Purpose: This study aimed to detect the M30 neoepitope on circulating tumor cells (CTC) as a tool for quantifying apoptotic CTC throughout disease course and treatment. Experimental Design: An automated sample preparation and analysis platform for computing CTC (CellSearch) was integrated with a monoclonal antibody (M30) targeting a neoepitope disclosed by caspase cleavage at cytokeratin 18 (CK18) in early apoptosis. The assay was validated using cell lines and blood samples from healthy volunteers and patients with epithelial cancer. Results: M30-positive CTC could be detected in >70% of CTC-positive carcinoma patients, which were free for both chemotherapy and radiologic treatments. The fraction of M30-positive CTC varied from 50% to 80%, depending on the histotype. To investigate the potential application of the M30 CTC assay for the evaluation of response in early phase trials, CTC and M30-positive CTC were enumerated in a small case series of breast cancer patients during treatment. Results indicate that changes in the balance of M30-negative/positive CTC may be used as a dynamic parameter indicating an active disease, as documented by consistent radiologic findings. Conclusions: M30 expression on CTC is detectable by immunofluorescence. The M30-integrated test has potential for monitoring dynamic changes in the quote of apoptotic CTC (in addition to CTC count) to evaluate response in clinical trials of molecularly targeted anticancer therapeutics as well as for translational research, in which there is a pressing need for informative circulating biomarkers. Clin Cancer Res; 16(21); 5233–43. ©2010 AACR.

B cell activation is a well known consequence of HIV-1 infection, and seropositive subjects show high numbers of spontaneously activated Ig-secreting cells in circulation. To better define the importance of the HIV-1-specific response in this phenomenon, we first studied whether in vitro spontaneous anti-HIV-1 antibody production was accompanied by reactivation of memory B lymphocytes. Unstimulated PBL from HIV-1-infected individuals with prior history of hepatitis B and/or EBV infection did not consistently show spontaneous in vitro synthesis of anti-hepatitis B core Ag or anti-EBV antibodies; in addition, PWM-induced synthesis of anti-hepatitis B virus and anti-EBV antibodies was decreased compared to HIV-1-seronegative subjects. Moreover, in comparing the frequencies of activated HIV-1-specific B cell precursors and activated Ig-secreting precursors in limiting dilution experiments, a sizable fraction (20 to 40%) of circulating cells spontaneously secreting Ig produced antibody against HIV-1 determinants. The ratio between the two frequencies fitted in very well with the amount of Ig removed from unstimulated culture supernatants after HIV-1-specific antibody absorption with solid-phase HIV-1. These findings indicate that B cell activation during HIV-1 infection is mainly oriented toward a specific response to HIV-1 determinants; the possible relevance of this phenomenon to lymphomagenesis in AIDS patients is discussed.

Tumor growth is currently viewed as a phenomenon associated with neovascularization and sustained production of angiogenic factors, but whether a transient angiogenic switch may trigger tumor growth remains unclear. Here, we report that leukemia cells (MOLT-3) were poorly angiogenic and remained dormant when injected s.c. into immunodeficient mice. However, progressive growth of lymphoid tumors was invariably recorded when irradiated angiogenic cells from Kaposi’s sarcoma (KS-IMM) were locally coinjected with MOLT-3 cells or administered later. The persistence of KS-IMM cells in vivo was tracked by flow cytometry and real-time PCR analysis, and it was limited to a few days, during which angiogenesis was induced and preceded tumor growth. The engraftment of other types of poorly tumorigenic cancer cells was also greatly improved by irradiated KS-IMM cells. Moreover, short-term treatment with angiogenic factors, including basic FGF or VEGF, either given as recombinant factors or delivered by retroviral vectors, also accelerated tumor growth. These findings may emphasize that tumor angiogenesis is a process requiring a higher amount of angiogenic factors for its induction than maintenance.