Cytotoxic T Lymphocytes to An Unmutated Tumor Rejection Antigen P1A: Normal Development but Restrained Effector Function In Vivo

Unmutated tumor antigens are chosen as primary candidates for tumor vaccine because of their expression on multiple lineages of tumors. A critical issue is whether unmutated tumor antigens are expressed in normal cells, and if so, whether such expression imposes special restrictions on cytotoxic T lymphocyte (CTL) responses. In this study, we use a transgenic approach to study the development and effector function of T cells specific for P1A, a prototypical unmutated tumor antigen. We report here that although P1A is expressed at low levels in normal tissues, including lymphoid tissues, the P1A-specific transgenic T cells develop normally and remain highly responsive to the P1A antigen. The fact that transgenic expression of P1A antigen in the thymus induces T cell clonal deletion demonstrates that normal hematopoietic cells can process and present the P1A antigen and that P1A-specific T cells are susceptible to clonal deletion. By inference, P1A-specific T cells must have escaped clonal deletion due to low expression of P1A in the thymus. Interestingly, despite the fact that an overwhelming majority of T cells in the T cell receptor for antigen (TCR)–transgenic mice are specific for P1A, these mice are no more resistant to a P1A-expressing plasmocytoma than nontransgenic littermates. Moreover, when the same TCR-transgenic mice were challenged simultaneously with B7-1+ and B7-1− tumors, only B7-1+ tumors were rejected. Therefore, even though P1A can be a tumor rejection antigen, the effector function of P1A-specific CTL is restrained in vivo. These results have important implications for the strategy of tumor immunotherapy.

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A Novel Bioluminescent Mouse Model for Optimization of Adoptive T Cell Therapy

Blood ◽

10.1182/blood.v128.22.2162.2162 ◽

2016 ◽

Vol 128 (22) ◽

pp. 2162-2162

Author(s):

Martin Szyska ◽

Stefanie Herda ◽

Stefanie Althoff ◽

Andreas Heimann ◽

Tra My Dang ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cell Therapy ◽

Tumor Model ◽

Effector Function ◽

Tumor Rejection ◽

Adoptive T Cell Therapy ◽

T Cell Therapy ◽

The Impact

Abstract Adoptive T cell therapy (ATT) is a promising option for the treatment of solid cancers. However, various defense mechanisms acquired by the tumor during evolution prevent transferred T cells (TC) to unfold their full potential. A combination of ATT with accessory therapeutic approaches including checkpoint inhibition and targeted therapy could lift TC inhibition and efficiently shift the immune balance towards tumor rejection. An in-vivo analysis of the impact of combination strategies on the outcome of ATT would greatly enhance the search for an optimal accessory to ATT therapy. We generated the transgenic mouse line BLITC (bioluminescence imaging of T cells) expressing an NFAT (nuclear factor of activated T cell)-dependent Click-beetle luciferase (Na et. al, 2010) and a constitutive Renilla Luciferase, allowing us to monitor migration and activation of transferred TCs in vivo. In order to analyze crucial ATT parameters in a clinically relevant tumor model, BLITC mice were crossed to the two HY-TCR transgenic mice Marilyn (CD4: H-2Ab-Dby) and MataHari (CD8: H-2Db-Uty) to generate TCs that could be monitored for in-vivo infiltration, local activation and rejection of established (> 0,5 cm x 0,5 cm / ≥10 days growth) H-Y expressing MB49 tumors. In order to better reflect the clinical situation, we lymphodepleted tumor-bearing immunocompetent albino B6 mice with fludarabine (FLu) and/or cyclophosphamide (CTX) prior to ATT. Transferred TCs were FACSorted and injected after an optional culture expansion phase. As shown before for freshly injected tumor cells (Perez-Diez, 2007), we observed a superior response of tumor-antigen specific CD4+ TCs compared to CD8+ TCs against established tumors. Whereas 5*106 CD8+ T cells hardly attenuated tumor growth, even as few as 5000 H-Y TCR-transgenic CD4+ T cells rejected tumors in most mice, depending on the lymphodepleting treatment (Figure A - remission rates in parentheses). Tumor infiltration and activation of adoptively transferred TCs was monitored in-vivo by the respective bioluminescent reporters. Around day 4 and 6, CD4+ TCs migrated from tumor-draining lymph nodes into the tumor environment and persisted until rejection. Interestingly, activation of CD4+ TCs was only transient (between days 4 and 7) in all mice, independent of therapy outcome (in Figure B shown for refractory tumor). Whereas loss of activation signal during remission was correlated with tumor clearance and decline of effector function, in refractory tumors it suggests a rapid inactivation of infiltrating TCs by the tumor microenvironment. Our data indicate that the failure of tumor rejection is not caused by impaired peripheral expansion or tumor homing but rather by inhibition of TC effector function. Responsible mechanisms and counter-acting therapeutic interventions are the focus of ongoing studies. In summary, the BLITC reporter system facilitates analysis of therapeutic parameters for ATT in a well-established solid tumor model. Using BLITC mice for transduction with TCR or CAR expression cassettes could allow rapid monitoring of on-target as well as undesired off-target effects in virtually any tumor setting. Future experiments will focus on the beneficial effects of combination treatments on the activation of adoptively transferred TCs. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

