Prolonged retention of liposomes in the pleural cavity of normal mice and high tumor distribution in mice with malignant pleural effusion, after intrapleural injection.

Background: Intrapleural administration of compounds is a lung targeted, innovative therapeutic strategy for mesothelioma, which can be refined as a route for drug delivery that minimizes the potential for systemic toxicity. However, little is currently known about the retention of liposomal drugs at the site, after such topical administration. Purpose: To evaluate the retention of liposomes in lungs following intrapleural injection, and how this might be modulated by liposome properties and disease progression. Methods: DiR-incorporating liposomes with various lipid compositions and sizes were prepared, characterized (for size distribution and zeta potential) and injected intrapleurally in normal mice and mice with malignant pleural effusion (MPE). DiR retention in pleural cavity was followed by biofluorescence imaging. Results: Experimental results demonstrate that liposome size and PEG-coating, have a significant effect on residence time in the pleural cavity; negative surface charge does not. More than 20% liposomal-DiR is retained 24 d post-injection (in some cases), indicating the high potential towards localized diseases. Ex-vivo liposomal-DiR signal in tumors of MPE mice was similar to signal in liver, suggesting high tumor targeting potential of intrapleurally injected liposomes. Finally, no difference was noticed in liposomal-DiR retention between tumor-inoculated (MPE) and healthy mice, indicating the stability of liposomes in the presence of effusion (in MPE mice). Conclusion: The current study provides novel insights for using liposomes by intrapleural administration for the treatment of lung diseases.

Cellular Vesicles: New Insights in Engineering Methods, Interaction with Cells and Potential for Brain Targeting.

Cellular vesicles (CVs) have been proposed as alternatives to exosomes for targeted drug delivery. CVs, prepared from human embryonic kidney 293 cells (HEK-293), C57BL/6 mouse B16F10 skin melanoma cells (B16F10), and immortalized human cerebral microvascular endothelial cells (hCMEC/D3) by liposome technology methods, were characterized for morphology, cytotoxicity, and cell uptake properties. CV brain-targeting potential was evaluated in vitro on the hCMEC/D3 blood-brain barrier (BBB) model, and in vivo/ex vivo. CV sizes were between 135 and 285 nm, and the ζ-potential was negative. The dehydration-rehydration method conferred highest calcein loading and latency to CVs compared with other methods. The increased calcein leakage from CVs when compared with liposomes indicated their poor integrity, which was increased by pegylation. The in vivo results confirmed lower liver uptake by PEG-CVs (compared with nonpegylated) proving that the calcein integrity test is useful for prediction of CV biodistribution, as used for liposomes. The cell uptake of homologous origin CVs was not always higher compared with that of non-homologous. Nevertheless, CVs from hCMEC/D3 demonstrated the highest BBB permeability (in vitro) compared with OX-26 targeted liposomes, and brain localization (in vivo). CVs from hCMEC/D3 cells grown in different media demonstrated decreased interaction with brain cells and brain localization. Significant differences in proteome of the two latter CV types were identified by proteomics, suggesting a potential methodology for identification of organotropism-determining CV components.

Mono and dually decorated nanoliposomes for brain targeting, in vitro and in vivo studies.

