Use of combination of monoclonal antibodies directed against three distinct epitopes of a tumor‐associated antigen: Analysis of cell binding and internalization

Casalini, Patrizia; Mezzanzanica, Delia; Canevari, Silvana; Torre, Gabriella Della; Miorn, Silvia; Colnaghi, Maria I.; Matzku, S.

doi:10.1002/ijc.2910480222

Cited by 19 publications

(8 citation statements)

References 18 publications

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“…The internalisation of NCAM-MAb 123C3 complex does not induce internalisation of a non-internalising MAb, which binds to another epitope (unpublished data). The absence of co-internalisation of MAbs is in analogy to the findings of other investigators Casalini et al, 1991).…”

Section: Antibody Labellingsupporting

confidence: 87%

Immunoscintigraphy of small-cell lung cancer xenografts with anti neural cell adhesion molecule monoclonal antibody, 123C3: improvement of tumour uptake by internalisation

Kwa

Wesseling²,

Verhoeven³

et al. 1996

Br J Cancer

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Summary The efficacy of three murine monoclonal antibodies (MAbs) for immunoscintigraphy of small-cell lung cancer (SCLC) xenografts was studied in a Balb/c nu/nu mouse model. These MAbs, 123C3, 123A8 and MOCl91, belong to cluster 1 of anti-SCLC MAbs and bind to the neural cell adhesion molecule (NCAM) with similar affinity. After intraperitoneal injection of these MAbs, labelled with 1251, the highest uptake in tumour tissue was obtained with MAb 123C3. Seven days after administration of this MAb 13.9% of the injected dose per gram of tumour tissue was retained in the tumour. The corresponding tumour to tissue ratios ranged from 3.97 for blood to 31.03 for colon. The imaging results and the tumour uptake were less favourable for the two other MAbs, 123A8 and MOCl91 (fractions of injected dose respectively 6.7% and 9.2%), although affinity, biological activity after labelling and uptake in non-tumour tissues were very similar for all three MAbs. These results may be explained by the differences in the interaction between the MAbs and the tumour cells. MAb 123C3 is internalised into tumour cells, whereas both other anti-NCAM MAbs are not. Internalisation into NCI H69 cells was demonstrated in vitro by a radioimmunoassay, confocal laser scanning microscopy and electron microscopy. The internalised fraction of MAb 123C3 was 22.3% after 24 h, whereas this fraction was only 7.5% for MAb 123A8. Although the internalised radiolabelled MAbs are usually degraded and dehalogenated intracellularly, the retained radioactivity is high. Apparently, intracellular degradation of radiolabelled MAb 123C3 and subsequent secretion of radioactive iodine did not prevent the accumulation of intracellular radioactivity. In conclusion, accumulation and retention of radioactivity in the tumour tissue, due to internalisation of radiolabelled MAbs, may improve the results of immunoscintigraphy.

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Section: Antibody Labellingsupporting

confidence: 87%

Immunoscintigraphy of small-cell lung cancer xenografts with anti neural cell adhesion molecule monoclonal antibody, 123C3: improvement of tumour uptake by internalisation

Kwa

Wesseling²,

Verhoeven³

et al. 1996

Br J Cancer

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“…Particularly, they are not coordinately present on olher ovarian carcinomas, e.g on the IGROVl cell line, nor on different types of adenocarcinoma, e.g. HT-29 colon carcinoma cell line [6].…”

Section: I121mentioning

confidence: 94%

“…The epitopes on OVCA432 were found to be co-expressed by a 38-kDa surface glycoprotein (gp38) [6], which is linked to the cell membrane by glycosyl-phosphatidylinositol (glycosylPtdIns) anchoring [7]. Recently, the protein has been revealed by cloning [8, 91 to belong to a highly homologous group of human folate-binding proteins (hFBPs) [lo, 111, in-volved in the uptake of extracellular 5-methyl tetrahydrofolate and delivered into the cells by a new endocytic pathway…”

Section: Bologna Italymentioning

confidence: 99%

Effect of hydrostatic pressure and cholesterol depletion on the expression of a tumor‐associated antigen