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Inhibitory Ligands CD200, CD270, CD274 and CD276 Are Expressed On Eμ-TCL1 Transgenic Mouse Splenocytes and Are of Potential Relevance to Impaired T-Cell Function in Vivo

Blood ◽

10.1182/blood.v120.21.313.313 ◽

2012 ◽

Vol 120 (21) ◽

pp. 313-313

Author(s):

Fabienne McClanahan ◽

Cristina Ghirelli ◽

Paul Greaves ◽

John C. Riches ◽

Rita Coutinho ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Transgenic Mice ◽

Synapse Formation ◽

Effector Function ◽

Immune Synapse ◽

Blocking Antibodies ◽

Cell Effector Function

Abstract Abstract 313 Background: We have previously demonstrated that CD4 and CD8 T-cells from CLL patients show profound dysfunctions in multiple gene pathways, including the actin cytoskeleton, which impairs the formation of functional immunologic synapses between T cells and APCs. Functional screening assays on Mec-1 cells have identified CD200, CD270, CD274, and CD276 as inhibitory ligands which induce impaired actin synapse formation in both allogeneic and autologous T cells. We also demonstrated that the Eμ-TCL1 transgenic mouse model of CLL closely resembles the T-cell defects observed in humans, validating it as a valuable preclinical tool to examine changes in the microenvironment alongside the development of leukaemia. The aim of the current study is to investigate the role of CD200, CD270, CD274, and CD276 in the Eμ-TCL1 model. Methods: We used multiparameter flow cytometry to establish the expression of inhibitory ligands on CD19+/CD5+ unpurified splenocytes from Eμ-TCL1 mice on both the C57Bl/6 (B6) and the C3HB6-F1 background and compared this to unpurified splenocytes from age matched wild-type (WT) controls of the respective coisogenic strain. Results: A total of 19 leukemic Eμ-TCL1 (n=10 C57Bl/6 and n=9 C3HB6-F1 background) and 11 WT mice (n=6 C57Bl/6 and n=5 C3HB6-F1 background) were examined. CD19+/CD5+ CLL cells constituted 92% (range 62–97%) of the DAPI-negative lymphocyte population. On CD19+/CD5+ CLL cells, CD274 (mean 98% ± SEM 0.4) and CD200 (mean 84% ± SEM 2.9 were uniformly strongly expressed, while CD270 (mean 74% ± SEM 4.7) and CD276 (mean 50% ± SEM 6.6) showed a weaker and more diverse expression, with no significant differences between the two backgrounds (all p>.05). Similar expression patterns were observed in Eμ-TCL1 mice with spontaneously occurring CLL and transplanted transgenic mice, with no differences between spontaneous and induced CLL (all p>.05). We then compared transgenic CD19+/CD5+ CLL cells to the WT CD19+ and the WT CD19+/CD5+ B1a-like cell population. Eμ-TCL1 CLL splenocytes showed a significant higher expression of CD274 and CD276 compared to expression on WT CD19+ (p<.0001, p=.00349) splenocytes. When compared to WT B1a-like splenocytes, only CD274 was significantly higher expressed (p<.0001). To clarify the impact of genetic strain, B6 and C3HB6-F1 were investigated separately: transgenic mice on the B6 background showed