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The use of co-processed materials for Orally Disintegrating Tablets (ODT) preparation by direct compression is well consolidated. However, the evaluation of their potential for ODT preparation by 3D printing technology remains almost unexplored. The present study aimed to estimate the use of commercially available co-processed excipients, conventionally applied in compression protocols, for the preparation of ODTs with binder jetting-3D printing technology. The latter was selected among the 3D printing techniques because the deposition of multiple powder layers allows for obtaining highly porous and easily disintegrating dosage forms. The influence of some process parameters, including layer thickness, type of waveform and spread speed, on the physical and mechanical properties of the prototypes printed were evaluated. Our results suggested that binder jetting-3D printing technology could benefit from the co-processed excipients for the preparation of solid dosage forms. The process optimization conducted with the experiments reported in this work indicated that additional excipients were needed to improve the physical properties of the resulting ODTs.

Nanomedicine provides various opportunities for addressing medical challenges associated with drug bioavailability, stability, and efficacy. In particular, oral nanoparticles (NPs) represent an alternative strategy to enhance the solubility and stability of active ingredients through the gastrointestinal tract. The nanocarriers could be used for both local and systemic targeting, enabling controlled release of encapsulated drugs. This approach allows more efficient therapies. In this work, we aim to develop reliable oral solid dosage forms incorporating NPs produced by either one pot synthesis or continuous production, following protocols that yield highly consistent outcomes, promoting their technology transfer and clinical use. Microfluidics technology was selected to allow an automated and highly productive synthetic approach suitable for the highly throughput production. In particular, innovative systems, which combine advantage of NPs and solid dosage formulation, were designed, developed, and characterized demonstrating the possibility to obtaining oral administration. The resulting NPs were thus carried on oral dosage forms, i.e., pellets and minitablets. NPs resulted stable after dosage forms manufacturing, leading to confidence also on protection of encapsulated drugs. Indomethacin was used as a tracer to test biopharmaceutical behaviour. Anti-inflammatories or cytotoxic chemotherapeutics could be vehiculated leading to a breakthrough in the treatment of severe diseases allowing the oral administration of these drugs. We believe that the advancement achieved with the results of our work paves the way for the progression of nanoproducts into clinical transition processes.

Il cancro è una delle principali cause di morte nel mondo e glioblastoma e tumore al seno triplo negativo sono due tipologie di tumore caratterizzate da una prognosi sfavorevole e da un alto rischio di recidive. I trattamenti farmacologici oggi a disposizione non sono sufficienti all’ottenimento di una guarigione completa sia a causa della bassa selettività che impedisce l’utilizzo di dosaggi più elevati, sia a causa dello sviluppo di meccanismi di resistenza da parte delle cellule tumorali. In questo contesto le nanoparticelle possono essere sviluppate sia come nuovi trattamenti terapeutici sia per superare alcune limitazioni delle terapie convenzionali. In questo progetto di tesi, ho scelto di utilizzare nanoparticelle di H-ferritina (HFn), una forma ricombinante di ferritina che possiede alcune caratteristiche che la rendono un buon candidato per migliorare i trattamenti terapeutici già disponibili per questi due tumori. L’HFn è infatti biocompatibile, la sua cavità interna può essere utilizzata per il caricamento di farmaci e molecole di detection e la sua superficie può essere facilmente modificata sfruttando i numerosi gruppi funzionali esposti verso l’esterno. La HFn è, inoltre, in grado di legarsi al recettore della transferrina (TfR1), over-espresso dal 98% dei tumori solidi. Tale recettore è over espresso anche dalle cellule endoteliali che compongono la barriera ematoencefalica, rendendo così la HFn in grado di attraversarla. Il mio lavoro di tesi può essere suddiviso in due progetti. Nel primo progetto i gruppi funzionali esposti sulla superficie della HFn sono stati utilizzati per la coniugazione dell’anticorpo monoclonale cetuximab al fine di studiare il mantenimento della sua attività antitumorale in modelli tumorali 3D e di paragonarla a quella del cetuximab non coniugato. Come modelli tumorali 3D sono stati utilizzati sferoidi ottenuti da una linea cellulare di glioblastoma e da una di tumore al seno triplo negativo. Sulla base dei prom, Cancer is a leading cause of disease worldwide and glioblastoma and triple negative breast cancer represent two types of tumors with poor prognosis and a high risk of recurrence. The therapeutic options currently in use are still subject to failure, due to (1) the poor selectivity of drugs, which compromises dose escalation to more therapeutically active levels and (2) the development of resistance by different mechanisms that cells put in place to defend themselves from the action of drugs. In this scenario, nanoparticles can be developed as new therapeutic strategies or to overcome the limitations of the treatments currently in use. In my PhD project, I exploited the unique features of a nanoparticle-based on recombinant heavy-chain ferritin cages (HFn) to improve the therapeutic treatments already available for these tumors. HFn can be a promising candidate for this study because 1) it is biocompatible; 2) it can be loaded with drugs; 3) it can be easily modified by exploiting the functional groups on its surface; 4) it exhibits tumor targeting by recognition of TfR1, overexpressed in 98% of human cancers and directly correlated with tumor stage or cancer progression; 5) HFn is able to cross the blood-brain barrier (BBB) exploiting the same TfR1 receptor, also found overexpressed on the BBB endothelial wall. My work is divided into two main subprojects, all sharing the use of HFn. In the first subproject (chapter one), the functional groups present on the surface of HFn were exploited for the conjugation of the monoclonal antibody cetuximab, studying the maintenance of its anticancer activity against a 3D tumor model in comparison with unconjugated cetuximab. Cancer spheroids of glioblastoma and triple negative breast cancer were selected and produced for this study. Then we focused on the documented ability of cetuximab to promote an indirect anticancer effect (ADCC) through the activation of immune systems cells. ADCC activation was evaluated both in live imagi

