elastin and Ovarian-Neoplasms

Chemotherapeutic drug testing of SCCOHT-1 and BIN-67 tumor cells revealed synergistic growth-inhibition of >95% in vitro with a combination of foretinib and FK228. Application of this drug combination in vivo in NODscid mice-induced SCCOHT-1GFP tumors was associated with ~6-fold reduction in tumor mass within 10 days, whereby synergistic effects of the two compounds remained undetectable compared to previous results with foretinib treatment alone. Histopathologic evaluation revealed a reduced vascularization and a lower amount of proliferating cells in the treated tumors. Surprisingly, a simultaneous significant accumulation of extracellular matrix structures with positive elastin-van Gieson staining was observed following foretinib/FK228 exposure. Expression analysis of treated animal tumors exhibited various changes including increased mouse transcript levels of elastin, laminin, and fibronectin. In parallel, markers for mesenchymal stroma/stem cells (MSC) including CD73 and CD90 were detectable in all mouse tumors suggesting a possible involvement of these cells in extracellular matrix restructure. Indeed, incubation of MSC with FK228 or foretinib/FK228 demonstrated morphologic alterations and enhanced expression of laminin and fibronectin. Moreover, a co-culture of MSC with lentiviral-labeled SCCOHT-1GFP cells contributed to protection of the tumor cells against FK228-mediated cytotoxicity. Furthermore, explant cultures of SCCOHT-1GFP-induced tumors acquired an increased resistance to FK228 and a combination of foretinib/FK228 in contrast to foretinib alone. Together, these data suggested that FK228-mediated extracellular matrix protein expression by MSC contributes to increased protection and enhanced resistance of SCCOHT tumors which could represent a more general mechanism of MSC during drug-induced alterations of a tumor microenvironment.

Topics: 5'-Nucleotidase; Anilides; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Depsipeptides; Drug Resistance, Neoplasm; Elastin; Extracellular Matrix; Female; Fibronectins; Humans; Laminin; Mesenchymal Stem Cells; Mice; Mice, Inbred NOD; Mice, SCID; Ovarian Neoplasms; Quinolines; Thy-1 Antigens; Tumor Microenvironment; Xenograft Model Antitumor Assays

Ovarian cancer has an extremely high mortality rate resulting from poor understanding of the disease. In order to aid understanding of disease etiology and progression, we identify the endogenous fluorophores present in a mouse model of ovarian cancer and describe changes in fluorophore abundance and distribution with age and disease.. A mouse model of ovarian cancer was created by dosing with 4-vinylcyclohexene diepoxide, which induces follicular apoptosis (simulating menopause), and 7,12-dimethylbenz[a]anthracene, a known carcinogen. Imaging of ovarian tissue was completed ex vivo with a multiphoton microscope using excitation wavelength of 780 nm and emission collection from 405 to 505 nm. Two-photon excited fluorescence images and corresponding histologic sections with selective stains were used to identify endogenous fluorophores.. The majority of collected fluorescence emission was attributed to NADH and lipofuscin, with additional contributions from collagen and elastin. Dim cellular fluorescence from NADH did not show observable changes with age. Changes in ovarian morphology with disease development frequently caused increased fluorescence contributions from collagen and adipose tissue-associated NADH. Lipofuscin fluorescence was much brighter than NADH fluorescence and increased as a function of both age and disease.. Our finding of NADH fluorescence patterns similar to that seen previously in human ovary, combined with the observation of lipofuscin accumulation with age and disease also seen in human organs, suggests that the findings from this model may be relevant to human ovarian disease. Increased lipofuscin fluorescence might be used as an indicator of disease in the ovary and this finding warrants further study.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Adenocarcinoma; Aging; Animals; Biomarkers, Tumor; Collagen; Cyclohexenes; Disease Progression; Elastin; Female; Image Interpretation, Computer-Assisted; Linear Models; Lipofuscin; Mice; Microscopy, Fluorescence, Multiphoton; NAD; Ovarian Neoplasms; Ovary; Vinyl Compounds

