Browsing by Author "Gaspar, Maria Manuela"
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- Liposomal delivery of repurposed antiviral drug saquinavir to macrophages as a host-directed therapy for tuberculosisPublication . Mandal, Manoj; Pires, David; Pinho, Jacinta; Catalão, Maria João; Almeida, António José; Azevedo-Pereira, José Miguel; Gaspar, Maria Manuela; Anes, ElsaMycobacterium tuberculosis (Mtb) latently infects approximately a quarter of the world’s population and 10 % of these will develop the disease tuberculosis. Mtb infects macrophages, manipulating the proteolytic mechanisms, particularly, by decreasing the expression and activity of lysosomal cathepsins. Consequently, Mtb survives and even replicates inside macrophages concomitant with poor priming of the adaptive immune response. Our group found that the protease inhibitor used in antiretroviral therapy for HIV infection, saquinavir (SQV), restores and further improves the overall activity of cathepsins in Mtb-infected macrophages and more specifcally, that of cathepsin S [1]. In this study, we tested the incorporation of SQV in liposomes to establish an improved delivery method for SQV to human monocyte-derived macrophages. Using fuorophore-tagged liposomes we demonstrated the effciency of SQV-loaded liposome internalization by human macrophages. Additionally, using a general fuorescent substrate of human cathepsins we could observe improved proteolytic activity in treated macrophages. When applying this treatment to Mtb-infected macrophages these effects resulted in better control of the infection. Furthermore, liposomal delivery of SQV reduced the cytotoxicity of the treatment and allowed the usage of higher concentrations without impacting cell viability. By using this strategy, we overcame the cathepsin activity blockade that is induced by the pathogen [2]. The results further demonstrate the effcacy of SQV-loaded liposomes to help control infections by Mtb clinical strains susceptible or resistant to the current antibiotic therapy. Our results suggest the use of liposomal delivery of SQV as a potential complementary therapy against Mtb infection. Human monocytes were isolated from buffy-coats of healthy human donors provided by the National Blood Institute (Instituto Português do Sangue e da Transplantação, IP, Lisbon, Portugal).
- Liposomal delivery of saquinavir to macrophages overcomes cathepsin blockade by Mycobacterium tuberculosis and helps control the phagosomal replicative nichesPublication . Pires, David; Mandal, Manoj; Pinho, Jacinta; Catalão, Maria João; Almeida, António José; Azevedo-Pereira, José Miguel; Gaspar, Maria Manuela; Anes, ElsaMycobacterium tuberculosis is able to establish a chronic colonization of lung macrophages in a controlled replication manner, giving rise to a so-called latent infection. Conversely, when intracellular bacteria undergo actively uncontrolled replication rates, they provide the switch for the active infection called tuberculosis to occur. Our group found that the pathogen is able to manipulate the activity of endolysosomal enzymes, cathepsins, directly at the level of gene expression or indirectly by regulating their natural inhibitors, cystatins. To provide evidence for the crucial role of cathepsin manipulation for the success of tuberculosis bacilli in their intracellular survival, we used liposomal delivery of saquinavir. This protease inhibitor was previously found to be able to increase cathepsin proteolytic activity, overcoming the pathogen induced blockade. In this study, we demonstrate that incorporation in liposomes was able to increase the efficiency of saquinavir internalization in macrophages, reducing cytotoxicity at higher concentrations. Consequently, our results show a significant impact on the intracellular killing not only to reference and clinical strains susceptible to current antibiotic therapy but also to multidrug- and extensively drug-resistant (XDR) Mtb strains. Altogether, this indicates the manipulation of cathepsins as a fine-tuning strategy used by the pathogen to survive and replicate in host cells.