Metastases from primary tumors are responsible for most cancer deaths. It has been shown that circulating tumor cells (CTCs) can be detected in the peripheral blood of patients with a variety of metastatic cancers and that the presence of these cells is associated with poor clinical outcomes. Characterization of CTCs in metastatic cancer patients could provide additional information to augment management of the disease. Here, we describe a novel approach for the identification of molecular markers to detect and characterize CTCs in peripheral blood. Using an integrated platform to immunomagnetically isolate and immunofluorescently detect CTCs, we obtained blood containing z100 CTCs from one metastatic colorectal, one metastatic prostate, and one metastatic breast cancer patient. Using the RNA extracted from the CTC-enriched portion of the sample and comparing it with the RNA extracted from the corresponding CTCdepleted portion, for the first time, global gene expression profiles from CTCs were generated and a list of cancerspecific, CTC-specific genes was obtained. Subsequently, samples immunomagnetically enriched for CTCs from 74 metastatic cancer patients and 50 normal donors were used to confirm by quantitative real-time reverse transcription-PCR CTC-specific expression of selected genes and to show that gene expression profiles for CTCs may be used to distinguish normal donors from advanced cancer patients as well as to differentiate among the three different metastatic cancers. Genes such as AGR2, S100A14, S100A16, FABP1, and others were found useful for detection of CTCs in peripheral blood of advanced cancer patients. (Cancer Res 2005; 65(12): 4993-7)

Malondialdehyde and a series of acrolein derivatives substituted in the beta-position with good leaving groups react with guanine and guanine nucleosides to form two different types of adducts. The reaction with guanosine is typical. One adduct exhibits ultraviolet absorbance maxima at 253, 319, and 348 nm and is fluorescent. Its NMR spectrum exhibits three new aromatic proton resonances derived from malondialdehyde. The mass spectrum exhibits an M + 1 at 320. The spectroscopic properties are consistent with the structure, 3-beta-D-erythro-pentofuranosyl-pyrimido[1,2-alpha]purin-10(3H)-one (PyP-ribose). The second guanosine adduct is an equal mixture of diastereomers that exhibit ultraviolet maxima at 217 and 244 nm and mirror image circular dichroism spectra. The NMR spectrum and mass spectrum (M + 1 = 392) indicate the addition of two molecules of MDA to one molecule of guanosine. Two-dimensional NMR (COSY) analysis reveals the presence of propano and enal functionalities. The spectroscopic and chemical properties suggest an oxadiazabicyclo[3.3.1]nonene structure that is confirmed by X-ray crystallography. Comparison of the deoxyguanosine adducts of malondialdehyde to those of the structurally related carbonyl compounds, methyl glyoxal and acrolein, provides a structural basis to explain the unique ability of malondialdehyde to induce frameshift mutations in bacterial mutagenesis systems.

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Background The prescence of circulating tumor cells (CTCs) in the peripheral blood of cancer patients and their frequency has been correlated with disease status. Methods In this study, CTCs were characterized by flow cytometry and fluorescence microscopy after immunomagnetic enrichment from 7.5‐ml blood samples collected from patients with prostate cancer in evacuated blood‐draw tubes that contained an anticoagulant and a preservative. Events were classified as tumor cell candidates if they expressed cytokeratin, lacked CD45, and stained with the nucleic acid dye 4,6‐diamidino‐2‐phenylindole. Results In the blood of prostate cancer patients, only few of these events were intact cells. Other CTC events appeared as damaged cells or cell fragments by microscopy. By flow cytometry, these events stained variably with 4,6‐diamidino‐2‐phenylindole and frequently expressed the apoptosis‐induced, caspase‐cleaved cytokeratin 18. Similar patterns of cell disintegration were observed when cells of the prostate line LNCaP were exposed to paclitaxel before spiking the cells into normal blood samples. Conclusions The different observed stages of tumor cell degradation or apoptosis varied greatly between patients and were not found in blood of normal donors. Enumeration of CTCs and identification of CTCs undergoing apoptosis may provide relevant information to evaluate the response to therapy in cancer patients. © 2004 Wiley‐Liss, Inc.

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