Current treatment of solid tumors is limited by normal tissue tolerance, resulting in a narrow therapeutic index. To increase drug specificity and efficacy and to reduce toxicity in normal tissues, we have developed a polypeptide carrier for a cell cycle inhibitory peptide, which has the potential to be thermally targeted to the tumor site. The design of this polypeptide is based on elastin-like polypeptide (ELP). The coding sequence of ELP was modified by the addition of the cell penetrating peptide Bac-7 at the N-terminus and a 23 amino acid peptide derived from p21 at the C-terminus (Bac-ELP1-p21). Bac-ELP1-p21 is soluble in aqueous solutions below physiological temperature (37 degrees C) but aggregates when the temperature is raised above 39 degrees C, making it a promising thermally responsive therapeutic carrier that may be actively targeted to solid tumors by application of focused hyperthermia. While Bac-ELP1-p21 at 37 degrees C did not have any effect on SKOV-3 cell proliferation, the use of hyperthermia increased the antiproliferative effect of Bac-ELP1-p21 compared with a thermally unresponsive control polypeptide. Bac-ELP1-p21 displayed both a cytoplasmic and nuclear distribution in the SKOV-3 cells, with nuclear-localized polypeptide enriched in the heated cells, as revealed by confocal microscopy. Using Western blotting, we show that Bac-ELP1-p21 caused a decrease in Rb phosphorylation levels in cells treated at 42 degrees C. The polypeptide also induced caspase activation, PARP cleavage, and cell cycle arrest in S-phase and G2/M-phase. These studies indicate that ELP is a promising macromolecular carrier for the delivery of cell cycle inhibitory peptides to solid tumors.

Topics: Amino Acid Sequence; Apoptosis; Blotting, Western; Caspases; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p21; Elastin; Female; Hot Temperature; Humans; Microscopy, Confocal; Molecular Sequence Data; Ovarian Neoplasms; Peptides; Phosphorylation; Poly(ADP-ribose) Polymerases; Recombinant Fusion Proteins; Retinoblastoma Protein; Transition Temperature

Tumor cell metastasis is a complex, multi-step process that is a major cause of death and morbidity amongst cancer patients. Cell adhesion plays a critical role in the development of metastatic cancer, and it is mediated by interactions between receptors on the cell surface and ligands of the extracellular matrix or other surfaces. Therefore, inhibition of the cell adhesion process appears to be an effective method of preventing metastasis. This work describes a genetically engineered polypeptide with the potential to prevent cell adhesion and inhibit metastasis. We have found that the cell penetrating peptide Tat, fused with elastin-like polypeptide (ELP) inhibited adhesion, spreading, invasion and migration of SKOV-3 ovarian cancer cells in cell culture. Furthermore, we have also confirmed that Tat-ELP has anti-metastatic potential in an experimental ovarian cancer metastasis model in vivo, causing approximately 80% reduction in the tumor burden. Since cell attachment is an important step in tumor cell invasion and metastasis, these results suggest a novel role of Tat-ELP as a therapeutic intervention in cancer metastasis.

Topics: Animals; Cell Adhesion; Cell Movement; Elastin; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Transplantation; Ovarian Neoplasms; Peptides; Recombinant Fusion Proteins; tat Gene Products, Human Immunodeficiency Virus; Transplantation, Heterologous

To determine whether elastin stains aid in classifying peritoneal implants associated with ovarian serous borderline tumours (SBT).. The study group comprised 80 implants (nine invasive and 71 non-invasive) from 28 patients with ovarian SBT. Elastin stains were performed using histochemical and immunohistochemical methods to demonstrate the peritoneal elastic lamina (PEL), and evaluated with regard to assessment of the subtype of implant. The elastin stains demonstrated the PEL in most anatomical sites other than the omentum and the bladder and were considered helpful in 44/80 (55%) cases. The stains were most useful in the assessment of poorly oriented or traumatized biopsy specimens and in confirming the superficial distribution of non-invasive implants. The staining was non-contributory in most of the remaining biopsies, because the PEL was not identified.. Demonstration of the PEL using elastin stains can be useful in the subclassification of implants associated with ovarian SBT and is of most value in confirming the superficial distribution of non-invasive lesions. However, evaluation is limited by the absence of a defined elastic layer in a proportion of biopsy specimens, possibly reflecting their superficial location, as well as absence of a distinct PEL in sites such as the omentum.

Topics: Adult; Aged; Aged, 80 and over; Cystadenocarcinoma, Serous; Elastin; Female; Humans; Immunohistochemistry; Middle Aged; Ovarian Neoplasms; Peritoneal Neoplasms; Peritoneum; Predictive Value of Tests

