Adoptive Cellular Therapies

Cassian Yee

Experiments performed more than 30 years ago, demonstrating that malignant cells could be eradicated in tumor-bearing mice following the transfer of splenocytes from an immune mouse, ignited the imagination of immunologists and oncologists who could observe for the first time that a tangible, measurable component—this bolus of immune cells—under appropriate conditions was necessary and sufficient to eliminate cancer in tumor-bearing hosts. Since that time, immunotherapy investigators have weathered numerous failures and obstacles attempting to recapitulate this success in patients with advanced malignancies, most notably metastatic melanoma. With the advent of molecular tools to dissect the immune response and a greater understanding of the extrinsic and intrinsic signals governing a productive T-cell response to cancer, the development of strategies to enrich, expand, and manipulate tumor-reactive T cells ex vivo has led to a renaissance in the field of adoptive cellular therapy.

In this chapter, early studies in adoptive therapy are first described and following this, two major sections are presented: strategies developed to elicit antigen-specific T cells in vitro and the clinical trials exploiting these strategies for the treatment of patients with cancer.

Early Studies

The field of adoptive therapy involves the use of in vitro-expanded immune effector cells. Early on, the availability of clinical-grade, recombinant interleukin-2 (IL-2), a lymphokine for natural killer (NK) cells and T cells, led to clinical trials using peripheral blood lymphocytes exposed to high doses of IL-2 to expand “activated killer” cells, that is, lymphokine-activated killer (LAK) cells. This approach has now been largely abandoned in favor of tumor-infiltrating lymphocytes or TILs when it was discovered that single cell suspensions of lymphocytes could be collected from tumor samples, expanded with IL-2, and generated as an antitumor response in murine models that was 50 to 100 times more effective on a percell basis than LAK cells. TIL therapy accompanied by high-dose IL-2 initially demonstrated a 50% response rate in patients with metastatic melanoma, but subsequent studies yielded a response rate of 22%.¹ Randomized clinical trials using the TIL regimen in patients with stage III renal cancer and melanoma revealed no differences in overall survival compared with IL-2 alone²,³; however, the subset of patients with Stage III melanoma and no more than one involved lymph node experienced a statistically significant increase in overall survival, a result that was confirmed in a follow-up report 7 years later.⁴ Taken together, these early results suggested that the therapeutic advantage in using TIL for adoptive therapy was marginalized in patients with increasing tumor burden (more advanced stage III disease or stage IV disease). Without the ability to more clearly define the tumor-rejecting T-cell population among the heterogeneous TIL product, the target antigens recognized mechanisms of immune resistance, and the means to overcome them, further advances in the field would be limited. Defining these parameters would require the development of strategies to generate antigen-specific T cells for adoptive therapy.

Generation of Antigen-Specific T Cells for Adoptive Therapy

The isolation and expansion of T cells require first that tumor-associated antigens recognized by T cells be identified. Following this, in vitro conditions to enrich for a population of antigen-specific T cells are presented. These T cells may be derived from the extant repertoire (endogenous specificity) (Fig 33-1) or genetically engineered to express the antigen-specific receptor of interest (redirected specificity).

Tumor Antigens Recognized by T Cells

The first human tumor antigen defined by T-cell recognition was identified by Thierry Boon and colleagues at the Ludwig Institute almost 20 years ago. A T-cell clone was generated that recognized an autologous melanoma cell line. The cDNA library of the tumor line was transfected into antigen-presenting cells (APCs) expressing the restricting allele and screened using the tumor-reactive T-cell clone. The cDNA of target cells sensitized to lysis by the antigen-specific T cell was recovered and sequenced. By this method, the first human T cell-defined tumor antigen was found to be MAGE-A1.⁵,⁶ It was later discovered that in addition to the MAGE-A1 antigen, a number of MAGE-like antigens, including GAGE, BAGE, LAGE, NY-ESO-1, and SSX-1, were also recognized by tumor-reactive T cells. Based on their restricted tissue expression, that is, tumor cells and germinal cells such as testis, fetal ovary, and placenta, these antigens were grouped together into the family of “cancer-testis” or CT antigens.⁷ There are now known to be over 80 CT antigens, several of which are immunogenic to T cells and are expressed in a wide variety of solid and liquid tumors including lung cancer, colorectal cancer, breast cancer, ovarian cancer, and leukemia.