PURPOSE: Mono- and dual-decorated (DUAL) liposomes (LIP) were prepared, by immobilization of MAb against transferrin (TfR[OX26 or RI7217]) and/or a peptide analogue of ApoΕ3 (APOe) -to target low-density lipoprotein receptor(LPR)-, characterized physicochemically and investigated for BBB-targeting, in-vitro and in-vivo. METHODS: Human microvascular endothelial cells (hCMEC/D3) were used as BBB model, and brain targeting was studied by in-vivo imaging of DiR-labelled formulations (at two doses and surface ligand densities), followed by ex-vivo organ imaging. RESULTS: LIP diameter was between 100 nm and 150 nm, their stability was good and they were non-cytotoxic. LIP uptake and transport across the hCMEC/D3 cell monolayer was significantly affected by decoration with APOe or MAb, the DUAL exerting an additive effect. Intact vesicle-transcytosis was confirmed by equal transport of hydrophilic and lipophilic labels. In-vivo and ex-vivo results confirmed MAb and DUAL-LIP increased brain targeting compared to non-targeted PEG-LIPs, but not for APOe (also targeting ability of DUAL-LIP was not higher than MAb-LIP). The contradiction between in-vitro and in-vivo results was overruled when in-vitro studies (uptake and monolayer transport) were carried out in presence of serum proteins, revealing their important role in targeted-nanoformulation performance. CONCLUSIONS: A peptide analogue of ApoΕ3 was found to target BBB and increase the targeting potential of TfR-MAb decorated LIP, in-vitro, but not in-vivo, indicating that different types of ligands (small peptides and antibodies) are affected differently by in-vivo applying conditions. In-vitro tests, carried out in presence of serum proteins, may be a helpful predictive "targetability" tool.

Cisplatin: process and future.

One of the most important anticancer agents is cisplatin (CDDP). Numerous studies with a CDDP-based combination have been reported over the last 30 years. The use of CDDP in the 1980s and 1990s showed responses in advanced stage non-small-cell lung cancer (NSCLC). Over the years it was found that the side effects of this agent (particularly nephrotoxicity) were a common problem. Agents such as carboplatin, taxanes, gemcitabine, irinotecan and pemetrexed proved to be effective in NSCLC with reduced or no nephrotoxicity. The administration of these newer agents improved several side effects, but without improving efficacy. When prophylactic (adjuvant) treatment for NSCLC was introduced, CDDP was the agent selected, which indicated the value of the drug. Recently, a novel formulation of CDDP, liposomal cisplatin, which has shown very low toxicity, no nephrotoxicity and equal effectiveness was produced; its importance is its higher effectiveness than standard CDDP in lung adenocarcinoma.

High-dose tamoxifen in breast cancer bone metastasis.

High-dose tamoxifen was used as a treatment for bone metastasis in 16 patients with breast cancer. All of them had been pretreated with hormonal therapy, including lowdose tamoxifen. The results were extremely positive with clinical amelioration and also disappearance of osteolysis in 5 patients.

Secreted phosphoprotein-1 directly provokes vascular leakage to foster malignant pleural effusion.

Secreted phosphoprotein-1 (SPP1) promotes cancer cell survival and regulates tumor-associated angiogenesis and inflammation, both central to the pathogenesis of malignant pleural effusion (MPE). Here, we examined the impact of tumor- and host-derived SPP1 in MPE formation and explored the mechanisms by which the cytokine exerts its effects. We used a syngeneic murine model of lung adenocarcinoma-induced MPE. To dissect the effects of tumor- versus host-derived SPP1, we intrapleurally injected wild-type and SPP1-knockout C57/BL/6 mice with either wild-type or SPP1-deficient syngeneic lung cancer cells. We demonstrated that both tumor- and host-derived SPP1 promoted pleural fluid accumulation and tumor dissemination in a synergistic manner (P<0.001). SPP1 of host origin elicited macrophage recruitment into the cancer-affected pleural cavity and boosted tumor angiogenesis, whereas tumor-derived SPP1 curtailed cancer cell apoptosis in vivo. Moreover, the cytokine directly promoted vascular hyper-permeability independently of vascular endothelial growth factor. In addition, SPP1 of tumor and host origin differentially affected the expression of proinflammatory and angiogenic mediators in the tumor microenvironment. These results suggest that SPP1 of tumor and host origin impact distinct aspects of MPE pathobiology to synergistically promote pleural fluid formation and pleural tumor progression. SPP1 may present an attractive target of therapeutic interventions for patients with MPE.

Combination of three cytotoxic agents in small-cell lung cancer.