Elastin-like polypeptides (ELPs) are biopolymers of the pentapeptide repeat Val-Pro-Gly-Xaa-Gly that undergo an inverse temperature phase transition. They are soluble in aqueous solutions below their transition temperature (T1) but hydrophobically collapse and aggregate at temperatures greater than T1. We hypothesized that ELPs conjugated to drugs would enable thermally targeted drug delivery to solid tumors if their T1 were between body temperature and the temperature in a locally heated region. To test this hypothesis, we synthesized a thermally responsive ELP with a T1 of 41 degrees C and a thermally unresponsive control ELP in Escherichia coli using recombinant DNA techniques. In vivo fluorescence videomicroscopy and radiolabel distribution studies of ELP delivery to human tumors (SKOV-3 ovarian carcinoma and D-54MG glioma) implanted in nude mice demonstrated that hyperthermic targeting of the thermally responsive ELP for 1 h provides a approximately 2-fold increase in tumor localization compared to the same polypeptide without hyperthermia. We observed aggregates of the thermally responsive ELP by fluorescence videomicroscopy within the heated tumor microvasculature but not in control experiments, which demonstrates that the phase transition of the thermally responsive ELP carrier can be engineered to occur in vivo at a specified temperature. By exploiting the phase transition-induced aggregation of these polypeptides, this method provides a new way to thermally target polymer-drug conjugates to solid tumors.

Topics: Amino Acid Sequence; Animals; Base Sequence; Drug Stability; Elastin; Female; Glioma; Humans; Hyperthermia, Induced; Mice; Mice, Inbred BALB C; Mice, Nude; Microscopy, Fluorescence; Microscopy, Video; Molecular Sequence Data; Ovarian Neoplasms; Peptides; Protein Engineering; Temperature; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Elastin-like polypeptides (ELPs) composed of a VPGXG repeat undergo a reversible phase transition in aqueous solution. They are hydrophilic and soluble in aqueous solution below their transition temperature (T(t)), but they become hydrophobic and aggregate when the temperature is raised above their T(t). In this study, we examine the quantitative uptake of a fluorescence-labeled, thermally responsive ELP as a function of ELP concentration between 5 and 15 microM in solution in response to hyperthermia by three cultured cancer cell lines. Flow cytometry of fluorescein-ELP conjugates showed that hyperthermia enhanced the cellular uptake of the thermally responsive ELP in human ovarian carcinoma cells (SKOV-3) and in HeLa cells at a concentration of 10 microM or higher, but not at a concentration of 5 microM, as compared with the uptake of a thermally inactive ELP control. In FaDu cells, hyperthermia stimulated uptake of the thermally responsive ELP at all solution concentrations of ELP between 5 and 15 microM. In particular, a >2-fold greater uptake of thermally responsive ELP compared with the thermally inactive control ELP was observed for FaDu cells at a solution concentration of 15 microM in heated cells. Confocal fluorescence microscopy of tumor cells incubated with a rhodamine conjugate of the thermally responsive ELP showed that the cytoplasm was uniformly stained below the T(t). Above the T(t), fluorescent particles were observed in the cytoplasm, suggesting that these particles are aggregates of the thermally responsive polypeptide resulting from the ELP phase transition. These studies demonstrate that the endocytotic uptake of a thermally responsive ELP is significantly enhanced by the thermally triggered phase transition of the polypeptide.

Topics: Carcinoma, Squamous Cell; Drug Carriers; Elastin; Female; Flow Cytometry; Fluorescent Dyes; Hot Temperature; Humans; Hyperthermia, Induced; Kinetics; Microscopy, Confocal; Ovarian Neoplasms; Peptides; Subcellular Fractions; Tumor Cells, Cultured

This report describes the collagen, glycosaminoglycans and elastin--composition of ovarian tumours. Rearrangement of extracellular matrix in ovarian tumour has been found. There was an increase of total collagen content in comparison to control. Type I collagen was found to be predominant in both tissues. Both tissues, normal and neoplastic one, contain all known glycosaminoglycans. Among them dermatan sulphate was found to be the most abundant. In ovarian tumour an increase of this glycosaminoglycan content was observed.

Topics: Adult; Collagen; Elastin; Female; Glycosaminoglycans; Humans; Middle Aged; Ovarian Neoplasms

Extracellular matrix components of benign ovarian tumours (cystadenoma, adenofibroma, cystadenofibroma) were analysed. The investigated tumours contained twice as much collagen than control ovarian tissues. Significant alterations in mutual quantitative relationships between collagens of various types were observed. The proportion of type I collagen decreased and that of type III collagen increased. The accumulation of collagen was accompanied by a reduction in sulphated glycosaminoglycan content whereas the amount of hyaluronic acid was not changed. Dermatan sulphate was the most abundant glycosaminoglycan component. It is suggested that the accumulation of collagen (natural barrier to the migration of tumour cells) and underexpression of glycosaminoglycans/proteoglycans (binding some growth factors and interleukins) may exert an inhibitory effect on tumour growth.

Topics: Adenofibroma; Adult; Collagen; Cystadenoma; Elastin; Extracellular Matrix; Female; Glycosaminoglycans; Humans; Middle Aged; Ovarian Neoplasms; Ovary