Using a similar approach, the melanoma-associated antigens, tyrosinase, gp100, and MART-1, were also identified as T-cell target antigens.⁸, ⁹, ¹⁰ In this case, their association with pigmentation pathways
found in normal melanocytes represented the first examples of nonmutated “differentiation” antigens serving as immunologic targets for human T cells.¹¹ Since then, other “differentiation” antigens for prostate cancer (PSMA, Kallikrein), colorectal cancer (CEA), and breast cancer (NY-BR-1, mammoglobin) have been identified. Along with differentiation antigens, other nonmutated self-proteins that are found to be over-expressed in tumor cells include Her-2-neu (breast cancer), adipophilin (renal cancer), mesothelin (ovarian cancer, pancreatic cancer), and more “universally” expressed antigens that confer a survival advantage to tumor cells, such as, survivin, telomerase, and WT-1.

FIGURE 33-1 Generation of Antigen-Specific T Cells for Adoptive Therapy. Antigen-specific T cells can be generated in vitro when cocultivated with naturally occurring or artificial APCs. Naturally occurring APCs: autologous DCs, CD40-activated B cells (CD40-B) or EBV-LCL can present endogenous tumor-associated viral antigens (EBV-LCL), peptide, or RNA/DNA-encoded tumor antigen (DC or CD40-B) in the context of native MHC. Artificial APCs include microbeads, insect cells (D. melanogaster), mouse 3T3 fibroblast, or K562 NK cell lines. MHC-Ig dimers bound to microbeads provide antigen-specific trigger for in vitro stimulation. For insect cells, 3T3 and K562 cell lines, plasmid or recombinant viral vectors encoding restricting MHC allele are used. To provide costimulatory signal, artificial APCs are engineered to present CD28 ligands using B7-Ig (microbeads), vector delivery of B7, ICAM, LFA3, OX40L and/or 4-1BBL, or FcR-mediated presentation of anti-CD28. Antigen-driven T cells expanded in the presence of γ-chain receptor cytokines (IL-2, IL-7, IL-15) with iterative cycles of in vitro stimulation may be used for adoptive transfer or further expanded using anti-CD3 antibody-based protocols.

Mutations in genes associated with tumorigenesis, such as those responsible for cell cycling (the cyclin-dependent kinase, CDK4), delivery of mitogenic signals (B-RAF), and apoptosis (CASP-8), represent attractive targets for immunotherapy because of the decreased likelihood for antigen-loss tumor variants developing. Unfortunately, most of these mutations are not highly prevalent among most tumors and appear to exhibit low immunogenicity.

T cells recognize peptide fragments of target antigens presented by self-MHC molecules on the surface of tumor or APCs such as dendritic cells (DCs), B cells, and monocytes. Those peptide fragments recognized by T cells in the context of the MHC complex are the result of internal processing by proteasomes (class I-restricted epitopes) or lyso/phagosome-associated enzymes (class II-restricted epitopes) followed by binding to their respective MHC complexes and surface presentation. The identification of epitopes recognized by tumor-reactive T cells has been a focus of considerable research since such epitope peptides can be used to elicit antigen-specific T cells and track T-cell responses and are amenable to clinical use as readily available GMP grade reagents to sort and collect antigen-specific T cells. For class I-restricted epitopes designed to elicit CD8 T-cell responses, these epitopes are generally nine to ten amino acids in length; for class II-restricted epitopes eliciting CD4 T-cell responses, the epitopes can be 14 to over 20 amino acids long since class II alleles are less stringent and can accommodate overhanging flanking regions.