significantly higher expression of CD274 compared to WT B6 CD19+ (p=.0015) and WT B6 B1a-like (p<.0001) splenocytes. In contrast, transgenic mice on the C3HB6-F1 background showed a significant higher expression of CD274 and CD276 compared to WT CD19+ (p=.0002, p=.00354) and WT B1a-like (p=.0005, p=.00384) splenocytes. These patterns substantiate differences of the expression of inhibitory ligands between the WT strains, but of note, these were not mirrored in TCL1 mice. In previous experiments, we used the Eμ-TCL1 model to investigate the polarization of F-actin and phosphotyrosine at the immune synapse between splenic autologous T-cells and APCs and subsequent effector function. Age-matched WT mice had a significantly higher accumulation than transgenic mice. To assess the functional role of inhibitory ligands, knock-down experiments using lentiviral shRNA and blocking antibodies are currently under way to assess if this restores immune synapse formation and T cell effector function in vivo. Conclusions: The inhibitory ligands CD200, CD270, CD274 and CD276 are expressed in vivo and appear to be of functional relevance for the anti-cancer immune response. They therefore represent attractive targets to restore T-cell effector function, which might be achieved by gene therapy approaches and blocking antibodies. Disclosures: Gribben: Celgene: Honoraria.

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WDFY4 is required for cross-presentation in response to viral and tumor antigens

Science ◽

10.1126/science.aat5030 ◽

2018 ◽

Vol 362 (6415) ◽

pp. 694-699 ◽

Cited By ~ 69

Author(s):

Derek J. Theisen ◽

Jesse T. Davidson ◽

Carlos G. Briseño ◽

Marco Gargaro ◽

Elvin J. Lauron ◽

...

Keyword(s):

T Cells ◽

Dendritic Cells ◽

Tumor Antigens ◽

Critical Role ◽

Tumor Rejection ◽

Interleukin 12 ◽

Cross Presentation ◽

Histocompatibility Complex ◽

Crispr Screen

During the process of cross-presentation, viral or tumor-derived antigens are presented to CD8+ T cells by Batf3-dependent CD8α+/XCR1+ classical dendritic cells (cDC1s). We designed a functional CRISPR screen for previously unknown regulators of cross-presentation, and identified the BEACH domain–containing protein WDFY4 as essential for cross-presentation of cell-associated antigens by cDC1s in mice. However, WDFY4 was not required for major histocompatibility complex class II presentation, nor for cross-presentation by monocyte-derived dendritic cells. In contrast to Batf3–/– mice, Wdfy4–/– mice displayed normal lymphoid and nonlymphoid cDC1 populations that produce interleukin-12 and protect against Toxoplasma gondii infection. However, similar to Batf3–/– mice, Wdfy4–/– mice failed to prime virus-specific CD8+ T cells in vivo or induce tumor rejection, revealing a critical role for cross-presentation in antiviral and antitumor immunity.

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Antigen-independent activation of naive and memory resting T cells by a cytokine combination.

