. However, through a combination of X-ray crystallography, NMR spectroscopy, FTIR experiments, and DFT calculations, it was determined that the folding behavior of ATC-containing hybrid peptides is much more complex. The homochiral α/(S)-ATC sequences were unable to adopt a stable conformation, whereas the heterochiral α/(R)-ATC peptides displayed novel ribbon structures stabilized by unusual C9/12 -bifurcated hydrogen bonds. These ribbon structures could be considered as a succession of pre-organized γ/α dipeptides and may provide the basis for designing original α-helix mimics.

Ribbon-like Foldamers for Cellular Uptake and Drug Delivery

Chembiochem 2017 Nov 2;18(21):2110-2114. doi: 10.1002/cbic.201700455. Epub 2017 Sep 22.

Vezenkov LL, Martin V, Bettache N, Simon M, Messerschmitt A, Legrand B, Bantignies JL, Subra G, Maynadier M, Bellet V, Garcia M, Martinez J, Amblard M.

Abstract

Different intracellular delivery systems of bioactive compounds have been developed, including cell-penetrating peptides. Although usually nontoxic and biocompatible, these vectors share some of the general drawbacks of peptides, notably low bioavailability and susceptibility to protease degradation, that limit their use. Herein, the conversion of short peptide sequences into poly-α-amino-γ-lactam foldamers that adopt a ribbon-like structure is investigated. This template is used to distribute critical cationic and/or hydrophobic groups on both sides of the backbone, leading to potent short, cell-permeable foldamers with a low positive-charge content. The lead compound showed dramatically improved protease resistance and was able to efficiently deliver a biologically relevant cargo inside cells. This study provided a simple strategy to convert short peptide sequences into efficient protease-resistant cell-penetrating foldamers.