Analysis of the interaction of monoclonal antibodies with surface IgM on neoplastic B-cells

In vitro studies identified three Burkitts lymphoma cell lines, Ramos, MUTU-I and Daudi, that were growth inhibited by anti-IgM antibody. However, only Ramos and MUTU-I were sensitive to monoclonal antibodies (mAb) recognizing the Fc region of surface IgM (anti-Fcμ). Experiments using anti-Fcμ mAb (single or non-crossblocking pairs), polyclonal anti-μ Ab, and hyper-crosslinking with a secondary layer of Ab, showed that growth inhibition of B-cell lines was highly dependent on the extent of IgM crosslinking. This was confirmed by using Fab′, F(ab′)2and F(ab′)3derivatives from anti-Fcμ mAb, where increasing valency caused corresponding increases in growth arrest and apoptosis, presumably as a result of more efficient BCR-crosslinking on the cell surface. The ability of a single mAb to induce growth arrest was highly dependent on epitope specificity, with mAb specific for the Fc region (Cμ2–Cμ4 domains) being much more effective than those recognizing the Fab region (anti-L chain, anti-Id and anti-Fdμ, or Cμ1). Only when hyper-crosslinked with polyclonal anti-mouse IgG did the latter result in appreciable growth inhibition. Binding studies showed that these differences in function were not related to differences in the affinity, but probably related to intrinsic crosslinking capacity of mAb. © 1999 Cancer Research Campaig

Molecular biology of breast cancer metastasis: Clinical implications of experimental studies on metastatic inefficiency

Recent technological advances have led to an increasing ability to detect isolated tumour cells and groups of tumour cells in patients' blood, lymph nodes or bone marrow. However, the clinical significance of these cells is unclear. Should they be considered as evidence of metastasis, necessitating aggressive treatment, or are they in some cases unrelated to clinical outcome? Quantitative experimental studies on the basic biology of metastatic inefficiency are providing clues that may help in understanding the significance of these cells. This understanding will be of use in guiding clinical studies to assess the significance of isolated tumour cells and micrometastases in cancer patients

In silico assessment of biomedical products: the conundrum of rare but not so rare events in two case studies

In silico clinical trials, defined as “The use of individualized computer simulation in the development or regulatory evaluation of a medicinal product, medical device, or medical intervention,” have been proposed as a possible strategy to reduce the regulatory costs of innovation and the time to market for biomedical products. We review some of the the literature on this topic, focusing in particular on those applications where the current practice is recognized as inadequate, as for example, the detection of unexpected severe adverse events too rare to be detected in a clinical trial, but still likely enough to be of concern. We then describe with more details two case studies, two successful applications of in silico clinical trial approaches, one relative to the University of Virginia/Padova simulator that the Food and Drug Administration has accepted as possible replacement for animal testing in the preclinical assessment of artificial pancreas technologies, and the second, an investigation of the probability of cardiac lead fracture, where a Bayesian network was used to combine in vivo and in silico observations, suggesting a whole new strategy of in silico-augmented clinical trials, to be used to increase the numerosity where recruitment is impossible, or to explore patients’ phenotypes that are unlikely to appear in the trial cohort, but are still frequent enough to be of concern

Functional Characterization of Circulating Tumor Cells with a Prostate-Cancer-Specific Microfluidic Device

Cancer metastasis accounts for the majority of cancer-related deaths owing to poor response to anticancer therapies. Molecular understanding of metastasis-associated drug resistance remains elusive due to the scarcity of available tumor tissue. Isolation of circulating tumor cells (CTCs) from the peripheral blood of patients has emerged as a valid alternative source of tumor tissue that can be subjected to molecular characterization. However, issues with low purity and sensitivity have impeded adoption to clinical practice. Here we report a novel method to capture and molecularly characterize CTCs isolated from castrate-resistant prostate cancer patients (CRPC) receiving taxane chemotherapy. We have developed a geometrically enhanced differential immunocapture (GEDI) microfluidic device that combines an anti-prostate specific membrane antigen (PSMA) antibody with a 3D geometry that captures CTCs while minimizing nonspecific leukocyte adhesion. Enumeration of GEDI-captured CTCs (defined as intact, nucleated PSMA+/CD45− cells) revealed a median of 54 cells per ml identified in CRPC patients versus 3 in healthy donors. Direct comparison with the commercially available CellSearch® revealed a 2–400 fold higher sensitivity achieved with the GEDI device. Confocal microscopy of patient-derived GEDI-captured CTCs identified the TMPRSS2:ERG fusion protein, while sequencing identified specific androgen receptor point mutation (T868A) in blood samples spiked with only 50 PC C4-2 cells. On-chip treatment of patient-derived CTCs with docetaxel and paclitaxel allowed monitoring of drug-target engagement by means of microtubule bundling. CTCs isolated from docetaxel-resistant CRPC patients did not show any evidence of drug activity. These measurements constitute the first functional assays of drug-target engagement in living circulating tumor cells and therefore have the potential to enable longitudinal monitoring of target response and inform the development of new anticancer agents