TAp73 represses NF-κB-mediated recruitment of tumor-associated macrophages in breast cancer.

Infiltration of tumor-promoting immune cells is a strong driver of tumor progression. Especially the accumulation of macrophages in the tumor microenvironment is known to facilitate tumor growth and to correlate with poor prognosis in many tumor types. TAp73, a member of the p53/p63/p73 family, acts as a tumor suppressor and has been shown to suppress tumor angiogenesis. However, what role TAp73 has in regulating immune cell infiltration is unknown. Here, we report that low levels of TAp73 correlate with an increased NF-κB-regulated inflammatory signature in breast cancer. Furthermore, we show that loss of TAp73 results in NF-κB hyperactivation and secretion of Ccl2, a known NF-κB target and chemoattractant for monocytes and macrophages. Importantly, TAp73-deficient tumors display an increased accumulation of protumoral macrophages that express the mannose receptor (CD206) and scavenger receptor A (CD204) compared to controls. The relevance of TAp73 expression in human breast carcinoma was further accentuated by revealing that TAp73 expression correlates negatively with the accumulation of protumoral CD163+ macrophages in breast cancer patient samples. Taken together, our findings suggest that TAp73 regulates macrophage accumulation and phenotype in breast cancer through inhibition of the NF-κB pathway.

MNK2 governs the macrophage antiinflammatory phenotype.

Tumor-associated macrophages (TAMs) continuously fine tune their immune modulatory properties, but how gene expression programs coordinate this immune cell plasticity is largely unknown. Selective mRNA translation, controlled by MNK1/MNK2 and mTOR pathways impinging on eIF4E, facilitates reshaping of proteomes without changes in abundance of corresponding mRNAs. Using polysome profiling developed for small samples we show that, during tumor growth, gene expression in TAMs is predominately modulated via mRNA-selective changes in translational efficiencies. These alterations in gene expression paralleled accumulation of antiinflammatory macrophages with augmented phosphorylation of eIF4E, a target of the MNK1 and MNK2 kinases, known to selectively modulate mRNA translation. Furthermore, suppression of the MNK2, but not the mTOR signaling pathway, reprogrammed antiinflammatory macrophages toward a proinflammatory phenotype with the ability to activate CD8+ T cells. Thus, selective changes of mRNA translation depending on MNK2 signaling represents a key node regulating macrophage antiinflammatory functions.

Screening and In Vitro Evaluation of Mucoadhesive Thermoresponsive System Containing Methylene Blue for Local Photodynamic Therapy of Colorectal Cancer.

PURPOSE: Photodynamic therapy (PDT) with methylene blue (MB) constitutes a potentially useful modality for colorectal cancer treatment. The limitations of the formulations containing MB are problems of administration and the inability to get the closeness contact at the site during the appropriate residence time. Present study aimed to develop and characterize mucoadhesive thermoresponsive system containing MB designed as platform for colorectal cancer therapy. METHODS: Formulations composed of different amounts of poloxamer 407 (Polox), Carbopol 934P (Carb), and MB were developed and characterized as rheological, compressional, mucoadhesive and syringeability properties, toxicity, photodynamic action, in vitro MB release profile, and ex vivo MB intestinal permeation. RESULTS: The different compositions resulted in formulations with distinctive macroscopic characteristics and wide range of gelation temperatures. The compressional flow, mucoadhesive, syringeability, and rheological properties were significantly influenced by temperature and/or composition. The MB release from formulation was governed by anomalous transport. In addition, it was observed that MB permeated the intestinal membrane; the formulation possesses photodynamic activity and low toxicity. CONCLUSIONS: The data obtained from the system composed of 20% Polox, 0.15% Carb, and 0.25% MB indicated a potentially functional role in PDT of the colorectal cancer and suggest it is worthy of clinical evaluation.

Microparticles containing propolis and metronidazole: in vitro characterization, release study and antimicrobial activity against periodontal pathogens.