Journal of Experimental Medicine ◽

10.1084/jem.180.3.1159 ◽

1994 ◽

Vol 180 (3) ◽

pp. 1159-1164 ◽

Cited By ~ 238

Author(s):

D Unutmaz ◽

P Pileri ◽

S Abrignani

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

Interleukin 2 ◽

Effector Function ◽

Necrosis Factor Alpha ◽

Naive T Cells ◽

Naïve T Cells

We investigated whether human resting T cells could be activated to proliferate and display effector function in the absence of T cell receptor occupancy. We report that combination of interleukin 2 (IL-2), tumor necrosis factor alpha, and IL-6 activated highly purified naive (CD45RA+) and memory (CD45RO+) resting CD4+ T cells to proliferate. Under this condition, memory resting T cells could also display effector function as measured by lymphokine synthesis and help for immunoglobulin production by B cells. This novel Ag-independent pathway of T cell activation may play an important role in vivo in recruiting effector T cells at the site of immune response and in maintaining the clonal size of memory T cells in the absence of antigenic stimulation. Moreover, cytokines can induce proliferation of naive T cells without switch to memory phenotype and this may help the maintenance of the peripheral pool of naive T cells.

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IL-7 and IL-21 are superior to IL-2 and IL-15 in promoting human T cell–mediated rejection of systemic lymphoma in immunodeficient mice

Blood ◽

10.1182/blood-2009-09-241398 ◽

2010 ◽

Vol 115 (17) ◽

pp. 3508-3519 ◽

Cited By ~ 118

Author(s):

John C. Markley ◽

Michel Sadelain

Keyword(s):

T Cells ◽

T Cell ◽

Adoptive Transfer ◽

Memory T Cells ◽

Tumor Rejection ◽

Effector Memory ◽

Transfer Model ◽

Human T Cell

Abstract The γc-cytokines are critical regulators of immunity and possess both overlapping and distinctive functions. However, comparative studies of their pleiotropic effects on human T cell–mediated tumor rejection are lacking. In a xenogeneic adoptive transfer model, we have compared the therapeutic potency of CD19-specific human primary T cells that constitutively express interleukin-2 (IL-2), IL-7, IL-15, or IL-21. We demonstrate that each cytokine enhanced the eradication of systemic CD19+ B-cell malignancies in nonobese diabetic/severe combined immunodeficient (NOD/SCID)/γcnull mice with markedly different efficacies and through singularly distinct mechanisms. IL-7– and IL-21–transduced T cells were most efficacious in vivo, although their effector functions were not as enhanced as IL-2– and IL-15–transduced T cells. IL-7 best sustained in vitro T-cell accumulation in response to repeated antigenic stimulation, but did not promote long-term T-cell persistence in vivo. Both IL-15 and IL-21 overexpression supported long-term T-cell persistence in treated mice, however, the memory T cells found 100 days after adoptive transfer were phenotypically dissimilar, resembling central memory and effector memory T cells, respectively. These results support the use of γc-cytokines in cancer immunotherapy, and establish that there exists more than 1 human T-cell memory phenotype associated with long-term tumor immunity.

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Therapeutic Antimyeloma Effect of Dendritic-Tumor Cells Hybrids Transduced with Adenovirus Encoding CD40L.

Blood ◽

10.1182/blood.v112.11.1709.1709 ◽

2008 ◽

Vol 112 (11) ◽

pp. 1709-1709

Author(s):

Eva Alvarez ◽

Esther Moga ◽

Jorge Sierra ◽

Javier Briones

Keyword(s):

