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Generation of Therapeutic T Cells for Prostate Cancer
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology, Clinical Immunology. (Gen och immunoterapigruppen)
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The work presented herein focuses on the activation of the adaptive immune system in order to develop T cell-based immunotherapy for viral infections and cancer. The main goal was to identify and activate viral or tumoral antigen-specific T cells by using different identification, isolation and stimulation techniques. One such approach was that we modified dendritic cells (DCs) with an adenoviral vector encoding the full length pp65 antigen from cytomegalovirus (CMV). Through strategic modification techniques we demonstrate that it is possible to obtain DCs presenting antigen-specific peptides both on major histocompatibility complex (MHC) class I and MHC class II molecules for simultaneous CD8+ and CD4+ T cell activation. We also demonstrate that it is possible to generate prostate antigen-specific CD8+ T cells from a naïve repertoire of T cells by using DCs and HLA-A2-restricted peptides derived from prostate tumor-associated antigens or by using an adenoviral vector encoding the full length prostate tumor-associated antigen STEAP. We further demonstrate that CD8+ T cells directed against several prostate-specific peptide epitopes can be found in peripheral blood and in the prostate tumor area of prostate cancer patients. Furthermore, we have characterized a number of prostate-derived cell lines in terms of HLA haplotype and tumor-association antigen expression. We concluded that our methods for generating T cells restricted to CMV antigen have the ability to be applied for adoptive T cell transfer to patients with CMV disease and that prostate antigen-specific T cells can be found within prostate cancer patients, which enables future development of immunotherapeutic strategies for prostate cancer.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2009. , p. 59
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 447
Keywords [en]
Prostate cancer, T cells, dendritic cells, peptides, cytomegalovirus, adenovirus, immunotherapy
National Category
Immunology in the medical area
Research subject
Clinical Immunology
Identifiers
URN: urn:nbn:se:uu:diva-100506ISBN: 978-91-554-7491-1 (print)OAI: oai:DiVA.org:uu-100506DiVA, id: diva2:210534
Public defence
2009-05-26, Rudbecksalen, Rudbecklaboratoriet C11, Dag Hammarskjölds väg 20, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2009-05-05 Created: 2009-04-01 Last updated: 2018-01-13Bibliographically approved
List of papers
1. Strategic use of an adenoviral vector for rapid and efficient ex vivo-generation of cytomegalovirus pp65-reactive cytolytic and helper T cells
Open this publication in new window or tab >>Strategic use of an adenoviral vector for rapid and efficient ex vivo-generation of cytomegalovirus pp65-reactive cytolytic and helper T cells
2008 (English)In: British Journal of Haematology, ISSN 0007-1048, E-ISSN 1365-2141, Vol. 141, no 2, p. 188-199Article in journal (Refereed) Published
Abstract [en]

Cytomegalovirus (CMV) reactivation can cause severe complications for transplant patients. Such patients can be protected against CMV-associated diseases through reconstitution of donor-derived CMV-reactive cytolytic and helper T cells. We have developed a strategic protocol for efficient simultaneous generation of CMV-reactive CD8+ and CD4+ T cells ex vivo. The protocol uses peripheral blood lymphocytes (PBLs), antigen-modified mature dendritic cells (DCs) generated in only 3 days and an adenoviral vector encoding the CMV pp65 antigen (Adpp65) both as an endogenous and exogenous source of antigen. PBLs stimulated once with Adpp65-transduced DCs (endogenously expressed pp65) resulted in preferential activation and expansion of pp65-specific CD8+ T cells while PBLs stimulated with DCs pulsed with cell lysate from Adpp65-transduced autologous monocytes (exogenously expressed pp65) yielded pp65-specific CD4+ T cells. Stimulation with double-modified DCs efficiently activated and expanded cytolytic and helper T cells simultaneously. The frequency of T cells producing interferon (IFN)-γ in response to pp65 increased after one stimulation on average 9.6-fold to 4.3% for CD8+ T cells and 25.8-fold to 6.5% for CD4+ T cells. This implies that sufficient number of pp65-specific cytolytic and helper T cells for adoptive transfer may be obtained in only two weeks.

