Liposomal hamycin in the control of experimental aspergillosis in mice: relative toxicity, therapeutic efficacy and tissue distribution of free and liposomal hamycin.

Therapeutic efficacy of liposomal Hamycin has been evaluated in an animal model system for aspergillosis in Balb/c mice. Hamycin was intercalated into soya phosphatidyl choline (SPC), SPC: choline (1:1, vol./vol.) and DMPC liposomes. A single dose of either 0.1 mg/kg, 0.25 mg/kg or 0.5 mg/kg of liposomal Hamycin and 0.1 mg/kg of free Hamycin was injected (i.v.) into animals infected with Aspergillus fumigatus. An increase in the survival rate of animals along with decrease in fungal count in various organs was observed with liposomal administration. Incorporation of cholesterol into liposomes decreased the in vivo toxicity of Hamycin in a dose dependent manner. However, antifungal activity both in the presence and absence of cholesterol showed marked variation as compared to that of non-aromatic polyenes, e.g. amphotericin B. Analysis of Hamycin distribution by HPLC in various tissues revealed higher blood concentration of this drug, when given in free form, compared to its liposomised form. These studies suggest that liposomal Hamycin is more effective than free Hamycin in controlling the experimental Aspergillosis.     

Design of liposomes to improve delivery of amphotericin-B in the treatment of aspergillosis

The efficacy and toxicity of free and liposome intercalated amphotericin-B (Amp-B) in controlling Aspergillosis, caused byAspergillus fumigatus in BALB/c mice were studied. Liposomal Amp-B had higher LD₅₀ (8.1 mg/kg) as compared to that of the free drug (1.2 mg/kg). An improvement in the therapeutic index of the drug was observed with liposomal formulation of the drug. We also focussed on the effect of lipid composition and surface sugar in modulating the therapeutic potency of Amp-B. The most effective liposomal preparation was composed of egg phosphatidylcholine (EPC) : L--phosphatidylethanolamine, dipalmitoyl (DPPE): cholesterol (Chol) in the molar ratio of 6:1:3. Amp-B intercalated into mannose grafted liposomes (LD₅₀ = 9.3 mg/kg) was more effective as compared to the other formulation tested.     

Liposomal muramyl dipeptide therapy of experimental M5076 liver metastases in mice

Summary The effectiveness ofN-acetylmuramyl-l-alanyl-d-isoglutamine (MDP) or of liposomes containing a lipophilic MDP derivative, MDP-glyceroyldipalmitate MDP-GDP in inhibiting the growth of M5076 reticulum cell sarcoma liver metastases in C57BL/6 mice has been determined. MDP (100 µg) or liposomal MDP-GDP (2.5 µmol containing 1 µg) were equally effective in inhibiting liver metastatic growth when given as a single treatment 3 days before tumor cell injection. Therapeutic treatment, initiated 3 days after tumor cell injection and continued for a period of 2 weeks, failed to inhibit metastatic growth. Activation of thioglycollate-elicited peritoneal macrophages or Kupffer cells in vitro with MDP or liposomal MDP-GDP resulted in the expression of tumoricidal activity against M5076 tumor cells. Adoptive cellular therapy with four injections of 2 × 10⁶ macrophages was ineffective: activation of the macrophages with either MDP or liposomal MDP-GDP prior to injection was effective in inhibiting liver metastatic growth. Incorporation of the macrophage toxin dichlorodimethylene diphosphonate within liposomes containing MDP-GDP abolished the ability of such liposomes to induce macrophage or Kupffer cell tumoricidal activity in vitro as well as the antitumor activity when administered 3 days before tumor cell challenge.     

