ABSTRACT
PURPOSE: In the initial PALOMA-2 (NCT01740427) analysis with median follow-up of 23 months, palbociclib plus letrozole significantly prolonged progression-free survival (PFS) in women with estrogen receptor-positive (ER+)/human epidermal growth factor receptor 2-negative (HER2-) advanced breast cancer (ABC) [hazard ratio (HR) 0.58; P < 0.001]. Herein, we report results overall and by subgroups with extended follow-up. METHODS: In this double-blind, phase 3 study, post-menopausal women with ER+/HER2- ABC who had not received prior systemic therapy for their advanced disease were randomized 2:1 to palbociclib-letrozole or placebo-letrozole. Endpoints include investigator-assessed PFS (primary), safety, and patient-reported outcomes (PROs). RESULTS: After a median follow-up of approximately 38 months, median PFS was 27.6 months for palbociclib-letrozole (n = 444) and 14.5 months for placebo-letrozole (n = 222) (HR 0.563; 1-sided P < 0.0001). All subgroups benefited from palbociclib treatment. The improvement of PFS with palbociclib-letrozole was maintained in the next 2 subsequent lines of therapy and delayed the use of chemotherapy (40.4 vs. 29.9 months for palbociclib-letrozole vs. placebo-letrozole). Safety data were consistent with the known profile. Patients' quality of life was maintained. CONCLUSIONS: With approximately 15 months of additional follow-up, palbociclib plus letrozole continued to demonstrate improved PFS compared with placebo plus letrozole in the overall population and across all patient subgroups, while the safety profile remained favorable and quality of life was maintained. These data confirm that palbociclib-letrozole should be considered the standard of care for first-line therapy in patients with ER+/HER2- ABC, including those with low disease burden or long disease-free interval. Sponsored by Pfizer; ClinicalTrials.gov: NCT01740427.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Breast Neoplasms/drug therapy , Letrozole/administration & dosage , Piperazines/administration & dosage , Pyridines/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Breast Neoplasms/metabolism , Breast Neoplasms/psychology , Double-Blind Method , Female , Humans , Letrozole/adverse effects , Piperazines/adverse effects , Postmenopause/psychology , Pyridines/adverse effects , Quality of Life/psychology , Receptor, ErbB-2/metabolism , Receptors, Estrogen/metabolism , Treatment OutcomeABSTRACT
Background: Patient-reported outcomes are integral in benefit-risk assessments of new treatment regimens. The PALOMA-2 study provides the largest body of evidence for patient-reported health-related quality of life (QOL) for patients with metastatic breast cancer (MBC) receiving first-line endocrine-based therapy (palbociclib plus letrozole and letrozole alone). Patients and methods: Treatment-naïve postmenopausal women with estrogen receptor-positive (ER+)/human epidermal growth factor receptor 2-negative (HER2-) MBC were randomized 2 : 1 to palbociclib plus letrozole (n = 444) or placebo plus letrozole (n = 222). Patient-reported outcomes were assessed at baseline, day 1 of cycles 2 and 3, and day 1 of every other cycle from cycle 5 using the Functional Assessment of Cancer Therapy (FACT)-Breast and EuroQOL 5 dimensions (EQ-5D) questionnaires. Results: As of 26 February 2016, the median duration of follow-up was 23 months. Baseline scores were comparable between the two treatment arms. No significant between-arm differences were observed in change from baseline in FACT-Breast Total, FACT-General Total, or EQ-5D scores. Significantly greater improvement in pain scores was observed in the palbociclib plus letrozole arm (-0.256 versus -0.098; P = 0.0183). In both arms, deterioration of FACT-Breast Total score was significantly delayed in patients without progression versus those with progression and patients with partial or complete response versus those without. No significant difference was observed in FACT-Breast and EQ-5D index scores in patients with and without neutropenia. Conclusions: Overall, women with MBC receiving first-line endocrine therapy have a good QOL. The addition of palbociclib to letrozole maintains health-related QOL and improves pain scores in treatment-naïve postmenopausal patients with ER+/HER2- MBC compared with letrozole alone. Significantly greater delay in deterioration of health-related QOL was observed in patients without progression versus those who progressed and in patients with an objective response versus non-responders. ClinicalTrials.gov: NCT01740427 (https://clinicaltrials.gov/ct2/show/NCT01740427).
