Search

Introduction. The clinical course of Coronavirus Disease 2019 (COVID-19) is highly heterogenous, ranging from asymptomatic to fatal forms. The identification of clinical and laboratory predictors of poor prognosis may assist clinicians in monitoring strategies and therapeutic decisions. Materials and Methods. In this study, we retrospectively assessed the prognostic value of a simple tool, the complete blood count, on a cohort of 664 patients ( F 260; 39%, median age 70 (56-81) years) hospitalized for COVID-19 in Northern Italy. We collected demographic data along with complete blood cell count; moreover, the outcome of the hospital in-stay was recorded. Results. At data cut-off, 221/664 patients (33.3%) had died and 453/664 (66.7%) had been discharged. Red cell distribution width (RDW) ( χ 2 10.4; p < 0.001 ), neutrophil-to-lymphocyte (NL) ratio ( χ 2 7.6; p = 0.006 ), and platelet count ( χ 2 5.39; p = 0.02 ), along with age ( χ 2 87.6; p < 0.001 ) and gender ( χ 2 17.3; p < 0.001 ), accurately predicted in-hospital mortality. Hemoglobin levels were not associated with mortality. We also identified the best cut-off for mortality prediction: a NL ratio > 4.68 was characterized by an odds ratio for in-hospital mortality OR = 3.40 (2.40-4.82), while the OR for a RDW > 13.7 % was 4.09 (2.87-5.83); a platelet count > 166,000 /μL was, conversely, protective (OR: 0.45 (0.32-0.63)). Conclusion. Our findings arise the opportunity of stratifying COVID-19 severity according to simple lab parameters, which may drive clinical decisions about monitoring and treatment.

The exact role of malignant ascites in the development of intraperitoneal metastases remains unclear, and the mechanisms by which extracellular vesicles (EVs) promote tumor progression in the pre‐metastatic niche have not been fully discovered. In this study, we characterized ascites from high‐grade epithelial ovarian cancer patients. Small‐EVs (30–150 nm) were isolated from two sources—the bulk ascites and the ascitic fluid‐derived tumor cell cultures—and assessed with a combination of imaging, proteomic profiling, and protein expression analyses. In addition, Gene Ontology and pathway analysis were performed using different databases and bioinformatic tools. The results proved that the small‐EVs derived from the two sources exhibited significantly different stiffness and size distributions. The bulk ascitic fluid‐derived small‐EVs were predominantly involved in the complement and coagulation cascade. Small‐EVs derived from ascites cell cultures contained a robust proteomic profile of extracellular matrix remodeling regulators, and we observed an increase in transforming growth factor‐β‐I (TGFβI), plasminogen activator inhibitor 1 (PAI‐1), and fibronectin expression after neoadjuvant chemotherapy. When measured in the two sources, we demonstrated that fibronectin exhibited opposite expression patterns in small‐EVs in response to chemotherapy. These findings highlight the importance of an ascites cell isolation workflow in investigating the treatment‐induced cancer adaption processes., The mechanisms by which extracellular vesicles (EVs) promote ovarian cancer progression in the pre‐metastatic niche have not been fully discovered. This study showed that sampling ascites before and after chemotherapy allows characterizing small‐EVs released by tumor cells, reflecting their biological status. Small‐EVs derived from ascitic fluid‐derived tumor cell cultures contained a robust proteomic profile of extracellular matrix remodeling regulators.

Endometrial cancer is the most common gynecologic malignancy arising from the endometrium. Identification of serum biomarkers could be beneficial for its early diagnosis. We have used 2D-Difference In Gel Electrophoresis (2D-DIGE) coupled with Mass Spectrometry (MS) procedures to investigate the serum proteome of 15 patients with endometrial cancer and 15 non-cancer subjects. We have identified 16 proteins with diagnostic potential, considering only spots with a fold change in %V ≥ 1.5 or ≤0.6 in intensity, which were statistically significant (p &lt, 0.05). Western blotting data analysis confirmed the upregulation of CLU, ITIH4, SERPINC1, and C1RL in endometrial and exosome cancer sera compared to those of control subjects. The application of the logistic regression constructed based on the abundance of these four proteins separated the controls from the cancers with excellent levels of sensitivity and specificity. After a validation phase, our findings support the potential of using the proposed algorithm as a diagnostic tool in the clinical stage.