In cases where a tumor-reactive antigen-specific T-cell clone has been isolated, such a clone can be used to probe overlapping peptides to identify the minimal epitope sequence. For common alleles, the target epitope may be deduced using algorithms that predict the sequence on the basis of consensus motifs based on known binding preferences as well as the predilection of the proteasome for certain cleavage sites. On occasion, splice variants, excision of intervening sequences, and even sequence reversal during antigen processing can lead to unexpected surface presentation of the cognate epitope.¹² Often, the definitive sequence can only be deduced by eluting peptides from surface MHC and subjecting the mix to mass spectrometry.

However, when the tumor-associated epitope is nonmutant, as is frequently the case, the naturally occurring peptide ligand may not engender robust and sustained antitumor CTL responses.¹³,¹⁴ This is a result of immune tolerance mechanisms that suppress or eliminate high-avidity autoreactive T cells.¹⁵ What remains is a low
frequency of tumor-specific T cells or T cells that bear low-avidity T-cell receptors for the cognate tumor antigen.¹⁶, ¹⁷, ¹⁸, ¹⁹ One method to activate and mobilize these rare and low-avidity tumor-specific T cells uses superagonist altered peptide ligands (APL)²⁰,²¹ that deviate from the native peptide sequence by one or more amino acids, to allow for enhanced binding to the restricting MHC molecule²⁰,²¹ or favorable interaction with the T-cell receptor (TCR) of a given tumor-specific T-cell subset. While, superagonist APLs have been identified that generate tumor-reactive T cells and have even been used to elicit desired immune responses in clinical studies,²²,²³ a comprehensive method for identifying superagonist ligands remains to be developed. Furthermore, the use of APLs must address the potential drawbacks, including cross-reactivity, of the induced T cell not only to the wild-type epitope but also to undesirable autoimmune targets and the possibility that APLinduced responses respond with lower avidity to endogenous target antigens. One advantage of adoptive therapy in this respect is the ability to choose ex vivo from among the population of APLinduced T cells that satisfy these criteria by screening against those that recognize normal tissue targets and selecting for those that recognize endogenously expressed tumor target antigens with high avidity.

Endogenous Specificity: Generating Antigen-Specific T Cells from Existing Repertoire

The in vitro isolation and expansion of antigen-specific T cells for adoptive therapy are in essence a recapitulation of the in vivo events of priming and expansion and involve at least four components: An effector population (source of T cells), stimulator cell, TCR ligand, and T-cell growth factor (i.e., lymphokines such as IL-2) (Table 33-1).

TABLE 33.1 Generating antigen-specific T cells for adoptive therapy from endogenous repertoire

Effector	Source of effector	Stimulator cell	TCR ligand	Growth signal
TIL	Infiltrating lymphocytes from tumor	Tumor cells, in situ APCs	Tumor antigen	IL-2 (high dose) required in vitro and in vivo
Antigen specific T-cell lines and clones	T cells from peripheral blood	Autologous DCs, monocytes, CD40-B cells	Peptide, RNA-/DNA-transfected gene product	IL-2
		Artificial APCs (insect cells, K562, beads)	Peptide, RNA-/DNA-transfected gene product, pMHC complex	IL-2, costimulatory receptors
EBV-specific T cells	T cells from peripheral blood	EBV-B cells, peptide, transduced APCs	EBV-B cells, peptides, adenoviral transgene	IL-2 or none
Infiltrating lymphocytes harvested from tumor sites (TIL) are stimulated by in situ tumor cells and APCs cross-presenting tumor antigens; TILs are expanded with high-dose IL-2 in vitro and require high-dose IL-2 in vivo to maintain survival. Antigen-specific T-cell lines and clones are generated by cocultivation with stimulator cells displaying the TCR ligand in the form of APCs pulsed with peptide, APCs expressing RNA- or DNA-transfected gene product, or artificial APCs decorated with pMHC complex. In the case of EBV-specific T cells, autologous EBV-transformed B cells (LCL) that express the immunodominant viral antigens (e.g., EBNA3) for treatment of PTLD serve as effective APCs; generation of T cells recognizing less immunogenic EBV-associated epitopes for the treatment of HD and NPC is stimulated with APCs infected with adenoviral vectors expressing subdominant epitopes (e.g., LMP2). Artificial APCs can be decorated with peptide-MHC complexes (pMHC) to elicit antigen-specific T cells and costimulatory receptors to facilitate T-cell activation and propagation.