PURPOSE: The established treatment for small-cell lung cancer has been a cisplatin-etoposide combination, as the most effective chemotherapy regimen. Paclitaxel has also been used in combination with cisplatin and etoposide but this has been unacceptable due to the toxicity. This toxicity could be attributed to the three consequent days of treatment with etoposide plus the doses of each of the three drugs. Our objectives were to determine an equal or longer survival and lower toxicity by administering all 3 drugs with low dosage on day one, compared to the established guideline of 3-day administration. METHODS: We tested the aforementioned three-drug combination and avoided the toxicity in the majority of patients by administering all 3 drugs on day one. Fifty-one patients (50 evaluable) were recruited from 4 oncology clinics. All patients had histologically or cytologically confirmed small-cell lung cancer with limited and extensive disease in 40 and 60 % of the patients, respectively. The treatment was: cisplatin 75 mg/m(2), etoposide 120 mg/m(2) (maximum 200 mg), and paclitaxel 135 mg/m(2). The agents were administered on day one and repeated every 3 weeks for 6 cycles. RESULTS: The median survival was 15 months (95 % CI 13.6-16.4) (mean 16 months). Forty-five (90 %) patients achieved a response: 20 (40 %) patients, a complete response and 25 (50 %), a partial response. Adverse reactions included grade 3 and 4 neutropenia in 12 and 2 % of the patients, respectively. Other side effects were of very low toxicity. CONCLUSION: The 1-day, three-agent (cisplatin-etoposide-paclitaxel) treatment of small-cell lung cancer is beneficial with respect to response rate and survival, and the toxicity is low and well-tolerated.

Two consecutive days of treatment with liposomal cisplatin in non-small cell lung cancer.

Liposomal cisplatin (Lipoplatin) is a new agent, a cisplatin formulation that has been investigated in a number of studies and compared with cisplatin with respect to toxicity and effectiveness. It has been administered once weekly and in combination with a second agent, once every two weeks. The main outcome of the studies was that lipoplatin has no renal toxicity and is as equally effective as cisplatin. The present study investigated toxicity and effectiveness when lipoplatin is administered on two consecutive days, repeated every two weeks. Between January 2011 and November 2011, a total of 21 patients with histologically- or cytologically-confirmed non-small cell lung cancer (NSCLC) were enrolled in the study. All but two patients, who had not been pretreated, had received one or two series of chemotherapy and some had undergone radiotherapy. Lipoplatin monotherapy was infused for 8 h the first and second days and repeated every 2 weeks with the aim of administering 6 cycles. The dose per day was 200 mg/m(2). Eight out of 21 (38.10%) patients had a partial response, 9 (42.86%) had stable disease and 4 (19.05%) had progressive disease. Results showed that there was no renal failure toxicity and no other adverse reactions apart from grade 1 myelotoxicity in only 2 patients who had been heavily pretreated, and grade 1 nausea/vomiting in 4 patients. Liposomal cisplatin is an agent with negligible toxicity and reasonably high effectiveness even when administered to pretreated patients with NSCLC.

Maintenance chemotherapy or not in ovarian cancer stages IIIA, B, C, and IV after disease recurrence.

PURPOSE: Ovarian cancer may have a high percentage of residual disease after chemotherapy. It is questionable whether second or more lines of chemotherapy are needed in patients with slow-growing residual disease. In the present trial we compared the median survival of patients with residual or recurrent disease who received 1-2 lines of chemotherapy with those who received 3-9 lines. METHODS: Two hundred and five patients with advanced stage IIIA, B, C and IV ovarian cancer were divided into two groups based on the number of chemotherapy lines they received. All patients had prior first-line chemotherapy; the criteria for recruitment in the study were: a) residual or recurrent disease and b) failure to respond to first-line therapy. Group A included patients who received 1 or 2 lines of chemotherapy and group B, 3-9 lines. RESULTS: The median survival of group A was 76 months and of group B 53 months (p<0.001). Complete response (CR) was observed in 80 out of the 193 7lpar;41.45%) evaluable patients, partial response (PR) in 37 (19.17%), stable disease (SD) in 54 (27.987percnt;) and progressive disease (PD) in 22 (11.40%) patients. CONCLUSION: In ovarian cancer patients with advanced disease, multiple chemotherapy lines (3=9) offer no advantage over 1 or 2 lines, with respect to overall survival.