Despite the advances in the treatment of rheumatoid arthritis (RA) achieved in the last few years, several patients are diagnosed late, do not respond to or have to stop therapy because of inefficacy and/or toxicity, leaving still a huge unmet need. Tissue-specific strategies have the potential to address some of these issues. The aim of the study is the development of a safe nanotechnology approach for tissue-specific delivery of drugs and diagnostic probes. CD34 + endothelial precursors were addressed in inflamed synovium using targeted biodegradable nanoparticles (tBNPs). These nanostructures were made of poly-lactic acid, poly-caprolactone, and PEG and then coated with a synovial homing peptide. Immunofluorescence analysis clearly demonstrated their capacity to selectively address CD34 + endothelial cells in synovial tissue obtained from human, mouse, and rat. Biodistribution studies in two different animal models of rheumatoid arthritis (antigen-induced arthritis/AIA and collagen-induced arthritis/CIA) confirmed the selective accumulation in inflamed joints but also evidenced the capacity of tBNP to detect early phases of the disease and the preferential liver elimination. The therapeutic effect of methotrexate (MTX)-loaded tBNPs were studied in comparison with conventional MTX doses. MTX-loaded tBNPs prevented and treated CIA and AIA at a lower dose and reduced administration frequency than MTX. Moreover, MTX-loaded tBNP showed a novel mechanism of action, in which the particles target and kill CD34 + endothelial progenitors, preventing neo-angiogenesis and, consequently, synovial inflammation. tBNPs represent a stable and safe platform to develop highly-sensitive imaging and therapeutic approaches in RA targeting specifically synovial neo-angiogenesis to reduce local inflammation.

The efficacy of antibody-based immunotherapy is due to the activation of apoptosis, the engagement of antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity (CDC). We developed a novel strategy to enhance CDC using bispecific antibodies (bsAbs) that neutralize the C-regulators CD55 and CD59 to enhance C-mediated functions. Two bsAbs (MB20/55 and MB20/59) were designed to recognize CD20 on one side. The other side neutralizes CD55 or CD59. Analysis of CDC revealed that bsAbs could kill 4-25 times more cells than anti-CD20 recombinant antibody in cell lines or cells isolated from patients with chronic lymphocytic leukemia. The pharmacokinetics of the bsAbs was evaluated in a human-SCID model of Burkitt lymphoma. The distribution profile of bsAbs mimics the data obtained by studying the pharmacokinetics of anti-CD20 antibodies, showing a peak in the tumor mass 3-4 days after injection. The treatment with bsAbs completely prevented the development of human/SCID lymphoma. The tumor growth was blocked by the activation of the C cascade and by the recruitment of macrophages, polymorphonuclear and natural killer cells. This strategy can easily be applied to the other anti-tumor C-fixing antibodies currently used in the clinic or tested in preclinical studies using the same vector with the appropriate modifications.

Cotyledonoid leiomyoma of the uterus is a rare variant of benign uterine leiomyoma. It has a favorable attitude, despite some ultrasound presentations. A bulky uterus with a heterogeneous mass with irregular margins, high vascularity, and infiltration of the myometrium can induce the suspicion of a malignant mesenchymal tumor and lead to a radical surgical treatment. If present, some imaging features may suggest this rare type of leiomyoma, thus avoiding extensive surgery, especially in young nulliparous women. We report 13 cases of cotyledonoid leiomyoma with clinical characteristics, imaging features, and a literature review.

