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Cetuximab Therapy in Head and Neck Cancer: Immune Modulation With Interleukin-12
ASSOCIATION FOR ACADEMIC SURGERY AND SOCIETY OF UNIVERSITY SURGEONS—ABSTRACTS
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in the presence of cetuximab-coated head and neck cancer cells leads to enhanced NK cell mediated ADCC and cytokine secretion independent of tumor cell HPV-status. Cytokine administration could be a useful adjuvant in the cetuximab treatment of HER1-pos. head and neck cancer.
41.3. The Plasma Fraction of Packed Red Blood Cells Increases Alternative Activation of Tumor Associated Macrophages. D. D. Benson,1,2 X. Meng,2 D. A. Fullerton,2 E. E. Moore,1,2 C. C. Silliman,2,3 C. C. Barnett1,2,3; 1Denver Health Medical Center, Denver, CO; 2University of Colorado Health Science Center, Aurora, CO; 3Bonfils Blood Center, Denver, CO
41.2. Cetuximab Therapy in Head and Neck Cancer: Immune Modulation With Interleukin-12. E. A. Luedke, N. Bhave, W. E. Carson; The Ohio State University, Columbus, OH Introduction: Squamous cell carcinoma of the head and neck (SCCHN) is the sixth most common cancer worldwide. Greater than 90% of SCCHN of the oropharynx overexpress the epidermal growth factor receptor (EGFR or HER1). Cetuximab (Erbitux) is a humanized anti-HER1 monoclonal antibody (mAb) that binds to HER1 overexpressing tumor cells. Cetuximab has a direct effect on HER1-pos. cancer cells, but it also can activate immune cells that bear receptors for the Fc (constant portion) of IgG such as natural killer (NK) cells. NK cells have an activating Fc receptor for IgG (FcgammaRIIIa), which mediates Ab dependent cellular cytotoxicity (ADCC) and enhances production of interferon-gamma (IFN-g) in response to Ab-coated targets. Interleukin-12 (IL-12) is a cytokine produced by antigen-presenting cells that stimulates IFN-g production from NK cells. We hypothesized that IL-12 would enhance the anti-tumor activity of cetuximab by activating the FcR effector mechanisms of NK cells. Methods: Expression of HER1 was measured on human papilloma virus (HPV)-pos. (UM-SCC2, UM-SCC47) and HPV-neg. (Cal27, UM-SCC74B) SCCHN cell lines by immunoblot analysis and flow cytometry. NK cells from normal donors were treated overnight with IL2 (100U), IL-12, IL-15 or IL-21 (all 10ng/ml) and tested for ADCC vs. cetuximab-coated cancer cells in a 4 hr 51Cr assay. Release of cytokines by NK cells in response to cetuximab-coated cells was measured by ELISA. Phosphorylation of the ERK transcription factor in NK cells was measured by flow cytometry. Results: All cell lines showed >99% expression of HER1 by flow cytometry and immunoblot analysis except UM-SCC74B (73%). Normal NK cells mediated 49.4% lysis of cetuximab-coated SCCHN cell lines as compared to 7.6% lysis of cells treated with control IgG (P¼0.0002). NK cell lysis of cetuximab-coated SCCHN cells was markedly enhanced by 12 hr treatment of NK cells with IL-12 (71.6% lysis, P¼0.005 vs. cetuximab alone). As a control, IL-12-activated NK cells were tested against IgG-treated cells. ADCC under these conditions was just 21.7%. Similar levels of lysis were noted for both HPV-pos. and HPV-neg. and cell lines. Other NK cell activating factors such as IL-2, IL-15 and IL-21 were also able to enhance NK cell ADCC. the stimulus of IL-12 and cetuximabcoated tumor cells induced the synergistic production of nanogram levels of IFN-g (>6-fold increase over controls) (P<0.001). A similar effect was seen for NK cell production of TNF-alpha, GMCSF and the chemokines RANTES and MIP-1alpha. Phosphorylation of ERK (which is critical for FcR-mediated ADCC and cytokine production) was enhanced in NK cells exposed to IL-12 and IgG as compared to control conditions. Conclusions: Cytokine stimulation of NK cells