Lipoplatin formulation review article.

Patented platform technologies have been used for the liposomal encapsulation of cisplatin (Lipoplatin) into tumor-targeted 110 nm (in diameter) nanoparticles. The molecular mechanisms, preclinical and clinical data concerning lipoplatin, are reviewed here. Lipoplatin has been successfully administered in three randomized Phase II and III clinical trials. The clinical data mainly include non-small-cell lung cancer but also pancreatic, breast, and head and neck cancers. It is anticipated that lipoplatin will replace cisplatin as well as increase its potential applications. For the first time, a platinum drug has shown superiority to cisplatin, at least in non-squamous non-small-cell lung cancer as reported in a Phase III study which documented a simultaneous lowering of all of the side effects of cisplatin.

Epithelial nuclear factor-κB signaling promotes lung carcinogenesis via recruitment of regulatory T lymphocytes.

The mechanisms by which chronic inflammatory lung diseases, particularly chronic obstructive pulmonary disease, confer enhanced risk for lung cancer are not well-defined. To investigate whether nuclear factor (NF)-κB, a key mediator of immune and inflammatory responses, provides an interface between persistent lung inflammation and carcinogenesis, we utilized tetracycline-inducible transgenic mice expressing constitutively active IκB kinase ß in airway epithelium (IKTA (IKKß trans-activated) mice). Intraperitoneal injection of ethyl carbamate (urethane), or 3-methylcholanthrene (MCA) and butylated hydroxytoluene (BHT) was used to induce lung tumorigenesis. Doxycycline-treated IKTA mice developed chronic airway inflammation and markedly increased numbers of lung tumors in response to urethane, even when transgene expression (and therefore epithelial NF-κB activation) was begun after exposure to carcinogen. Studies using a separate tumor initiator/promoter model (MCA+BHT) indicated that NF-κB functions as an independent tumor promoter. Enhanced tumor formation in IKTA mice was preceded by increased proliferation and reduced apoptosis of alveolar epithelium, resulting in increased formation of premalignant lesions. Investigation of inflammatory cells in lungs of IKTA mice revealed a substantial increase in macrophages and lymphocytes, including functional CD4+/CD25+/FoxP3+ regulatory T lymphocytes (Tregs). Importantly, Treg depletion using repetitive injections of anti-CD25 antibodies limited excessive tumor formation in IKTA mice. At 6 weeks following urethane injection, antibody-mediated Treg depletion in IKTA mice reduced the number of premalignant lesions in the lungs in association with an increase in CD8 lymphocytes. Thus, persistent NF-κB signaling in airway epithelium facilitates carcinogenesis by sculpting the immune/inflammatory environment in the lungs.

Metastatic lung disease treated with pemetrexed-docetaxel combination chemotherapy.

PURPOSE: According to the ASCO guidelines 4 cytotoxic agents are used in colorectal cancer. Numerous agents have not been used or tested and thus their effectiveness is not known. Herein we present the preliminary data of a trial whose objective was to investigate the effectiveness to 2 combined agents in patients with heavily pretreated adenocarcinomas. METHODS: Ten patients have been recruited up until now. Colorectal adenocarcinoma was the primary disease in 7 patients, gastric cancer in 2, and endometrial adenocarcinoma in one. All patients had undergone at least 2 lines of previous chemotherapy during 2-3 years before the trial. The lungs were the only or main site of metastases in all patients. Chemotherapy agents used were pemetrexed 500 mg/m(2) and docetaxel 75 mg/m(2), repeated every 3 weeks, with a plan to administer 6 cycles. RESULTS: The mean number of cycles were 3 (range 1-6). Total number of cycles 33. No serious toxicity was detected. One patient had complete response, 3 partial response, 5 stable disease, 1 disease progression. Median survival was 6 months (range 2-12+ months). CONCLUSION: The effectiveness of the pemetrexed-docetaxel combination in heavily pretreated adenocarcinoma patients justifies their use when no other alternative exists.

Granulocyte colony-stimulating factor expression as a prognostic biomarker in non-small cell lung cancer.