Ethylcellulose microparticles containing metronidazole and propolis extractive solution were prepared and evaluated in vitro against periodontal pathogens. Scanning electron microscopy, particle size analysis, drug entrapment efficiency and drug release of microparticles were determined. The antimicrobial activity of microparticles was evaluated against microorganisms of periodontal importance (Enterococcus faecalis, Streptococcus pyogenes, Streptococcus mutans, Staphylococcus aureus, Klebsiella pneumoniae and Escherichia coli). It was obtained particles with regular morphology, mean diameter of 1.23 µm, and entrapment efficiency for propolis and metronidazole were 91.41% and 22.23%, respectively. In vitro release studies of propolis and metronidazole from microparticles showed prolonged drug release and controlled by Fickian diffusion. Both propolis and metronidazole displayed activity against the tested strains. Moreover, the results showed that the strains of E. faecalis, S. pyogenes and S. mutans were more susceptible to the propolis and E. faecalis to the metronidazole. It was also observed that the amount of metronidazole to inhibit the microorganism strains in the physical mixture with propolis was smaller than in the metronidazole alone, suggesting potentiation effect between propolis and metronidazole. These microparticles would be useful for developing intermediary or eventual dosage form to be administered into the periodontal pocket more easily and safely.

Design as strategy for evaluation of the mechanical properties of binary mixtures composed of poly(methyl vinyl ether-alt-maleic anhydride) and Pluronic F127 for biomedical applications.

The synergism between thermoresponsive and bioadhesive polymers can lead to the optimization of materials with enhanced mechanical and bioadhesive properties. Quality by Design can assure the understanding and control of formulation variables. In this approach, Design of Experiment has been widely utilized as an important strategy. Poly(methyl vinyl ether-alt-maleic anhydride) (PVMMA) is a bioadhesive polymer and Pluronic F127 (PF127) shows thermoresponsiveness. The association of these two polymers has been poorly investigated. The aim of this work was to study the mechanical, bioadhesive and rheological properties of polymer mixtures composed of PVMMA and PF127, in order to select the best conditions and formulations for biomedical applications. Textural properties (hardness, compressibility, adhesiveness, cohesiveness and elasticity), softness index, bioadhesion and rheological characteristics (flow and viscoelasticity) showed that 17.5-20% (w/w) PF127-polymer mixtures displayed improved values of the parameters. However, the rheological interaction parameter showed low synergism, due to the polymers' characteristics and system organization. The formulations displayed gelation temperatures suitable for administration, with improved bioadhesive properties mainly at 34 °C and suggests the formulations can be used for biomedical applications. DoE constituted an important tool to investigate these systems showing the main effects that significantly influence the binary mixtures.

VEGF-C-expressing TAMs rewire the metastatic fate of breast cancer cells.

The expression of pro-lymphangiogenic VEGF-C in primary tumors is associated with sentinel lymph node metastasis in most solid cancer types. However, the impact of VEGF-C on distant organ metastasis remains unclear. Perivascular tumor-associated macrophages (TAMs) play a crucial role in guiding hematogenous spread of cancer cells by establishing metastatic pathways within the tumor microenvironment. This process supports breast cancer cell intravasation and metastatic dissemination. We show here that VEGF-C-expressing TAMs reduce the dissemination of mammary cancer cells to the lungs while concurrently increasing lymph node metastasis. These TAMs express podoplanin and interact with normalized tumor blood vessels expressing VEGFR3. Moreover, clinical data suggest inverse association between VEGF-C-expressing TAMs and breast cancer malignancy. Thus, our study elucidates the paradoxical role of VEGF-C-expressing TAMs in redirecting cancer cells to preferentially disseminate to lymph nodes rather than to lungs, partially achieved by normalizing tumor blood vessels and promoting lymphangiogenesis.

Tumor MHC class I expression alters cancer-associated myelopoiesis driven by host NK cells.

Downregulation of MHC class I (MHCI) molecules on tumor cells is recognized as a resistance mechanism of cancer immunotherapy. Given that MHCI molecules are potent regulators of immune responses, we postulated that the expression of MHCI by tumor cells influences systemic immune responses. Accordingly, mice-bearing MHCI-deficient tumor cells showed reduced tumor-associated extramedullary myelopoiesis in the spleen. Depletion of natural killer (NK) cells abrogated these differences, suggesting an integral role of immune-regulatory NK cells during tumor progression. Cytokine-profiling revealed an upregulation of TNF-α by NK cells in tumors and spleen in mice-bearing MHCI expressing tumors, and inhibition of TNF-α enhanced host myelopoiesis in mice receiving adoptive transfer of tumor-experienced NK cells. Our study highlights a critical role of NK cells beyond its identity as a killer lymphocyte and more importantly, the potential host responses to a localized tumor as determined by its MHCI expression.