La nanomedicina offre differenti opzioni per ovviare ad alcune problematiche riguardanti la biodisponibilità del farmaco, la sua stabilità o la scarsa efficienza. Ne sono un esempio le nanoparticelle (NPs) somministrate per via orale, che possono rappresentare un’alternativa per migliorare la stabilità o la solubilità di principi attivi nel tratto gastrointestinale. Infatti, le NPs possono favorire un assorbimento sistemico o locale, permettere un rilascio controllato del farmaco riducendo la frequenza di somministrazione e aumentando la compliance del paziente. In questo scenario, lo scopo di questo lavoro è stato quello di sviluppare un protocollo altamente riproducibile per la sintesi di NPs, grazie all’utilizzo di tecnologie innovative così da incentivare il loro trasferimento clinico-tecnologico e la loro messa in commercio. La possibilità di lavorare con queste tecnologie è avvenuta in seguito alla creazione di una piattaforma, chiamata NanoCosPha, che comprende una serie di laboratori all’interno dell’Università Bicocca, che hanno come scopo generale quello di tentare di colmare il divario tra la ricerca accademica e l’industria farmaceutica e cosmetica La microfluidica e il sistema di filtrazione tangenziale sono state scelte per la sintesi e la purificazione di nanoparticelle e, in seguito dell'inserimento del nanoformulato in mini compresse e pellets, ne sono stati analizzati i parametri dimensionali e di monodispersione per confermare la loro stabilità. Questo importante risultato, porta a supporre come la stabilità del carrier possa indicare la stabilità del farmaco incapsulato, aprendo quindi la possibilità di veicolare anche molecole biologiche per via orale Un'altra tecnologia innovativa inserita in questo progetto è rappresentata dalla stampante 3D, che permette di sviluppare formulazioni orali per lo sviluppo nel campo della medicina personalizzata. Prima di investigare il suo uso in combinazione con le NPs, sono stati studiati e valut, The nanomedicine field offers different options to overcome medical issues linked to the low bioavailability of drugs, their stability or low efficiency; for example, oral nanoparticles (NPs) could be selected as an alternative approach to improve the solubility and the stability of active ingredients in the gastrointestinal tract. The nanocarriers could reach both local and systemic drug targeting; allow a controlled release of encapsulated drugs reducing the frequency of administration and ameliorating patient compliance. In this scenario, part of this work aims to develop a highly reproducible protocol to produce NPs from single synthesis to continuous production, thanks to innovative technologies, to promote their technological transfer and their clinical use. These technologies are included in NanoCosPha, a platform composed by several laboratories whose only purpose is to increase nanotechnology academic research to support the pharmaceutical and cosmetics industry. Microfluidics and tangential flow filtration are the technologies choose to enhance the automated synthesis and purification of NPs; then, after nano delivery in oral devices systems, like pellets or minitablets, the size and polydispersity index of NPs are investigated, and the stability is confirmed. These important results provide confidence on protection of encapsulated drugs and lay the foundation for the following future steps concerning the replacement of tracer drug, used for this preliminary data, with biological molecules for their oral administration. Moreover, an innovative formulation, represented in this project, is 3D printed tablet. The advantages of this technique are linked to the possibility to produce almost a single oral device personalized and composed by the ingredients desired for a specific medical need. Before investigating their combination with NPs, the study of instrument parameters, combined with a different mix of powders, is explored in order to better understand Bin

Ribosome-inactivating proteins, including Saporin toxin, have found application in the search for innovative alternative cancer therapies to conventional chemo- and radiotherapy. Saporin's main mechanism of action involves the inhibition of cytoplasmic protein synthesis. Its strong theoretical efficacy is counterbalanced by negligible cell uptake and diffusion into the cytosol. In this work, we demonstrate that by immobilizing Saporin on iron oxide nanoparticles coated with an amphiphilic polymer, which promotes nanoconjugate endosomal escape, a strong cytotoxic effect mediated by ribosomal functional inactivation can be achieved. Cancer cell death was mediated by apoptosis dependent on nanoparticle concentration but independent of surface ligand density. The cytotoxic activity of Saporin-conjugated colloidal nanoparticles proved to be selective against three different cancer cell lines in comparison with healthy fibroblasts.

