In vivooptical imaging of human lymphoma xenograft using a library-derived peptidomimetic against α4β1 integrin

Li Peng,Ruiwu Liu,Mirela Andrei,Kit S Lam,Wenwu Xiao

doi:10.1158/1535-7163.mct-07-0575

Li Peng, Ruiwu Liu + Show 3 more

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https://doi.org/10.1158/1535-7163.mct-07-0575

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Abstract

Increasing literature suggests that cell adhesion molecule alpha4beta1 integrin plays a pivotal role in autoimmune diseases and cancer development. Noninvasive visualization of alpha4beta1 integrin in vivo will facilitate the understanding of its involvement in disease progression and development of targeted therapies. Due to the lack of high-affinity targeting ligands, molecular imaging of alpha4beta1 integrin is much less explored than that of alphavbeta3 and alphavbeta5 integrins. We have recently reported using the one bead-one compound combinatorial library method to identify a peptidomimetic, LLP2A, that preferentially binds to activated alpha4beta1 integrin. Here, we described the use of LLP2A-Cy5.5 conjugate as an in vivo optical imaging probe in a human lymphoma xenograft model. This univalent LLP2A-Cy5.5 conjugate retained the binding activity and specificity to alpha4beta1 integrin as shown by cell binding assays using alpha4beta1-positive Molt-4 T-leukemia cells. The subcutaneous Molt-4 tumor was clearly visualized from 1 to 24 h after tail vein injection of the conjugate. Direct imaging and confocal microscopic examination of excised tumors and organs confirmed the accumulation of LLP2A in tumors and revealed very little or no uptake in normal organs except for lymph nodes. Kidney uptake was high when the whole organ was scanned but it was negative when examined microscopically, suggesting that LLP2A bound to the renal tubules loosely. Tumor uptake of LLP2A-Cy5.5 conjugate was blocked by excess unlabeled LLP2A. This study showed that the combinatorial chemical library-derived peptidomimetic LLP2A can be easily developed into an optical imaging probe for noninvasively monitoring of activated alpha4beta1 integrin in vivo.

Highlights

Integrins are heterodimeric transmembrane proteins crucial for cell-cell, cell-matrix, and cell-pathogen interactions and play important roles in autoimmune diseases, cancer metastasis, tumor angiogenesis, and viral infections (1 – 3)
Specificity of LLP2A-Cy5.5 to A4B1 integrin To determine whether Cy5.5 conjugation had any effect on the binding characteristics of LLP2A, we measured the IC50 of LLP2A-Cy5.5 in a competitive Molt-4 cell adhesion assay
LLP2A-Cy5.5 blocked the interaction between a4h1 integrin on Molt-4 cells and CS-1 peptide of fibronectin, which is the natural ligand for a4h1 integrin, in a concentration-dependent manner

Summary

Introduction

Integrins are heterodimeric transmembrane proteins crucial for cell-cell, cell-matrix, and cell-pathogen interactions and play important roles in autoimmune diseases, cancer metastasis, tumor angiogenesis, and viral infections (1 – 3). Integrin a4h1 is found to express on leukemias, lymphomas, melanomas, and sarcomas [16]. It facilitates tumor cell extravasation [16, 17], prevents apoptosis of malignant B-chronic lymphocytic leukemia cells [19], plays important roles in the drug resistance of both multiple myeloma [20] and acute myelogenous leukemia [21], and is an excellent therapeutic target for malignant lymphoid cancers [24, 26]. A4h1 integrin is much less explored in in vivo imaging due to the lack of high-affinity and highspecificity targeting ligands for the receptor

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