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Breast Neoplasms/drug therapy , Letrozole/administration & dosage , Piperazines/administration & dosage , Pyridines/administration & dosage , Quality of Life , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Breast Neoplasms/metabolism , Breast Neoplasms/physiopathology , Double-Blind Method , Female , Humans , Middle Aged , Placebos , PostmenopauseABSTRACT
Background: This report assesses the efficacy and safety of palbociclib plus endocrine therapy (ET) in women with hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer (ABC) with or without visceral metastases. Patients and methods: Pre- and postmenopausal women with disease progression following prior ET (PALOMA-3; N = 521) and postmenopausal women untreated for ABC (PALOMA-2; N = 666) were randomized 2 : 1 to ET (fulvestrant or letrozole, respectively) plus palbociclib or placebo. Progression-free survival (PFS), safety, and patient-reported quality of life (QoL) were evaluated by prior treatment and visceral involvement. Results: Visceral metastases incidence was higher in patients with prior resistance to ET (58.3%, PALOMA-3) than in patients naive to ET in the ABC setting (48.6%, PALOMA-2). In patients with prior resistance to ET and visceral metastases, median PFS (mPFS) was 9.2 months with palbociclib plus fulvestrant versus 3.4 months with placebo plus fulvestrant [hazard ratio (HR), 0.47; 95% confidence interval (CI), 0.35-0.61], and objective response rate (ORR) was 28.0% versus 6.7%, respectively. In patients with nonvisceral metastases, mPFS was 16.6 versus 7.3 months, HR 0.53; 95% CI 0.36-0.77. In patients with visceral disease and naive to ET in the advanced disease setting, mPFS was 19.3 months with palbociclib plus letrozole versus 12.9 months with placebo plus letrozole (HR 0.63; 95% CI 0.47-0.85); ORR was 55.1% versus 40.0%; in patients with nonvisceral disease, mPFS was not reached with palbociclib plus letrozole versus 16.8 months with placebo plus letrozole (HR 0.50; 95% CI 0.36-0.70). In patients with prior resistance to ET with visceral metastases, palbociclib plus fulvestrant significantly delayed deterioration of QoL versus placebo plus fulvestrant, whereas patient-reported QoL was maintained with palbociclib plus letrozole in patients naive to endocrine-based therapy for ABC. Conclusions: Palbociclib plus ET prolonged mPFS in patients with visceral metastases, increased ORRs, and in patients previously treated for ABC, delayed QoL deterioration, presenting a standard treatment option among patients with visceral metastases amenable to endocrine-based therapy. Clinical trial registration: NCT01942135, NCT01740427.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Breast Neoplasms/drug therapy , Neoplasm Metastasis/drug therapy , Piperazines/administration & dosage , Pyridines/administration & dosage , Adult , Aged , Aged, 80 and over , Antineoplastic Agents, Hormonal/therapeutic use , Breast Neoplasms/mortality , Breast Neoplasms/pathology , Female , Fulvestrant/administration & dosage , Humans , Letrozole/administration & dosage , Middle Aged , Progression-Free Survival , Quality of Life , VisceraABSTRACT
BACKGROUND: In a phase III trial in patients with advanced, well-differentiated, progressive pancreatic neuroendocrine tumors, sunitinib 37.5 mg/day improved investigator-assessed progression-free survival (PFS) versus placebo (11.4 versus 5.5 months; HR, 0.42; P < 0.001). Here, we present PFS using retrospective blinded independent central review (BICR) and final median overall survival (OS), including an assessment highlighting the impact of patient crossover from placebo to sunitinib. PATIENTS AND METHODS: In this randomized, double-blind, placebo-controlled study, cross-sectional imaging from patients was evaluated retrospectively by blinded third-party radiologists using a two-reader, two-time-point lock, followed by a sequential locked-read, batch-mode paradigm. OS was summarized using the Kaplan-Meier method and Cox proportional hazards model. Crossover-adjusted OS effect was derived using rank-preserving structural failure time (RPSFT) analyses. RESULTS: Of 171 randomized patients (sunitinib, n = 86; placebo, n = 85), 160 (94%) had complete scan sets/time points. By BICR, median (95% confidence interval [CI]) PFS was 12.6 (11.1-20.6) months for sunitinib and 5.8 (3.8-7.2) months for placebo (HR, 0.32; 95% CI 0.18-0.55; P = 0.000015). Five years after study closure, median (95% CI) OS was 38.6 (25.6-56.4) months for sunitinib and 29.1 (16.4-36.8) months for placebo (HR, 0.73; 95% CI 0.50-1.06; P = 0.094), with 69% of placebo patients having crossed over to sunitinib. RPSFT analysis confirmed an OS benefit for sunitinib. CONCLUSIONS: BICR confirmed the doubling of PFS with sunitinib compared with placebo. Although the observed median OS improved by nearly 10 months, the effect estimate did not reach statistical significance, potentially due to crossover from placebo to sunitinib. TRIAL REGISTRATION NUMBER: NCT00428597.
