In vitro drug responses in primary and metastatic colorectal cancers.

Objective: Treatment of primary and metastatic colorectal carcinoma (CRC) based on 5-fluorouracil and folinic acid (5FU + FA), combined with irinotecan (FOLFIRI) or oxaliplatin (FOLFOX), provides response rates approaching 50% and a 20-month overall survival. Approximately 50% of CRC patients fail to respond to one or more drugs in either regimen, in many cases due to inherent or acquired drug resistance. We therefore characterized in vitro drug response and cross-resistance in primary and metastatic CRC lesions.

Detection of the MDR1 P-glycoprotein expression and function.

Acquired and intrinsic multidrug resistance is the major reason for the failure of anticancer chemotherapy. The most important component of clinical multidrug resistance is mediated by P-glycoprotein (Pgp), an ABC transporter encoded by the MDR1 gene and expressed on the membrane of tumor and normal cells. Sensitive and reproducible detection of Pgp expression and function are critical for the development of new MDR1 drugs and clinical protocols aimed at modulating Pgp-mediated multidrug resistance.

Development and characterization of mouse hybridomas.

Cell fusion protocols that were developed by Kohler and Milstein in the mid-1970s and aimed at producing and characterization of mouse monoclonal antibodies (MAbs) remain the gold standard of hybridoma development. Despite tremendous progress in using MAbs in multiple research, diagnostic, and therapeutic areas, major experimental flaws in designing and carrying out hybridoma experimentation often result in the production of hybridomas exhibiting poor growth parameters and secreting low-specificity and low-affinity antibodies. This methodology chapter is built around the conventional hybridoma protocol, with a special emphasis on tissue culture and biochemical techniques aimed at producing truly monospecific and highly active mouse MAbs.

Chemotherapy resistance and oncogene expression in non-small cell lung cancer.

Objectives: Empiric chemotherapy for patients with non-small cell lung cancer who have undergone resection is recommended without knowledge of the tumor's specific biologic characteristics, and many patients may not benefit. In vitro chemotherapy resistance is associated with clinical unresponsiveness in some tumors, and in lung cancer, chemotherapy resistance is prevalent. Multiple-agent chemotherapy resistance and association of chemotherapy resistance with molecular markers are described.

Improved immunohistochemical method for detecting hypoxia gradients in mouse tissues and tumors.

We describe an improved immunohistochemical procedure for detecting regions of hypoxia in normal organs and tumors in mice. The method employs a primary fluorescein-conjugated mouse monoclonal antibody directed against pimonidazole protein adducts that are created in hypoxic tissues and a secondary mouse anti-fluorescein antibody that is conjugated to horseradish peroxidase. Using these reagents, we clearly visualized the regions of relative hypoxia in implanted tumors in mice as well as in normal organs such as liver and kidney.

Correlation of dynamic contrast enhancement MRI parameters with microvessel density and VEGF for assessment of angiogenesis in breast cancer.

Purpose: To investigate the association between parameters obtained from dynamic contrast enhanced MRI (DCE-MRI) of breast cancer using different analysis approaches, as well as their correlation with angiogenesis biomarkers (vascular endothelial growth factor and vessel density).

Materials And Methods: DCE-MRI results were obtained from 105 patients with breast cancer (108 lesions). Three analysis methods were applied: 1) whole tumor analysis, 2) regional hot-spot analysis, and 3) intratumor pixel-by-pixel analysis.

Analysis of P-glycoprotein-mediated membrane transport in human peripheral blood lymphocytes using the UIC2 shift assay.

Background: During transport-associated adenosine triphosphate hydrolysis, P-glycoprotein (Pgp) undergoes conformation transitions detected by UIC2, a functional anti-Pgp monoclonal antibody. A newly developed UIC2 shift assay is based on increased UIC2 reactivity in the presence of Pgp substrates. All peripheral blood leukocytes express low Pgp levels.

Effects of ATP depletion and phosphate analogues on P-glycoprotein conformation in live cells.

P-glycoprotein (Pgp), a membrane pump often responsible for the multidrug resistance of cancer cells, undergoes conformational changes in the presence of substrates/modulators, or upon ATP depletion, reflected by its enhanced reactivity with the UIC2 monoclonal antibody. When the UIC2-shift was elicited by certain modulators (e.g.