Introduction: Perioperative blood transfusion has been linked to decreased survival in patients with pancreatic adenocarcinoma, although a causative mechanism has not been elucidated. Recent work, suggests that increased tumor associated macrophages (TAMs) also correlate with poor clinical outcomes wherein it is believe that tumors cells coapt macrophages to augment tumor growth by taking on an alternatively activated phenotype, which promotes angiogenesis and extracellular remodeling functions. Our lab has recently shown significant TAM expression of alternative activation markers within metastatic murine pancreatic adenocarcinoma. We hypothesize that transfusion of the plasma fraction of packed red blood cells increases TAM expression of alternative activation. Methods: C57/BL6 mice, age 8-9 weeks, underwent splenic inoculation with 2.5 x105 PanO2 murine pancreatic adenocarcinoma cells. at 7 days, mice were transfused via tail vein injection with the plasma fraction of packed red blood cells (pRBCs) at a doses of 1ml/kg (equivalent of 2 units), or saline control (NS). Necropsy was performed at day 35 or earlier if clinically indicated, and metastatic liver tumors collected. Tumors were homogenized and evaluated for the expression of the alternative activation markers CD206 and CD163 by immunoblotting. Mice not inoculated with tumor were used for control expression. Data are presented as fold-expression over control +/- standard error of the mean, N6 per group. Results: Evaluating hepatic metastases from tumor bearing mice receiving either NS or pRBCs, expression of both CD206 and CD163 were increased compared to control livers (p<0.001 for both). Comparing mice receiving NS compared to mice receiving pRBCs, transfused mice had an increased expression of CD206, 4.2560.35 fold for pRBCs vs. 2.9460.44 for NS (p¼0.044), and had increased expression of CD163, 7.7760.80 for pRBCs vs. 4.3060.53 for NS (p¼0.017). Conclusions: Clinically, the detrimental effects of perioperative blood have been recognized for some time. These data suggest blood transfusion affects tumor microenvironmental changes in TAMs that may promote tumor progression as alternative activation is known to correlate with angiogenic and remodeling functions of macrophages.
311
in the presence of cetuximab-coated head and neck cancer cells leads to enhanced NK cell mediated ADCC and cytokine secretion independent of tumor cell HPV-status. Cytokine administration could be a useful adjuvant in the cetuximab treatment of HER1-pos. head and neck cancer.
41.3. The Plasma Fraction of Packed Red Blood Cells Increases Alternative Activation of Tumor Associated Macrophages. D. D. Benson,1,2 X. Meng,2 D. A. Fullerton,2 E. E. Moore,1,2 C. C. Silliman,2,3 C. C. Barnett1,2,3; 1Denver Health Medical Center, Denver, CO; 2University of Colorado Health Science Center, Aurora, CO; 3Bonfils Blood Center, Denver, CO
41.2. Cetuximab Therapy in Head and Neck Cancer: Immune Modulation With Interleukin-12. E. A. Luedke, N. Bhave, W. E. Carson; The Ohio State University, Columbus, OH Introduction: Squamous cell carcinoma of the head and neck (SCCHN) is the sixth most common cancer worldwide. Greater than 90% of SCCHN of the oropharynx overexpress the epidermal growth factor receptor (EGFR or HER1). Cetuximab (Erbitux) is a humanized anti-HER1 monoclonal antibody (mAb) that binds to HER1 overexpressing tumor cells. Cetuximab has a direct effect on HER1-pos. cancer cells, but it also can activate immune cells that bear receptors for the Fc (constant portion) of IgG such as natural killer (NK) cells. NK cells have an activating Fc receptor for IgG (FcgammaRIIIa), which mediates Ab dependent cellular cytotoxicity (ADCC) and enhances production of interferon-gamma (IFN-g) in response to Ab-coated targets. Interleukin-12 (IL-12) is a cytokine produced by antigen-presenting cells that stimulates IFN-g production from NK cells. We hypothesized that IL-12 would enhance the anti-tumor activity of cetuximab by activating the FcR effector mechanisms of NK cells. Methods: Expression of HER1 was measured on human papilloma virus (HPV)-pos. (UM-SCC2, UM-SCC47) and HPV-neg. (Cal27, UM-SCC74B) SCCHN cell lines by immunoblot analysis and flow cytometry. NK cells from normal donors were treated overnight with IL2 (100U), IL-12, IL-15 or IL-21 (all 10ng/ml) and tested for ADCC vs. cetuximab-coated cancer cells in a 4 hr 51Cr assay. Release of cytokines by NK cells in response to cetuximab-coated cells was measured by ELISA. Phosphorylation of the ERK transcription factor in NK cells was measured by flow cytometry. Results: All cell lines showed >99% expression of HER1 by flow cytometry and immunoblot analysis except UM-SCC74B (73%). Normal NK cells mediated 49.4% lysis of cetuximab-coated SCCHN cell lines as compared to 7.6% lysis of cells treated with control IgG (P¼0.0002). NK cell lysis of cetuximab-coated SCCHN cells was markedly enhanced by 12 hr treatment of NK cells with IL-12 (71.6% lysis, P¼0.005 vs. cetuximab alone). As a control, IL-12-activated NK cells were tested against IgG-treated cells. ADCC under these conditions was just 21.7%. Similar levels of lysis were noted for both HPV-pos. and HPV-neg. and cell lines. Other NK cell activating factors such as IL-2, IL-15 and IL-21 were also able to enhance NK cell ADCC. the stimulus of IL-12 and cetuximabcoated tumor cells induced the synergistic production of nanogram levels of IFN-g (>6-fold increase over controls) (P<0.001). A similar effect was seen for NK cell production of TNF-alpha, GMCSF and the chemokines RANTES and MIP-1alpha. Phosphorylation of ERK (which is critical for FcR-mediated ADCC and cytokine production) was enhanced in NK cells exposed to IL-12 and IgG as compared to control conditions. Conclusions: Cytokine stimulation of NK cells
Introduction: Perioperative blood transfusion has been linked to decreased survival in patients with pancreatic adenocarcinoma, although a causative mechanism has not been elucidated. Recent work, suggests that increased tumor associated macrophages (TAMs) also correlate with poor clinical outcomes wherein it is believe that tumors cells coapt macrophages to augment tumor growth by taking on an alternatively activated phenotype, which promotes angiogenesis and extracellular remodeling functions. Our lab has recently shown significant TAM expression of alternative activation markers within metastatic murine pancreatic adenocarcinoma. We hypothesize that transfusion of the plasma fraction of packed red blood cells increases TAM expression of alternative activation. Methods: C57/BL6 mice, age 8-9 weeks, underwent splenic inoculation with 2.5 x105 PanO2 murine pancreatic adenocarcinoma cells. at 7 days, mice were transfused via tail vein injection with the plasma fraction of packed red blood cells (pRBCs) at a doses of 1ml/kg (equivalent of 2 units), or saline control (NS). Necropsy was performed at day 35 or earlier if clinically indicated, and metastatic liver tumors collected. Tumors were homogenized and evaluated for the expression of the alternative activation markers CD206 and CD163 by immunoblotting. Mice not inoculated with tumor were used for control expression. Data are presented as fold-expression over control +/- standard error of the mean, N6 per group. Results: Evaluating hepatic metastases from tumor bearing mice receiving either NS or pRBCs, expression of both CD206 and CD163 were increased compared to control livers (p<0.001 for both). Comparing mice receiving NS compared to mice receiving pRBCs, transfused mice had an increased expression of CD206, 4.2560.35 fold for pRBCs vs. 2.9460.44 for NS (p¼0.044), and had increased expression of CD163, 7.7760.80 for pRBCs vs. 4.3060.53 for NS (p¼0.017). Conclusions: Clinically, the detrimental effects of perioperative blood have been recognized for some time. These data suggest blood transfusion affects tumor microenvironmental changes in TAMs that may promote tumor progression as alternative activation is known to correlate with angiogenic and remodeling functions of macrophages.