Subject(s)
Indoles/administration & dosage , Neuroendocrine Tumors/drug therapy , Pancreatic Neoplasms/drug therapy , Pyrroles/administration & dosage , Antineoplastic Agents/administration & dosage , Cross-Sectional Studies , Disease-Free Survival , Double-Blind Method , Humans , Kaplan-Meier Estimate , Neuroendocrine Tumors/diagnostic imaging , Pancreatic Neoplasms/diagnostic imaging , Proportional Hazards Models , Sunitinib , Survival RateABSTRACT
BACKGROUND: The X-box binding protein 1 (XBP1) is a critical transcription factor in the endoplasmic reticulum stress response, which is essential for the maintenance of cellular homeostasis. Here, we investigated whether the regulatory variant rs2269577 of the XBP1 gene influences clinical outcome in advanced non-small cell lung cancer (NSCLC) patients undergoing platinum-based chemotherapy. PATIENTS AND METHODS: We recruited 663 Chinese patients with advanced NSCLC treated with platinum-based regimens and assessed the association between rs2269577 and clinical outcome. Subsequent functional analyses, including real-time quantitative PCR and dual-luciferase assays, were performed to explore possible molecular mechanisms. RESULTS: The G/G genotype of rs2269577 was significantly associated with severe gastrointestinal toxicity compared with the homozygous C/C genotype (P=0.012, odds ratio=2.755), particularly in the female, performance status 0-1, and adenocarcinoma subgroups. No significant relevance was found between rs2269577 and treatment efficacy. In gastric epithelial cells, in vitro molecular analyses demonstrated that XBP1 mRNA expression levels decreased after treatment with cisplatin and the G allele of rs2269577 weakened the transcriptional activity of the XBP1 promoter. CONCLUSION: This is the first study to evaluate the effect of XBP1 polymorphism on severe chemotherapy-related adverse outcomes in platinum-treated advanced NSCLC patients using both pharmacogenomics and functional molecular analyses.
Subject(s)
Carcinoma, Non-Small-Cell Lung/drug therapy , DNA-Binding Proteins/genetics , Gastrointestinal Tract/pathology , Lung Neoplasms/drug therapy , Platinum/administration & dosage , Transcription Factors/genetics , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Carcinoma, Non-Small-Cell Lung/pathology , China , Drug-Related Side Effects and Adverse Reactions , Female , Gastrointestinal Tract/drug effects , Genotype , Humans , Lung Neoplasms/pathology , Male , Middle Aged , Neoplasm Staging , Polymorphism, Genetic , Promoter Regions, Genetic , Regulatory Factor X Transcription Factors , X-Box Binding Protein 1ABSTRACT
Valosin-containing protein (VCP), or p97, is a member of the ATP-binding protein family, and is involved in numerous cellular events, such as, protein degradation, membrane fusion, and chaperone activity. VCP has been demonstrated playing a critical role in non-small-cell lung cancer (NSCLC) pathogenesis and progression recently. We investigated the association between VCP polymorphisms and clinical outcome in advanced NSCLC patients undergoing platinum-based chemotherapy. We recruited 663 Chinese advanced NSCLC patients who were treated with platinum-based regimens, and using their clinical data, we assessed the efficacy and side effects of their treatment. Three tag-single nucleotide polymorphisms (SNPs) of VCP were genotyped. SNP rs2074549 showed a significant association with severe neutropenia. Its G/G genotype increased the risk of grade 3 or 4 neutropenia compared with wild-type homozygotes A/A (P = .001, odds ratio = 2.975). Haplotype association analysis revealed that CGA was associated with the increased incidence of severe neutropenia (P = .041, odds ratio = 1.439). However, no significant relationship was found between the presence of VCP polymorphisms and treatment efficacy when objective response, progression-free survival, and overall survival (OS) were evaluated. Our study is the first to provide evidence that VCP polymorphisms are associated with a severe chemotherapy-related adverse outcome in platinum-treated advanced NSCLC patients.
