In vivo molecular imaging with cetuximab, an anti-EGFR antibody, for prediction of response in xenograft models of human colorectal cancer

Background and study aims: Molecular imaging has mainly been studied for detection of lesions using diagnostic probes. The aim of the current trial was to evaluate in vivo confocal laser endomicroscopy (CLE) with cetuximab, an antibody targeting the epidermal growth factor receptor (EGFR), for detection and moreover early prediction of response to molecular chemotherapy in models of human colorectal cancer (CRC).

Methods: Xenografts with cetuximab-sensitive (HT29) and cetuximab-resistant (SW620) human CRC cell lines were induced in 44 mice. CLE was performed 48 h after injection of a fluorescently labelled cetuximab test dose, and compared with isotype antibody or untreated controls on d0, and d30 (HT29) or d15 (SW620). Initial fluorescence intensity was examined in relation to clinical readouts (tumor growth, thriving, mortality) during cetuximab treatment vs. controls. Results were validated in vivo with wide-field molecular imaging in three HT29 mice and ex vivo using fluorescence-activated cell sorting (FACS) and immunohistochemistry.

Results: All HT29 xenografts showed specific fluorescence in vivo after cetuximab injection on d0 and d30. Fluorescence at d0 was significantly stronger in cetuximab-treated HT29 tumors than in HT29 controls (P = 0.0017) or cetuximab-treated SW620 tumors (P = 0.0027), and accorded with significantly slower tumor progression (P = 0.0009), better overall survival (P = 0.02), and better physical condition (P < 0.0001). Cetuximab sensitivity could be predicted from fluorescence intensity at d0 with high positive predictive value.

Conclusions: Molecular CLE was for the first time linked to early prediction of response to targeted therapy in models of human CRC. Therapeutic antibodies can be used as molecular beacons in CLE and wide-field techniques. These results may indicate a promising principle for early patient stratification.

^* The first two authors contributed equally.