Effects of endotoxin (lipopolysaccharides) on experimental aspergillosis in leukemic mice

In an attempt to evaluate effects of bacterial endotoxin on systemic fungal infection, experimental systemic Aspergillus infection was induced in leukemic mice (AKR strain) with (Group B) or without (Group A) preceding intraperitoneal administration of Pseudomonas aeruginosa derived lipopolysaccharides. All mice in group A died within 4 days after intravenous inoculation of Aspergillus fumigatus spores and developed extensive and disseminated fungal lesions without inflammatory reactions. In contrast, 5 of 10 mice in group B were alive at day 4 and 1 mouse in group B was alive when the experiment was terminated on the 14th day. These mice showed less extensive fungal lesions with definite, albeit minimal, inflammatory reactions which were composed of macrophages and neutrophils. In addition, serum iron levels and iron saturation rates were significantly lower in mice in group B than in group A. These results indicate that P. aeruginosa endotoxin has a deterring effect on systemic Aspergillus infection in leukemic mice.     

Folate receptor targeted NIR cleavable liposomal delivery system augment penetration and therapeutic efficacy in breast cancer

BackgroundLiposomes are predominantly used sorts of nanocarriers for active a targeted delivery through surface functionalization using targeting ligand. The folate receptors are overexpressed in various cancers including breast cancer and because of its binding aptitude specifically to folate receptors, folic acid became the attractive ligand.MethodsIn this research, we have developed a folate and Poly-l-Lysine conjugate and coated this conjugate onto the liposomes. The prepared liposomes were characterized using DLS, FTIR, NMR, SEM, TEM, XRD, AFM, stability and drug release studies. Furthermore, in vitro studies were carried out on FR overexpressed breast cancer cell line.ResultsThe FA-LUT-ABC-Lip have diameter of 183 ± 3.17 nm with positive surface charge +33.65 ± 3 mV and the drug release studies confirm the NIR responsive payload cleavage. The coated formulation (in presence of NIR light) effectively reduced the IC₅₀ values and kills breast cancer cells through FR mediated internalization and accelerated drug release. Moreover, LUT Formulation shows anticancer effect due to significant inhibition of cell migration and proliferation by regulating VEGF expression and induced apoptosis through the caspase-3 up-regulation.ConclusionIt is evident from the in vitro studies that the formulation was found to be very effective and can be explored for triggered and targeted delivery of the substances through active targeting.General significanceCombining receptor mediated drug delivery with triggered release aid in more amounts of drug reaching the target site and achieving enhanced therapeutic activity.     

Preventive and therapeutic euphol treatment attenuates experimental colitis in mice

Background

The tetracyclic triterpene euphol is the main constituent found in the sap of Euphorbia tirucalli. This plant is widely known in Brazilian traditional medicine for its use in the treatment of several kinds of cancer, including leukaemia, prostate and breast cancers. Here, we investigated the effect of euphol on experimental models of colitis and the underlying mechanisms involved in its action.

Methodology/Principal Findings

Colitis was induced in mice either with dextran sulfate sodium (DSS) or with 2,4,6-trinitrobenzene sulfonic acid (TNBS), and the effect of euphol (3, 10 and 30 mg/kg) on colonic injury was assessed. Pro-inflammatory mediators and cytokines were measured by immunohistochemistry, enzyme-Linked immunoabsorbent assay (ELISA), real time-polymerase chain reaction (RT-PCR) and flow cytometry. Preventive and therapeutic oral administration of euphol attenuated both DSS- and TNBS-induced acute colitis as observed by a significant reduction of the disease activity index (DAI), histological/microscopic damage score and myeloperoxidase (MPO) activity in colonic tissue. Likewise, euphol treatment also inhibited colon tissue levels and expression of IL-1β, CXCL1/KC, MCP-1, MIP-2, TNF-α and IL-6, while reducing NOS2, VEGF and Ki67 expression in colonic tissue. This action seems to be likely associated with inhibition of activation of nuclear factor-κB (NF-κB). In addition, euphol decreased LPS-induced MCP-1, TNF-α, IL-6 and IFN-γ, but increased IL-10 secretion from bone marrow-derived macrophages in vitro. Of note, euphol, at the same schedule of treatment, markedly inhibited both selectin (P- and E-selectin) and integrin (ICAM-1, VCAM-1 and LFA-1) expression in colonic tissue.

