Panitumumab Modified with Metal-Chelating Polymers (MCP) Complexed to 111In and 177Lu-An EGFR-Targeted Theranostic for Pancreatic Cancer.

Abstract

A metal-chelating polymer (MCP) with a polyglutamide (PGlu) backbone presenting on average 13 DOTA (tetraazacyclododecane-1,4,7,10-tetraacetic acid) chelators for complexing 111In or 177Lu and 10 polyethylene glycol (PEG) chains to minimize liver and spleen uptake was conjugated to antiepidermal growth factor receptor (EGFR) monoclonal antibody (mAb), panitumumab. Because panitumumab-MCP may be dual-labeled with 111In and 177Lu for SPECT, or radioimmunotherapy (RIT) exploiting the Auger electrons or β-particle emissions, respectively, we propose that panitumumab-MCP could be a useful theranostic agent for EGFR-positive PnCa. Bioconjugation was achieved by reaction of a hydrazine nicotinamide (HyNIC) group on the MCP with an aryl aromatic aldehyde introduced into panitumumab by reaction with succinimidyl-4-formylbenzamide (S-4FB). The conjugation reaction was monitored by measurement of the chromophoric bis-aryl hydrazone bond formed (ε350nm = 24 500 M-1 cm-1) to achieve two MCPs/panitumumab. Labeling of panitumumab-MCP with 111In or 177Lu demonstrated that masses as small as 0.1 μg were labeled to >90% labeling efficiency (L.E.) and a specific activity (SA) of >70 MBq/μg. Panitumumab-DOTA incorporating two DOTA per mAb was labeled with 111In or 177Lu to a maximum SA of 65 MBq/μg and 46 MBq/μg, respectively. Panitumumab-MCP-177Lu exhibited saturable binding to EGFR-overexpressing MDA-MB-468 human breast cancer cells. The Kd for binding of panitumumab-MCP-177Lu to EGFR (2.2 ± 0.6 nmol/L) was not significantly different than panitumumab-DOTA-177Lu (1.0 ± 0.4 nmol/L). 111In and 177Lu were stably complexed to panitumumab-MCP. Panitumumab-MCP-111In exhibited similar whole body retention (55-60%) as panitumumab-DOTA-111In in NOD-scid mice up to 72 h postinjection (p.i.) and equivalent excretion of radioactivity into the urine and feces. The uptake of panitumumab-MCP-111In in most normal tissues in NOD-scid mice with EGFR-positive PANC-1 human pancreatic cancer (PnCa) xenografts at 72 h p.i. was not significantly different than panitumumab-DOTA-111In, except for the liver which was 3-fold greater for panitumumab-MCP-111In. Tumor uptake of panitumumab-MCP-111In (6.9 ± 1.3%ID/g) was not significantly different than panitumumab-DOTA-11In (6.6 ± 3.3%ID/g). Tumor uptake of panitumumab-MCP-111In and panitumumab-DOTA-111In were reduced by preadministration of excess panitumumab, suggesting EGFR-mediated uptake. Tumor uptake of nonspecific IgG-MCP (5.4 ± 0.3%ID/g) was unexpectedly similar to panitumumab-MCP-111In. An increased hydrodynamic radius of IgG when conjugated to an MCP may encourage tumor uptake via the enhanced permeability and retention (EPR) effect. Tumor uptake of panitumumab-DOTA-111In was 3.5-fold significantly higher than IgG-DOTA-111In. PANC-1 tumors were imaged by microSPECT/CT at 72 h p.i. of panitumumab-MCP-111In or panitumumab-DOTA-111In. Tumors were not visualized with preadministration of excess panitumumab to block EGFR, or with nonspecific IgG radioimmunoconjugates. We conclude that linking panitumumab to an MCP enabled higher SA labeling with 111In and 177Lu than DOTA-conjugated panitumumab, with preserved EGFR binding in vitro and comparable tumor localization in vivo in mice with s.c. PANC-1 human PnCa xenografts. Normal tissue distribution was similar except for the liver which was higher for the polymer radioimmunoconjugates.