Subject(s)
Adenosine Triphosphatases/genetics , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Carcinoma, Non-Small-Cell Lung/mortality , Cell Cycle Proteins/genetics , Lung Neoplasms/mortality , Organoplatinum Compounds/adverse effects , Polymorphism, Single Nucleotide/genetics , Adenocarcinoma/drug therapy , Adenocarcinoma/genetics , Adenocarcinoma/mortality , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/mortality , DNA/blood , DNA/genetics , Female , Follow-Up Studies , Haplotypes/genetics , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lymphatic Metastasis , Male , Middle Aged , Neoplasm Staging , Neutropenia/chemically induced , Polymerase Chain Reaction , Prognosis , Survival Rate , Valosin Containing ProteinABSTRACT
AIMS/HYPOTHESIS: Hepatic glucokinase (GCK) is a key enzyme in glucose utilisation. Downregulation of its activity is associated with insulin resistance and type 2 diabetes mellitus. However, it is unknown whether hepatic Gck expression is influenced by age and is involved in ageing-mediated diabetes, and whether the degree of methylation of the hepatic Gck promoter is correlated with the transcription of Gck. To address the question, we evaluated hepatic Gck transcription and promoter methylation in young (14 weeks), adult (40 weeks) and aged (80 weeks) rats. METHODS: Hepatic glycogen, Gck expression and the kinase activity of GCK were measured in three age groups. The CpG methylation status was determined by both bisulphite direct sequencing and clone sequencing of the PCR amplificates of Gck promoter. The causal relationship between Gck methylation and mRNA expression was confirmed by treating rat primary hepatocytes with 5-aza-2'-deoxycytidine (5-Aza-CdR). RESULTS: We have shown an age-associated decline in hepatic glycogen, Gck expression levels and the kinase activity of hepatic GCK. The eleven CpG sites studied displayed age-related progressive methylation changes in hepatic Gck promoter, which were confirmed by two methods: direct and clone sequencing. After 5-Aza-CdR treatment of rat primary hepatocytes, there was a fourfold increase in Gck expression. CONCLUSIONS/INTERPRETATION: Our results demonstrate that an age-related increase in methylation is negatively associated with hepatic Gck expression, suggesting that DNA methylation could be involved in increasing age-dependent susceptibility to hepatic insulin resistance and diabetes. Thus, the epigenetic modification of the hepatic Gck promoter may represent an important marker for diabetogenic potential during the ageing process.
Subject(s)
DNA Methylation , Diabetes Mellitus/genetics , Glucokinase/genetics , Insulin Resistance/genetics , Liver/enzymology , Promoter Regions, Genetic , Aging , Animals , Base Sequence , DNA Primers , Dinucleoside Phosphates , Genetic Predisposition to Disease , Liver/growth & development , Liver Glycogen/metabolism , Male , Polymerase Chain Reaction , RNA, Messenger/genetics , Rats , Rats, WistarABSTRACT
Quantum dots (QDs), as novel fluorescence probes, have shown a great potential for bio-molecular labeling and cellular imaging. To stain cellular targets, the sufficient intracellular delivery of QDs is required. In this work the tat, a typical membrane-permeable carrier peptide, was conjugated with thiol-capped CdTe QDs to form CdTe Tat-QDs, and the intracellular deliveries of CdTe QDs or CdTe Tat-QDs were compared in human hepatocellular carcinoma (QGY) cells and human breast cancer (MCF7) cells in vitro by means of confocal laser scanning microscopy. Added into the cell dishes, both QDs and Tat-QDs adhered to the outer leaflet of the plasma membrane of cells within a few minutes, but the binding amount of Tat-QDs was obviously higher than that of QDs. Then both QDs and Tat-QDs can penetrate into cells, and their cellular contents increased with incubation time but both saturated after 3 hours incubation. However the cellular levels of Tat-QDs were higher than those of QDs, with the ratio of 2.1 (+/-0.3) times in QGY cells and 1.5 (+/-0.2) times in MCF7 cells, demonstrating the enhancing effect of Tat conjugation on the intracellular delivery of QDs.
Subject(s)
Cadmium/metabolism , Fluorescent Dyes/metabolism , Quantum Dots , Tellurium/metabolism , Cadmium/chemistry , Cadmium Compounds/chemistry , Fluorescent Dyes/chemistry , Gene Products, tat/chemistry , Humans , Tellurium/chemistry , Tumor Cells, CulturedABSTRACT
A new cholesterol-carborane conjugate (BCH) has been synthesized as a potential targeting agent for boron neutron capture therapy (BNCT) of cancers. The compound is extremely water insoluble and was formulated in two liposomal formulations to determine if the compound could be adequately taken up by 9L rat glioma cells in cell culture. Several factors potentially affecting the cellular uptake were evaluated, such as concentration of BCH in the incubation medium, incubation time, cell confluence, and the addition of polyethylene glycol (PEG) phospholipids to the liposomal formulation. The studies indicated that the cellular uptakes of BCH in the conventional and PEG liposomal formulations were 49.1 and 45.9 microg boron/g cells, respectively. Therefore, this compound, formulated in both liposomal formulations, delivered sufficient levels of boron to cancer cells in vitro, indicating that BCH is a promising approach for use in BNCT. The uptake appeared to depend upon BCH concentration in the media as well as the confluence of the cells. The greater boron uptake by nonconfluent cells indicated that active growth of cells was a factor in the uptake of this compound.