Background: Surfactant protein-D (SP-D) is a lung-resident protein that has emerged as a potential biomarker for COVID-19. Previous investigations on acute respiratory distress syndrome patients demonstrated a significant increment of SP-D serum levels in pathological conditions. Since SP-D is not physiologically permeable to alveoli-capillary membrane and poorly expressed by other tissues, this enhancement is likely due to an impairment of the pulmonary barrier caused by prolonged inflammation. Methods: A retrospective study on a relatively large cohort of patients of Hospital Pio XI of Desio was conducted to assess differences of the hematic SP-D concentrations among COVID-19 patients and healthy donors and if SP-D levels resulted a risk factor for disease severity and mortality. Results: The first analysis, using an ANOVA-model, showed a significant difference in the mean of log SP-D levels between COVID-19 patients and healthy donors. Significant variations were also found between dead vs survived patients. Results confirm that SP-D concentrations were significantly higher for both hospitalized COVID-19 and dead patients, with threshold values of 150 and 250 ng/mL, respectively. Further analysis conducted with Logistic Mixed models, highlighted that higher SP-D levels at admission and increasing differences among follow-up and admission values resulted the strongest significant risk factors of mortality (model predictive accuracy, AUC = 0.844). Conclusions: The results indicate that SP-D can be a predictive marker of COVID-19 disease and its outcome. Considering its prognostic value in terms of mortality, the early detection of SP-D levels and its follow-up in hospitalized patients should be considered to direct the therapeutic intervention.

The role of cosmetic products is rapidly evolving in our society, with their use increasingly seen as an essentialcontribution to personal wellness. This suggests the necessity of a detailed elucidation of the use of nanoparticles(NPs) in cosmetics. The aim of the present work is to offer a critical and comprehensive review discussing the im-pact of exploiting nanomaterials in advanced cosmetic formulations, emphasizing the beneficial effects of theirextensive use in next-generation products despite a persisting prejudice around the application of nanotechnol-ogy in cosmetics. The discussion here includes an interpretation of the data underlying generic information re-ported on the product labels of formulations already available in the marketplace, information that often lacksdetailsidentifying specific components of the product, especially when nanomaterials are employed. The empha-sis of this review is mainly focused on skincare because it is believed to be the cosmetics market sector in whichthe impact of nanotechnology is being seen most significantly. To date, nanotechnology has been demonstratedto improve the performance of cosmetics in a number of different ways: 1) increasing both the entrapment effi-ciency and dermal penetration of the active ingredient, 2) controlling drug release, 3) enhancing physical stabil-ity, 4) improving moisturizing power, and 5) providing better UV protection. Specific attention is paid to theeffect of nanoparticles contained in semisolid formulations on skin penetration issues. In light of the emergingconcerns about nanoparticle toxicity, an entire section has been devoted to listing detailed examples ofnanocosmetic products for which safety has been investigated

Paolo Rainone,1– 3 Antonella De Palma,4 Francesco Sudati,5 Valentina Roffia,4 Valentina Rigamonti,6 Lucia Salvioni,6 Miriam Colombo,6 Marilena Ripamonti,2 Antonello Enrico Spinelli,7 Davide Mazza,7 Pierluigi Mauri,4 Rosa Maria Moresco,1,2,7 Davide Prosperi,6 Sara Belloli2,7 1Department of Medicine and Surgery, University of Milano-Bicocca, Monza, 20900, Italy; 2Institute of Molecular Bioimaging and Physiology of CNR, Segrate, 20090, Italy; 3Doctorate School of Molecular and Translational Medicine, University of Milan, Milan, Italy; 4Institute of Biomedical Technologies of CNR, Segrate, 20090, Italy; 5PET and Nuclear Medicine Unit, San Raffaele Scientific Institute, Milan, 20132, Italy; 6NanoBioLab, Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milan, 20126, Italy; 7Experimental Imaging Center, San Raffaele Scientific Institute, Milan, 20132, ItalyCorrespondence: Sara BelloliInstitute of Molecular Bioimaging and Physiology of CNR, Via Fratelli Cervi 93, Segrate, 20090, ItalyTel +39 02 26433640Fax +39 02 26432717Email belloli.sara@hsr.itIntroduction: The overexpression of Human Epidermal Growth Factor Receptor 2 (HER2) is usually associated with aggressive and infiltrating breast cancer (BC) phenotype, and metastases. Functionalized silica-based nanocarriers (SiNPs) can be labeled for in vivo imaging applications and loaded with chemotherapy drugs, making possible the simultaneous noninvasive diagnosis and treatment (theranostic) for HER2-positive BC.Methods: Firstly, FITC-filled SiNPs, were engineered with two different amounts of Hc-TZ (trastuzumab half-chain) per single nanoparticle (1:2 and 1:8, SiNPs to Hc-TZ ratio), which was 99mTc-radiolabeled at histidine residues for ex vivo and in vivo biodistribution evaluations. Secondly, nanoparticles were loaded with DOX and their in vitro and ex vivo/in vivo delivery was assessed, in comparison with liposomal Doxorubicin (Caelyx). Finally, the treatment efficacy of DOX-SiNPs-TZ (1