Date of Award

2-23-2009

Document Type

Open Access Thesis

Degree Name

Medical Doctor (MD)

First Advisor

Richard Edelson

Abstract

Transimmunization (TI), a novel modification of the widely used immunotherapy extracorporeal photopheresis (ECP), induces conversion of processed monocytes into cells expressing phenotypic and functional features of dendritic antigen presenting cells (DC). To further characterize TI-induced DC, we analyzed differential gene expression in the monocyte/DC population after TI treatment. Because ECP, the therapy upon which TI is based, has the unique capacity to induce both anti-cancer immune responses in cutaneous T cell lymphoma (CTCL) patients and tolerogenic responses in graft-versus-host disease (GVHD), we studied TI-induced gene expression changes in both of these patient populations as well as in healthy normal control individuals with the goal of fully characterizing the gene expression profile(s) induced by TI. Peripheral blood leukocytes from 6 patients (3 patients with CTCL and 3 patients with GVHD) were procured prior to ECP, immediately after ECP, and following TI processing, and were then enriched for monocytes/DC. RNA was extracted and gene expression compared using Affymetrix total human genome microarrays to analyze 39,000 genes. Differential gene expression was considered as a ≥2-fold change and P-value ≤0.05. TI induced significant upregulation of genes associated with DC maturation including: DC-LAMP, CD80, CD40, and Decysin. In addition, TI induced down-regulation of monocyte genes such as CD33 and CD36. These changes in gene expression were seen in both CTCL and GVHD patients, suggesting that TI is capable of mediating DC differentiation regardless of disease process. However, some genes (e.g. IL-19, Tryptophan 2,3-dioxygenase) were differentially expressed after TI only in GVHD patients, while others, (e.g. heat-shock proteins 70, 27, and 40) were differentially expressed only in CTCL patients. Our microarray findings were confirmed by quantitative realtime PCR on patient samples as well as on samples from healthy normal controls that underwent the TI procedure. Analysis of the microarray data using GeneGo pathway analysis software demonstrated that the chemokines and adhesion signaling pathway was significantly involved in the mechanism of both ECP and TI, suggesting a crucial role for cell adhesion in these therapies. Taken together, our gene expression and pathway data suggest that TI activates specific signaling cascades that lead to activation and up-regulation of mature DC genes. Our results support the use of TI as a method of generating mature dendritic cells for immunotherapy.

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This is an Open Access Thesis.

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