Uterine leiomyoma presents the highest incidence among benign tumors of the female reproductive tract. The present study compared the proteome of leiomyoma treated with ulipristal acetate with that of untreated leiomyoma to investigate protein expression patterns in relation to oxidative stress. Paired tissue samples from seven treated and untreated leiomyomas were collected and the proteome was analyzed by two‑dimensional gel electrophoresis (2‑DE). Western blotting was used to validate the results of 2‑DE, and mass spectrom‑ etry was used to identify proteins. The tissue expression of 30 proteins was markedly affected by treatment with ulipristal acetate. Bioinformatics analysis revealed that several of the differentially expressed proteins were involved in the degrada‑ tion of hydrogen peroxide and the synthesis of reactive oxygen species. The present study suggested the involvement of oxida‑ tive stress as a novel mechanism of action of ulipristal acetate. These findings require further investigations to understand the role of ulipristal acetate in the treatment of the leiomyoma.

Abnormal uterine bleeding (AUB) is a common symptom in the female population, with an estimated prevalence of 10 to 30% in fertile age and up to 90% in perimenopausal women. In most cases, AUB is due to a benign cause. However, it can also be a symptom of atypical endometrial hyperplasia or endometrial cancer, a more common disease during menopause which can also affect women in their reproductive age. Considering the high prevalence of this symptom an appropriate diagnostic algorithm is needed. Concerns about the risks, pain, and stress associated with an endometrial biopsy and its impact on the healthcare system make the choice of AUB diagnostic strategy extremely relevant. Even if the scientific community agrees on the definition of AUB, International Guidelines show some differences in the management of women of reproductive age with AUB, especially regarding the age cut-off as an independent indication for endometrial biopsy. This study compared different diagnostic strategies to identify a diagnostic pathway with high sensitivity and specificity but low impact on the health system’s resources. The analysis was based on three diagnostic algorithms defined as part of the guidelines of leading scientific societies. Women of reproductive age with AUB (n = 625) and without risk of endometrial cancer were included in the study. Results showed that the best criterion to investigate AUB in women at low risk of endometrial cancer is not age cut-off but the presence or absence of focal endometrial pathology at the ultrasound and the response to the progestin therapy. This approach makes it possible to perform fewer outpatient hysteroscopic biopsies without excluding positive cases from the examination.

BackgroundEndometriosis is a debilitating gynecologic disease characterized by the implantation of endometrial tissue in ectopic locations, with signs of severe and chronic inflammation. The new knowledge on endometriosis has highlighted the value of secondary prevention through the early diagnosis and treatment of lesions to reduce serious consequences, first of all, infertility and chronic pelvic pain. The purpose of this study is to assess the reliability and validity of the questionnaire, as a tool to precociously identify women with endometriosis, to prevent the progression of symptoms.MethodWe reviewed the literature and selected risk factors, symptoms, and phenotypic traits of the women affected by endometriosis to create the questionnaire divided into 8 modules, with 47 questions. A total of 151 women completed the questionnaires: 51 patients who have endometriosis (the cases) and 100 matched women without endometriosis (the controls). After data collection, bivariate and multivariate analyses were conducted.ResultsWe retained four of the significant variables from a step-down logistic regression, namely chronic pelvic pain, dyspareunia with VAS≥3, painful defecation, and acne, to develop a final "predictive" logistic model achieving 90.2% sensitivity and 75% specificity.ConclusionOur pilot study demonstrated that the questionnaire provides a powerful tool for the secondary prevention of endometriosis.