Naturally Occurring Antigen-Presenting Cells as Stimulator Cells

Although DCs, under specific conditions, can be tolerogenic, they are generally considered “professional” APCs with specialized stimulatory features, for example, their capacity to up-regulate expression of costimulatory ligands, and Th1-type cytokines such as IL-12. They represent a robust in vitro stimulator population. In one embodiment, an enriched population of DCs can be generated by treatment of adherent mononuclear cells with GM-CSF and IL-4. Following maturation with an immunomodulatory cocktail or conditioned media, DCs can then be loaded with peptide, transfected with RNA or expression plasmid or transduced with recombinant viral vectors expressing the target antigen of interest. When cocultivated with autologous T cells, and a source of cytokines such as IL-2, antigen-specific T cells can be expanded in vitro for downstream applications. Other approaches for generating human dendritic cells include the use of cytokines such as IL-15²⁴,²⁵, toll receptor (TLR) agonists, or CD40 to enhance DC function and the addition of FLT3 ligand to expand DCs in vitro or in vivo prior to peripheral blood mononuclear cell harvest.²²,²⁶

A more readily available source of autologous APCs has also been developed using CD40 ligand to activate and facilitate long-term growth of human B cells.²⁷ These CD40-activated B cells express high levels of costimulatory molecules and when transfected with the target antigen of interest or epitope peptide serve as robust APCs in vivo and in vitro. Furthermore, CD40 activation of malignant B cells can render them effective APCs and provide a direct means of stimulating leukemia-reactive CD4 and CD8 T cells.²⁸

Whatever the source, when stimulator cells are cocultivated with autologous T cells, in vitro expansion of antigen-specific cells
requires a γ-chain receptor cytokine, such as IL-2, and iterative cycles of in vitro restimulation. In contrast to more nonspecific strategies for expanding effectors such as LAK and TIL where high doses of IL-2 (upwards of 6,000 U/mL) are required, low doses of IL-2 (10 to 50 U/mL) are generally sufficient to induce antigen-driven expansion due to the up-regulation of high-affinity IL-2Rα coreceptor following TCR engagement. Other γ-chain receptor cytokines, such as IL-7 and IL-15, can also be used to augment expansion and have been shown to augment a population of memory effector cells. The addition of IL-21 is unique in its capacity to generate antigen-specific T cells displaying elevated levels of CD28 and, in some cases, CD62L, with effector function, and may represent a central memory-like helper-independent CD8⁺ T cells with features of arrested differentiation and enhanced replicative potential.²⁹

Artificial Antigen-Presenting Cells as Stimulator Cells

The use of artificial APCs addresses some of the obstacles to using autologous mononuclear cells by providing a convenient ex vivo source of stimulator cells, a product that is more likely to be uniform in its physical and functional properties and the flexibility of expressing desired antigen-specific and/or costimulatory receptors.

Mouse fibroblast (3T3) lines and insect cells (D. melanogaster) can be engineered to express the appropriate HLA (usually the prevalent HLA-A2 allele that presents many of the known tumor-associated antigenic epitopes) as well as costimulatory receptors, B7.1, ICAM-1, and LFA-3, found to be necessary for optimal CD8 T-cell stimulation.³⁰ Pulsed with the desired peptide, these APCs could be used to generate human antigen-specific CTL responses in vitro. Using HLA-A2+ insect cells expressing B7.1 and ICAM, Mitchell et al.³¹ were able to enrich for and expand tyrosinase-specific CTL from peripheral blood of patients with metastatic melanoma to more than 10⁹ cells in the presence of IL-2 and IL-7 for use in a clinical trial of adoptive therapy.