Hematologic paraneoplastic alternations may not be very common, but they have been observed in a small number of patients. Granulocytosis has been described in several malignancies and its common mechanism may be associated with granulocyte colony-stimulating factor (G-CSF) production by the tumor. High neutrophilia (150,000-240,000 white blood cell count) observed in two patients with non-small cell lung cancer led us to run the present trial. The purpose of this study was to compare the white blood cell counts and the G-CSF plasma levels of the patients and classify the results into groups, as well as to determine the survival rates of the patients in each of the groups. The histological specimens and plasma of two patients as well as the plasma of another 87 patients with several malignancies, were examined. The plasma G-CSF levels were determined using a quantitative sandwich immunoassay technique (Quantikine; RySD systems) according to the manufacturer's instructions and all samples were tested in triplicate. It was found that 12 patients had a white blood cell count increased beyond normal as well as a high G-CSF plasma level and the survival of these patients was shorter as compared to the rest of the patients. We assume that these findings may indicate that increased G-CSF levels may function as a biomarker of survival.

Comparison of liposomal cisplatin versus cisplatin in non-squamous cell non-small-cell lung cancer.

PURPOSE: Liposomal cisplatin was developed to reduce the systemic toxicity of cisplatin, particularly the nephrotoxicity, and it has been used in combination with other agents in pancreatic and head and neck cancers and non-small-cell lung cancer (NSCLC). Our objective was to compare the effectiveness of lipoplatin combined with paclitaxel versus cisplatin with paclitaxel in advanced non-squamous NSCLC. METHODS: During 2007-2010, 202 patients with non-squamous NSCLC (stage IIIB and IV) were recruited from the two participating institutions and divided into two arms: Arm A was treated with liposomal cisplatin 200 mg/m(2) combined with paclitaxel 135 mg/m(2) and Arm B with cisplatin 75 mg/m(2) in combination with paclitaxel 135 mg/m(2), repeated every 2 weeks. The number of cycles administered was 632 (Arm A) and 640 (Arm B), totaling 1,272. RESULTS: A partial response was achieved by 59.22% of Arm A patients versus 42.42% of Arm B, and the difference was statistically significant (P 0.036). The median survival time in months was 10 for Arm A and 8 for Arm B (P 0.1551). After 18 months, the number of surviving patients was double for Arm A versus Arm B. CONCLUSION: Liposomal cisplatin in combination with paclitaxel produces a statistically significantly higher response rate than cisplatin combined with paclitaxel in non-squamous NSCLC.

Survival of untreated advanced colorectal cancer patients.

Colorectal cancer has specific biological characteristics that distinguish it from other malignancies. One such characteristic is its slow growth in patients in advanced stages. For the past 15 years, no effective systemic treatment has been available in clinical practice. The present study involved a retrospective evaluation of patients with advanced colorectal cancer in order to assess the median and overall survival of patients. Concurrently, the study aimed to describe the biological characteristics of this slow-growing disease and the quality of life of the patients. The key characteristic of this patient group was the lack of any systemic treatment. The study included 40 patients (25 male and 15 female, median age 67 years) who were evaluated between 1993 and 1996. Only supportive treatment was provided. One patient underwent 2 cycles of chemotherapy. Liver surgery was unsuccessfully performed on 3 patients. Two patients underwent radiofrequency once and 2 had intra-arterial treatment, also once. The results showed the median survival of patients to be 24 months (range 16-42). One-year survival was found to be 65% while the 2-year survival was found to be 25%. A satisfactory quality of life was also observed. In conclusion, colorectal cancer is a slow-going malignancy, as indicated by the long-term survival of patients and the biological characteristics of the tumor.

Treatment of colorectal cancer with and without bevacizumab: a phase III study.