Subject(s)
Boron/pharmacokinetics , Cholesterol Esters/pharmacokinetics , Polyethylene Glycols/pharmacokinetics , Animals , Cell Culture Techniques/methods , Cholesterol Esters/chemistry , Liposomes , Polyethylene Glycols/chemistry , Rats , Tumor Cells, CulturedABSTRACT
In order to produce large amounts of protein for vaccine trials, a synthetic msp1-42 gene was inserted into Pichia pastoris expression vector and the plasmid was introduced into Pichia pastoris SMD1168 by electroporation. The expressed MSP1-42 was secreted into the protein-free medium. To measure the conformational properties of MSP1-42,16 monoclonal antibodies (11 recognizing conformational epitopes) were allowed to interact with the Pichia-derived MSP1-42, and all antibodies specific for conserved and K1 protype interacted with the protein. Interestingly, three monoclonal antibodies (e.g. 9.8, 13.1 and 7.3), that were shown not to interact with CHO-derived MSP1, could interact with the Pichia-derived MSP1-42. Rabbits were immunized with recombinant MSP1-42 formulated with CFA adjuvant four times. The rabbits were bled on the day 3 after last immunization, and total IgG isolated by protein A column from the immunized rabbits was shown to strongly inhibit the parasite growth in vitro dose-dependently, whereas IgG from rabbit with adjuvant had no inhibition.
Subject(s)
Antibodies, Monoclonal/pharmacology , Antiprotozoal Agents/pharmacology , Merozoite Surface Protein 1/immunology , Plasmodium falciparum/drug effects , Animals , Antibody Specificity , Parasitic Sensitivity Tests , Peptide Fragments/immunology , Pichia/genetics , Plasmodium falciparum/growth & development , Plasmodium falciparum/immunology , Rabbits , Recombinant Proteins/immunologyABSTRACT
The aim of this study was to investigate therapeutic efficacy of adenovirus-mediated E1a gene therapy for ovarian cancer in vitro and in vivo. Recombinant replication-deficient adenoviral vectors were prepared by superinfection of 293 cells, and then purified. The efficacy of the adenovirus vector system to infect ovarian cells was tested using different multiplicity of infection (MOI) and different times (1-4) of Ad.RSVlacZ. SKOV-3 cells (10(3) per well) were infected once with 2 x 10(4) adenovirus. The cells were harvested and counted on different days for 7 days to generate the in vitro growth curve. Tumor-bearing mice were injected intraperitoneally with ovarian cancer cells and treated by intraperitoneal injection of 100 microl (2.5 x 10(8) PFU) viral solution containing either replication-deficient Ad.E1a(+); control virus Ad.E1a(-) which is the same adenovirus as Ad.E1a(+) except for E1a deletion, or just phosphate buffered solution. The transduction efficacy increased with higher MOI and reached a plateau at the 20:1 ratio. When Ad.E1a(+) was used to transduce the HER-2/neu overexpressing human ovarian cancer cell line SKOV-3, tumor cell growth in vitro was greatly inhibited by E1a transduction. Also, Ad.E1a+ greatly inhibited tumor growth of SKOV-3-bearing mice. Immunohistochemistry analysis indicated that Ad.E1a protein was expressed in tumor tissue and expression of HER-2/neu p185 protein was suppressed. Very strong beta-gal staining was detected in tumors, and beta-gal activity in small intestine, lung, heart, stomach, liver, and kidney was detected. No beta-gal activity was detected in the tumor and other organs in control mice injected with Ad.E1a(-) or PBS. Adenovirus-type 5 E1a gene can efficaciously inhibit HER-2/neu-overexpressing ovarian cancer, and this promising procedure could greatly benefit ovarian cancer patients with high expression of HER-2/neu.
