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A novel fully human antitumour immunoRNase targeting ErbB2-positive tumours.

Borriello M, Laccetti P, Terrazzano G, D'Alessio G, De Lorenzo C - Br. J. Cancer (2011)

Bottom Line: Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer with success, can engender cardiotoxicity and a high fraction of patients is resistant to Trastuzumab treatment.Erb-hcAb-RNase retains the enzymatic activity of HP-RNase and specifically binds to ErbB2-positive cells with an affinity comparable with that of the parental Erb-hcAb.Its antitumour activity is more potent than that of the parental Erb-hcAb as the novel immunoconjugate has acquired RNase-based cytotoxicity in addition to the inhibitory growth effects, antibody-dependent and complement-dependent cytotoxicity of Erb-hcAb.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia Strutturale e Funzionale, Università Federico II, via Cinthia, Napoli 80126, Italy.

ABSTRACT

Background: ErbB2 is an attractive target for immunotherapy, as it is a tyrosine kinase receptor overexpressed on tumour cells of different origin, with a key role in the development of malignancy. Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer with success, can engender cardiotoxicity and a high fraction of patients is resistant to Trastuzumab treatment.

Methods: A novel human immunoRNase, called anti-ErbB2 human compact antibody-RNase (Erb-hcAb-RNase), made up of the compact anti-ErbB2 antibody Erbicin-human-compact Antibody (Erb-hcAb) and human pancreatic RNase (HP-RNase), has been designed, expressed in mammalian cell cultures and purified. The immunoRNase was then characterised as an enzymatic protein, and tested for its biological actions in vitro and in vivo on ErbB2-positive tumour cells.

Results: Erb-hcAb-RNase retains the enzymatic activity of HP-RNase and specifically binds to ErbB2-positive cells with an affinity comparable with that of the parental Erb-hcAb. Moreover, this novel immunoRNase is endowed with an effective and selective antiproliferative action for ErbB2-positive tumour cells both in vitro and in vivo. Its antitumour activity is more potent than that of the parental Erb-hcAb as the novel immunoconjugate has acquired RNase-based cytotoxicity in addition to the inhibitory growth effects, antibody-dependent and complement-dependent cytotoxicity of Erb-hcAb.

Conclusion: Erb-hcAb-RNase could be a promising candidate for the immunotherapy of ErbB2-positive tumours.

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Related in: MedlinePlus

SDS–PAGE and western blotting analyses of purified Erb-hcAb-RNase. Erb-hcAb-RNase was run under reducing (lane B) or non-reducing (lane C) conditions; molecular weight standards are in lane A; western blotting analyses of the purified sample with an anti-human IgG1 (Fc specific) (lane D) or with the anti-HP-RNase antibody (lane E); Zymogram of Erb-hcAb-RNase using yeast RNA as a substrate in lane F.
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fig2: SDS–PAGE and western blotting analyses of purified Erb-hcAb-RNase. Erb-hcAb-RNase was run under reducing (lane B) or non-reducing (lane C) conditions; molecular weight standards are in lane A; western blotting analyses of the purified sample with an anti-human IgG1 (Fc specific) (lane D) or with the anti-HP-RNase antibody (lane E); Zymogram of Erb-hcAb-RNase using yeast RNA as a substrate in lane F.

Mentions: When Erb-hcAb-RNase was analysed by SDS–PAGE (Figure 2), it was found to migrate under reducing conditions with the expected molecular size of about 70 kDa (Figure 2, lane B), and as a dimer of about 140 kDa under non-reducing conditions (Figure 2, lane C). This result indicates that the fusion protein is expressed as a disulphide-linked dimer. Western blotting analyses performed with either an anti-human Fc or an anti-HP-RNase antibody demonstrated immunoreactivity of the purified, dimeric protein with a molecular size of 140 kDa (Figure 2, lanes D and E).


A novel fully human antitumour immunoRNase targeting ErbB2-positive tumours.

Borriello M, Laccetti P, Terrazzano G, D'Alessio G, De Lorenzo C - Br. J. Cancer (2011)

SDS–PAGE and western blotting analyses of purified Erb-hcAb-RNase. Erb-hcAb-RNase was run under reducing (lane B) or non-reducing (lane C) conditions; molecular weight standards are in lane A; western blotting analyses of the purified sample with an anti-human IgG1 (Fc specific) (lane D) or with the anti-HP-RNase antibody (lane E); Zymogram of Erb-hcAb-RNase using yeast RNA as a substrate in lane F.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3111160&req=5

fig2: SDS–PAGE and western blotting analyses of purified Erb-hcAb-RNase. Erb-hcAb-RNase was run under reducing (lane B) or non-reducing (lane C) conditions; molecular weight standards are in lane A; western blotting analyses of the purified sample with an anti-human IgG1 (Fc specific) (lane D) or with the anti-HP-RNase antibody (lane E); Zymogram of Erb-hcAb-RNase using yeast RNA as a substrate in lane F.
Mentions: When Erb-hcAb-RNase was analysed by SDS–PAGE (Figure 2), it was found to migrate under reducing conditions with the expected molecular size of about 70 kDa (Figure 2, lane B), and as a dimer of about 140 kDa under non-reducing conditions (Figure 2, lane C). This result indicates that the fusion protein is expressed as a disulphide-linked dimer. Western blotting analyses performed with either an anti-human Fc or an anti-HP-RNase antibody demonstrated immunoreactivity of the purified, dimeric protein with a molecular size of 140 kDa (Figure 2, lanes D and E).

Bottom Line: Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer with success, can engender cardiotoxicity and a high fraction of patients is resistant to Trastuzumab treatment.Erb-hcAb-RNase retains the enzymatic activity of HP-RNase and specifically binds to ErbB2-positive cells with an affinity comparable with that of the parental Erb-hcAb.Its antitumour activity is more potent than that of the parental Erb-hcAb as the novel immunoconjugate has acquired RNase-based cytotoxicity in addition to the inhibitory growth effects, antibody-dependent and complement-dependent cytotoxicity of Erb-hcAb.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia Strutturale e Funzionale, Università Federico II, via Cinthia, Napoli 80126, Italy.

ABSTRACT

Background: ErbB2 is an attractive target for immunotherapy, as it is a tyrosine kinase receptor overexpressed on tumour cells of different origin, with a key role in the development of malignancy. Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer with success, can engender cardiotoxicity and a high fraction of patients is resistant to Trastuzumab treatment.

Methods: A novel human immunoRNase, called anti-ErbB2 human compact antibody-RNase (Erb-hcAb-RNase), made up of the compact anti-ErbB2 antibody Erbicin-human-compact Antibody (Erb-hcAb) and human pancreatic RNase (HP-RNase), has been designed, expressed in mammalian cell cultures and purified. The immunoRNase was then characterised as an enzymatic protein, and tested for its biological actions in vitro and in vivo on ErbB2-positive tumour cells.

Results: Erb-hcAb-RNase retains the enzymatic activity of HP-RNase and specifically binds to ErbB2-positive cells with an affinity comparable with that of the parental Erb-hcAb. Moreover, this novel immunoRNase is endowed with an effective and selective antiproliferative action for ErbB2-positive tumour cells both in vitro and in vivo. Its antitumour activity is more potent than that of the parental Erb-hcAb as the novel immunoconjugate has acquired RNase-based cytotoxicity in addition to the inhibitory growth effects, antibody-dependent and complement-dependent cytotoxicity of Erb-hcAb.

Conclusion: Erb-hcAb-RNase could be a promising candidate for the immunotherapy of ErbB2-positive tumours.

Show MeSH
Related in: MedlinePlus