Keywords
Dendritic cells, T lymphocytes, Cytomegalovirus, Adenoviral vector, pp65
National Category
Microbiology in the medical area Immunology in the medical area
Research subject
Clinical Immunology
Identifiers
urn:nbn:se:uu:diva-99908 (URN)10.1111/j.1365-2141.2008.07026.x (DOI)000254189100005 ()18307565 (PubMedID)
Available from: 2009-03-24 Created: 2009-03-23 Last updated: 2018-01-13Bibliographically approved
2. Characterization of human prostate and breast cancer cell lines for experimental T cell-based immunotherapy
Open this publication in new window or tab >>Characterization of human prostate and breast cancer cell lines for experimental T cell-based immunotherapy
Show others...
2007 (English)In: The Prostate, ISSN 0270-4137, E-ISSN 1097-0045, Vol. 67, no 4, p. 389-395Article in journal (Refereed) Published
Abstract [en]

BACKGROUND. In order to develop experimental immunotherapy for prostate and breast cancer it is of outmost importance to have representative target cell lines that through human leukocyte antigen (HLA) class I molecules present relevant levels of peptides from tumor-associated antigens for cytotoxic T lymphocyte (CTL) recognition. METHODS. We sequenced the HLA-A and HLA-B loci of eight commonly used prostate and breast cancer cell lines and analyzed the surface expression of HLA-ABC, HLA-DR, CD40, CD80, CD86, and CD54 by flow cytometry. We also analyzed the cell lines for mRNA expression from 25 genes reported to be specifically or preferentially expressed by prostate cells. RESULTS. Among the analyzed cell lines we found that LNCaP, PC-346C and MCF-7 are HLA-A*0201 positive. However, the HLA-A2 expression level is low and only MCF-7 upregulates HLA-A2 in response to IFN-γ stimulation. MCF-7 also expresses high levels of CD54, which further improve its value as a CTL target cell line. On the other hand, LNCaP and PC-346C express 25 and 23 out of 25 prostate-related genes, respectively, while MCF-7 expresses 16 out of 25 genes. CONCLUSIONS. None of the analyzed prostate cancer cell lines are optimal CTL target cells. However, MCF-7 could in many cases be used as a complement to HLA-A*0201 positive prostate cancer cells. The LNCaP and PC-346C cell lines are rich sources of prostate-related antigens that may be valuable for cancer vaccine development.

Keywords
Cell line, Tumor associated antigens, HLA, immunotherapy
National Category
Medical and Health Sciences
Research subject
Clinical Immunology
Identifiers
urn:nbn:se:uu:diva-99914 (URN)10.1002/pros.20498 (DOI)000244510000006 ()17219382 (PubMedID)
Available from: 2009-03-24 Created: 2009-03-23 Last updated: 2017-12-13Bibliographically approved
3. Identification of prostate infiltrating lymphocytes and activation of prostate antigen-specific T cells isolated from prostate cancer patients
Open this publication in new window or tab >>Identification of prostate infiltrating lymphocytes and activation of prostate antigen-specific T cells isolated from prostate cancer patients
(English)Manuscript (Other academic)
Abstract [en]

Although prostate antigen-specific CD8+ T lymphocytes have been found both in peripheral blood and in tumor area of prostate cancer patients there has not yet been any adequate adoptive T lymphocyte transfer trial for prostate cancer patients. Methods for efficient generation of large number of prostate antigen-specific T cells are still lacking. In the present study we isolate and expand prostate infiltrating lymphocytes (PILs) from resected prostates by fine needle aspiration of patients with localized prostate cancer. By using HLA-A*0201-restricted tetramers, specific for the prostate tumor-associated antigen epitopes TARP(P5L)4-13, STEAP262-270 and PSA53-61, we were able to identify prostate antigen-specific CD8+ PILs in 5 out of 5 patients. Most frequent were STEAP262-270-specific T cells (5/5). Furthermore, by using fast-matured dendritic cells (DCs) transduced with an adenoviral vector encoding the STEAP antigen we were able to generate CD8+ T cells from peripheral blood of prostate cancer patients, for three HLA-A*0201-restricted STEAP epitopes simultaneously by one stimulation only. If combined, an effective protocol for generation of prostate antigen-specific T cells may be developed.

National Category
Immunology in the medical area
Research subject
Clinical Immunology
Identifiers
urn:nbn:se:uu:diva-100510 (URN)
Available from: 2009-04-07 Created: 2009-04-01 Last updated: 2018-01-13Bibliographically approved

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