OBJECTIVE: To study incidence, type, etiology, risk factors, and impact on outcome of nosocomial infections during extracorporeal membrane oxygenation. DESIGN: Retrospective analysis of prospectively collected data. SETTING: Italian tertiary referral center medical-surgical ICU. PATIENTS: One hundred five consecutive patients who were treated with extracorporeal membrane oxygenation from January 2010 to November 2015. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Ninety-two patients were included in the analysis (48.5 [37-56] years old, simplified acute physiology score II 37 [32-47]) who underwent peripheral extracorporeal membrane oxygenation (87% veno-venous) for medical indications (78% acute respiratory distress syndrome). Fifty-two patients (55%) were infected (50.4 infections/1,000 person-days of extracorporeal membrane oxygenation). We identified 32 ventilator-associated pneumonia, eight urinary tract infections, five blood stream infections, three catheter-related blood stream infections, two colitis, one extracorporeal membrane oxygenation cannula infection, and one pulmonary-catheter infection. G+ infections (35%) occurred earlier compared with G- (48%) (4 [2-10] vs. 13 [7-23] days from extracorporeal membrane oxygenation initiation; p < 0.001). Multidrug-resistant organisms caused 56% of bacterial infections. Younger age (2-35 years old) was independently associated with higher risk for nosocomial infections. Twenty-nine patients (31.5%) died (13.0 deaths/1,000 person-days of extracorporeal membrane oxygenation). Infected patients had higher risk for death (18 vs. 8 deaths/1,000 person-days of extracorporeal membrane oxygenation; p = 0.037) and longer ICU stay (32.5 [19.5-78] vs. 19 [10.5-27.5] days; p = 0.003), mechanical ventilation (36.5 [20-80.5] vs. 16.5 [9-25.5] days; p < 0.001), and extracorporeal membrane oxygenation (25.5 [10.75-54] vs. 10 [5-13] days; p < 0.001). Older age (> 50 years old), reason for connection different from acute respiratory distress syndrome, higher simplified acute physiology score II, diagnosis of ventilator-associated pneumonia, and infection by multidrug-resistant bacteria were independently associated to increased death rate. CONCLUSIONS: Infections (especially ventilator-associated pneumonia) during extracorporeal membrane oxygenation therapy are common and frequently involve multidrug-resistant organisms. In addition, they have a negative impact on patients' outcomes.

Background: Invasive mold infections in children with hematological malignancies are associated with high mortality rates. The use of combination antifungal therapy in cases with a severe clinical course is increasing, although information on the efficacy and safety of this approach is limited. Methods: We present a case series of 13 children affected by hemato-oncological disorders who received combination antifungal therapy for invasive mold infections at our center (Pediatric Hematology, San Gerardo Hospital, Monza, Italy) from 2011 to 2016, with the aim of describing their clinical characteristics, types of infections, treatment regimens, clinical outcomes, and treatment safety. Medical records were retrospectively reviewed in order to describe patients’ characteristics. Results: Combination antifungal therapy included liposomal amphotericin associated with caspofungin (5/13, 38.4%), voriconazole (5/13, 38.4%), or posaconazole (3/13, 23.1%). The 12-week treatment response rate was 69.2% (6/13 patients showed complete response, 3/13 partial response). The crude mortality was 30.7% (4/13): half was related to invasive mold infections (2/13, 15.38%) and half to disease progression (2/13, 15.38%). Overall, treatment was well tolerated, and we did not observe any permanent discontinuation of antifungals due to related side effects. Conclusions: In our experience, combination antifungal therapy seems to be a safe option in immunocompromised children with invasive mold infections. Well-designed studies are needed to confirm the safety of this approach and to better understand its efficacy in the pediatric setting. Keywords: antifungal therapy; mold; hematological malignancies

Extracorporeal membrane oxygenation (ECMO) is a life support technique used in patients with respiratory and/or cardiac failure. The ECMO circuit consists of vascular cannulae, a pump and an artificial lung. Infections are among the most common complications associated with ECMO and have a significant impact on the mortality rate. Here we present a narrative literature review regarding the epidemiology, risk factors, pathogenesis and prevention of infectious complications during ECMO support. The prevalence of hospital-acquired infections during ECMO is 10-12% and their occurrence is likely to be more frequent compared with other critically ill patients. Coagulase-negative staphylococci, Candida spp., Enterobacteriaceae and Pseudomonas aeruginosa are the most frequently involved pathogens. A high incidence of ventilator-associated pneumonia was reported (24.4 cases/1000 ECMO days), with a major role unexpectedly played by Enterobacteriaceae. The infectious risk was shown to increase along the duration of the ECMO run, which represents the most important risk factor for the development of infections. Other ECMO-specific factors predisposing to infections include the severity of illness in ECMO patients, the high risk of bacterial translocation from the gut, and ECMO-related impairment of the immune system. Another important issue could be microbial colonisation of catheters, ECMO cannulae and the oxygenator, which is consistent with most commonly observed aetiologies.

