Nanoconjugation modulates the trafficking and mechanism of antibody induced receptor endocytosis

Abstract

Treatment with monoclonal antibody (mAbs) is a viable therapeutic option in cancer. Recently, these mAbs such as cetuximab, herceptin, etc., have been used as targeting agents to selectively deliver chemotherapeutics to cancerous cells. However, mechanisms of nanoparticles-mAbs interactions with the target cells and its effect on intracellular trafficking and mechanism are currently unknown. In this paper, we demonstrate that the distinct patterning and dynamics of anti-EGFR (epidermal growth factor receptor) antibody cetuximab (C225)-induced EGFR internalization in pancreatic cancer cells with variable receptor expression is altered upon nanoconjugation. Nanoconjugation uniformly enhanced C225-induced EGFR endocytosis in PANC-1, AsPC-1, and MiaPaca-2 cells, influenced its compartmentalization and regulated the involvement of dynamin-2 in the endocytic processes. Receptor endocytosis and its intracellular trafficking were monitored by confocal microscopy and transmission electron microscopy. The role of dynamin-2 in EGFR endocytosis was determined after overexpressing either wild-type dynamin-2 or mutant dynamin-2 in pancreatic cancer cells followed by tracking the receptor-antibody complex internalization by confocal microscopy. Significantly, these findings demonstrate that the nanoconjugation cannot be construed as an innocuous reaction involved in attaching the targeting agent to the nanoparticle, instead it may distinctly alter the cellular processes at the molecular level, at least antibody induced receptor endocytosis. This information is critical for successful design of a nanoparticle-based targeted drug delivery system for future clinical translation.EGFR | nanoparticle | dynamin | pancreatic cancer N anotechnology provides opportunities for biomedical applications that may improve human health care in the future (1-13). Engineered nanoparticles are evolving as promising candidates for various biological applications due to their tunable biophysical and biochemical properties (14-16). However, fundamental studies to understand the mechanism of cell-nanomaterial interactions are still lacking. To advance the successful development of biomedical nanotechnology for clinical use, vivid understanding of such mechanisms is essential.Epidermal growth factor receptor (EGFR) represents a unique target as over expression of EGFR has been implicated in the pathogenesis of many cancers (17, 18). Cetuximab (C225), a monoclonal anti-EGFR antibody, has been approved by the FDA for treatment of colon and head and neck cancers (19). Cetuximab is also in different phases of clinical trials for other cancers (20,21). However, the mechanism and pattern of C225 induced endocytosis of EGFR in pancreatic cancer cells of variable EGFR expression is not fully understood (20,21). Furthermore, the effect of nanoconjugation on the mechanism of C225 induced endocytosis remains to be investigated. Here, we demonstrate the mechanism and endocytic pattern of C225 and gold conjugated-C225 induced internalization of EGFR in pancreatic ...

Inhibition of tumor growth and metastasis by a self-therapeutic nanoparticle

“…Imaging was carried on a LSM780 confocal microscope, and the images were processed using ImageJ (National Institutes of Health) software (51).…”

Cetuximab-conjugated iron oxide nanoparticles for cancer imaging and therapy

“…This finding demonstrates the rapid internalization and degradation of EGFR, which is consistent with the result reported by Bhattacharyya et al who demonstrated that conjugation of cetuximab to gold nanoparticles enhanced EGFR endocytosis in pancreatic cancer cell lines with variable EGFR expression. 34 Their results using confocal microscopy and transmission electron microscopy confirmed rapid internalization and predominantly endosomal localization of these antibody-nanoparticle conjugates in EGFR-expressing cells via receptor-mediated endocytosis. The ability of cetuximab-SPION conjugates to downregulate surface EGFR is clinically significant because attenuation of the membrane presentation of the receptor implies reduced signaling through its corresponding ligand.…”

“…Nanoconjugation has been reported to enhance cetuximabinduced EGFR endocytosis and to influence intracellular trafficking of EGFR in pancreatic cancer cells. 34 To examine the ability of cet-PEG-dexSPIONs to induce degradation of EGFR by enhancing internalization of the receptor, the kinetics of cet-PEG-dexSPION-induced EGFR downregulation were compared with those of cetuximab-induced EGFR Abbreviations: cet, cetuximab; dex, dextran; egFr, epidermal growth factor receptor; Peg, polyethylene glycol; sD, standard deviation; sPIONs, superparamagnetic iron oxide nanoparticles.…”

Multiple cues on the physiochemical, mesenchymal, and intracellular trafficking interactions with nanocarriers to maximize tumor target efficiency

“…In this regard, nanoconjugation (eg, antibody) can enhance localization to the Golgi and Ly through EE. 17 Specifically, EGFRs induced greater endocytosis when conjugated with Au NP and highlighted the importance of conjugation for cellular transport. 17 In addition, cholesterol and phospholipid liposome NPs were localized into the ER and Golgi apparatus after endosome/Ly stages, and were eventually effluxed by permeability glycoprotein and ABCG1 efflux pumps ( Figure 5).…”

“…6,[10][11][12][13][14][15][16] Specifically, epidermal growth factor receptor (EGFR) is the most well-known cancer receptor, and EGFR-antibody interactions of cancer drugs were previously reported. 6,10,17 However, a more revolutionary approach than current antibody receptor targeting is necessary to further increase targeting efficacy. For example, the utilization of selective organ accumulation with respect to shape and size dimensions of nanomaterials would be advantageous in targeting specific organs (ie, in the case of source of tumor tissues) ( Figure 1).…”