OBJECTIVE: The objective of this phase III trial was to compare chemotherapy combined with bevacizumab versus chemotherapy alone in the treatment of patients with advanced colorectal cancer. METHODS: From September 2004 till September 2008, 222 treatment-naive patients were enrolled and divided into 2 arms: 114 arm A patients were treated with leucovorin, 5-fluorouracil plus irinotecan in combination with bevacizumab, and 108 arm B patients were treated as above without bevacizumab. All patients were stage IV with histologically confirmed adenocarcinoma. RESULTS: The median overall survival of arm A patients was 22.0 months (95% CI: 18.1-25.9) and 25.0 months (CI: 18.1-31.9) for arm B patients. There was no statistically significant difference between the 2 arms (p = 0.1391). No statistically significant difference between the 2 arms regarding the response rate was observed: partial response, 42 patients (36.8%) and 38 patients (35.2%) for arms A and B, respectively. Hematologic toxicity did not differ in the comparison of the 2 arms. Nonhematologic toxicity in arm A involved hypertension in 23 (20.2%) of the patients and proteinuria in 7 (6.1%); 3 patients experienced hemorrhage and 1 patient intestinal perforation. None of these side effects was observed in arm B patients. CONCLUSION: No statistically significant difference in median overall survival in patients with advanced colorectal cancer treated with bevacizumab plus a combination therapy (arm A) and those treated with the combination only, without bevacizumab (arm B), was observed.

Liposomal cisplatin dose escalation for determining the maximum tolerated dose and dose-limiting toxicity: a phase I study.

BACKGROUND: The aim of the present trial was to determine the maximum tolerated dose (MTD) and dose-limiting toxicity (DLT) of liposomal cisplatin (lipoplatin) using nephrotoxicity, gastrointestinal toxicity and myelotoxicity as the main adverse reactions. PATIENTS AND METHODS: Lipoplatin, a liposomal formulation of cisplatin was first tested as monotherapy starting at a dose of 125 mg/m(2) and escalating up to 350 mg/m(2). Lipoplatin was then escalated in combination with paclitacel starting at a dose of 100 mg/m(2) escalating up to 250 mg/m(2) for the former and 100 mg/m(2) escalating up to 175 mg/m(2) for the latter. RESULTS AND CONCLUSION: The present trial determined the DLT for lipoplatin monotherapy at 350 mg/m(2) and the MTD at 300 mg/m(2); for lipoplatin-paclitaxel combination therapy, the DLT was 250 mg/m(2) for lipoplatin and 175 mg/m(2) for paclitaxel whereas the MTD was 200 mg/m(2) for lipoplatin and 175 mg/m(2) for paclitaxel.

Liposomal cisplatin combined with paclitaxel versus cisplatin and paclitaxel in non-small-cell lung cancer: a randomized phase III multicenter trial.

BACKGROUND: Liposomal cisplatin is a new formulation developed to reduce the systemic toxicity of cisplatin while simultaneously improving the targeting of the drug to the primary tumor and to metastases by increasing circulation time in the body fluids and tissues. The primary objectives were to determine nephrotoxicity, gastrointestinal side-effects, peripheral neuropathy and hematological toxicity and secondary objectives were to determine the response rate, time to tumor progression (TTP) and survival. PATIENTS AND METHODS: Two hundred and thirty-six chemotherapy-naive patients with inoperable non-small-cell lung cancer were randomly allocated to receive either 200 mg/m² of liposomal cisplatin and 135 mg/m² paclitaxel (arm A) or 75 mg/m² cisplatin and 135 mg/m² paclitaxel (arm B), once every 2 weeks on an outpatient basis. Two hundred and twenty-nine patients were assessable for toxicity, response rate and survival. Nine treatment cycles were planned. RESULTS: Arm A patients showed statistically significant lower nephrotoxicity, grade 3 and 4 leucopenia, grade 2 and 3 neuropathy, nausea, vomiting and fatigue. There was no significant difference in median and overall survival and TTP between the two arms; median survival was 9 and 10 months in arms A and B, respectively, and TTP was 6.5 and 6 months in arms A and B, respectively. CONCLUSIONS: Liposomal cisplatin in combination with paclitaxel has been shown to be much less toxic than the original cisplatin combined with paclitaxel. Nephrotoxicity in particular was negligible after liposomal cisplatin administration. TTP and survival were similar in both treatment arms.