So far, 29 clinical trials were registered around the NIH website (clinicaltrials.gov). and FaDu head and neck carcinomas (p 0.05) and amplified the radioresponse of FaDu xenografts in a dose-dependent manner with enhancement factors ranging from 1.2 to 1 1.8 (p 0.01). Immunohistochemical analysis of FaDu xenografts exhibited that A12 inhibited tumor PF-543 Citrate cell proliferation (P 0.05) and VEGF expression. When A12 was combined PF-543 Citrate with radiation, this resulted in apoptosis induction that persisted till 6 days from the start of treatment and in increased necrosis at day 1 (p 0.01, respectively). Combined treatment with A12 and radiation resulted in additive or sub-additive growth delay in H460 or A549 xenografts, respectively. Conclusions The results of this study strengthen the evidence for investigating how anti-IGF-1R strategies can be integrated into radiation and radiation-cetuximab regimen in the treatment of cancer of the upper aero-digestive tract cancers. in non-small cell lung cancer cells included radiation-induced activation of the IGF-1R as a cell-protective stress response because its impairment enhanced lung cancer cell radiosensitivity (24). Several preclinical studies were conducted using monoclonal anti-IGF-1R antibody A12 (Imclone Systems Incorporated, NY) (20, 28). A12 exhibited activity against a wide range of human tumor types as well as in xenograft and orthotopic tumor models. The effects of A12 PF-543 Citrate were initially evaluated in a series of studies involving human MCF7 breast, BxPC-3 pancreas and Colo205 colon carcinomas (28). In these tumors, A12 exhibited significant inhibition of growth based on antiproliferative and proapoptotic effects (28). A12 also exhibited PF-543 Citrate potency to enhance the effects of cytotoxic brokers. In myeloma models, A12 enhanced the effects of melphalan or bortezomib thereby prolonging survival (21). In an androgen-independent prostate cancer model, the combination of A12 and docetaxel resulted in greater anticancer activity than docetaxel alone (22). A recent study by Allen and colleagues showed that A12 enhances the effect of radiation in different lung cancer cell Rabbit polyclonal to EGFL6 lines (23). In H460 lung cancer xenografts, the combination of 1.5 Gy given once a week with twice-weekly A12 (1 mg per mouse) for a total period of 4 weeks was shown to significantly inhibit tumor growth (23). Based on preclinical results an extensive clinical research program has been initiated testing A12 (cixutumumab) alone or in combination with other agents in various cancers, including non-small cell lung cancer (NSCLC) and head and neck carcinoma (HNC). So far, 29 clinical trials were registered around the NIH website (clinicaltrials.gov). However, none of these trials tested A12 in combination with radiation. The present study was undertaken to first investigate whether A12 potentiates the response of a human HNC and NSCLC models to radiation and to quantify the magnitude of enhancement achievable and its dependence on IGF-1R expression level. MATERIALS AND METHODS Cell culture Human HNC cell line HN-5 (kindly provided by Dr. Zhen Fan, University of Texas M. D. Anderson Cancer Center, Houston, TX) and NSCLC cell lines H460 and A549 (ATCC; Manassas, Va) were maintained in DMEM/F-12 medium supplemented with 10% fetal calf serum and 10,000 U/ml of penicillin-streptomycin. A human HNC, FaDu (ATCC; Manassas, Va.) was maintained in MEM medium supplemented with 10% fetal calf serum, 10,000 U/ml penicillin-streptomycin and 1% non-essential amino acids. A12 monoclonal antibody PF-543 Citrate The fully human IgG1 monoclonal antibody IMC-A12 (A12) was provided by ImClone Systems Incorporated (NY). This antibody was designed to selectively bind to the IGF-1R and was developed by screening a human Fab phage display library to specifically yield a high-affinity monoclonal antibody (4.11 10?11 mol/L; IC50, 0.6-1 nmol/L). A12 readily cross-reacts with the mouse IGF-1R (20). Clonogenic cell survival determination Between 50 and 400 cells were plated in 6 cm dishes in triplicate. The next day, the cells were exposed to A12 (100 nM), and 5 h later they were irradiated with graded doses (2 or 4 Gy) of -rays using a 137Cs source (3.7 Gy/min). The cells were left in the incubator with A12 in the medium. The medium was changed 67h after radiation (total 72 h of A12 treatment) and the cells were incubated in drug free medium. Cells were stained after 14 days with 0.5% crystal violet in absolute ethanol, and colonies with more than 50 cells were counted under a dissection microscope. For treatment with IGF1 with or without A12,.