Subject(s)
Adenovirus E1A Proteins/genetics , Carcinoma/genetics , Carcinoma/therapy , Genetic Therapy , Ovarian Neoplasms/genetics , Ovarian Neoplasms/therapy , Adenoviridae/pathogenicity , Animals , Down-Regulation , Female , Genetic Vectors , Humans , Immunohistochemistry , Injections, Intraperitoneal , Mice , Mice, Inbred BALB C , Neoplasms, Experimental , Peptide Fragments/biosynthesis , Random Allocation , Receptor, ErbB-2/biosynthesis , Transduction, Genetic , Tumor Cells, CulturedABSTRACT
The efficacy of a liposomal formulation for intracerebral delivery of borocaptate (BSH) to brain tumor cells has been investigated using cell culture to study BSH uptake and persistence and using tumor-bearing rats to determine BSH distribution in the brain. During a 16-hr incubation, cellular uptake of BSH solution or BSH liposomal formulation was similar. However, the cellular persistence of BSH greatly increased when BSH was present in liposome. The differences in cellular persistence for BSH solution and BSH-loaded liposomes were significant both in 12-hr and 24-hr incubation experiments (p < 0.05 and p < 0.01, respectively). For the studies involving tumor-bearing rats, BSH level in tumor tissue was significantly higher than that in normal brain tissue at 2 hr and 6 hr after intracerebral injection of BSH-loaded liposomes (p < 0.01). Our study indicated that the liposomal formulation enhanced cellular persistence of BSH in tumor cells and therefore favored the boron accumulation in the cells. With the prolonged physical retention of liposomes at the local injection site and the cellular retention of BSH enhanced by the liposomes, the intracerebral delivery of BSH using liposomal formulation may provide an effective boron delivery approach for boron neutron capture therapy of brain tumors.
Subject(s)
Borohydrides/pharmacokinetics , Boron Neutron Capture Therapy , Brain Neoplasms/metabolism , Drug Delivery Systems , Glioma/metabolism , Sulfhydryl Compounds/pharmacokinetics , Animals , Borohydrides/pharmacology , Brain/metabolism , Brain Neoplasms/drug therapy , Glioma/drug therapy , Injections , Injections, Intraventricular , Liposomes , Male , Rats , Rats, Inbred F344 , Sulfhydryl Compounds/pharmacology , Tumor Cells, Cultured/drug effectsABSTRACT
Sequence of MSP1-31 of Plasmodium falciparum was constructed into eukaryotic expression vector pTRE, which could be repressed by tetracycline (Tc) and resulted in recombinant plasmid pTRE-31. The plasmid was injected into the quadriceps muscle of BALB/c mice with Tc responsive plasmid pTet-off to measure specific antibodies. The MSP1-31 prokaryotic expressed protein was used as antigen in ELISA. Results showed that mice orally administered by Tc had a seroconversion rate of 7.1% (1/14) 4 weeks after injection, whereas the control mice had a seroconversion rate of 100% and the titers of antibody were raised continusly within 12 weeks. The study suggested that the recombinant plasmids pTRE-31/pTet-off could efficiently induce humoral response against MSP1-31 of malaria. Moreover this immune response was controlled by Tc and was reversible after withdrawal of Tc dilivery. The induction of antibody by removing Tc at the fourth week after injection indicated that DNA vaccine could remain in mice and capable of expressing antigen for at least 4 weeks.
Subject(s)
Antigens, Protozoan/genetics , Malaria Vaccines/immunology , Promoter Regions, Genetic , Tetracycline/pharmacology , Vaccines, DNA/immunology , Animals , Antibodies, Protozoan/blood , Female , Gene Expression Regulation , Immunization , Mice , Mice, Inbred BALB CABSTRACT
The blood brain barrier (BBB) and the systemic toxicity of conventional chemotherapy present obstacles to the success of future blood-borne drug therapies of brain tumors. The work with polymer-encapsulated cancer drugs suggests an alternative and more focused treatment approach. Our experimental strategy integrates direct intracerebral drug delivery, sustained drug release from liposomes or polymer implants, and increased targeting of the drug either by chemically modifying the drug or by using tumor-specific carriers. This review will present some of the recent work on targeted drug delivery for brain cancer treatment.
