Selectively Targeting Breast Cancer Stem Cells by 8-Quinolinol and Niclosamide

Cancer stem cells; Combination therapy; NiclosamideCélulas madre cancerosas; Terapia combinada; NiclosamidaCèl·lules mare cancerígenes; Teràpia combinada; NiclosamidaCancer maintenance, metastatic dissemination and drug resistance are sustained by cancer stem cells (CSCs). Triple negative breast cancer (TNBC) is the breast cancer subtype with the highest number of CSCs and the poorest prognosis. Here, we aimed to identify potential drugs targeting CSCs to be further employed in combination with standard chemotherapy in TNBC treatment. The anti-CSC efficacy of up to 17 small drugs was tested in TNBC cell lines using cell viability assays on differentiated cancer cells and CSCs. Then, the effect of 2 selected drugs (8-quinolinol -8Q- and niclosamide -NCS-) in the cancer stemness features were evaluated using mammosphere growth, cell invasion, migration and anchorage-independent growth assays. Changes in the expression of stemness genes after 8Q or NCS treatment were also evaluated. Moreover, the potential synergism of 8Q and NCS with PTX on CSC proliferation and stemness-related signaling pathways was evaluated using TNBC cell lines, CSC-reporter sublines, and CSC-enriched mammospheres. Finally, the efficacy of NCS in combination with PTX was analyzed in vivo using an orthotopic mouse model of MDA-MB-231 cells. Among all tested drug candidates, 8Q and NCS showed remarkable specific anti-CSC activity in terms of CSC viability, migration, invasion and anchorage independent growth reduction in vitro. Moreover, specific 8Q/PTX and NCS/PTX ratios at which both drugs displayed a synergistic effect in different TNBC cell lines were identified. The sole use of PTX increased the relative presence of CSCs in TNBC cells, whereas the combination of 8Q and NCS counteracted this pro-CSC activity of PTX while significantly reducing cell viability. In vivo, the combination of NCS with PTX reduced tumor growth and limited the dissemination of the disease by reducing circulating tumor cells and the incidence of lung metastasis. The combination of 8Q and NCS with PTX at established ratios inhibits both the proliferation of differentiated cancer cells and the viability of CSCs, paving the way for more efficacious TNBC treatments.This work was supported by the Instituto de Salud Carlos III (ISCiii), through Networking Research Center on Bioengineering, Biomaterials, and Nanomedicine (CIBER-BBN), an initiative that also counts with the assistance from the European Regional Development Fund (ERDF), specifically in the PENTRI-2 Project and by the “Fundació Marató TV3” (337/C/2013) to I.A., M.R. and E.V. Our laboratories were also supported by the Fondo de Investigaciones Sanitarias (FIS, grants PI20/1474 to S.S.J. and PI18/00871 and PI21/00936), co-financed by the ERDF and the 2017-SGR-638 of the Catalan Government to S.S.J. and EvoNano Project (GA800983), funded by European Union’s Horizon 2020 FET Open Programme. N.G.-A. was supported by grants from Pla Estratègic de Recerca i Innovació en Salut (PERIS) of Catalonia (SLT006/17/00270 270)

Expression and Role of MicroRNAs from the miR-200 Family in the Tumor Formation and Metastatic Propensity of Pancreatic Cancer

MicroRNAs from the miR-200 family are commonly associated with the inhibition of the metastatic potential of cancer cells, following inhibition of ZEB transcription factors expression and epithelial-to-mesenchymal transition. However, previous studies performed in pancreatic adenocarcinoma revealed a more complex picture challenging this canonical model. To gain better insights into the role of miR-200 family members in this disease, we analyzed the expression of miR-200a, miR- 200b, miR-200c, miR-141, miR-429, and miR-205, and ZEB1, ZEB2, and CDH1 in pancreatic tumors and matching normal adjacent parenchyma and patient-derived xenografts. We found that miR-200a, miR-429, and miR-205 are frequently overex- pressed in pancreatic tumors, whereas CDH1 is downregulated, and ZEB1 and ZEB2 levels remain unchanged. Furthermore, we measured a positive correlation between miR-200 family mem- bers and CDH1 expression, and a negative correlation between ZEB1 and miR-200c, miR-141, and miR-205 expression, respec- tively. Interestingly, we identified significant changes in expres- sion of epithelial-to-mesenchymal transition regulators and miR-200 members in patient-derived xenografts. Lastly, func- tional studies revealed that miR-141 and miR-429 inhibit the tumorigenic potential of pancreatic cancer cells. Taken together, this comprehensive analysis strongly suggests that miRNAs from the miR-200 family, and in particular miR-429, may act as a tu- mor suppressor gene in pancreatic cancer

