We have made an anti-mesothelin immunotoxin in which the major human B cell epitopes have been silenced or removed. Roche has taken a license to this invention and manufactured a new immunotoxin named RG7787 that has an anti-mesothelin Fab fused to a mutated form of PE. RG7787 has good anti-tumor activity in animal models and when combined with a Taxane produces complete remissions in these models. We have also initiated a program to identify and remove human specific T cell epitopes. We have identified all the major T cell epitopes in PE38 and using alanine-scanning mutagenesis identified amino acids essential for T cell activation. Using this information we have constructed LMB-T20, which contains six mutations in domain III (R427A, F443A, L477H, R494A, R505A and L552E) and a deletion of all the epitopes in domain II. LMB-T20 has low immunogenicity, is very active on mesothelin-expressing cancer cells and has good anti-tumor activity in mice. It is a good candidate for clinical development We are attempting to make a new immunotoxin with mutations that will eliminate both B and T cell epitopes. In collaboration with D. FitzGerald we have begun to study how protein synthesis arrest leads to apoptosis and have identified BAK and Mcl-1 as critical players in this process. We have also begun to study how protein phosphorylation regulates sensitivity of cells to immunotoxins by doing knock down studies of tyrosine and serine/threonine kinases and measuring the effect of knock down on immunotoxin mediated cell death. In collaboration with Alan Wayne POB, we have analyzed the response of cells directly isolated from patients with ALL to HA22 killing and are trying to set up an assay that may help predict if patients will respond to treatment. We have also isolated HA22 resistant cell lines and are using these to understand the basis of clinical drug resistance. One of the mechanisms is the loss of ability to synthesize DPH4 due to methylation of the the promoter of that gene. A significant advance is described in Liu, Onda et al in PNAS this year. This paper shows the identification and removal of human B cell epitopes from PE based immunotoxins producing a new immunotoxin predicted to have very low immunogenicity in humans.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC008753-33
Application #
9153500
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
33
Fiscal Year
2015
Total Cost
Indirect Cost
Name
Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
Kaplan, Gilad; Mazor, Ronit; Lee, Fred et al. (2018) Improving the In Vivo Efficacy of an Anti-Tac (CD25) Immunotoxin by Pseudomonas Exotoxin A Domain II Engineering. Mol Cancer Ther 17:1486-1493
Bera, T K; Abe, Y; Ise, T et al. (2018) Recombinant immunotoxins targeting B-cell maturation antigen are cytotoxic to myeloma cell lines and myeloma cells from patients. Leukemia 32:569-572
Wei, Junxia; Bera, Tapan K; Liu, Xiu Fen et al. (2018) Recombinant immunotoxins with albumin-binding domains have long half-lives and high antitumor activity. Proc Natl Acad Sci U S A 115:E3501-E3508
Mazor, Ronit; King, Emily M; Pastan, Ira (2018) Strategies to Reduce the Immunogenicity of Recombinant Immunotoxins. Am J Pathol 188:1736-1743
Müller, Fabian; Cunningham, Tyler; Beers, Richard et al. (2018) Domain II of Pseudomonas Exotoxin Is Critical for Efficacy of Bolus Doses in a Xenograft Model of Acute Lymphoblastic Leukemia. Toxins (Basel) 10:
Mazor, Ronit; King, Emily M; Onda, Masanori et al. (2018) Tolerogenic nanoparticles restore the antitumor activity of recombinant immunotoxins by mitigating immunogenicity. Proc Natl Acad Sci U S A 115:E733-E742
King, Emily M; Mazor, Ronit; Çuburu, Nicolas et al. (2018) Low-Dose Methotrexate Prevents Primary and Secondary Humoral Immune Responses and Induces Immune Tolerance to a Recombinant Immunotoxin. J Immunol 200:2038-2045
Müller, Fabian; Cunningham, Tyler; Stookey, Stephanie et al. (2018) 5-Azacytidine prevents relapse and produces long-term complete remissions in leukemia xenografts treated with Moxetumomab pasudotox. Proc Natl Acad Sci U S A 115:E1867-E1875
Liu, Xiu-Fen; Zhou, Qi; Hassan, Raffit et al. (2017) Panbinostat decreases cFLIP and enhances killing of cancer cells by immunotoxin LMB-100 by stimulating the extrinsic apoptotic pathway. Oncotarget 8:87307-87316
Mazor, Ronit; Addissie, Selamawit; Jang, Youjin et al. (2017) Role of HLA-DP in the Presentation of Epitopes from the Truncated Bacterial PE38 Immunotoxin. AAPS J 19:117-129

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