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Abstract LB-163: Development of an immunoPET tracer for imaging human CD8+ T cells
Background: Infiltration of CD8+ T cells into human tumors is associated with an increased disease-specific survival in many cancers. A better understanding of T cell trafficking and characterization of the subset of CD8+ T cells with the highest localized activity would improve the ability to sched...
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Published in: | Cancer research (Chicago, Ill.) Ill.), 2014-10, Vol.74 (19_Supplement), p.LB-163-LB-163 |
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Main Authors: | , , , , , , , , , |
Format: | Article |
Language: | English |
Online Access: | Get full text |
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Summary: | Background: Infiltration of CD8+ T cells into human tumors is associated with an increased disease-specific survival in many cancers. A better understanding of T cell trafficking and characterization of the subset of CD8+ T cells with the highest localized activity would improve the ability to schedule and select patients for chemo- and immuno-therapy regimens. To facilitate this objective, we have engineered a humanized anti-CD8 antibody fragment and used it to image human CD8+ T cells in a tumor xenograft and humanized mouse model.
Methods: The VH and VL sequences of a murine anti-CD8 antibody were cloned by RT-PCR, engineered into minibody (Mb) fragments (scFv-CH3 dimer) of 80 kDa in size and humanized by CDR grafting onto a human germline framework. Multiple humanized variants were evaluated and the lead Mb candidate was selected based on ELISA, flow cytometry and SPR binding properties. The lead Mb, IAb22G3M1 was transiently expressed in CHO cells and purified by Protein L chromatography. PET imaging was performed with desferrioxamine (Df) conjugated IAb22G3M1 radiolabeled with Zr-89 (T1/2 3.3 d). SCID mice bearing subcutaneous HPB-ALL (CD8+ve) or Daudi (CD8-ve) xenografts were serially imaged at 4, 24 and 41 h after i.v. administration of 89Zr-IAb22G3M2 and tissues harvested and counted to determine the biodistribution at the time of sacrifice. The Mb was also evaluated in NOD-SCID-Gamma (NSG) mice that were engrafted with 20 million human PBMCs. In this latter study IAb22G3M1 was radiolabeled with Cu-64 (T1/2 12.7 hrs) following NODAGA conjugation. Mice were imaged at 4 and 7 hrs followed by tissue collection for biodistribution. NSG mice that were not grafted with PBMCs served as experimental controls.
Results: Purification of IAb22G3M1 yielded a product that migrated at the expected MW of 80 kDa with very low ( |
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ISSN: | 0008-5472 1538-7445 |
DOI: | 10.1158/1538-7445.AM2014-LB-163 |