Fine Control of In Vivo Magnetic Hyperthermia Using Iron Oxide Nanoparticles with Different Coatings and Degree of Aggregation

Cancer; Magnetic hyperthermia; NanoparticlesCáncer; Hipertermia magnética; NanopartículasCàncer; Hipertèrmia magnètica; NanopartículesThe clinical implementation of magnetic hyperthermia has experienced little progress since the first clinical trial was completed in 2005. Some of the hurdles to overcome are the reliable production of magnetic nanoparticles with controlled properties and the control of the temperature at the target tissue in vivo. Here, forty samples of iron oxide superparamagnetic nanoparticles were prepared by similar methods and thoroughly characterized in terms of size, aggregation degree, and heating response. Selected samples were intratumorally administered in animals with subcutaneous xenografts of human pancreatic cancer. In vivo experiments showed that it is possible to control the rise in temperature by modulating the field intensity during in vivo magnetic hyperthermia protocols. The procedure does not require sophisticated materials and it can be easily implemented by researchers or practitioners working in magnetic hyperthermia therapies.This research was funded by European Commission H2020 programme (NoCanTher project, grant agreement no. 685795), the Ministerio de Ciencia e Innovación (PID2019-106301RB-I00), Comunidad de Madrid (Consejería de Educación e Investigación, NANOMAGCOST-CM, ref. P2018/NMT-4321), COST actions MyWave (CA17115) and Nano2Clinic (CA17140). MICINN “Redes de Investigación” (RED2018-102626-T). IMDEA Nanociencia acknowledges support from the ‘Severo Ochoa’ Programme for Centres of Excellence in R&D (MINECO, CEX2020-001039-S)

Targeting antitumoral proteins to breast cancer by local administration of functional inclusion bodies

Biofabrication; Cancer therapy; Functional amyloidsBiofabricación; Terapia contra el cáncer; Amiloides funcionalesBiofabricació; Teràpia contra el càncer; Amiloides funcionalsTwo structurally and functionally unrelated proteins, namely Omomyc and p31, are engineered as CD44-targeted inclusion bodies produced in recombinant bacteria. In this unusual particulate form, both types of protein materials selectively penetrate and kill CD44+ tumor cells in culture, and upon local administration, promote destruction of tumoral tissue in orthotropic mouse models of human breast cancer. These findings support the concept of bacterial inclusion bodies as versatile protein materials suitable for application in chronic diseases that, like cancer, can benefit from a local slow release of therapeutic proteins.This study has been supported by La Fundacio Marato TV3 and NanoCanTri (CIBER-BBN) to E.V. and I.A., and partially by ISCIII (PI15/00272 and PI1702242 co-founded by Fondo Europeo de Desarrollo Regional (FEDER), to E.V. and S.S., respectively), and Agencia Estatal de Investigacion (AEI) and FEDER (BIO2016-76063-R, AEI/FEDER, UE), AGAUR (2017SGR-229) and CIBER-BBN (VENOM4CANCER) granted to A.V. Protein production and DLS have been partially performed by the ICTS "NANBIOSIS," more specifically by the Protein Production Platform of CIBER-BBN/IBB () and the Biomaterial Processing and Nanostructuring Unit (), respectively. Biodistribution and immunohistochemistry assays were performed at the ICTS "NANBIOSIS," specifically by U20/FVPR (). L.S.-G. was supported by predoctoral fellowship from AGAUR (2018FI_B2_00051). L.S. was supported by the European Research Council (CoG #617473) and the Instituto de Salud Carlos III (FIS #PI16/01224). J.S.-F. was supported by an AECC post-doctoral fellowship. A.V. received an ICREA ACADEMIA awar

Targeting antitumoral proteins to breast cancer by local administration of functional inclusion bodies