Conclusions/Significance

Together, these results clearly demonstrated that orally-administered euphol, both preventive or therapeutic treatment were effective in reducing the severity of colitis in two models of chemically-induced mouse colitis and suggest this plant-derived compound might be a potential molecule in the management of inflammatory bowel diseases.     

Tissue distribution evaluation of stealth pH-sensitive liposomal cisplatin versus free cisplatin in Ehrlich tumor-bearing mice

Multidrug resistance and drug toxicity represent major obstacles to cancer chemotherapy. Drug delivery systems, such as liposomes, offer improved chemical stability of encapsulated drugs, enhanced accumulation in tumors and decreased toxicity. The aim of this study was to evaluate the tissue distribution of stealth pH-sensitive liposomes containing cisplatin (SpHL-CDDP), compared with free cisplatin (CDDP), in solid Ehrlich tumor-bearing mice. After administering a 6 mg/kg single intravenous bolus injection of either free radiolabeled cisplatin or SpHL containing radiolabeled cisplatin, blood and tissues were analyzed for cisplatin content by determining radioactivity using an automatic scintillation apparatus. The area under the CDDP concentration-time curve (AUC) obtained for blood after SpHL-CDDP administration was 2.1 fold larger when compared with free CDDP treatment. The longer circulation of SpHL-CDDP led to a higher tumor AUC, and the determination of the ratio between AUC in each tissue and that in blood (Kp) showed a higher accumulation of CDDP in SpHL-CDDP administrated tumors. The SpHL-CDDP was also significantly uptaken by the liver and spleen. The distribution of SpHL-CDDP in these organs was extensive, revealing a high extravasation of CDDP to the tissues. The SpHL-CDDP kidney uptake was also greater than that of free CDDP; however, the Kp value found was lower. This indicates that the SpHL-CDDP led to a reduction of CDDP retention by renal tissue. Thus, these results indicate that the SpHL-CDDP may indeed be useful in alleviating renal damage induced by CDDP and thus represents a promising delivery system for cancer treatment through CDDP.     

Amniotic fluid-derived extracellular vesicles: characterization and therapeutic efficacy in an experimental model of bronchopulmonary dysplasia

Background aimsExtracellular vesicles (EVs) are being tested for their use as novel therapeutics. However, the optimal source of EVs is currently under investigation. Amniotic fluid (AF) is a natural source of EVs that can be easily obtained for use in regenerative medicine, yet AF-EV characterization has not been fully explored.MethodsHere the authors demonstrate AF as a rich source of EVs and identify the microRNA and proteomic cargo. Bioinformatics analysis of this cargo revealed multiple pathway targets, including immunomodulatory, anti-inflammatory and free radical scavenging networks. The authors further demonstrated the therapeutic potential of this EV product as a novel preventative agent for bronchopulmonary dysplasia (BPD).ResultsIntra-tracheal administration of AF-EVs preserved alveolar development, attenuated vascular remodeling and pulmonary hypertension, decreased lung pro-inflammatory cytokine expression and reduced macrophage infiltration in an experimental BPD model.ConclusionsThe authors’ results suggest that AF is a viable biological fluid for EV harvest and that AF-EVs have strong therapeutic potential for pulmonary diseases, such as BPD, warranting further development to transition this novel EV product into the clinic.     