The human NK-susceptible chronic myeloid leukemia (CML) cell line, K562, has also been explored for use as an artificial APC. In the absence of MHC expression, it can be used as a “blank” slate for decorating with the desired TCR ligand and costimulatory molecules. June et al. stably transduced Fc receptors to allow for display of anti-CD3 and anti-CD28 antibodies. When 4-1BBL (CD137L) was coexpressed, optimal, nonspecific in vitro expansion of T cells was achieved. By transfecting K562 with the relevant HLA allele and pulsing with the desired epitope peptide, tumor-associated antigen-specific CD8⁺ T cells could be reliably generated in vitro.³² Engineering aAPCs to express γ-chain receptor cytokines, such as IL-21, led to further enhancement in the qualitative and quantitative expansion of antigen-specific CTL.³³

Acellular products used as artificial APCs include magnetic beads, liposomes, and exosomes. Magnetic beads covalently linked to anti-CD3 and anti-CD28 provide a means to rapidly expand a population of polyclonal T cells and have been used in a number of clinical trials: anti-CD3-/anti-CD28-activated donor lymphocyte infusions (DLI) have led to increased responses posttransplant for CML, non-Hodgkin’s lymphoma (NHL), and myeloma³⁴, ³⁵, ³⁶ compared to untreated DLI. For generating antigen-specific T cells, class I and class II Ig dimers can be attached to the beads to permit exogenous loading of peptides for in vitro stimulation and, as a vaccine reagent, for in vivo expansion of adoptively transferred CTL.³⁷

Ex Vivo Selection and Expansion

By whatever means a population of antigen-specific T cells is generated, the question facing immunotherapy investigators is whether the population will require further expansion or in vitro selection before infusion. For some antigens, by virtue of a preexisting high endogenous frequency³⁸ or an elevated frequency that might be predicted in patients due to a concomitant serologic response,³⁹ application of the above methods to generate antigen-specific T cells in culture may be sufficient to produce a population with adequate antitumor activity. Further expansion may involve the use of ligands or agonist antibodies to TCR (CD3) and/or costimulatory receptors (CD28, 4-1BB, etc.) coupled with irradiated feeder cells and cytokines. In some cases, the population of T cells can be expanded from 100- to 1,000-fold over 2 to 3 weeks.⁴⁰, ⁴¹, ⁴²

If selection is desirable, then reagents to nondestructively identify antigen-specific T cells, for example, peptide-MHC multimers, can be used together with a separation method (i.e., flow cytometric cell sorting). Since the natural ligand for the TCR, the peptide-MHC complex, cannot be used singly as a staining reagent for antigen-specific T cells because of its high dissociation rate, multimerizing pMHC complex to a fluorophore-conjugated molecule in a method first pioneered by the Davis lab permits a robust and sensitive means for not only detecting antigen-specific T cells (in a nondestructive manner) but also for isolating tumor-reactive T cells for downstream analysis or adoptive therapy.⁴³,⁴⁴ Later developments in this field include the novel use of Ig fragments to multimerize pMHC complexes for ex vivo detection,⁴⁵ (and as described above, for in vitro as well as in vivo stimulation) and the creation of class II tetramers to identify and select for antigen-specific CD4 T cells.⁴⁶ Global approaches to the selection of antigen-specific T cells exploit functional antigen-driven properties such as cytokine production and surface marker up-regulation. A bispecific antibody binding to a constitutively expressed T-cell surface marker (e.g., CD45) linked to an antibody that captures a secreted cytokine, such as IFN-γ, permits selection of viable antigen-activated IFN-γ+T cells when a second detection antibody to IFN-γ is used.⁴⁷ Alternatively, among surface markers that are up-regulated during antigen recognition (CD25, CD69, 4-1BB/CD137), CD137 surface up-regulation correlates most strongly with TCR ligation and can be used to identify and sort for antigen-specific CD8 T cells.⁴⁸

