Structure and Molecular Interactions of a Unique Antitumor Antibody Specific for N-Glycolyl GM3

Krengel, Ute; Olsson, Lars‐Inge; Martínez, Carlos Enrique Ibarra; Talavera, Ariel; Rojas, Gertrudis; Mier, Elin S; Ångström, Jonas; Moreno, Ernesto

doi:10.1074/jbc.m311693200

Cited by 40 publications

(36 citation statements)

References 39 publications

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“…Interestedly, it was found a motif RXRR in the H-CDR3 of 14F7 mAb (23). Modeling studies together with the crystal structure of 14F7 mAb supported the importance of R98 and R100a in the recognition of NGcGM3 oligosaccharide by this antibody (46). It is possible that these arginines could also be relevant for cytotoxic activity.…”

Section: Discussionmentioning

confidence: 84%

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Anti-ganglioside antibody-induced tumor cell death by loss of membrane integrity

Roque-Navarro

Chakrabandhu

León

et al. 2008

Molecular Cancer Therapeutics

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Gangliosides have been involved in multiple cellular processes such as growth, differentiation and adhesion, and more recently as regulators of cell death signaling pathways. Some of these molecules can be considered as tumor-associated antigens, in particular, N-glycolyl sialic acid -containing gangliosides, which are promising candidates for cancer-targeted therapy because of their low expression in normal human tissues. In this study, we provided the molecular and cellular characterization of a novel cell death mechanism induced by the anti-NGcGM3 14F7 monoclonal antibody (mAb) in L1210 murine tumor cell line but not in mouse normal cells (B and CD4 + T lymphocytes) that expressed the antigen. Impairment of ganglioside synthesis in tumor cells abrogated the 14F7 mAb cytotoxic effect; however, exogenous reincorporation of the ganglioside did not restore tumor cell sensitivity to 14F7 mAb-induced cytotoxicity. 14F7 F(ab ¶) 2 but not Fab fragments retained the cytotoxic capacity of the whole mAb. By contrary, other mAb, which recognizes N-glycolylated gangliosides, did not show any cytotoxic effect. These mAbs showed quite different capacities to bind NGcGM3-positive cell lines measured by binding inhibition experiments. Interestingly, this complementindependent cell death mechanism did not resemble apoptosis, because no DNA fragmentation, caspase activation, or Fas mediation were observed. However, NGcGM3 ganglioside-mediated 14F7 mAb-induced cell death was accompanied by cellular swelling, membrane lesion formation, and cytoskeleton activation, suggesting an oncosislike phenomenon. This novel mechanism of cell death lets us to support further therapeutic approaches using NGcGM3 as a molecular target for antibody-based cancer immunotherapy.

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Section: Discussionmentioning

confidence: 84%

“…P3 mAb is restricted to the N-glycolyl sialic acid residue (48), whereas the epitope recognized by 14F7 mAb involved the disaccharide N-glycolyl sialic acid-galactose (46). Also, P3 mAb was encoded by a germ-line VH region (47), whereas 14F7 variable regions were heavily mutated (23), arguing an affinity maturation process.…”

Section: Discussionmentioning

confidence: 99%

Anti-ganglioside antibody-induced tumor cell death by loss of membrane integrity

Roque-Navarro

Chakrabandhu

León

et al. 2008

Molecular Cancer Therapeutics

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“…Fab fragments were prepared from purified mAb (produced at the Center of Molecular Immunology) by papain digestion (21). Further purification of the isoforms was performed as described by Krengel and colleagues (21).…”

Section: Purification Of Nimotuzumab (H-r3) Fab Fragmentsmentioning

confidence: 99%

“…Further purification of the isoforms was performed as described by Krengel and colleagues (21). The pH gradient was created in this case by mixing 50 mmol/L DEtA (pH 9.6) and 50 mmol/L DEtA (pH 8.6).…”

Section: Purification Of Nimotuzumab (H-r3) Fab Fragmentsmentioning

confidence: 99%

Nimotuzumab, an Antitumor Antibody that Targets the Epidermal Growth Factor Receptor, Blocks Ligand Binding while Permitting the Active Receptor Conformation

Friemann²,

et al. 2009

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Overexpression of the epidermal growth factor (EGF) receptor (EGFR) in cancer cells correlates with tumor malignancy and poor prognosis for cancer patients. For this reason, the EGFR has become one of the main targets of anticancer therapies. Structural data obtained in the last few years have revealed the molecular mechanism for ligand-induced EGFR dimerization and subsequent signal transduction, and also how this signal is blocked by either monoclonal antibodies or small molecules. Nimotuzumab (also known as h-R3) is a humanized antibody that targets the EGFR and has been successful in the clinics. In this work, we report the crystal structure of the Fab fragment of Nimotuzumab, revealing some unique structural features in the heavy variable domain. Furthermore, competition assays show that Nimotuzumab binds to domain III of the extracellular region of the EGFR, within an area that overlaps with both the surface patch recognized by Cetuximab (another anti-EGFR antibody) and the binding site for EGF. A computer model of the Nimotuzumab-EGFR complex, constructed by docking and molecular dynamics simulations and supported by mutagenesis studies, unveils a novel mechanism of action, with Nimotuzumab blocking EGF binding while still allowing the receptor to adopt its active conformation, hence warranting a basal level of signaling. [Cancer Res 2009;69(14):5851-9]

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“…64 14F7 mAb was able to recognize breast and melanoma tumors by immunohistochemistry, 53 and also breast cancer by radioimmunoscintigraphy, in a Phase 1/2 diagnostic clinical study. 55 14F7 mAb displayed anti-tumor properties in vivo against a GM3(Neu5Gc)-expressing murine myeloma.…”

confidence: 99%

Lonely killers

Fernández-Marrero

López‐Requena²

2011

mAbs

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Structure and Molecular Interactions of a Unique Antitumor Antibody Specific for N-Glycolyl GM3

Cited by 40 publications

References 39 publications

Anti-ganglioside antibody-induced tumor cell death by loss of membrane integrity

Anti-ganglioside antibody-induced tumor cell death by loss of membrane integrity

Nimotuzumab, an Antitumor Antibody that Targets the Epidermal Growth Factor Receptor, Blocks Ligand Binding while Permitting the Active Receptor Conformation

Lonely killers

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