We report the use of switchable lipids to improve the endosomal escape and cytosolic delivery of ... more We report the use of switchable lipids to improve the endosomal escape and cytosolic delivery of cell-impermeable compounds. The system is based on a conformational reorganization of the lipid structure upon acidification, as demonstrated by NMR spectroscopic studies. When incorporated in a liposome formulation, the switchable lipids triggered bilayer destabilization through fusion even in the presence of poly(ethylene glycol). We observed 88 % release of sulforhodamine B in 15 min at pH 5, and the liposome formulations demonstrated high stability at pH 7.4 for several months. By using sulforhodamine B as a model of a highly polar drug, we demonstrated fast cytosolic delivery mediated by endosomal escape in HeLa cells, and no toxicity.
Taken to the molecular level, the concept of &amp... more Taken to the molecular level, the concept of "tweezers" opens a rich and fascinating field at the convergence of molecular recognition, biomimetic chemistry and nanomachines. Composed of a spacer bridging two interaction sites, the behaviour of molecular tweezers is strongly influenced by the flexibility of their spacer. Operating through an "induced-fit" recognition mechanism, flexible molecular tweezers select the conformation(s) most appropriate for substrate binding. Their adaptability allows them to be used in a variety of binding modes and they have found applications in chirality signalling. Rigid spacers, on the contrary, display a limited number of binding states, which lead to selective and strong substrate binding following a "lock and key" model. Exquisite selectivity may be expressed with substrates as varied as C(60) , nanotubes and natural cofactors, and applications to molecular electronics and enzyme inhibition are emerging. At the crossroad between flexible and rigid spacers, stimulus-responsive molecular tweezers controlled by ionic, redox or light triggers belong to the realm of molecular machines, and, applied to molecular tweezing, open doors to the selective binding, transport and release of their cargo. Applications to controlled drug delivery are already appearing. The past 30 years have seen the birth of molecular tweezers; the next many years to come will surely see them blooming in exciting applications.
Nonviral gene delivery is limited to a large extent by the cationic nature of most of the chemica... more Nonviral gene delivery is limited to a large extent by the cationic nature of most of the chemical vector. We have shown that lipopolythioureas interact with DNA. However, lipopolythioureas were not very efficient at transfecting cells, probably due to reduced interaction between the noncationic synthetic lipid and the cell membrane. Here, we report that liposomes made from a new thiourea lipid, DPPC, and a lipid bearing an RGD ligand allowed very efficient entry of the lipopolythioureas into integrin alpha(v)beta(3) expressing cells. In addition, we show that a stable interaction between DNA and lipopolythiourea could be obtain with two thiourea groups. Moreover, the addition of a hydrophilic terminus improves the formulation of these new DNA binding agents.
Advanced-glycation end products (AGEs) were recently implicated in vascular calcification, throug... more Advanced-glycation end products (AGEs) were recently implicated in vascular calcification, through a process mediated by RAGE (receptor for AGEs). Although a correlation between AGEs levels and vascular calcification was established, there is no evidence that reducing in vivo AGEs deposition or inhibiting AGEs-RAGE signaling pathways can decrease medial calcification. We evaluated the impact of inhibiting AGEs formation by pyridoxamine or elimination of AGEs by alagebrium on diabetic medial calcification. We also evaluated if the inhibition of AGEs-RAGE signaling pathways can prevent calcification. Rats were fed a high fat diet during 2 months before receiving a low dose of streptozotocin. Then, calcification was induced with warfarin. Pyridoxamine was administered at the beginning of warfarin treatment while alagebrium was administered 3 weeks after the beginning of warfarin treatment. Results demonstrate that AGEs inhibitors prevent the time-dependent accumulation of AGEs in femoral arteries of diabetic rats. This effect was accompanied by a reduced diabetes-accelerated calcification. Ex vivo experiments showed that N-methylpyridinium, an agonist of RAGE, induced calcification of diabetic femoral arteries, a process inhibited by antioxidants and different inhibitors of signaling pathways associated to RAGE activation. The physiological importance of oxidative stress was demonstrated by the reduction of femoral artery calcification in diabetic rats treated with apocynin, an inhibitor of reactive oxygen species production. We demonstrated that AGE inhibitors prevent or limit medial calcification. We also showed that diabetes-accelerated calcification is prevented by antioxidants. Thus, inhibiting the association of AGE-RAGE or the downstream signaling reduced medial calcification in diabetes.