Redirected Specificity: T-Cell Receptor and Chimeric Antigen Receptor Gene Therapy

Factors limiting the application of adoptive T-cell therapy to a broader pool of patients and diseases include the difficulty in reproducibly isolating high-affinity T cells recognizing tumor-associated antigens and the time and resource-intensive process required to generate such cells. These obstacles may be addressed by genetically modifying T cells to express the cognate TCR or a chimeric antibody receptor (CAR) comprised of the extracellular variable chains recognizing antigen fused to a cytoplasmic T cell signaling domain (usually the TCR zeta chain). TCR and CAR gene therapy enable targeting a wider range of tumor-rejection antigens as well as a means to rapidly produce effector cells for adoptive transfer.⁴⁹,⁵⁰

TCR Gene Therapy

For TCR gene therapy, a T-cell clone of desired specificity and high affinity for the presented target epitope must first be isolated so
that its TCR α– and β-chains can be sequenced and cloned into viral vectors for transgene expression.⁵¹ However, forced expression of specific TCR α– and β-chains in T cells can present mispairing opportunities with the endogenous TCR chains. This can lead to decreased expression of the properly paired chains and lower surface levels of functional TCRs on gene-modified T cells, as well as the possibility that mispaired α– and β-chains can give rise to TCRs recognizing undesired autoimmune targets. To reduce mispairing with endogenous chains, the constant region domains of the transferred TCR α– and β– chains can be substituted with their murine counterparts, thereby eliminating pairing with any human constant regions expressed by endogenous chains.⁵² Other strategies include disulfide linkage of the expressed TCR by the addition of a cysteine residue in the α– and β– chain constant domains,⁵³ the engineering of “knob-in-hole” interaction,⁵⁴ fusing of the α– and β– chains directly to CD3 zeta, and silencing of endogenous TCR.⁵⁵,⁵⁶

In addition to the increased flexibility in antigen targeting that TCR gene therapy offers, functional avidity may be achieved by enhancing TCR mobility⁵⁷ or TCR affinity. For the latter, TCR gene sequences were mutagenized and screened using yeast or phage display technology to identify sequences that increase affinity by three logs or more (equivalent to that seen with antibody-antigen interactions).⁵⁸,⁵⁹ While increased TCR affinity has been associated with increased functional T-cell avidity for tumor targets, at least one study has demonstrated that this is not necessarily the case as other factors, such as downstream signaling events may attenuate functional avidity.⁶⁰ Strategically, this may be addressed by modulating T cell signaling pathways in a manner that decreases the activation threshold, the functional equivalent of enhancing T-cell avidity; T cells engineered to down-regulate expression of Cbl-b or SHP-1 display enhanced efficacy when transferred into tumor-bearing mice. Augmenting target interaction to such a degree allows for a robust response to the desired tumor-associated antigens but also warrants heightened awareness of the autoimmune consequences of targeting “over-expressed” self-antigens.

Target antigen redirection with either TCR or CAR gene therapy will require a means to ensure efficient delivery to effector cells. This is usually achieved through the use of retroviral or lentiviral vectors. Retroviral vectors are more commonly available and well suited for large-scale production. However, their propensity to integrate near transcriptional start sites can lead to the potential for insertional mutagenesis⁶¹; in addition, quiescent T cells do not maintain expression of retrovirally driven transgenes which can be a problem since ligation of the TCR is required to initiate the process of cell cycling. Lentiviral vectors hold several advantages in this regard; they have a lowered likelihood of insertional mutagenesis and an increased infection and expression efficiency among quiescent T cells. A nonviral strategy that utilizes transposon technology has been shown to be highly efficient (>40%) in transfecting resting primary T cells and is potentially scalable for clinical use.⁶² Ultimately, the choice of a delivery vector will depend on reproducibility of high-efficiency transfection and cognate TCR function, as well as regulatory and safety considerations including integration site bias and transgene copy number.