Objective To show that a new recombinant protein (MT07) obtained by fusing a synovial-homing peptide to a neutralizing antibody to C5 can be selectively delivered to inflamed synovium and can effectively control joint inflammation in experimental models of arthritis. Methods Binding of MT07 to human, rat, and mouse synovial tissue was evaluated in vitro by immunofluorescence, and selective localization in the inflamed joints of rats was documented in vivo using time-domain optical imaging. The antiinflammatory effect of MT07 was tested in a rat model of antigen-induced arthritis (AIA) and in a mouse model of collagen antibody–induced arthritis (CAIA). Results MT07 was able to bind to samples of inflamed synovium from humans, mice, and rats while failing to recognize uninflamed synovium as well as inflamed mouse lung or rat kidney. In vivo analysis of the biodistribution of MT07 confirmed its preferential homing to inflamed joints, with negligible inhibition of circulating C5 levels. MT07 prevented and resolved established inflammation in a rat model of AIA, as demonstrated by changes in joint swelling, polymorphonuclear cell counts in synovial washes, release of interleukin-6 and tumor necrosis factor α, and tissue damage. A similar therapeutic effect was obtained testing MT07 in a CAIA model. Conclusion Our findings show that the novel recombinant molecule MT07 has the unique ability to selectively target inflamed joints and to exert local control of the inflammatory process by neutralizing the complement system without interfering with circulating C5 levels. We believe that this approach can be extended to other antiinflammatory drugs currently used to treat patients with rheumatoid arthritis.

Sara Capolla,1 Chiara Garrovo,2 Sonia Zorzet,1 Andrea Lorenzon,3 Enrico Rampazzo,4 Ruben Spretz,5 Gabriele Pozzato,6 Luis Núñez,7 Claudio Tripodo,8 Paolo Macor,1,9 Stefania Biffi2 1Department of Life Sciences, University of Trieste, 2Institute for Maternal and Child Health – IRCCS “Burlo Garofolo”, Trieste, 3Animal Care Unit, Cluster in Biomedicine (CBM scrl), Trieste, Italy; 4Department of Chemistry “G. Ciamician”, University of Bologna, Bologna, Italy; 5LNK Chemsolutions LLC, Lincoln, NE, USA; 6Department of Medical, Surgery and Health Sciences, University of Trieste, Trieste, Italy; 7Bio-Target, Inc., University of Chicago, Chicago, IL, USA; 8Department of Human Pathology, University of Palermo, Palermo, Italy; 9Callerio Foundation Onlus, Institutes of Biological Researches, Trieste, Italy Abstract: The expectations of nanoparticle (NP)-based targeted drug delivery systems in cancer, when compared with convectional therapeutic methods, are greater efficacy and reduced drug side effects due to specific cellular-level interactions. However, there are conflicting literature reports on enhanced tumor accumulation of targeted NPs, which is essential for translating their applications as improved drug-delivery systems and contrast agents in cancer imaging. In this study, we characterized biodegradable NPs conjugated with an anti-CD20 antibody for in vivo imaging and drug delivery onto tumor cells. NPs’ binding specificity mediated by anti-CD20 antibody was evaluated on MEC1 cells and chronic lymphocytic leukemia patients’ cells. The whole-body distribution of untargeted NPs and anti-CD20 NPs were compared by time-domain optical imaging in a localized human/mouse model of B-cell malignancy. These studies provided evidence that NPs’ functionalization by an anti-CD20 antibody improves tumor pharmacokinetic profiles in vivo after systemic administration and increases in vivo imaging of tumor mass compared to non-targeted NPs. Together, drug delivery and imaging probe represents a promising theranostics tool for targeting B-cell malignancies. Keywords: active targeting, optical imaging, tumor accumulation

