ABSTRACT
The L-methioninase-annexin V/selenomethionine enzyme prodrug system, designed to target the tumor vasculature and release the methylselenol anticancer drug in the tumor, was tested in mice with implanted MBA-MB-231 breast tumors. This therapy was able to cause a reduction in the size of the tumors during the treatment period. It was shown that L-methioninase-annexin V was uniformly bound at the blood vessel surface in the tumor and also that there was a substantial cutoff of blood flowing through the treated tumor, consistent with the therapy's design. This new approach for enzyme prodrug therapy of breast cancer appears promising.
Subject(s)
Annexin A5/metabolism , Antineoplastic Agents/therapeutic use , Carbon-Sulfur Lyases/metabolism , Mammary Neoplasms, Animal/drug therapy , Methanol/analogs & derivatives , Organoselenium Compounds/therapeutic use , Selenomethionine/metabolism , Animals , Breast Neoplasms/blood supply , Breast Neoplasms/drug therapy , Cell Line, Tumor , Enzyme Therapy , Female , Humans , Mammary Neoplasms, Animal/blood supply , Methanol/therapeutic use , Mice , Mice, SCID , Neoplasm Transplantation , Prodrugs/metabolism , Prodrugs/therapeutic useABSTRACT
This paper focuses on the targeting of single-walled carbon nanotubes (SWNTs) for the treatment of breast cancer with minimal side effects using photothermal therapy. The human protein annexin V (AV) binds specifically to anionic phospholipids expressed externally on the surface of tumour cells and endothelial cells that line the tumour vasculature. A 2 h incubation of the SWNT-AV conjugate with proliferating endothelial cells followed by washing and near-infrared (NIR) irradiation at a wavelength of 980 nm was enough to induce significant cell death; there was no significant cell death with irradiation or the conjugate alone. Administration of the same conjugate i.v. in BALB/c female mice with implanted 4T1 murine mammary at a dose of 0.8 mg SWNT kg(-1) and followed one day later by NIR irradiation of the tumour at a wavelength of 980 nm led to complete disappearance of implanted 4T1 mouse mammary tumours for the majority of the animals by 11 days since the irradiation. The combination of the photothermal therapy with the immunoadjuvant cyclophosphamide resulted in increased survival. The in vivo results suggest the SWNT-AV/NIR treatment is a promising approach to treat breast cancer.
Subject(s)
Hyperthermia, Induced , Mammary Neoplasms, Animal/therapy , Nanotubes, Carbon/chemistry , Phototherapy , Animals , Annexin A5/isolation & purification , Annexin A5/metabolism , Biotinylation , Cell Line, Tumor , Cell Proliferation , Cell Survival , Endothelial Cells/cytology , Endothelial Cells/metabolism , Female , Fluorescein-5-isothiocyanate/metabolism , Humans , Lung Neoplasms/pathology , Lung Neoplasms/secondary , Maleimides/chemistry , Mammary Neoplasms, Animal/pathology , Mice , Mice, Inbred BALB C , Microscopy, Atomic Force , Microscopy, Fluorescence , Phosphatidylethanolamines/chemistry , Polyethylene Glycols/chemistry , Recombinant Proteins/isolation & purification , Spectroscopy, Near-Infrared , Staining and Labeling , SuspensionsABSTRACT
A new approach for targeting carbon nanotubes to the tumor vasculature was tested using human endothelial cells and MCF-7 breast cancer cells in vitro. Single-walled carbon nanotubes were functionalized with the F3 peptide using a polyethylene glycol linker to target nucleolin, a protein found on the surface of endothelial cells in the vasculature of solid tumors. Confocal microscopy and Raman analysis confirmed that the conjugate was internalized by actively dividing endothelial cells. Dividing endothelial cells were used to mimic these cells in the tumor vasculature. Incubation with the conjugate for 8 h or more caused significant cell death in both actively dividing endothelial cells and MCF-7 breast cancer cells, an effect that is hypothesized to be due to the massive uptake of the conjugate. This targeted cell killing was further enhanced when coupled with near-infrared laser treatment. For confluent (non-dividing) endothelial cells, no cytotoxic effect was seen for incubation alone or incubation coupled with laser treatment. These results are promising and warrant further studies using this conjugate for cancer treatment in vivo.
Subject(s)
Endothelial Cells/metabolism , Infrared Rays , Nanotubes, Carbon/chemistry , Neoplasms/drug therapy , Phototherapy/instrumentation , Analysis of Variance , Cell Line, Tumor , Cell Survival/drug effects , Drug Carriers , Humans , Microscopy, Confocal , Peptides/chemistry , Peptides/metabolism , Peptides/pharmacokinetics , Peptides/pharmacology , Phosphoproteins/metabolism , Photochemotherapy/methods , Polyethylene Glycols , Protein Binding , RNA-Binding Proteins/metabolism , Spectrum Analysis, Raman , NucleolinABSTRACT
A fusion protein, consisting of cytosine deaminase (CD) linked to human annexin V, was created for use in an enzyme prodrug therapy targeted to the tumor vasculature and associated cancer cells in the primary tumor and distant metastases. The major finding of this study is that the CD-annexin V fusion protein in combination with the prodrug 5-fluorocytosine has significant cytotoxic activity against endothelial cells and two breast cancer cells lines in vitro that expose phosphatidylserine on their surface. The cytotoxicity experiments verified this novel enzyme prodrug system has the ability to produce therapeutic levels of 5-fluorouracil and thus appears promising.
Subject(s)
Annexin A5/genetics , Breast Neoplasms/therapy , Cytosine Deaminase/genetics , Endothelium, Vascular/physiology , Flucytosine/pharmacology , Genetic Therapy , Prodrugs/pharmacology , Antimetabolites/pharmacology , Aorta/cytology , Breast Neoplasms/enzymology , Breast Neoplasms/genetics , Cell Survival/drug effects , Cell Survival/physiology , Cells, Cultured , Combined Modality Therapy , Cytosine Deaminase/metabolism , Endothelium, Vascular/cytology , Female , Fluorouracil/metabolism , Humans , Phosphatidylserines/metabolism , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolismABSTRACT
A new approach for enzyme prodrug therapy for cancer was tested using human endothelial cells and two breast cancer cell lines in vitro. The concept is to use the human annexin V protein to selectively target the enzyme L-methioninase to the tumor vasculature. The major finding was that enzyme prodrug treatment using the L-methioninase-annexin V fusion protein and selenomethionine as the prodrug over 3 days was shown to be lethal to the endothelial cells and the cancer cells, while having little or no effect with the prodrug but with no fusion protein present. Thus, this new approach appears promising.