Subject(s)
Antineoplastic Agents/therapeutic use , Brain Neoplasms/drug therapy , Drug Delivery Systems , Animals , Antineoplastic Agents/administration & dosage , Blood-Brain Barrier , Delayed-Action Preparations , Drug Implants , HumansABSTRACT
To explore whether human umbilical cord blood CD34+ cells transduced with human aldehyde dehydrogenase class-1 (ALDH1) and multidrug resistance gene (MDR1) increase resistance to 4-Hyaroxycyclophosphophamide (4-HC) and P-Glycoprotein Effluxed Drugs, a bicistronic Retroviral vector G1Na-ALDH1-IRES-MDR1 was constructed. The vector was transduced into the packaging cell lines GP + E86 and PA317 by LipofectAMINE. Using the medium containing VCR and 4-HC for cloning selection and pingponging supernatant infection between ecotropic producer clone and amphotropic producer clone, we obtained high titer amphotropic PA317 producer clone with the highest titer up to 5.6 x 10(5) CFU/ml. Cord blood CD34+ cells were transfectced repeatedly with supernatant of retrovirus containing human ALDH1 and MDR1cDNA under stimulation of hemopoietie growth factors. PCR, RT-PCR, Southern blot, Northern blot, FACS and MTT method analyses show that dual drug resistance genes have been integrated into the genomic DNA of cord blood CD34+ cells and expressed efficiently. The transgenes recipient cells confered 4- to 7.2-folds stronger resistance to cyclophospsphamede and P-Glycoprotein Effluxes drug in comparison with the nontransduced cells. This study provided a foundation for the application of combination chemotherapy in tumor clinical trial.
Subject(s)
Aldehyde Dehydrogenase/genetics , Antigens, CD34/analysis , Fetal Blood/cytology , Genes, MDR , Hematopoietic Stem Cells/metabolism , Isoenzymes/genetics , Retroviridae/genetics , ATP Binding Cassette Transporter, Subfamily B, Member 1/biosynthesis , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Aldehyde Dehydrogenase 1 Family , Cells, Cultured , Gene Expression , Gene Transfer Techniques , Genetic Therapy , Genetic Vectors , Humans , Retinal DehydrogenaseABSTRACT
The purpose of this study is to prepare and characterize injectable carboplatin-loaded poly(D,L,-lactic-co-glycolic) acid copolymer (PLGA) microspheres for the intracerebral treatment of malignant glioma. The microspheres were prepared by an acetone/mineral oil emulsion and solvent evaporation method. Preparation variables were optimized and the following processing conditions resulted in the highest drug loading and best yields of the microspheres compared with those prepared with the other variables: the PLGA concentration was 8% (w/w) in the internal phase; the emulsifier (Span 80) concentration was 8% (w/w) in the external phase; the ratio of the internal phase: the external phase was 1:8; the stirring speed was 1500 rpm; the emulsion time was 15 min; the solvent evaporation time was 3.75 hr. Microspheres so prepared were analysed for size distribution, drug loading, in vitro release and morphological characteristics. The drug release in phosphate buffer solution started with a 10-day slow release period, followed by a fast near zero order release period from 12 to 22 days. The carboplatin release in brain homogenate was slower than in phosphate buffer solution. The morphological changes of the microspheres during the in vitro degradation correlated with the drug relase profile. In conclusion, the carboplatin-loaded PLGA microspheres were specifically prepared to meet the specification as an injectable and biodegradable brain implant.
Subject(s)
Antineoplastic Agents/chemistry , Biocompatible Materials/chemistry , Carboplatin/chemistry , Lactic Acid/chemistry , Polyglycolic Acid/chemistry , Polymers/chemistry , Animals , Antineoplastic Agents/administration & dosage , Biocompatible Materials/administration & dosage , Brain/metabolism , Carboplatin/administration & dosage , Delayed-Action Preparations , Emulsions , Injections, Intraventricular , Lactic Acid/administration & dosage , Microscopy, Electron, Scanning , Microspheres , Molecular Weight , Polyglycolic Acid/administration & dosage , Polylactic Acid-Polyglycolic Acid Copolymer , Polymers/administration & dosage , Rats , Solubility , VolatilizationABSTRACT
The purpose of this study was to assess the effect of gastrointestinal proteins on the in vitro release of zidovudine (AZT) from ethylcellulose microspheres, and to investigate protein adsorption as a possible mechanism that mediates this effect. AZT release from ethylcellulose microspheres was tested in the presence of different gastrointestinal proteins, both dietary (casein and albumin) and endogenous (pepsin, pancreatin, and mucin) in simulated gastric fluid and/or simulated intestinal fluid. The resulting release profiles were compared with those produced in the corresponding release media without the presence of proteins. Protein adsorption on AZT-loaded ethylcellulose microspheres was studied for the five proteins under investigation. The amounts of adsorbed proteins were determined by fluorescent spectrometry after the protein solution was reacted with fluoraldehyde reagent. All of the investigated proteins were found to slow the release of AZT from ethylcellulose microspheres. At gastric pH, ovalbumin and casein had the maximum effect on AZT release. Mucin exerted a more pronounced effect at gastric pH compared with that at intestinal pH. The negative effect of pancreatin on AZT release increased when its concentration was increased. The five proteins were found to adsorb on AZT-loaded ethylcellulose microspheres with varying quantities. The observed protein adsorption is believed to cause blockage of the small pores and channels in the microsphere structure, and consequently slow the release of AZT.
