Synergistic Combination of Calcium and Citrate in Mesoporous Nanoparticles Targets Pleural Tumors

•Synthesis of mesoporous, amorphous calcium-phosphate citrate nanoparticles•Induction apoptosis selectively in cancer cells•Excellent growth inhibition pleural tumors with minimal adverse effects Pleural are often highly aggressive and rapidly growing. Their proximity to the lung requires selective anticancer agents avoid effects. Such a agent is an urgent, yet unmet clinical need. A promising strategy realize this selectivity may be based on material that degrades cells releases high amounts ions into these cells. Calcium generally nontoxic, but they can induce cell death when released at concentrations. However, triggering their efficient release has not been achieved yet. Here, we developed nanoparticles. These features key intracellular degradation, allowing for ion toxicity toward Successful mouse experiments show potential meet need therapeutic against tumors. Conventional chemotherapy leads severe since it involves systemic administration toxic drugs dosage. Unlike traditional chemotherapeutics, calcium phosphate have both discussed as very inherently toxic. Yet, breakthrough hampered by lack approach overcomes strict regulatory mechanisms cell. present combinatorial calcium, phosphate, colloidal, nanoparticles (CPCs) kill without involvement drugs. The particles neither before endosomal nor after degradation. This allowed us successfully treat two different mice, reducing size about 40% 70% only local applications. Safety assessment studies over 2 months no signs except slightly enhanced thickening eight Calcium-phosphate (CPC) replacement conventional chemotherapeutics. chemotherapeutics toxic, extracellular well tolerated organisms. Moreover, even necessary body concentrations, e.g., main inorganic component bone teeth.1Dorozhkin S.V. Epple M. Biological medical significance phosphates.Angew. Chem. Int. Ed. Engl. 2002; 41: 3130-3146Crossref PubMed Scopus (1566) Google Scholar, 2Mycielska M.E. Patel A. Rizaner N. Mazurek M.P. Keun H. 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At same time, allows co-delivery citrate. hybrid CPCs acidifying followed cytosol. For purpose, synthesized CPC reacting PO43? citric molar ratio 5:3:5. Citric complexes Ca2+, thus, important role reaction kinetics during formation phosphates.18Davies Müller K.H. Wong W.C. Pickard C.J. Reid D.G. Duer bridges between mineral platelets bone.Proc. Natl. Acad. Sci. USA. 111: E1354-E1363Crossref (180) Precipitation induced rapid pH change upon triethanolamine, structure surfactant templates cetyltrimethylammonium chloride (CTAC) Pluronic® F127 (Figure S1). Following extraction S2) yielded desired nanoparticles, depicted Figure 1A. successful synthesis 1B) was verified XRD (amorphous nature) IR-, ssNMR-, TGA-measurements (incorporation phosphate): IR-spectrum CPC, 1C, exhibits some characteristics (such apatite). Additionally, shows strong vibrations 1,414 1,590 cm?1, attributed symmetric antisymmetric stretching modes COO? groups incorporated structure.19Cifuentes I. González-Díaz P.F. 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Acidification 0.1 M HCl led dramatic dye, neutralization 7.4 1F; see Most importantly, preserved S10). Next, investigated lipid-coated calcein-loaded HeLa Lipid-coated were internalized 2A. Time-resolved weak within first 2–4 incubation 20 ?g mL?1 Video S3). accompanied temporary drop NADPH (and reduction equivalent) consequences ATP visible impact viability S11A S11B, black lines). release,23Varkouhi A.K. Scholte Storm G. Haisma H.J. Endosomal escape BioLogicals.J. 151: 220-228Crossref (995) small (CTAC), destabilize membranes. While adsorbed particle, attraction negatively prevents leakage destabilization particles. Once turns acidic,24Gruenberg endocytic pathway: mosaic domains.Nat. 2001; 721-730Crossref (570) dissolves CTAC molecules longer retained particle. When incubating calcein/CTAC-loaded S1) along permanent decrease 2D; S11A–S11D, red lines, associated S12) ultimately via marker caspase 3/7 2C; S4C, attribute observation rupture containing calcein. subsequent content including all components Scholar,25Hansen E.L. Sozer E.B. Romeo Frandsen S.K. Vernier P.T. Gehl Dose-dependent