Insulin Modulates Paracoccidioides brasiliensis-Induced Inflammation by Restoring the Populations of NK Cells, Dendritic Cells, and B Lymphocytes in Lungs.

Paracoccidioidomycosis, a key issue for Brazilian health service, can be aggravated in patients with impaired immunological responses, such as diabetic patients. We evaluated the role of insulin in inflammatory parameters in diabetic and nondiabetic mice using a systemic mycosis Paracoccidioides brasiliensis (Pb) model. Diabetic C57BL-6 mice and controls were infected with Pb18 and treated with insulin for 12 days prior to experiments. After 55 days, infected diabetic mice exhibited fewer leukocytes in both peritoneal lavage fluid (PeLF) and bronchoalveolar lavage fluid and reduced secretion of interleukin- (IL-) 6 in lungs. In addition, diabetic mice presented a reduced influx of TCD4+ cells, TCD8+ cells, B lymphocytes, NK cells, and dendritic cells compared to control infected groups. Insulin treatment restored the leukocyte number in PeLF and restored the presence of B lymphocytes, dendritic cells, and NK cells in lungs of diabetic animals. The data suggest that diabetic mice present impaired immunological response to Pb18 infection and insulin modulates inflammation by reducing IL-6 levels in lung and CINC-1 levels in spleen and liver homogenates, restoring leukocyte concentrations in PeLF and also restoring populations of dendritic cells and B lymphocytes in lungs of diabetic mice, permitting the host to better control the infection.

The importance of the relationship between mechanical analyses and rheometry of mucoadhesive thermoresponsive polymeric materials for biomedical applications.

Pluronic F127® was associated with a carbomer homopolymer type B, as a model polymer blend to evidence the information provided by rheological and mechanical analyses on the development of bioadhesive thermoresponsive systems. The mechanical analysis enabled to observe that 20% (w/w) Pluronic F127®-polymer blends were harder, more adhesive, more mucoadhesive, more compressive and less soft. In addition, continuous flow rheometry demonstrated that the systems were plastic with rheopexy (15%, w/w, Pluronic F127®) or thixotropic (20%, w/w, Pluronic F127®). Oscillatory rheometry exhibited the increase of temperature, and the polymeric concentration increases the elasticity of the formulations. Moreover, correlation index showed that softness and textural analysis can be correlated and complementary, whereas adhesiveness cannot be correlated to mucoadhesion and is less specific. Rheological interaction parameter and gelation temperature showed that 15/0.25-polymer blend is suitable for pharmaceutical and biomedical application, since it can be administered in the liquid form and be gelled in the application site with proper mucoadhesion that can suggest an improved clinical efficacy. Therefore, the mechanical and rheological analyses are useful to characterize and select the best bioadhesive thermoresponsive formulation for the proposed treatment with improved performance.

Linear correlation between rheological, mechanical and mucoadhesive properties of polycarbophil polymer blends for biomedical applications.

Polycarbophil is widely used in a variety of pharmaceutical formulations, mainly for their strong ability to adhere to the epithelial and mucous barriers (bio/mucoadhesion). On the other hand, its association with the thermoresponsive polymer (poloxamer 407) has been poorly explored. This work investigates the rheological, mechanical and mucoadhesive properties of polymer blends containing polycarbophil and poloxamer 407, in order to select the best formulations for biomedical and pharmaceutical applications. Mechanical (hardness, compressibility, adhesiveness, softness, and mucoadhesion) and rheological characteristics (consistency index, yield value and hysteresis area) showed that 20% (w/w) poloxamer 407- polymer blends exhibited higher values parameters. However, the rheological interaction parameter, which was more sensible than the mechanical interaction parameter, revealed higher synergism for systems comprising 15% (w/w) poloxamer 407, due to the system organization and polymers' properties. Furthermore, gelation temperatures were appropriated, suggesting that polymer blends can be used as biomedical materials, and displaying easy administration, enhanced retention and prolonged residence time at the site of application. Therefore, rheological, mechanical and mucoadhesive characterization provided a rational basis for selecting appropriated systems, useful for mucoadhesive drug delivery systems and biomedical applications.