Gesmundo

Calonghi

Sartor

et al. 1993

European Journal of Biochemistry

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The molecular events related to the expression of three tumor‐associated epitopes, Ca‐MOv17, Ca‐MOv18 and Ca‐MOv19 have been addressed. The epitopes are carried by a 38‐kDa glycoprotein (gp38), recently cloned and identified as a human folate‐binding protein. They were found to be coexpressed on the surface of the ovarian carcinoma cell line OVCA432, while they are not coordinately expressed on other adenocarcinoma cell lines (IGROV1, HT‐29). This lack of coexpression was investigated from a molecular point of view. We studied three carcinoma cell lines, characterized by a different reactivity with the three relevant monoclonal antibodies MOv17, MOv18 and MOv19. The epitope expression was examined after modifying the membrane properties by using hydrostatic pressure and/or the variation of cholesterol content. Measurement of the expression after cell labelling by mAbs was performed by indirect immunofluorescence, using both fluorescence microscopy and flow cytometry. At variance with HT‐29 cells, treatment of ovarian carcinoma IGROV1 cells with hydrostatic pressure failed to exert any effect. On IGROV1, instead, cholesterol depletion affected the expression Ca‐MOv17, increasing, in the indirect immunofluorescence tests, the proportion of positive cells from 0 to 66 ± 9%. Moreover, restoring the cholesterol content of the plasma membrane did not reverse the induced epitope expression. In parallel, ìmmunoprecipitation experiments confirmed that, on IGROV1 surface, gp38 was recognized by all three mAbs. The data presented suggest that in IGROV1 cells the selective lacking of the epitope expression is related to the physical state of the plasma membrane. An explanation is provided by the model of membrane microdomains in which epitope expression may be influenced by the cholesterol level of different plasma membrane regions.

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“…Protein-G-purified (MAb Trap G, Pharmacia, Sweden) anti-saporin mAbs were labelled by the lactoperoxidase-catalysed iodination method (Casalini et al, 1991) to a specific activity of 52 TBq/ mmol. The binding assay was performed as previously described (Sforzini et al, 1995a).…”

Section: Methodsmentioning

confidence: 99%

Targeting of saporin to Hodgkin's lymphoma cells by anti‐CD30 and anti‐CD25 bispecific antibodies

et al. 1998

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Summary. CD25 and CD30 represent suitable target molecules for bispecific antibody (bimAb)-driven toxin delivery to lymphoid tumour cells. We describe two new anti-CD30/anti-saporin bimAbs (termed CD30 × sap1 and CD30 × sap2), produced by hybrid hybridomas, which react against non-cross-reactive epitopes of the saporin molecule, and compared their effect with a bimAb reacting with saporin and with CD25 (CD25 × sap1). In a protein synthesis inhibition assay these bimAbs were able to enhance saporin toxicity (IC 50 8·5 × 10 ¹9 M in the absence of mAbs) with a similar activity: in the presence of 10 ¹9 M CD30 × sap1 bimAb the IC 50 was 2·75 × 10 ¹11 M, whereas with CD30 × sap2 bimAb the IC 50 was 6·5 × 10 ¹11 M and CD25 × sap1 bimAb displayed an IC 50 of 3 × 10 ¹11 M (as saporin). The combined use of the two anti-CD30 bimAbs further increased cytotoxicity by 100-fold, resulting in an IC 50 of 1·9 × 10 ¹13 M. A slightly less efficient improvement was obtained by combining the CD25 × sap1 bimAb with the CD30 × sap2 bimAb directed against a different toxin epitope (saporin IC 50 to 7 × 10 ¹13 M). In contrast, no synergistic effect was observed using the combination of the anti-CD25 bimAb with the anti-CD30 bimAb reacting with the same epitope of saporin (IC 50 ¼ 4·5 × 10 ¹11 M). Analysis of FITC-saporin binding to L540 cells by flow cytometry demonstrated that the appropriate combinations of the two anti-CD30/antisaporin bimAbs or of the anti-CD30/anti-saporin and anti-CD25/anti-saporin bimAbs had a cooperative effect on the binding of the ribosome-inactivating protein (RIP) to the cells, when compared with single bimAbs.

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Use of combination of monoclonal antibodies directed against three distinct epitopes of a tumor‐associated antigen: Analysis of cell binding and internalization

Cited by 19 publications

References 18 publications

Immunoscintigraphy of small-cell lung cancer xenografts with anti neural cell adhesion molecule monoclonal antibody, 123C3: improvement of tumour uptake by internalisation

Immunoscintigraphy of small-cell lung cancer xenografts with anti neural cell adhesion molecule monoclonal antibody, 123C3: improvement of tumour uptake by internalisation

Effect of hydrostatic pressure and cholesterol depletion on the expression of a tumor‐associated antigen

Targeting of saporin to Hodgkin's lymphoma cells by anti‐CD30 and anti‐CD25 bispecific antibodies

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