HA22 (R490A) Is a Recombinant Immunotoxin with Increased Antitumor Activity without an Increase in Animal Toxicity

Bang, Su Sik; Nagata, Satoshi; Onda, Masanori; Kreitman, Robert J.; Pastan, Ira

doi:10.1158/1078-0432.ccr-04-1939

Cited by 74 publications

(56 citation statements)

References 19 publications

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“…The pharmacokinetics and cytotoxic and antitumor activities were measured as previously described (13,16,21) using mAb IP12 for the blood levels. Animal experiments were performed under National Cancer Institute Animal Care and Use Committee-approved protocols.…”

Section: Methodsmentioning

confidence: 99%

Recombinant immunotoxin engineered for low immunogenicity and antigenicity by identifying and silencing human B-cell epitopes

Liu

Onda

Lee

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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143

140

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Recombinant immunotoxins (RITs) are hybrid proteins used to treat cancer. These proteins are composed of an Fv that reacts with cancer cells joined to a portion of Pseudomonas exotoxin A, which kills the cell. Because the toxin is a foreign protein, it can induce neutralizing antibodies and thereby limit the number of doses a patient can receive. We previously identified seven major mouse B-cell epitopes in the toxin, and subsequently silenced them using point mutations that converted large hydrophilic amino acids to alanine, yet retained full antitumor activity. Here we present results in which we identify and silence human B-cell epitopes in the RIT HA22. We obtained B cells from patients with antibodies to RITs, isolated the corresponding variable fragments (Fvs), and constructed a phage-display library containing Fvs that bind to the RITs. We then used alanine scanning mutagenesis to locate the epitopes. We found that human and mouse epitopes frequently overlap but are not identical. Most mutations that remove mouse epitopes did not remove human epitopes. Using the epitope information, we constructed a variant immunotoxin, HA22-LR-LO10, which has low reactivity with human antisera, yet has high cytotoxic and antitumor activity and can be given to mice at high doses without excess toxicity. The toxin portion of this RIT (LR-LO10) can be used with Fvs targeting other cancer antigens and is suitable for clinical development.cancer treatment | immunotherapy | leukemia | mesothelioma | protein engineering

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Section: Methodsmentioning

confidence: 99%

Recombinant immunotoxin engineered for low immunogenicity and antigenicity by identifying and silencing human B-cell epitopes

Liu

Onda

Lee

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

143

140

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“…A431/K5 cells expressing the mesothelin protein were maintained as previously described in DMEM containing 10% fetal bovine serum and 750 Ag/mL G418 (17).…”

Section: Methodsmentioning

confidence: 99%

Synergistic Antitumor Activity of Taxol and Immunotoxin SS1P in Tumor-Bearing Mice

Zhang

Xiang

Hassan

et al. 2006

Clinical Cancer Research

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Purpose:To investigate the combined antitumor activity in mice of immunotoxin SS1P and Taxol.Methods: Immunodeficient mice were implanted with A431/K5 tumors expressing mesothelin. Established tumors were treated i.v. with immunotoxin SS1P alone, i.p. with Taxol alone, or with the two agents together. SS1P was radiolabeled with 111 In and used to study the effect of Taxol on its uptake by A431/K5 tumors. Results: Using doses at which either agent alone caused stabilization of tumor growth, the combination was synergistic causing long-lasting complete remissions in many animals. In contrast, synergy was not observed when the same cells were treated with these agents in vitro. Tumor uptake of 111In-SS1P was not affected by treatment withTaxol. Conclusion: The combination of Taxol and SS1P exerts a synergistic antitumor effect in animals but not in cell culture. This effect is not secondary to increased tumor uptake of the immunotoxin. Synergy could be due to improved immunotoxin distribution within the tumor or could involve factors released by other cell types in the tumors.

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“…The emergence of clinically relevant, nonspecific toxicity has limited the therapeutic potential of numerous immunotoxins (40)(41)(42) and is a significant limitation to the clinical development of this class of molecules in general. Therefore, it is of paramount importance to design immunotoxins with minimal toxicity to normal tissues.…”

Section: Discussionmentioning

confidence: 99%

Single-Chain Antibody-Based Immunotoxins Targeting Her2/neu: Design Optimization and Impact of Affinity on Antitumor Efficacy and Off-Target Toxicity

Cao

Marks

Huang

et al. 2012

Molecular Cancer Therapeutics

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Recombinant immunotoxins, consisting of single-chain variable fragments (scFv) genetically fused to polypeptide toxins, represent potentially effective candidates for cancer therapeutics. We evaluated the affinity of various anti-Her2/neu scFv fused to recombinant gelonin (rGel) and its effect on antitumor efficacy and off-target toxicity. A series of rGel-based immunotoxins were created from the human anti-Her2/neu scFv C6.5 and various affinity mutants (designated ML3-9, MH3-B1, and B1D3) with affinities ranging from 10 À8 to 10 À11 mol/L. Against Her2/neu-overexpressing tumor cells, immunotoxins with increasing affinity displayed improved internalization and enhanced autophagic cytotoxicity. Targeting indices were highest for the highest affinity B1D3/rGel construct. However, the addition of free Her2/neu extracellular domain (ECD) significantly reduced the cytotoxicity of B1D3/rGel because of immune complex formation. In contrast, ECD addition had little impact on the lower affinity constructs in vitro. In vivo studies against established BT474 M1 xenografts showed growth suppression by all immunotoxins. Surprisingly, therapy with the B1D3-rGel induced significant liver toxicity because of immune complex formation with shed Her2/neu antigen in circulation. The MH3-B1/rGel construct with intermediate affinity showed effective tumor growth inhibition without inducing hepatotoxicity or complex formation. These findings show that while high-affinity constructs can be potent antitumor agents, they may also be associated with mistargeting through the facile formation of complexes with soluble antigen leading to significant off-target toxicity.

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HA22 (R490A) Is a Recombinant Immunotoxin with Increased Antitumor Activity without an Increase in Animal Toxicity

Cited by 74 publications

References 19 publications

Recombinant immunotoxin engineered for low immunogenicity and antigenicity by identifying and silencing human B-cell epitopes

Recombinant immunotoxin engineered for low immunogenicity and antigenicity by identifying and silencing human B-cell epitopes

Synergistic Antitumor Activity of Taxol and Immunotoxin SS1P in Tumor-Bearing Mice

Single-Chain Antibody-Based Immunotoxins Targeting Her2/neu: Design Optimization and Impact of Affinity on Antitumor Efficacy and Off-Target Toxicity

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