Environmentally Responsive Systems for Drug Delivery.

BACKGROUND: In recent decades, the development of the environmentally responsive systems for drug delivery has been well regarded, with enormous potential in different applications. <P><P> Methods: These environmentally sensitive, smart, intelligent formulations have the ability to alter their physical properties in response to small changes in physical or chemical conditions, such as temperature, glucose, pH, ultrasound, light, electric field and redox potential with a huge potential in drug delivery systems. The use of formulations containing smart materials enables to carry the drug to the target tissue, cells and release in a triggered way. Consequently, they have demonstrated several advantages like decreased dose frequency, ease of preparation and administration, prolonged release with reduced side effects, as well as, reduced costs when compared to conventional processes for industrial applications. In this sense, many patents have deposited, displaying different pharmaceutical devices using responsive systems. <P><P> Results: There are more than twenty-five patents deposited about thermoresponsive systems. Furthermore, a few number of patents within glucose responsive, ultrasound responsive and light responsive deposited. There also are about eight patents that are pH-responsive, four as electric-field responsive. Most of them cover more than one type of stimuli. <P><P> Conclusion: Therefore, in this review, since 1975 to 2016, we have categorized, reviewed and discussed the patents, applications, pharmaceutical dosage forms, the importance and perspectives of this environmentally responsive approach as potentially useful therapeutic modality.

Preparation and characterization of bioadhesive system containing hypericin for local photodynamic therapy.

Hypericin (Hyp) is a natural photoactive pigment utilized in the treatment of different types of cancer and antimicrobial inactivation using photodynamic therapy (PDT). Hyp is poorly soluble in water leading to problems of administration, getting close contact with the site, and bio-availability. Therefore, this study aimed to develop bioadhesive thermoresponsive system containing Hyp for local PDT. Carbomer 934P, poloxamer 407, and Hyp were used to prepare the thermoresponsive bioadhesive formulations. They were characterized for sol-gel transition temperature, mechanical, mucoadhesive, rheological (continuous flow and oscillatory) and dielectric properties, syringeability, in vitro Hyp release kinetics, ex vivo permeability, and photodynamic activity. The formulations displayed suitable gelation temperature and rheological characteristics. The compressional, mechanical and mucoadhesive properties, as well the syringeability showed the easiness of administration and the permanence of the system adhered to the mucosa or skin. The dielectric analysis helped to understand the Hyp availability, and its release presented an anomalous behavior. The system did not permeate the pig skin nor rat intestine and showed good biological photodynamic activity. Therefore, data obtained from the bioadhesive system indicate a potentially useful role as a platform for local hypericin delivery in PDT, suggesting it is worthy of in vivo evaluation.

Rheological, mucoadhesive and textural properties of thermoresponsive polymer blends for biomedical applications.

The development of binary polymeric mixtures (polymer blends) containing bioadhesive and thermoresponsive polymers can provide new materials for biomedical applications, with higher contact, increased adhesion, prolonged residence time, protection, and in determined cases, secured absorption of an active agent from the site of application. Mixtures were prepared using a wide range of poloxamer 407 and Carbopol 971P(®) amounts. The rheological (flow and oscillatory), sol-gel transition temperature, mechanical (hardness, compressibility, adhesiveness, cohesiveness and elasticity), softness, and mucoadhesive properties of formulations were investigated. Moreover, the interaction between the different proportions of polymers was also analyzed. Continuous shear and oscillatory rheometry identified the plastic flow with various degrees of thixotropy, besides the viscoelastic behavior of formulations. The determination of gelation temperature displayed values ranged from 27.17 to 41.09°C. It was also found that low carbomer concentrations were enough to provide positive interaction parameter. However, the highest values were obtained for the polymeric blends with higher concentration of poloxamer 407. The mucoadhesion and softness index were greater in preparations containing 20% (w/w) poloxamer 407. The rheological, mechanical and mucoadhesive properties of the polymeric blends can be manipulated by changing the concentrations of the polymers and they suggest the blends are worthy of biomedical applications.