Generation from tumor-bearing mice of lymphocytes with in vivo therapeutic efficacy

Systemic transfer of sensitized lymphocytes can effectively mediate the regression of established tumors. However, virtually all prior experimental applications of this approach have utilized lymphocytes from animals that have been immunized to reject tumor challenge. A similar source of cells is not available in the human. With the use of a weakly immunogenic murine tumor, MCA 105, we demonstrate here that following in vitro sensitization (IVS) with viable tumor cells and interleukin 2, the nontherapeutic lymphoid cells from mice bearing a progressively growing tumor acquired antitumor reactivity capable of mediating the regression of established pulmonary metastases. Although the IVS system induced nonspecific lymphokine-activated killer-like cytotoxic activity from lymphoid cells of normal as well as tumor-bearing mice, therapeutically active cells could only be generated from cultures initiated with lymphoid cells from tumor-bearing animals, indicating that the IVS was a secondary in vitro immune response. Without other treatment, the IVS cells could mediate antitumor effects. However, low doses of exogenous interleukin 2 administration could enhance their therapeutic efficacy. By in vivo T cell subset depletion with monoclonal antibodies, the primary effector cells were identified as belonging to cytotoxic/suppressor T cell lineage expressing the Lyt-2 phenotype. In addition, these therapeutic effector cells could be further expanded in numbers in vitro with continuous stimulation by tumor cells in the presence of interleukin 2. Compared to the number of cells initiating the culture, as many as 126 times the number of cells were obtained after 9 days of IVS followed by in vitro expansion for an additional 5 days. Studies on the kinetics of the occurrence of the pre-effector lymphocytes during tumor growth revealed that they were readily obtained from draining lymph nodes of mice with a broad range of tumor burdens as well as durations of tumor growth. The ability to generate and expand, in vitro, therapeutically active lymphocytes from tumor-bearing hosts has important implications for cellular therapy of human cancers.     

Effect of cholesterol in various liposomal compositions on the in vivo toxicity, therapeutic efficacy, and tissue distribution of amphotericin B

The effect of cholesterol in neutral, positively and negatively charged liposomes on the toxicity, therapeutic efficacy, and alteration in the tissue distribution pattern of amphotericin B (Amp-B) in normal and infected mice was studied. It was observed that inclusion of cholesterol (CHOL) into egg phosphatidylcholine (EPC) liposomes increased the LD50 of Amp-B from 5.3 to 8.5 mg/kg body weight. In the case of phosphatidylserine (PS) liposomes as well as stearylamine (SA) liposomes, cholesterol incorporation had no effect in altering the toxicity of the drug. The survival pattern of animals with all types of liposomal formulation of Amp-B was similar. The tissue distribution studies indicated that in the case of normal mice, cholesterol inclusion in all types of liposomes increased the organ concentration of the drug in various tissues. In infected animals, the concentration of Amp-B in all organs was increased when cholesterol was included in EPC and EPC/PS liposomes. The organ concentration of Amp-B in lung and liver after 1 h of injection was the same in the case of EPC/SA and EPC/SA/CHOL liposomes. Considering the observations on toxicity, therapeutic efficacy, and tissue distribution, it was suggested that cholesterol had a beneficial therapeutic effect on neutral EPC liposomes.     

Polyene and cytokine treatment of experimental aspergillosis

The aim of the present study was to assess the efficacy of amphotericin B (AMB), AMB-intralipid admixture (AMB-IL) or nystatin-IL formulation (Ny-IL) in combination with granulocyte colony stimulating factor (G-CSF) in treatment of experimental murine aspergillosis. ICR mice were immunosuppressed by cyclophosphamide and 3 days later inoculated intravenously (i.v.) with Aspergillus fumigatus. Treatment was initiated 2 h later and administered for 5 consecutive days (polyenes, i.v.; G-CSF, intraperitoneally). Combination therapy, particularly G-CSF with AMB or AMB-IL, significantly increased the survival rate (up to 87.3%) and prolonged the mean survival time (MST) (up to 28.8 days) in comparison to untreated controls (0% survival, MST 6.7 days) and to treatment with polyenes alone (up to 51.5% survival, MST 18.4 days). These data indicate that combination therapy could be beneficial for management of disseminated aspergillosis in humans.     

Investigation of luromarin efficacy on mice with experimental tick-borne encephalitis

Oral use of Luromarin, extracted from sea algae Zostera asiatica, was efficient in protection of mice from lethal infection induced by highly pathogenic strain of TBE virus, by extending their average lifespan. Luromarin demonstrated potentiating action in combination with ribavirin and cycloferon.     