Subject(s)
Anti-HIV Agents/administration & dosage , Cellulose/analogs & derivatives , Zidovudine/administration & dosage , Adsorption , Cellulose/administration & dosage , Hydrogen-Ion Concentration , Microscopy, Electron, Scanning , Microspheres , Mucins/pharmacology , Pancreatin/pharmacology , Pepsin A/pharmacology , Zidovudine/chemistryABSTRACT
There has been an increasing interest in intracerebral delivery of anticancer agents using biodegradable polymers for the treatment of malignant glioma. This approach circumvents the blood-brain barrier (BBB) to achieve a high local drug concentration in the brain tumor sites and minimize side effects associated with a high systemic dose. It could also control local tumor recurrence and improve survival. In order to deliver anticancer drugs intracerebrally from polymers to tumor sites with minimal surgery, injectable poly(d,l-lactic-coglycolic acid) (PLGA) microspheres impregnated with carboplatin have been prepared. In the current studies, the brain tissue reaction to blank or carboplatin-loaded PLGA microspheres was investigated in rats. The PLGA micro-spheres were well tolerated by all of the rats. The brain tissue reaction to the blank microspheres was accompanied by mild edema, and macrophage/astrocyte/microglia proliferation. The tissue reaction to the carboplatin microspheres was characterized as edema, necrosis, and a more pronounced phagocytic inflammatory reaction. The observed inflammatory reactions decreased remarkably after 1 month. Carboplatin microspheres were then implanted intracerebrally in the rat glioma models. Higher drug concentrations were achieved in the brain tumor than in normal tissues. The survival and weight loss of the rats receiving carboplatin microspheres were compared with those of the rats receiving systemic doses. The local treatment was more effective in controlling the weight loss of the tumor-bearing rats, and was as effective as the systemic treatment in prolonging survival.
ABSTRACT
To study the role of intron in the expression of hFIX, retroviral vectors with intron containing hFIX were constructed. It is fundamental for the intron study whether the intron constructed in retroviral vector can be steadily transferred into target cell. First, we constructed two forward-orientation retroviral vectors: G1NaC-i-IX contains the exogenous intron from IL-2, and G1NaC-i'-IX contains the truncated intron I from hFIX gene, covering the splicing donor and acceptor sequences. RT-PCR result indicated that intron in the forward-orientation retroviral vector was spliced after packaging in PA317. Then, reverse-orientation retroviral vectors G1NaC-i'-IXR and G1NaPAIXi' BAM were constructed, in which the reverse and complimentary sequences of hFIX gene with intron appeared in retroviral RNA. RT-PCR assay combined with ELISA test indicated that intron was retained after packaging and hFIX gene with intron constructed in the reverse-orientation retroviral vector can be transduced intact and expressed hFIX at a high level in vitro.
Subject(s)
Factor IX/genetics , Genetic Vectors , Introns , RNA Splicing , Retroviridae/genetics , Humans , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
The purpose of this study was to evaluate the in vivo performance of sustained-release zidovudine (AZT) microspheres after oral administration in Beagle dogs, and to establish an in vitro-in vivo correlation. Two AZT microsphere formulations as well as AZT powder were administered to four Beagle dogs. Plasma samples were analyzed by HPLC. The plasma concentration-time data was analyzed by both compartmental and noncompartmental pharmacokinetic analyses. Based on the calculated pharmacokinetic parameters, in vivo release profiles were simulated and compared with in vitro release profiles in three different release media. Significantly longer mean residence time (MRT) was observed after administration of the sustained-release microspheres compared with AZT powder. Significantly lower maximum (Cmax) concentration values and longer times to Cmax (tmax) values were also observed. Formulation I showed the longest MRT (4.4 h). AZT plasma concentration was maintained above the minimum effective concentration for approximately 10 h after administration of Formulation I. The relative bioavailability of the microsphere formulations with respect to AZT powder was not significantly different from 1. The in vitro release of the three formulations was slower in simulated gastric fluid compared with simulated intestinal fluid. The addition of enzymes and mucin to the release media significantly lowered the in vitro release rate of AZT from the microspheres formulations, but not from AZT powder. A good level of in vitro-in vivo correlation (Level A correlation) was achieved with a release medium that was composed of simulated gastric fluid with pepsin and mucin for 2 h followed by simulated intestinal fluid with pancreatin and mucin for 8 h. This in vitro model may be used to predict the in vivo release of AZT, in the further development of controlled-release AZT formulations.