ABSTRACT The positive charge of cationic-lipid/DNA cationic polymer/DNA complexes renders them hi... more ABSTRACT The positive charge of cationic-lipid/DNA cationic polymer/DNA complexes renders them highly susceptible to interactions with the biological milieu, leading to aggregation and destabilization, and rapid clearance from the blood circulation. To overcome such drawback we have investigated the possibility to deliver DNA using non-cationic synthetic vectors. We have previously demonstrated that neutral lipopolythioureas DT3TU bind to DNA leading to small particles. These new DNA binding agents lack non-specific accumulation in the lung and increasing circulation time in the bloodstream compared to cationic liposomes. However DT3TU is highly hydrophobic and are inefficient for gene delivery.To improve lipopolythiourea efficiency we have synthesized a series of new lipopolythioureas (LPT) build around two di-thiourea scaffolds derived from (2-amino-3-thiocarbamoylamino- propyl)thiourea or from 2,6-bis(thiocarbamoylamino)hexanoic acid. Variation of hydrophobic anchor size and linker has also been investigated. We have found that derivative ended with an N-diol could be formulated without helper lipid. On the other hand LPT with methyl terminus yielded also small particles when formulated with DPPC. Ten compounds were evaluated for their transfection efficiency against B16 or Eahycell line in the presence of serum. The results revealed that (2-amino-3-thiocarbamoylamino-propyl)thioureas, elicited transfection activity with the same level as cationic lipids. Intratumoral injection of thiourea lipoplex was used to evaluate gene expression on 3LL tumor in C57BL\6 mice. The luciferase reporter gene revealed that LPT express the protein with a similar level as cationic lipid. Finally, pharmacokinetics and biodistribution in mice showed that 10 % of the complexes were still circulating after two hours (3% for other cationic lipid vectors). To the best of our knowlege these new LPT are the first examples of non-cationic transfecting lipids.
We report the use of switchable lipids to improve the endosomal escape and cytosolic delivery of ... more We report the use of switchable lipids to improve the endosomal escape and cytosolic delivery of cell-impermeable compounds. The system is based on a conformational reorganization of the lipid structure upon acidification, as demonstrated by NMR spectroscopic studies. When incorporated in a liposome formulation, the switchable lipids triggered bilayer destabilization through fusion even in the presence of poly(ethylene glycol). We observed 88 % release of sulforhodamine B in 15 min at pH 5, and the liposome formulations demonstrated high stability at pH 7.4 for several months. By using sulforhodamine B as a model of a highly polar drug, we demonstrated fast cytosolic delivery mediated by endosomal escape in HeLa cells, and no toxicity.
Taken to the molecular level, the concept of &amp... more Taken to the molecular level, the concept of "tweezers" opens a rich and fascinating field at the convergence of molecular recognition, biomimetic chemistry and nanomachines. Composed of a spacer bridging two interaction sites, the behaviour of molecular tweezers is strongly influenced by the flexibility of their spacer. Operating through an "induced-fit" recognition mechanism, flexible molecular tweezers select the conformation(s) most appropriate for substrate binding. Their adaptability allows them to be used in a variety of binding modes and they have found applications in chirality signalling. Rigid spacers, on the contrary, display a limited number of binding states, which lead to selective and strong substrate binding following a "lock and key" model. Exquisite selectivity may be expressed with substrates as varied as C(60) , nanotubes and natural cofactors, and applications to molecular electronics and enzyme inhibition are emerging. At the crossroad between flexible and rigid spacers, stimulus-responsive molecular tweezers controlled by ionic, redox or light triggers belong to the realm of molecular machines, and, applied to molecular tweezing, open doors to the selective binding, transport and release of their cargo. Applications to controlled drug delivery are already appearing. The past 30 years have seen the birth of molecular tweezers; the next many years to come will surely see them blooming in exciting applications.
