Le cancer de la prostate est le cancer le plus fréquent chez les hommes et la capacité des tumeurs à développer une résistance face aux thérapies anti-androgéniques vient souvent compromettre le pronostic des patients. Le développement de... more
Le cancer de la prostate est le cancer le plus fréquent chez les hommes et la capacité des tumeurs à développer une résistance face aux thérapies anti-androgéniques vient souvent compromettre le pronostic des patients. Le développement de nouvelles approches thérapeutiques afin de circonvenir à la progression de ces tumeurs représente un besoin important la gestion de ce type de cancer. Plusieurs démonstrations récentes établissent l’implication de la famille des proprotéines convertases dans la progression tumorale. Ces enzymes ont pour fonctions biologiques de cliver une variété de précurseurs protéiques jouant des rôles importants dans la tumorigénèse. Dans le cancer de la prostate, la proprotéine convertase PACE4 est fortement surexprimée dans les cellules cancéreuses et joue un rôle dans la prolifération et la capacité à former des tumeurs, ce qui en fait une cible thérapeutique d’intérêt. En ce sens, des inhibiteurs peptidomimétiques ont été développés dans l’optique de la thé...
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Bacteriocins are receiving increased attention as potent candidates in food preservation and medicine. Although the inhibitory activity of bacteriocins has been studied widely, little is known about their gastrointestinal stability and... more
Bacteriocins are receiving increased attention as potent candidates in food preservation and medicine. Although the inhibitory activity of bacteriocins has been studied widely, little is known about their gastrointestinal stability and toxicity toward normal human cell lines. The aim of this study was to evaluate the gastrointestinal stability and activity of microcin J25, pediocin PA-1, bactofencin A and nisin using in vitro models. In addition cytotoxicity and hemolytic activity of these bacteriocins were investigated on human epithelial colorectal adenocarcinoma cells (Caco-2) and rat erythrocytes, respectively. Pediocin PA-1, bactofencin A, and nisin were observed to lose their stability while passing through the gastrointestinal tract, while microcin J25 is only partially degraded. Besides, selected bacteriocins were not toxic to Caco-2 cells, and integrity of cell membrane was observed to remain unaffected in presence of these bacteriocins at concentrations up to 400 μg/mL. In...
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Paired basic amino acid cleaving enzyme 4 (PACE4), a serine endoprotease of the proprotein convertases family, has been recognized as a promising target for prostate cancer. We previously reported a selective and potent peptide-based... more
Paired basic amino acid cleaving enzyme 4 (PACE4), a serine endoprotease of the proprotein convertases family, has been recognized as a promising target for prostate cancer. We previously reported a selective and potent peptide-based inhibitor for PACE4, named the multi-Leu peptide (Ac-LLLLRVKR-NH2 sequence), which was then modified into a more potent and stable compound named C23 with the following structure: Ac-dLeu-LLLRVK-Amba (Amba: 4-amidinobenzylamide). Despite improvements in both in vitro and in vivo profiles of C23, its selectivity for PACE4 over furin was significantly reduced. We examined other Arg-mimetics instead of Amba to regain the lost selectivity. Our results indicated that the replacement of Amba with 5-(aminomethyl)picolinimidamide increased affinity for PACE4 and restored selectivity. Our results also provide a better insight on how structural differences between S1 pockets of PACE4 and furin could be employed in the rational design of selective inhibitors.
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The serine protease, PACE4, is a proprotein convertase that plays a substantial role in malignancy of prostate cancer. Our initial selective PACE4 inhibitor (Ac-LLLLRVKR-NH) has evolved to the current lead compound C23... more
The serine protease, PACE4, is a proprotein convertase that plays a substantial role in malignancy of prostate cancer. Our initial selective PACE4 inhibitor (Ac-LLLLRVKR-NH) has evolved to the current lead compound C23 (Ac-dLeu-LLLRVK-Amba), which is active both in vitro and in vivo. By screening natural residues, except Cys, in C-terminal P1' position, it was established that increasing hydrophobicity was improving cell permeability, which was directly translated into PCa cells antiproliferative activity. This cell antiproliferation enhancement seems independent from effect of P1' residue on PACE4 affinity. Replacement of P1-Amba of C23 by Acpa (( S)-2-amino-3-(4-carbamimidoylphenyl)propanoic acid) followed by addition of tryptamine in P1' resulted in compound 32 exhibiting superior PCa cells antiproliferative activity over the reference compound C23 (3-fold). This study sheds light on key factors that improve cell penetrating property and antiproliferative activity of ...