T Cell ◽

Immune Responses ◽

Tumor Cells ◽

Antitumor Effect ◽

Tumor Antigens ◽

Recombinant Adenovirus ◽

Lymphoid Tissues ◽

Term Survival ◽

Mhc Ii

Abstract Dendritic cells (DCs) are the main antigen presenting cells and play a pivotal role in the stimulation of T-cell immune responses. DCs cultured in the presence of a single tumor antigen can elicit an immune response against tumor cells expressing that antigen. However, simultaneous use of several tumor antigens may be advantageous since polyclonal activation of T cells against different tumor antigens may be a better approach to eradicate tumor cells. In this sense, fusions of dendritic and tumor cells (FCs) show a broad spectrum of tumor antigens, both known and unidentified, to be presented by class I and II MHC. Although prophylactic vaccines were successful in murine models, the results in the therapeutic setting have been unsatisfactory. We hypothesised that enhancing costimulation of FCs would help to break tumor tolerance once the tumor is established. To this purpose, we transduced FCs with a recombinant adenovirus encoding CD40L (AdvCD40L or AdvGFP as control) and we studied the therapeutic antitumoral effect of the administration of FC-CD40L in a murine model of myeloma. DCs obtained from day 7-bone marrow cultures of Balb/c mice were fused with tumor cells, a syngeneic murine myeloma cell line (4TOO). FCs hybrids were generated with PEG and selected after culturing in HAT medium plus GM-CSF for 7 days. FC were quantified by determining the percentage of cells that coexpress specific DC (CD11c) and tumor markers (CD138). Mean fusion efficiency was 30% (20–40%) and FCs expressed moderate levels of CD80, CD83, CD86, CD54, CD40 and MHC II and did not express CD40L. FC-CD40L showed a significant increase of expression of costimulatory molecules (CD80, CD86, CD54, and MHC II) compared to FC-GFP (p=0.011). Moreover, in a syngeneic mixed lymphocyte reaction, FC-CD40L induced a two-fold higher T-cell proliferation than FC-GFP or FC alone. In addition, FC-CD40L had improved migration to lymphoid tissues, preferentially to spleen, in comparison with FC-GFP (2.8% versus 1.6%). The antitumor effect of FC-CD40L was analyzed in vivo. Mice (n=10 per group) were injected i.v. with 2.5×105 tumor cells and treated with irradiated FC, FC-GFP or FC-CD40L (1×106 cells each) on days 2, 6 and 10 after tumor challenge. 40% of mice treated with FC-CD40L had long-term survival (>120 days). In contrast, all of mice treated with FC or FC-GFP died between days 25 and 35 (p=0.012). In parallel, treatment with mixed cells (not fused DC+ tumor cells), mix transduced with AdvGFP, or mix transduced with AdvCD40L did not provide any significant antitumor effect. We conclude that FCs transduced with AdvCD40L better stimulate in vitro and in vivo immune responses than FC alone and may provide a new strategy for treating patients with multiple myeloma or lymphoma.

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Enforced Expression of Lin28b Drives Development of Peripheral T Cell Lymphoma In Vivo

Blood ◽

10.1182/blood.v118.21.1392.1392 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1392-1392 ◽

Cited By ~ 1

Author(s):

Sarah H Beachy ◽

Masahiro Onozawa ◽

Yang Jo Chung ◽

Christopher Slape ◽

Sven Bilke ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Transgenic Mice ◽