Rituximab is a chimeric monoclonal antibody directed against human CD20 antigen, which is expressed on B-cell lymphocytes and on the majority of B-cell lymphoid malignancies. Herein we report the conjugate of rituximab with the near-infrared (NIR) fluorophore Cy5.5 (RI-Cy5.5) as a tool for in vitro, in vivo, and ex vivo NIR time-domain (TD) optical imaging. In vitro, RI-Cy5.5 retained biologic activity and led to elevated cell-associated fluorescence on tumor cells. In vivo, TD optical imaging analysis of RI-Cy5.5 injected into lymphoma-bearing mice revealed a slow tumor uptake and a specific long-lasting persistence of the probe within the tumor. Biodistribution studies after intraperitoneal and endovenous administration were undertaken to evaluate differences in the tumor uptake. RI-Cy5.5 concentration in the organs after intraperitoneal injection was not as high as after endovenous injection. Ex vivo analysis of biologic tissues and organs by both TD optical imaging and immunohistochemistry confirmed the probe distribution, as demonstrated by imaging experiment in vivo, showing that RI-Cy5.5 selectively accumulated in the tumor tissue and major excretion organs. In summary, the study indicates that NIR TD optical imaging is a powerful tool for rituximab-targeting investigation, furthering understanding of its administration outcome in lymphoma treatment.

In vitro studies have documented β2 glycoprotein I (β2GPI) binding to endothelial cells (ECs) and trophoblast using antiphospholipid antibodies. The in vivo binding of β2GPI to these cells and the conditions that favor their interaction have not been investigated. We analyzed the in vivo distribution of cyanine 5.5-labeled β2GPI in mice and evaluated the effect of pregnancy and circulating antibodies on its tissue localization. The signal was detected in the liver by whole body scan and ex vivo analysis. The β2GPI failed to bind to the vascular endothelium and reacted only with the ECs of uterine vessels. In pregnant mice the protein was localized on ECs and trophoblast at the embryo implantation sites. Immunized mice showed a similar β2GPI biodistribution to naive mice but the immunized pregnant animals exhibited a significant increase in fetal loss associated with C3 and C9 deposition at the implantation sites. Treatment of mice with LPS after β2GPI-Cy5.5 injection promoted protein localization on gut and brain ECs associated with IgG, C1q, and C9 deposition in immunized mice. These findings indicate that β2GPI binding to EC requires priming with pro-inflammatory factors which is not needed for uterine and placental localization probably dependent on hormonal changes.

Background Bac7 is a proline-rich peptide with a potent in vitro antimicrobial activity against Gram-negative bacteria. Here we investigated its activity in biological fluids and in vivo using a mouse model of S. typhimurium infection. Results The efficacy of the active 1-35 fragment of Bac7 was assayed in serum and plasma, and its stability in biological fluids analyzed by Western blot and mass spectrometry. The ability of the peptide to protect mice against Salmonella was assayed in a typhoid fever model of infection by determination of survival rates and bacterial load in liver and spleen of infected animals. In addition, the peptide's biodistribution was evaluated by using time-domain optical imaging. Bac7(1-35) retained a substantial in vivo activity showing a very low toxicity. The peptide increased significantly the number of survivors and the mean survival times of treated mice reducing the bacterial load in their organs despite its rapid clearance. Conclusions Our results provide a first indication for a potential development of Bac7-based drugs in the treatment of salmonellosis and, eventually, other Gram-negative infections. The in vivo activity for this peptide might be substantially enhanced by decreasing its excretion rate or modifying the treatment schedule.

The design of new photosensitizers with enhanced phototoxicity and pharmacokinetic properties remains a central challenge for cancer photodynamic therapy (PDT). In this study, Pheophorbide a (Pba) has been pegylated to methoxypolyethylene glycol (mPE G-Pba) to produce a soluble photosensitizer that exhibits a higher tissue distribution than free Pba. In vitro studies have shown that mPE G-Pba promotes a fairly strong photosensitizing effect in cancer cells, as previously observed for the unpegylated molecule. mPE G-Pba targets the mitochondria where, following photoactivation, ROS are produced which cause a cellular injury by lipid peroxidation. The effect of pegylation on the photosensitizer biodistribution has been examined in different selected organs of female mice, at different time points after intraperitoneal administration of the drug (50 μmol/Kg body weight). Other than free Pba, which showed a low tissue accumulation, mPE G-Pba has been detected in significant amounts (8 to 16 μg/ml) in liver, spleen, duodenum and kidney and, 3-5 hours after intraperitoneal injection, in moderate amounts (3 to 8 μg/ml) in brain and lung. In vivo optical imaging performed on living female C57/BL6 mice bearing a subcutaneous melanoma mass, showed that injected mPEG-Pba distributes all over the body, with an higher uptake in the tumor respect to free Pba. Our results indicate that although pegylation somewhat decreases the phototoxicity, it significantly increases the drug solubility and tissue distribution and tumor uptake of mPE G-Pba, making the conjugate an interesting photosensitizer for PDT.