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Breast cancer is the most frequent and deadly malignancy in women worldwide. Despite national screening programs combined with new treatments relapse rate remain high and new therapies are needed. From previous work, we identified PACE4,... more
Breast cancer is the most frequent and deadly malignancy in women worldwide. Despite national screening programs combined with new treatments relapse rate remain high and new therapies are needed. From previous work, we identified PACE4, a member of the proprotein convertase (PCs) family of endoproteases, as a novel therapeutic target in prostate cancer. In the present study we asked the question if PACE4 could also be a potential target in breast cancer. In clinical samples of breast adenocarcinoma, we observed a specific overexpression of PACE4 in the estrogen-receptor (ER) positive subtype. We therefore looked for a breast cancer cell line model which would be representative and thus focused on the ZR-75-1 since it both expresses PACE4 and is estrogen-receptor positive. We compared stable knockdowns of furin, PACE4 and PC7 in the estrogen-receptor-positive cell line ZR-75-1 to evaluate their respective contribution to cell growth and tumor progression. PACE4 was the only PC displ...
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PACE4, one of the seven members of the proprotein convertase family, plays an important role in the progression of prostate cancer. Therefore, its inhibition has become an attractive target to develop new therapies against this disease.... more
PACE4, one of the seven members of the proprotein convertase family, plays an important role in the progression of prostate cancer. Therefore, its inhibition has become an attractive target to develop new therapies against this disease. Recently, we have developed a highly potent and selective PACE4 inhibitor, known as the multi-Leu peptide with the following sequence Ac-LLLLRVKR-NH2. Herein, with the aim of improving the stability profile of this inhibitor for potential in vivo application, we investigated the impact of different cyclization strategies. The inhibitory activity of new peptides was tested and compared to their linear counterparts. The potent analogues were further selected for stability evaluation. Our results showed that the cyclization involving a C-terminal carboxylic acid (head-to-tail or side chain-to-tail) led to compounds with significantly diminished inhibitory potency towards PACE4, indicating that an appropriate balance between rigidity and flexibility of t...
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The proprotein convertase family of enzymes includes seven endoproteases with significant redundancy in their cleavage activity. We previously described the peptide Ac-LLLLRVK-Amba that displays potent inhibitory effects on both PACE4 and... more
The proprotein convertase family of enzymes includes seven endoproteases with significant redundancy in their cleavage activity. We previously described the peptide Ac-LLLLRVK-Amba that displays potent inhibitory effects on both PACE4 and prostate cancer cell lines proliferation. Herein, the molecular determinants for PACE4 and furin inhibition were investigated by positional scanning using peptide libraries that substituted its leucine core with each natural amino acid. We determined that the incorporation of basic amino acids led to analogues with improved inhibitory potency toward both enzymes, whereas negatively charged residues significantly reduced it. All the remaining amino acids were in general well tolerated, with the exemption of the P6 position. However, not all of the potent PACE4 inhibitors displayed antiproliferative activity. The best analogues were obtained by the incorporation of the Ile residue at the P5 and P6 positions. These substitutions led to inhibitors with...
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PACE4 plays important roles in prostate cancer cell proliferation. The inhibition of this enzyme has been shown to slow prostate cancer progression and is emerging as a promising therapeutic strategy. In previous work, we developed a... more
PACE4 plays important roles in prostate cancer cell proliferation. The inhibition of this enzyme has been shown to slow prostate cancer progression and is emerging as a promising therapeutic strategy. In previous work, we developed a highly potent and selective PACE4 inhibitor, the multi-Leu (ML) peptide, an octapeptide with the sequence Ac-LLLLRVKR-NH2 . Here, with the objective of developing a useful compound for in vivo administration, we investigate the effect of N-terminal modifications. The inhibitory activity, toxicity, stability, and cell penetration properties of the resulting analogues were studied and compared to the unmodified inhibitor. Our results show that the incorporation of a polyethylene glycol (PEG) moiety leads to a loss of antiproliferative activity, whereas the attachment of a lipid chain preserves or improves it. However, the lipidated peptides are significantly more toxic when compared with their unmodified counterparts. Therefore, the best results were achieved not by the N-terminal extension but by the protection of both ends with the d-Leu residue and 4-amidinobenzylamide, which yielded the most stable inhibitor, with an excellent activity and toxicity profile.
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There is increasing interest in the therapeutic targeting of proteases for the treatment of important diseases. Additionally new protein-based therapeutic strategies have the potential to widen the available treatments against these... more
There is increasing interest in the therapeutic targeting of proteases for the treatment of important diseases. Additionally new protein-based therapeutic strategies have the potential to widen the available treatments against these pathologies. In the last decade, accumulated evidence has confirmed that the family of proteases known as proprotein convertases (PCs) are potential targets for viral infections, osteoarthritis, cancer and cardiovascular disease, among others. Nevertheless, there are still many unanswered questions about the relevance of targeting PCs in a therapeutic context, especially regarding the anticipated secondary effects of treatment, considering the observed embryonic lethality of some PC knockout mice. In this review, the benefits of PCs as pharmacological targets will be discussed, with focus on concepts and strategies, as well as on the state of advancement of actual and future inhibitors.