Nonviral gene delivery is limited to a large extent by the cationic nature of most of the chemica... more Nonviral gene delivery is limited to a large extent by the cationic nature of most of the chemical vector. We have shown that lipopolythioureas interact with DNA. However, lipopolythioureas were not very efficient at transfecting cells, probably due to reduced interaction between the noncationic synthetic lipid and the cell membrane. Here, we report that liposomes made from a new thiourea lipid, DPPC, and a lipid bearing an RGD ligand allowed very efficient entry of the lipopolythioureas into integrin alpha(v)beta(3) expressing cells. In addition, we show that a stable interaction between DNA and lipopolythiourea could be obtain with two thiourea groups. Moreover, the addition of a hydrophilic terminus improves the formulation of these new DNA binding agents.
Advanced-glycation end products (AGEs) were recently implicated in vascular calcification, throug... more Advanced-glycation end products (AGEs) were recently implicated in vascular calcification, through a process mediated by RAGE (receptor for AGEs). Although a correlation between AGEs levels and vascular calcification was established, there is no evidence that reducing in vivo AGEs deposition or inhibiting AGEs-RAGE signaling pathways can decrease medial calcification. We evaluated the impact of inhibiting AGEs formation by pyridoxamine or elimination of AGEs by alagebrium on diabetic medial calcification. We also evaluated if the inhibition of AGEs-RAGE signaling pathways can prevent calcification. Rats were fed a high fat diet during 2 months before receiving a low dose of streptozotocin. Then, calcification was induced with warfarin. Pyridoxamine was administered at the beginning of warfarin treatment while alagebrium was administered 3 weeks after the beginning of warfarin treatment. Results demonstrate that AGEs inhibitors prevent the time-dependent accumulation of AGEs in femoral arteries of diabetic rats. This effect was accompanied by a reduced diabetes-accelerated calcification. Ex vivo experiments showed that N-methylpyridinium, an agonist of RAGE, induced calcification of diabetic femoral arteries, a process inhibited by antioxidants and different inhibitors of signaling pathways associated to RAGE activation. The physiological importance of oxidative stress was demonstrated by the reduction of femoral artery calcification in diabetic rats treated with apocynin, an inhibitor of reactive oxygen species production. We demonstrated that AGE inhibitors prevent or limit medial calcification. We also showed that diabetes-accelerated calcification is prevented by antioxidants. Thus, inhibiting the association of AGE-RAGE or the downstream signaling reduced medial calcification in diabetes.
ABSTRACT The positive charge of cationic-lipid/DNA cationic polymer/DNA complexes renders them hi... more ABSTRACT The positive charge of cationic-lipid/DNA cationic polymer/DNA complexes renders them highly susceptible to interactions with the biological milieu, leading to aggregation and destabilization, and rapid clearance from the blood circulation. To overcome such drawback we have investigated the possibility to deliver DNA using non-cationic synthetic vectors. We have previously demonstrated that neutral lipopolythioureas DT3TU bind to DNA leading to small particles. These new DNA binding agents lack non-specific accumulation in the lung and increasing circulation time in the bloodstream compared to cationic liposomes. However DT3TU is highly hydrophobic and are inefficient for gene delivery.To improve lipopolythiourea efficiency we have synthesized a series of new lipopolythioureas (LPT) build around two di-thiourea scaffolds derived from (2-amino-3-thiocarbamoylamino- propyl)thiourea or from 2,6-bis(thiocarbamoylamino)hexanoic acid. Variation of hydrophobic anchor size and linker has also been investigated. We have found that derivative ended with an N-diol could be formulated without helper lipid. On the other hand LPT with methyl terminus yielded also small particles when formulated with DPPC. Ten compounds were evaluated for their transfection efficiency against B16 or Eahycell line in the presence of serum. The results revealed that (2-amino-3-thiocarbamoylamino-propyl)thioureas, elicited transfection activity with the same level as cationic lipids. Intratumoral injection of thiourea lipoplex was used to evaluate gene expression on 3LL tumor in C57BL\6 mice. The luciferase reporter gene revealed that LPT express the protein with a similar level as cationic lipid. Finally, pharmacokinetics and biodistribution in mice showed that 10 % of the complexes were still circulating after two hours (3% for other cationic lipid vectors). To the best of our knowlege these new LPT are the first examples of non-cationic transfecting lipids.
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Papers by Jeanne Leblond