We report a 12-year-old patient with growth retardation, exercise intolerance, lactic acidosis (increasing after exercise) and autoimmune polyendocrinopathy type 2. Muscle biopsy shows abundant COX-negative fibers, subsarcolemmal mitochondrial aggregates and markedly reduced activities of all respiratory chain complexes. Genetic analysis identified two new cosegregating mutations in Met-tRNA (m.4415A>G) and Cox III (m.9922A>C), located in highly conserved regions of MtDNA. Both the mutations are heteroplasmics in multiple patients' tissues. Single-muscle fiber analysis showed significantly higher levels of both the mutations in COX-negative than in normal fibers. In addition, a possible link between the mitochondrial dysfunction and the autoimmune disease is suggested.

Current B-cell disorder treatments take advantage of dose-intensive chemotherapy regimens and immunotherapy via use of monoclonal antibodies. Unfortunately, they may lead to insufficient tumor distribution of therapeutic agents, and often cause adverse effects on patients. In this contribution, we propose a novel therapeutic approach in which relatively high doses of Hydroxychloroquine and Chlorambucil were loaded into biodegradable nanoparticles coated with an anti-CD20 antibody. We demonstrate their ability to effectively target and internalize in tumor B-cells. Moreover, these nanoparticles were able to kill not only p53 mutated/deleted lymphoma cell lines expressing a low amount of CD20, but also circulating primary cells purified from chronic lymphocitic leukemia patients. Their safety was demonstrated in healthy mice, and their therapeutic effects in a new model of Burkitt's lymphoma. The latter serves as a prototype of an aggressive lympho-proliferative disease. In vitro and in vivo data showed the ability of anti-CD20 nanoparticles loaded with Hydroxychloroquine and Chlorambucil to increase tumor cell killing in comparison to free cytotoxic agents or Rituximab. These results shed light on the potential of anti-CD20 nanoparticles carrying Hydroxychloroquine and Chlorambucil for controlling a disseminated model of aggressive lymphoma, and lend credence to the idea of adopting this therapeutic approach for the treatment of B-cell disorders.

Steroids are the standard therapy for autoimmune hepatitis (AIH) but the long-lasting administration is hampered by severe side effects. Methods to improve the tropism of the drug toward the liver are therefore required. Among them, conjugation to nanoparticles represents one possible strategy. In this study, we exploited the natural liver tropism of Avidin-Nucleic-Acid-Nano-Assemblies (ANANAS) to carry dexamethasone selectively to the liver in an AIH animal model. An acid-labile biotin-hydrazone linker was developed for reversible dexamethasone loading onto ANANAS. The biodistribution, pharmacokinetics and efficacy of free and ANANAS-linked dexamethasone (ANANAS-Hz-Dex) in healthy and AIH mice were investigated upon intraperitoneal administration. In ANANAS-treated animals, the free drug was detected only in the liver. Super-resolution microscopy showed that nanoparticles segregate inside lysosomes of liver immunocompetent cells, mainly involved in AIH progression. In agreement with these observational results, chronic low-dose treatment with ANANAS-Hz-Dex reduced the expression of liver inflammation markers and, in contrast to the free drug, also the levels of circulating AIH-specific autoantibodies. These data suggest that the ANANAS carrier attenuates AIH-related liver damage without drug accumulation in off-site tissues. The safety and biodegradability of the ANANAS carrier make this formulation a promising tool for the treatment of autoimmune liver disorders.