Cellular Therapy Of Cancer: Development Of Gene Therapy Based Approaches

Development of Gene Therapy Based Approaches

Robert E Hawkins

• Publisher: WSPC
• Year: 2014
• eBook ISBN: 9789814618533
• Topic: Biological Sciences
• Subtopic: Science General
• Index: Biological Sciences

Cancer research has progressed enormously in recent years. This review volume will address recent findings in the area of T-cell therapy for cancer, including use of tumour infiltrating lymphocytes (TILs) as a therapy for melanoma, choice of target antigens, advances in engineered receptors, methods of gene transfer to T cells, review of cell processing methods and clinical trial design. Written by leadings scientists in the field, this up-to-date review on cancer research will be an important reference source to the researchers and healthcare professionals in the field.

Contents:

• Challenges of T-Cell Therapy (Naomi Taylor, Anna Mondino and Balbino Alarcon)
• Gene Transfer into T Cells (David Gilham and Hinrich Abken)
• TCR-Engineered T Cells (Reno Debets and Ton Schumacher)
• T-Bodies: Antibody-Based Engineered T-Cell Receptors (John Bridgeman, Andreas A Hombach, David Gilham, Zelig Eshhar and Hinrich Abken)
• T-Cell Engineering and Expansion — GMP Issues (Cor Lamers and Ryan Guest)
• Clinical Trial Design (Robert Hawkins, John Haanen and Fiona Thistlethwaite)

Readership: Cancer researchers, oncologists and healthcare professionals.

Chapter 1

Challenges of T-Cell Therapy

Naomi Taylor, Anna Mondino and Balbino Alarcon

1.1 T-Cell Homeostasis

The size of the lymphocyte pool is critical for an efficient adaptive immune system; it must be sufficiently diverse to detect and destroy a wide range of potential pathogens, yet there is only limited physical space in the body to house all of these cells. Throughout life, the peripheral T-cell pool is constantly submitted to transient fluctuations in cell numbers and subset composition, yet it has long been known that regulatory mechanisms are at work to maintain a stable equilibrium. The clonal expansion of antigen-specific T cells during an immune response is subsequently followed by mass apoptosis of effector cells; only a few survive to become long-lived memory cells. Moreover, when the T-cell pool is severely depleted, the remaining T cells sense the increased availability of peripheral “space” and undergo proliferation to reconstitute steady-state T-cell numbers.¹ The overall size and composition of the peripheral T-cell pool remains fairly constant and is regulated at several levels. Thymic export seeds the peripheral tissues with newly generated T cells. Survival and proliferation through contact with self-antigens and cytokines subsequently allows their maintenance; finally, mechanisms to induce T-cell death are necessary to provide a stable equilibrium. Homeostasis, from the Greek words for “same” and “steady”, refers to the self-regulating process that maintains the stability of a biological system; in this case, the preservation of T-cell numbers and composition over time.

In patients with tumors, the presence of a “full” lymphocyte compartment can have negative consequences for T-cell-mediated tumor immunity in that the tumor-specific T cells are not able to expand in an optimal manner. This problem is further compounded by the nature of the tumor antigen. Indeed, most identified tumor antigens represent proteins that are also expressed in non-transformed tissues and as such, mechanisms responsible for self-tolerance have ample opportunity to negatively shape the T-cell repertoire available for tumor rejection. Moreover, optimal responses by the available T cells may be further affected by tumor-induced tolerance.^2,3

However, it appears that some of these problems can be surmounted under conditions of lymphopenia, a state in which there is a reduced number of circulating lymphocytes. In lymphopenic conditions, there is an increased homeostatic proliferation of T lymphocytes which results in the expansion of conventional T cells, in the apparent absence of antigenic stimulation.⁴ Notably, this non-specific proliferation has been shown to enhance the reactivity of T cells and this effect has been exploited to augment the responsiveness of T lymphocytes in cancer patients. Indeed, the ability of adoptively-transferred autologous tumor-infiltrating lymphocytes to mount an effective response in melanoma patients has been shown to be significantly enhanced by first rendering the patients lymphopenic.⁵ The mechanisms by which T cells differentiate and bypass the mechanisms of peripheral tolerance have not yet been completely clarified.

1.2 Lymphopenia and Immune Responsiveness to Self-Antigens

How does lymphopenia trigger anti-self-responses? The response is likely to be multifaceted but it is important to note that lymphopenia may result in an imbalance between effector and regulatory T cells (Tregs), with a preferential loss of the latter.^6,7 However, the loss of Tregs cannot in itself explain self-reactivity since their absence in lymphoreplete adult animals does not result in autoimmunity.⁸ It is also important to note that conditioning regimens, including irradiation or chemotherapy, as well as the absence of Tregs may result in the generalized activation of antigen presenting cells (APCs), and specifically dendritic cells (DCs).^9,10

Lymphopenia also induces the antigen-independent activation of potentially autoreactive T cells. Naïve T cells proliferate under acute lymphopenic conditions in response to the same factors that promote their survival, the interleukin-7 (IL-7) cytokine and T-cell receptor (TCR) engagement with self-peptide/major histocompatibility complex (MHC) complexes. The homeostatic proliferation of both naïve CD4 and CD8 T cells requires low affinity interactions with self-peptide/MHC complexes.^11–13 This proliferation is accompanied by a direct differentiation into memory-like T cells in the apparent absence of antigenic stimulation. Indeed, these cells are functionally and phenotypically similar to bonafide memory cells. Interestingly, it has been shown that memory-like T cells are less prone to tolerization than naïve cells, most likely due to their less stringent requirements for activation (reviewed in Refs. 14 and 15).

