Low response rates in solid tumors including head and neck cancers (HNCs) have been attributed to failure of the drug to reach its intended target. However, investigation of drug delivery has been limited due to difficulties in measuring concentrations in the tumor and the ability to localizing drugs in human tissues. Factors determining intratumoral antibody distribution in primary tumor and metastatic lymph nodes have not been well-studied in human patients. To address this challenge, we propose to leverage fluorescently labeled antibodies to investigate antibody delivery into HNCs.
To this end, we have conducted a first-in-human clinical trial to assess the delivery of panitumumab-IRDye800 in HNCs. Twenty-two patients enrolled in this study received intravenous administration of panitumumab-IRDye800 at multiple subtherapeutic doses: (1) 0.06mg/kg, (2) 0.5 mg/kg, (3) 1 mg/kg, (4) 50 mg flat dose, (5) 25 mg flat dose. To quantify the antibody delivery, fresh tumor samples were procured and the amount of antibody in the tumor was quantified as ng/mg of tissue, which was then correlated with tumor characteristics. Immunohistochemistry of multiple protein markers, including EGFR, ERG, cytokeratin, Ki67, alpha-smooth muscle actin, etc., have been implemented in serial sections of primary tumors and metastatic lymph nodes. A quantitative image analysis pipeline was developed to analyze these IHC images and score the staining on both global and local scale. A predictive model was built to identify the most important predictors for antibody penetration from pharmacological factors, tumor pathophysiological factors, and tumor microenvironmental factors.
Introduction: Glioblastoma (GBM) is the most common and devastating primary brain tumor. The recurrence rate remains high with a median survival of 15 months. GBM’s infiltrative nature results in ill-defined margins that makes maximal tumor resection with minimal morbidity a challenge. Epidermal growth factor receptor (EGFR) is the most frequently amplified gene in GBM (35-45% of tumors) and is associated with overexpression in about 40-98% of cases, a characteristic of more aggressive phenotypes. We hypothesize that fluorescence labeled anti-EGFR monoclonal antibodies (mAb), panitumumab-IRDye800 (pan800) and cetuximab-IRDye800 (cet800), could be leveraged to enhance tumor contrast during surgical resection and improve patient outcome.
Methods: 50mg fluorescently labeled corresponding study drugs, pan800 and cet800 respectively, were administered 1-2 days in glioblastoma patients with contrast enhancing (CE) tumors prior to surgery following 100 mg loading dose of unlabeled cetuximab or panitumumab. Near-infrared fluorescence imaging of tumor and histologically negative peri-tumoral tissue was performed intraoperatively and ex vivo. Fluorescence was measured as mean fluorescence intensity (MFI), and tumor-to-background ratios (TBRs) were calculated by comparing MFIs of tumor and histologically uninvolved tissue.
Results: Despite heterogeneous drug uptake across all resected brain tissues, mean fluorescence intensity (MFI) correlated strongly (R^2=0.97) with tumor volume among histologically confirmed tumor tissues. The smallest detectable tumor size in a closed-field setting was 4.2 x 2.7 mm^2 (8.2 mg) for pan800 and 8.5 x 6.6 mm^2 (70mg) for cet800. Tumor tissues from pan800 infusion had significantly higher mean TBR (8.1 ± 4.6) than cet800 infused ones in intraoperative imaging (3.3 ± 2.7; P = 0.004). NIR fluorescence from both test drugs provided high contrast to identify as few as a cluster of (5 ± 1) tumor cells in macroscopic imaging of whole sections of paraffin embedded tissues. Sensitivity and specificity of MFI for viable tumor detection was calculated and fluorescence was found to be highly sensitive (64.4% for pan800, 73.0% for cet800) and specific (98.0% for pan800, 66.3% for cet800) for viable tumor tissue while normal peri-tumoral tissue showed minimal fluorescence. No related grade-2 adverse events were observed 30 days beyond the infusion of either study drugs.
Conclusion: EGFR antibody based imaging for contrast-enhanced glioblastomas proved safe in human patients and specific intratumoral delivery of NIR fluorescence provided high optical contrast and resolution for intraoperative image-guided resection. Fully humanized panitumumab-IRDye800 demonstrated superior detection sensitivity and tumor specificity over the chimeric cetuximab-IRDye800.