Genetic control of susceptibility to experimental allergic encephalomyelitis in mice

The genetic control of susceptibility to experimental allergic encephalomyelitis (EAE) was studied in mice. The results indicate that sensitivity to disease is not inherited in a simple Mendelian dominant way. Susceptibility to EAE is governed by genes located outside of the major histocompatibility complex and not byH-2-linkedIr genes. No correlation was observed between susceptibility to disease and the cellular immune response toward the small encephalitogenic protein.     

Protective efficacy of combined administration of the multicomponent vaccine and the immunomodulator myelopid in experimental infections in mice

The influence of myelopid (MP) on the protective activity of polycomponent vaccine VP-4 prepared from the antigens of opportunistic bacteria was studied on experimental infections of mice, caused by Klebsiella pneumoniae, Salmonella typhimurium and Staphylococcus aureus. In staphylococcal and Klebsiella infections the joint administration of vaccine VP-4 and MP produced more pronounced protective effect than each of these preparations, introduced alone. The protective action of vaccine VP-4 was specially enforced by MP in cases of local staphylococcal infection. Recommendations on the joint use of two or more immunomodulating agents are possible only on the basis of the experimental substantiation of their effect in definite infections.     

Boron neutron capture therapy (BNCT) for liver metastasis: therapeutic efficacy in an experimental model

Boron neutron capture therapy (BNCT) was proposed for untreatable colorectal liver metastases. The present study evaluates tumor control and potential radiotoxicity of BNCT in an experimental model of liver metastasis. BDIX rats were inoculated with syngeneic colon cancer cells DHD/K12/TRb. Tumor-bearing animals were divided into three groups: BPA-BNCT, boronophenylalanine (BPA) + neutron irradiation; Beam only, neutron irradiation; Sham, matched manipulation. The total absorbed dose administered with BPA-BNCT was 13 ± 3 Gy in tumor and 9 ± 2 Gy in healthy liver. Three weeks post-treatment, the tumor surface area post-treatment/pre-treatment ratio was 0.46 ± 0.20 for BPA-BNCT, 2.7 ± 1.8 for Beam only and 4.5 ± 3.1 for Sham. The pre-treatment tumor nodule mass of 48 ± 19 mg fell significantly to 19 ± 16 mg for BPA-BNCT, but rose significantly to 140 ± 106 mg for Beam only and to 346 ± 302 mg for Sham. For both end points, the differences between the BPA-BNCT group and each of the other groups were statistically significant (ANOVA). No clinical, macroscopic or histological normal liver radiotoxicity was observed. It is concluded that BPA-BNCT induced a significant remission of experimental colorectal tumor nodules in liver with no contributory liver toxicity.     

Investigation of remaxol efficacy in complex therapy of experimental generalized tuberculosis on mice

Efficacy of remaxol in complex chemotherapy of generalized drug resistant tuberculosis was studied on mice. Mycobacterium tuberculosis 5419 SPBNIIF isolated from a patient with freshey diagnosticated pulmonary tuberculosis resistant to isoniazid (10 mcg/ml), rifampicin (40 mcg/ml), streptomycin (10 mcg/ml) and ethionamide (30 mcg/ml) was used in the experiments. The main polychemotherapy included 4 antituberculosis drugs in the highest therapeutic doses: isoniazid, amikacin, ethambutol and tavanic, the treatment course was 8 weeks. Remaxol was administered in a dose of 25 ml/kg intraperitoneally 5 times a week (14 injections). Significant activating effect of remaxol on the tension of the lung tissue local immunity was revealed by changes in the granuloma cell composition (from mainly epitheliod to mainly lymphoid) and by more frequent large lymphohistiocytic infiltrates. The use of remaxol also greatly increased the absorptive and digestive activity of the peritoneal macrophages phagocytosis, inhibited in the process of the experimental tuberculosis development.