depletion following electroporation, relative effect concentration electric field strength.PLoS One. 2015; e0122973PubMed propose our S13) nature thin walls area. unique create conditions fast dissolution Ca2+. https://www.cell.com/cms/asset/7cf6f118-5dc3-4c60-8c0d-017273ca10f5/mmc2.mp4Loading ... Download .mp4 (117.02 MB) Help files S1. Cells Incubation Therapeutically Active ReleaseOverlay transmission channels. green dots mark CPC. Every 10 min transmittance- GFP-channels recorded stacked video fps. During various fluorescence visualized. Afterwards, morphology undergoes shock order study possible bystander substances surrounding treatment, incubated did loss S14C). Hence, seems sudden preventing undesired damage neighboring above results suggest avoiding studied nontumorigenic line MCF10A. observe S2). Accordingly, MCF10A decreased below 50% 4E). Based result, tested wide variety lines. performed MTT assays measure 72 h. Neither uncoated specific up 100 lines 2E 2F; S14 S15, strongly less 30 2B Interestingly, inhibitory values increasing malignancy reaching lowest Lewis carcinoma 2B; With unusual property, belong group cells.26Gupta P.B. Onder T.T. Jiang Tao Kuperwasser Weinberg Lander E.S. Identification inhibitors high-throughput screening.Cell. 138: 645-659Abstract (1855) Strikingly, none benign, noncancerous significantly affected 2E; S14, efficiency CPCs, demonstrating new generation agents. https://www.cell.com/cms/asset/d03567cc-ad54-453d-ab0d-cbda9996c224/mmc3.mp4Loading (71.43 S2. MCF 10A 5 detected. https://www.cell.com/cms/asset/57a195f4-b76d-4b9a-9a0d-45a884bc285a/mmc4.mp4Loading (135.85 S3. Lipid-Coated Are Not Toxic CellsOverlay 12 few detectable (blue arrows). far therapeutically non-toxic validate promise vivo. chose model adenocarcinoma injection LLC B16F10 syngeneic C57BL/6 mice27Stathopoulos G.T. Everhart Kalomenidis Lawson W.E. Bilaceroglu Peterson T.E. Mitchell Yull F.E. Light R.W. et al.Nuclear factor-?B affects progression malignant effusion.Am. Respir. 2006; 34: 142-150Crossref recapitulates disease date therapy available.28Stathopoulos Malignant effusion: tumor–host interactions unleashed.Am. Crit. Care Med. 186: 487-492Crossref (108) employs bioluminescent (called pNGL) enabling noninvasive imaging mass background full immunocompetence.27Stathopoulos intrapleural therapies currently delivered patients cancers, tumor-cell killing pulmonary tissues remains need.29Lee Y.C.G. Idell Stathopoulos Translational research infection beyond.Chest. 2016; 150: 1361-1370Abstract Scholar,30Debnath Muthuswamy Brugge J.S. Morphogenesis oncogenesis MCF-10A mammary acini grown three-dimensional basement membrane cultures.Methods. 256-268Crossref (1480) whether address applying mg suspended ?L simulated fluid (SBF) intrapleurally days 7 post 3A), foci had already established (intervention group; n = 8). Control consisted SBF (control 9 8, respectively). On day 11/13 injection, mice sacrificed (primary end point, 3B 3D, left panels). Secondary points serial measurements chest bioluminescence, indicator 3C 3E) right panels) number nucleated (an inflammation). Both thoracic luminescence markedly ?40% ?70%, respectively, CPC-treated (for statistical information Supplemental Information) compared controls. groups, indicating inflammation caused process S16). safety further, long-term experiments. experiment, injected weekly alone, respectively. After 8 injections 4A–4C, S17, S18). Visual inspection specifically cavity difference. Also, macrophages might result influenced difference found resulting pleurodesis (obstruction space), actually desirable malignancies.28Stathopoulos second environment. end, xenografts PANO2 system. form slowly growing allow study. would untreated group. Tumors offer additional aggressive, hence much vitro S19). provide comparable environment control. ?l 4 weeks starting injection. Mice tumor-injection experiment. Again, 4D), inflammatory total lavage reduced presence tumor, predicted experiments, microenvironment Importantly, survival tumor-bearing four injections. 16 tumor-free mice. minor effects, combined applications short window days, confirms reduce efficiency. presents option mostly lethal believe composed nontoxic turn composite microenvironment, where act