Once transfected, a high level of transgene expression and translation in human T cells must be achieved. For viral vectors, empirical studies demonstrate that promoters such as MSCV U3, EF-1 alpha, and SSFV offer favorable transcriptional profiles, especially when combined with other elements, for example, a nuclear translocation sequence, codon optimization, and strategies that enhance RNA stabilization.⁵⁵ Persistence of expression in quiescent and activated cells must also be evaluated when comparing among these different delivery vectors and flanking sequences.

Chimeric Antibody Receptor

CARs are comprised of an extracellular portion, the variable fragment (scFv) of a tumor-antigen specific antibody coupled to an intracellular domain delivering a T-cell activation signal (CD3 zeta). These CARs allow the targeting of surface proteins, generally with much higher affinity than TCRs (in the nanomolar vs micromolar range), and, in an MHC-unrestricted fashion, activate effector function through the intracellular signaling domain.⁶³ CARs have been developed targeting antigens expressed by several epithelial tumors⁶⁴; however, first-generation CARs signaling through CD3 zeta alone are relatively short lived with limited proliferative capacity. In clinical trials, first-generation CARs for the treatment of neuroblastoma targeting CD171,⁶⁵ B-cell lymphomas targeting CD20,⁶⁶ ovarian cancer targeting the folate receptor⁶⁷, and renal cell cancer targeting carbonic anhydrase⁶⁸ have provided modest responses limited by short duration of in vivo persistence and an absent second signal.

Subsequent generations incorporated dual and triple signaling domains fused to the antibody receptor so that costimulatory signals such as CD137, ICOS, CD28, and OX40⁶⁹ could be delivered solely through antibody engagement without requiring tumors to express the corresponding ligand on its cell surface. These double and triple fusion receptors exhibited increased IL-2 production, enhanced proliferative capacity, and in vivo effector function.

Adoptive T Cell Therapy for the Treatment of Malignant Diseases

Hematopoietic Malignancies

Allogeneic hematopoietic stem cell transplantation (HSCT) represents the earliest evidence of T cell-mediated antitumor immunity in humans when studies documented that a graft versus leukemia (GVL) effect was greatest in patients developing graft versus host disease (GVHD) and weakest in those receiving T cell-depleted marrow cells.⁷⁰ This observation was ultimately exploited in the development of nonmyeloablative HSCT as a means of eliminating regimen-related toxicities of host conditioning by establishing donor T-cell chimerism to mediate engraftment and eradicate residual leukemic cells.⁷¹ DLI early on were shown to be instrumental in mediating a GVL effect and remain in use as effective therapy for specific indications in the allogeneic transplant setting.

Donor Lymphocyte Infusions

DLI can induce durable complete remissions in the majority of patients (60% to 75%) with CML relapsing after allogeneic stem cell transplantation and for a subset of patients relapsing with acute lymphoblastic leukemia, lymphoma, and myeloma although results were somewhat less encouraging.⁷², ⁷³, ⁷⁴, ⁷⁵ Strategies to augment DLI for posttransplant relapse include the use of anti-CD3/CD28 beads to activate T cells before transfer.³⁶ By comparison to standard DLI, activated donor lymphocytes were significantly more effective against acute leukemias and lymphoma (>50% long-term
remissions), without excessive GVHD. For patients receiving cord blood transplants, anti-CD3/CD28 provides a feasible means to expand the small cord blood T-cell population for DLI.⁷⁶,⁷⁷ For patients with myeloma receiving an allogeneic HSCT, immunizing the donor against patient-specific myeloma idiotype, which was predicted by preclinical studies in murine models to enhance transferred immunity, has shown promising results in clinical trials.⁷⁸, ⁷⁹, ⁸⁰

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