Biofabrication; Cancer therapy; Functional amyloidsBiofabricación; Terapia contra el cáncer; Amiloides funcionalesBiofabricació; Teràpia contra el càncer; Amiloides funcionalsTwo structurally and functionally unrelated proteins, namely Omomyc and p31, are engineered as CD44-targeted inclusion bodies produced in recombinant bacteria. In this unusual particulate form, both types of protein materials selectively penetrate and kill CD44+ tumor cells in culture, and upon local administration, promote destruction of tumoral tissue in orthotropic mouse models of human breast cancer. These findings support the concept of bacterial inclusion bodies as versatile protein materials suitable for application in chronic diseases that, like cancer, can benefit from a local slow release of therapeutic proteins.This study has been supported by La Fundacio Marato TV3 and NanoCanTri (CIBER-BBN) to E.V. and I.A., and partially by ISCIII (PI15/00272 and PI1702242 co-founded by Fondo Europeo de Desarrollo Regional (FEDER), to E.V. and S.S., respectively), and Agencia Estatal de Investigacion (AEI) and FEDER (BIO2016-76063-R, AEI/FEDER, UE), AGAUR (2017SGR-229) and CIBER-BBN (VENOM4CANCER) granted to A.V. Protein production and DLS have been partially performed by the ICTS "NANBIOSIS," more specifically by the Protein Production Platform of CIBER-BBN/IBB () and the Biomaterial Processing and Nanostructuring Unit (), respectively. Biodistribution and immunohistochemistry assays were performed at the ICTS "NANBIOSIS," specifically by U20/FVPR (). L.S.-G. was supported by predoctoral fellowship from AGAUR (2018FI_B2_00051). L.S. was supported by the European Research Council (CoG #617473) and the Instituto de Salud Carlos III (FIS #PI16/01224). J.S.-F. was supported by an AECC post-doctoral fellowship. A.V. received an ICREA ACADEMIA awar

Extracellular vesicles from recombinant cell factories improve the activity and efficacy of enzymes defective in lysosomal storage disorders

N-sulfoglucosamina sulfohidrolasa; Teràpia de reemplaçament enzimàtic; Trastorns d’emmagatzematge lisosomalN‐sulfoglucosamina sulfohidrolasa; Terapia de reemplazo enzimático; Trastornos de almacenamiento lisosomalN‐sulfoglucosamine sulfohydrolase; Enzyme replacement therapy; Lysosomal storage disordersIn the present study the use of extracellular vesicles (EVs) as vehicles for therapeutic enzymes in lysosomal storage disorders was explored. EVs were isolated from mammalian cells overexpressing alpha-galactosidase A (GLA) or N-sulfoglucosamine sulfohydrolase (SGSH) enzymes, defective in Fabry and Sanfilippo A diseases, respectively. Direct purification of EVs from cell supernatants was found to be a simple and efficient method to obtain highly active GLA and SGSH proteins, even after EV lyophilization. Likewise, EVs carrying GLA (EV-GLA) were rapidly uptaken and reached the lysosomes in cellular models of Fabry disease, restoring lysosomal functionality much more efficiently than the recombinant enzyme in clinical use. In vivo, EVs were well tolerated and distributed among all main organs, including the brain. DiR-labelled EVs were localized in brain parenchyma 1 h after intra-arterial (internal carotid artery) or intravenous (tail vein) administrations. Moreover, a single intravenous administration of EV-GLA was able to reduce globotriaosylceramide (Gb3) substrate levels in clinically relevant tissues, such kidneys and brain. Overall, our results demonstrate that EVs from cells overexpressing lysosomal enzymes act as natural protein delivery systems, improving the activity and the efficacy of the recombinant proteins and facilitating their access to organs neglected by conventional enzyme replacement therapies.This study has been supported by ISCIII (PI18_00871 co-founded by Fondo Europeo de Desarrollo Regional (FEDER)), and CIBER-BBN (EXPLORE) granted to IA. Different CIBER-BBN units of ICTS ‘NANBIOSIS’ have participated in this work (https://www.nanbiosis.es/platform-units/), more specifically the U1/Protein Production Platform for protein purification, Unit 6 for NTA analysis and TFF purification and U20/FVPR for in vivo assays