PACE4 plays an important role in the progression of prostate cancer and is an attractive target for the development of novel inhibitor-based tumor therapies. We previously reported the design and synthesis of a novel, potent, and... more
PACE4 plays an important role in the progression of prostate cancer and is an attractive target for the development of novel inhibitor-based tumor therapies. We previously reported the design and synthesis of a novel, potent, and relatively selective PACE4 inhibitor known as a Multi-Leu (ML) peptide. In the present work, we examined the ML peptide through detailed structure-activity relationship studies. A variety of ML-peptide analogues modified at the P8-P5 positions with leucine isomers (Nle, DLeu, and DNle) or substituted at the P1 position with arginine mimetics were tested for their inhibitory activity, specificity, stability, and antiproliferative effect. By incorporating d isomers at the P8 position or a decarboxylated arginine mimetic, we obtained analogues with an improved stability profile and excellent antiproliferative properties. The DLeu or DNle residue also has improved specificity toward PACE4, whereas specificity was reduced for a peptide modified with the arginine...
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Prostate cancer is the leading cancer in North American men. Current pharmacological treatments are limited to anti-androgen strategies and the development of new therapeutic approaches remains a challenge. As a fundamentally new... more
Prostate cancer is the leading cancer in North American men. Current pharmacological treatments are limited to anti-androgen strategies and the development of new therapeutic approaches remains a challenge. As a fundamentally new approach, we propose the inhibition of PACE4, a member of the proprotein convertases family of enzymes, as a therapeutic target in prostate cancer. We developed an inhibitor named the Multi-Leu peptide, with potent in vitro anti-proliferative effects. However, the Multi-Leu peptide has not been tested under in vivo conditions and its potency under such conditions is most likely limited, due to the labile characteristics of peptides in general. Using a peptidomimetic approach, we modified the initial scaffold, generating the analog Ac-[DLeu]LLLRVK-Amba, which demonstrates increased inhibitory potency and stability. The systemic administration of this peptidomimetic significantly inhibits tumor progression in the LNCaP xenograft model of prostate cancer by in...
Research Interests: Apoptosis, Humans, Peptidomimetics, Mice, Animals, and 3 moreMale, Disease Progression, and Cell Proliferation
Better understanding of the distinct and redundant functions of the proprotein convertase (PC) enzyme family within pathophysiological states has a great importance for potential therapeutic strategies. In this study, we investigated the... more
Better understanding of the distinct and redundant functions of the proprotein convertase (PC) enzyme family within pathophysiological states has a great importance for potential therapeutic strategies. In this study, we investigated the functional redundancy of PCs in prostate cancer in the commonly used androgen-sensitive LNCaP and the androgen-independent DU145 human cell lines. Using a lentiviral-based shRNA delivery system, we examined in vitro and in vivo cell proliferation characteristics of knockdown cell lines for the endogenous PCs furin, PACE4, and PC7 in both cell lines. Of the three PCs, only PACE4 was essential to maintain a high-proliferative status, as determined in vitro using XTT proliferation assays and in vivo using tumor xenografts in nude mice. Furin knockdowns in both cell lines had no effects on cell proliferation or tumor xenograft growth. Paradoxically, PC7 knockdowns reduced in vitro cellular proliferation but had no effect in vivo. Because PCs act within ...
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Gene silencing strategies targeting mRNA are suitable methods to validate the functions of specific genes. In this chapter, we sought to compare two knockdown strategies for the study of proprotein convertases and proliferation in... more
Gene silencing strategies targeting mRNA are suitable methods to validate the functions of specific genes. In this chapter, we sought to compare two knockdown strategies for the study of proprotein convertases and proliferation in prostate cancer cells. We used both SOFA-HDV ribozyme and lentiviral-mediated shRNA delivery system to reduce PACE4 mRNA levels and validate its implication in the proliferation of DU145 prostate cancer cells. The cellular effects of PACE4 knockdown were assessed (1) in vitro using two tetrazolium salts (MTT and XTT assays) and (2) in vivo using a tumor xenograft approach in immunodeficient mice (Nu/Nu). Our results confirm the unique role of the proprotein convertase PACE4 in prostate cancer cell proliferation while demonstrating advantages and disadvantages of each approach. Achieving target validation in an effective manner is critical, as further development using a drug development approach is highly laborious and requires enormous resources.