• References

• 1 Jemal A, Siegel R, Ward E et al. Cancer statistics, 2008. CA Cancer J Clin 2008; 58: 71-96
• 2 Kahi CJ, Imperiale TF, Juliar BE et al. Effect of screening colonoscopy on colorectal cancer incidence and mortality. Clin Gastroenterol Hepatol 2009; 7: 770-775 ; quiz 711
• 3 Brenner H, Altenhofen L, Hoffmeister M. Eight years of colonoscopic bowel cancer screening in Germany: initial findings and projections. Dtsch Arztebl Int 2010; 107: 753-759
• 4 Rex DK, Cutler CS, Lemmel GT et al. Colonoscopic miss rates of adenomas determined by back-to-back colonoscopies. Gastroenterology 1997; 112: 24-28
• 5 Heresbach D, Barrioz T, Lapalus MG et al. Miss rate for colorectal neoplastic polyps: a prospective multicenter study of back-to-back video colonoscopies. Endoscopy 2008; 40: 284-290
• 6 Kiesslich R, Burg J, Vieth M et al. Confocal laser endoscopy for diagnosing intraepithelial neoplasias and colorectal cancer in vivo. Gastroenterology 2004; 127: 706-713
• 7 Polglase AL, McLaren WJ, Skinner SA et al. A fluorescence confocal endomicroscope for in vivo microscopy of the upper- and the lower-GI tract. Gastrointest Endosc 2005; 62: 686-695
• 8 Wallace MB, Fockens P. Probe-based confocal laser endomicroscopy. Gastroenterology 2009; 136: 1509-1513
• 9 Mahmood U. Optical molecular imaging approaches in colorectal cancer. Gastroenterology 2010; 138: 419-422
• 10 Pierce MC, Javier DJ, Richards-Kortum R. Optical contrast agents and imaging systems for detection and diagnosis of cancer. Int J Cancer 2008; 123: 1979-1990
• 11 Weissleder R, Pittet MJ. Imaging in the era of molecular oncology. Nature 2008; 452: 580-589
• 12 Goetz M, Wang TD. Molecular imaging in gastrointestinal endoscopy. Gastroenterology 2010; 138: 828-833 e821
• 13 Goetz M. Molecular imaging in GI endoscopy. Gastrointest Endosc 2012; 76: 1207-1209
• 14 Barrett T, Koyama Y, Hama Y et al. In vivo diagnosis of epidermal growth factor receptor expression using molecular imaging with a cocktail of optically labeled monoclonal antibodies. Clin Cancer Res 2007; 13: 6639-6648
• 15 Goetz M, Ziebart A, Foersch S et al. In vivo molecular imaging of colorectal cancer with confocal endomicroscopy by targeting epidermal growth factor receptor. Gastroenterology 2010; 138: 435-446
• 16 Foersch S, Kiesslich R, Waldner MJ et al. Molecular imaging of VEGF in gastrointestinal cancer in vivo using confocal laser endomicroscopy. Gut 2010; 59: 1046-1055
• 17 Hsiung PL, Hardy J, Friedland S et al. Detection of colonic dysplasia in vivo using a targeted heptapeptide and confocal microendoscopy. Nat Med 2008; 14: 454-458
• 18 Liu Z, Miller SJ, Joshi BP et al. In vivo targeting of colonic dysplasia on fluorescence endoscopy with near-infrared octapeptide. Gut 2013; 62: 395-403
• 19 Alencar H, Funovics MA, Figueiredo J et al. Colonic adenocarcinomas: near-infrared microcatheter imaging of smart probes for early detection--study in mice. Radiology 2007; 244: 232-238
• 20 Van Cutsem E, Kohne CH, Lang I et al. Cetuximab plus irinotecan, fluorouracil, and leucovorin as first-line treatment for metastatic colorectal cancer: updated analysis of overall survival according to tumor KRAS and BRAF mutation status. J Clin Oncol 2011; 29: 2011-2019
• 21 Cunningham D, Humblet Y, Siena S et al. Cetuximab monotherapy and cetuximab plus irinotecan in irinotecan-refractory metastatic colorectal cancer. N Engl J Med 2004; 351: 337-345
• 22 Engstrom PF, Arnoletti JP, Benson AB et al. NCCN Clinical Practice Guidelines in Oncology: colon cancer. J Natl Compr Canc Netw 2009; 7: 778-831
• 23 Italiano A, Follana P, Caroli FX et al. Cetuximab shows activity in colorectal cancer patients with tumors for which FISH analysis does not detect an increase in EGFR gene copy number. Ann Surg Oncol 2008; 15: 649-654
• 24 Chung KY, Shia J, Kemeny NE et al. Cetuximab shows activity in colorectal cancer patients with tumors that do not express the epidermal growth factor receptor by immunohistochemistry. J Clin Oncol 2005; 23: 1803-1810
• 25 De Roock W, Jonker DJ, Di Nicolantonio F et al. Association of KRAS p.G13D mutation with outcome in patients with chemotherapy-refractory metastatic colorectal cancer treated with cetuximab. JAMA 2010; 304: 1812-1820
• 26 Bohanes P, LaBonte MJ, Winder T et al. Predictive molecular classifiers in colorectal cancer. Semin Oncol 2011; 38: 576-587
• 27 Schrag D. The price tag on progress – chemotherapy for colorectal cancer. N Engl J Med 2004; 351: 317-319
• 28 Helman EE, Newman JR, Dean NR et al. Optical imaging predicts tumor response to anti-EGFR therapy. Cancer Biol Ther 2010; 10: 166-171
• 29 Goetz M, Fottner C, Schirrmacher E et al. In vivo confocal real-time mini-microscopy in animal models of human inflammatory and neoplastic diseases. Endoscopy 2007; 39: 350-356
• 30 Hoetker MS, Kiesslich R, Diken M et al. Molecular in vivo imaging of gastric cancer in a human-murine xenograft model: targeting epidermal growth factor receptor (EGFR). Gastrointest Endosc 2012; 76: 612-620
• 31 Manning HC, Merchant NB, Foutch AC et al. Molecular imaging of therapeutic response to epidermal growth factor receptor blockade in colorectal cancer. Clin Cancer Res 2008; 14: 7413-7422
• 32 Liu Z, Miller SJ, Joshi BP et al. In vivo targeting of colonic dysplasia on fluorescence endoscopy with near-infrared octapeptide. Gut 2013; 62: 395-403 Epub 17 March 2012
• 33 Fottner C, Mettler E, Goetz M et al. In vivo molecular imaging of somatostatin receptors in pancreatic islet cells and neuroendocrine tumors by miniaturized confocal laser-scanning fluorescence microscopy. Endocrinology 2010; 151: 2179-2188
• 34 Miller SJ, Lee CM, Joshi BP et al. Targeted detection of murine colonic dysplasia in vivo with flexible multispectral scanning fiber endoscopy. J Biomed Opt 2012; 17 021103
• 35 Li M, Anastassiades CP, Joshi B et al. Affinity peptide for targeted detection of dysplasia in Barrett's esophagus. Gastroenterology 2010; 139: 1472-1480
• 36 van Dam GM, Themelis G, Crane LM et al. Intraoperative tumor-specific fluorescence imaging in ovarian cancer by folate receptor-alpha targeting: first in-human results. Nat Med 2011; 17: 1315-1319
• 37 Maughan TS, Adams RA, Smith CG et al. Addition of cetuximab to oxaliplatin-based first-line combination chemotherapy for treatment of advanced colorectal cancer: results of the randomised phase 3 MRC COIN trial. Lancet 2011; 377: 2103-2114
• 38 Kwekkeboom DJ, Krenning EP, Lebtahi R et al. ENETS consensus guidelines for the standards of care in neuroendocrine tumors: peptide receptor radionuclide therapy with radiolabeled somatostatin analogs. Neuroendocrinology 2009; 90: 220-226
• 39 Boudreaux JP, Klimstra DS, Hassan MM et al. The NANETS consensus guideline for the diagnosis and management of neuroendocrine tumors: well-differentiated neuroendocrine tumors of the jejunum, ileum, appendix, and cecum. Pancreas 2010; 39: 753-766