1.3 CD4 and CD8 T-Cell Differentiation States

Adoptive cellular immunotherapy aims to eradicate malignancies by the transfer of reactive T cells. These T cells can be derived from the tumor-bearing host and all of the following have been tested: (i) tumor-infiltrating lymphocytes, (ii) tumor-primed lymph node cells, (iii) in vitro-sensitized peripheral blood lymphocytes, (iv) and tumor-specific T cells generated in vitro by TCR/CAR (chimeric antigen receptor) gene transfer.^16–18 One important question, that had been heatedly debated until recently, revolved around the relative persistence and efficacies of naïve, central memory, and effector memory T cells for adoptive cell therapy (ACT). Central memory cells, with high proliferative and reconstituting capacity, maintain the ability to relocate to secondary lymphoid organs and are involved in recall responses. In contrast, terminally differentiated effectors, present primarily in peripheral tissues, are endowed with immediate effector function upon antigen reencounter but show poor proliferative and reconstituting abilities.¹⁹ This is of importance as terminally differentiated effector cells might exert potent, but transient anti-tumor activity, while central memory T cells may confer a more durable T-cell immunity required for long-lasting immunosurveillance. Increasing evidence indicates that while anti-tumor effector T cells, obtained after multiple rounds of ex vivo stimulation, possess highly effective in vitro cytotoxic activity, they are less effective than naïve or memory-like T cells in vivo.^20–22

Notably, it has recently been found, in mice as well as in macaques, that antigen-specific CD8+ T cell derived from central memory T cells (T_CM) show significantly higher long-term persistence/survival than effector memory T cells (T_EM).^20,22 Moreover, these T_CM mediate superior anti-tumor immunity than T_EM upon adoptive transfer into tumor-bearing mice^20,22 and that stem cell memory subsets result in a more pronounced tumor regression.²³ It is also important to note that under conditions where a tumor-specific TCR is introduced ex vivo into either naïve or central memory T cells (by retroviral-mediated gene transfer), infusion of the former results in a significantly more robust anti-tumor response.²¹ Finally, it has recently been shown that a long-lived human memory T-cell population, characterized as CD45RO(−), CCR7(+), CD45RA(+), CD62L(+), CD27(+), CD28(+), and IL-7Ralpha(+), has a significantly enhanced capacity to reconstitute immunodeficient hosts and mediate an anti-tumor response in a humanized mouse model.²⁴

Considerable research efforts have thus been devoted to the development of ex vivo activation strategies that preserve a naïve- or central memory-“like” phenotype. While prolonged IL-2 signaling promotes the terminal effector differentiation of CD8 T cells,^25,26 homeostatic gamma chain cytokines (IL-7, IL-15, IL-21) have proven efficacious in sustaining T-cell proliferation and in vivo antitumor function without favoring terminal differentiation.^27,28 Of note, it has been shown that IL-7, IL-15, and IL-21 support the generation of CD8+ cells with superior therapeutic activity as compared to IL-2 alone.^29–33 IL-7 and IL-15 have also been exploited for ex vivo gene transfer; these cytokines sustain sufficient activation of lymphocytes in the absence of TCR stimulation, rendering them susceptible to lentiviral infection without favoring cell differentiation.^34–37 As an alternative to the use of recombinant cytokines, genetic modification of T cells with a vector expressing a homeostatic cytokine such as IL-15 or IL-21, may favor the generation of lymphocytes with a central memory phenotype.³⁸

CD8 T cells clearly play a critical role in anti-tumor immunity, but it is important to note that there is also an important function for CD4 T cells.^39–41 Moreover, CD4 lymphocytes have recently been shown to play a direct role in adoptive immunotherapy.^42,43 Upon retroviral transfer of a tumor-specific receptor into T lymphocytes, it was demonstrated that gene-modified CD8 as well as CD4 T cells are required for an efficient immune response. The most potent antitumor responses were observed when the ratio of gene-modified CD4:CD8 T cells was 1:1.⁴⁴ Moreover, we and others have found that adoptively transferred memory-like CD8 T cells are subject to peripheral cross-tolerance; breakdown of this tolerance and differentiation of CD8 T cells into effectors require CD4 T cell help.^45,46 Thus, the cooperativity between CD4 and CD8 T cells needs to be taken into account in ACT protocols.

The independent role of CD4 T cells in tumor eradication was demonstrated in a model...

Table of contents

• Cover Page
• Title Page
• Copyright Page
• Contents
• Chapter 1 Challenges of T-Cell Therapy
• Chapter 2 Gene Transfer into T Cells
• Chapter 3 TCR-Engineered T cells
• Chapter 4 T-Bodies: Antibody-Based Engineered T-Cell Receptors
• Chapter 5 T-Cell Engineering and Expansion — GMP Issues
• Chapter 6 Clinical Trial Design