Cell Lymphoma ◽

Regulatory Elements ◽

T Cell Lymphoma ◽

Effector Memory ◽

Peripheral T Cell Lymphoma ◽

Chronic Inflammatory Condition

Abstract Abstract 1392 Lin-28 was first identified as a heterochronic gene in C. elegans and is expressed at high levels during early larval stages with dramatically decreased expression in subsequent developmental stages. Interestingly, recent work demonstrated that induced expression of LIN28 can reprogram human fibroblasts to acquire pluripotency (in combination with NANOG, OCT4 and SOX2), providing additional evidence for a positive correlation between LIN28 expression and maintenance of a more immature and stem cell-like developmental state. Lin28a and Lin28b, the mammalian homologues of lin-28, have been implicated in oncogenic transformation in a variety of tumor types, in part by their ability to promote the degradation of the let-7 family of microRNAs (miRs), which are known to target oncogenes such as Myc and Ras. We recently noted that Lin28b was markedly overexpressed in hematopoietic tissues of NUP98-HOXD13 (NHD13) transgenic mice that develop a myelodysplastic syndrome (MDS) that subsequently transforms to acute myeloid leukemia (AML) or pre-T lymphoblastic leukemia/lymphoma (pre-T LBL). In order to elucidate the contribution of Lin28b overexpression to the differentiation block and malignant phenotype observed in the NHD13 mice, we designed a Lin28b transgenic mouse by targeting the expression of the transgene to hematopoietic tissues with Vav regulatory elements. In this model, clinically healthy Lin28b mice exhibited aberrant thymic architecture and retention of thymocytes that was correlated with peripheral blood lymphopenia (a 2.6-fold decrease in circulating lymphocytes). The lymphopenia was principally due to decreased numbers of CD4+ and CD8+ cells, although there was a significant increase in the number of CD4 and CD8 effector memory T cells (CD44hiCD62Llo) compared with wild type mice. Additionally, deep sequencing of thymic miRs from clinically healthy transgenic mice revealed a 2–5-fold downregulation of let-7 family members, including let-7d, g, f, i and miR-98. Importantly, with age, the Lin28b mice developed an aggressive, lethal, peripheral T cell lymphoma (PTCL), characterized by widespread infiltration of parenchymal organs with a mixed infiltrate of inflammatory cells and malignant CD4+ T cells. Clonal Tcrb gene rearrangements were observed in the lymphomas and the malignant cells engrafted and formed tumors in immunodeficient Scid mice. The Lin28b transgenic mice also had clinical signs consistent with a chronic inflammatory condition, such as eosinophilia, anemia, pleural effusions and ascites. The lymphomas overexpressed Il6 and Myc, and activated Nfκb, demonstrating in vivo involvement of a previously reported pathway that links Lin28b expression with inflammation and malignant transformation. Analysis of a publically available dataset indicated that Lin28b was overexpressed by 8-fold in a set of PTCL patient samples compared with activated CD4+ cells. Taken together, these findings demonstrate in vivo evidence for an oncogenic function of Lin28b and provide a model for further study of both the biology and identification of new therapeutic targets for PTCL, a heterogenous disease with poor prognosis. Disclosures: No relevant conflicts of interest to declare.

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New differentiation pathway for double-negative regulatory T cells that regulates the magnitude of immune responses

Blood ◽

10.1182/blood-2006-10-050625 ◽

2006 ◽

Vol 109 (9) ◽

pp. 4071-4079 ◽

Cited By ~ 64

Author(s):

Dong Zhang ◽

Wei Yang ◽

Nicolas Degauque ◽

Yan Tian ◽

Allison Mikita ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Cell Surface ◽

Immune Responses ◽

Lymphoid Tissues ◽

Double Negative ◽

Healthy Humans

Abstract Recent studies have demonstrated that in peripheral lymphoid tissues of normal mice and healthy humans, 1% to 5% of αβ T-cell receptor–positive (TCR+) T cells are CD4−CD8− (double-negative [DN]) T cells, capable of down-regulating immune responses. However, the origin and developmental pathway of DN T cells is still not clear. In this study, by monitoring CD4 expression during T-cell proliferation and differentiation, we identified a new differentiation pathway for the conversion of CD4+ T cells to DN regulatory T cells. We showed that the converted DN T cells retained a stable phenotype after restimulation and that furthermore, the disappearance of cell-surface CD4 molecules on converted DN T cells was a result of CD4 gene silencing. The converted DN T cells were resistant to activation-induced cell death (AICD) and expressed a unique set of cell-surface markers and gene profiles. These cells were highly potent in suppressing alloimmune responses both in vitro and in vivo in an antigen-specific manner. Perforin was highly expressed by the converted DN regulatory T cells and played a role in DN T-cell–mediated suppression. Our findings thus identify a new differentiation pathway for DN regulatory T cells and uncover a new intrinsic homeostatic mechanism that regulates the magnitude of immune responses. This pathway provides a novel, cell-based, therapeutic approach for preventing allograft rejection.

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Melanocyte Destruction after Antigen-Specific Immunotherapy of Melanoma

Journal of Experimental Medicine ◽

10.1084/jem.192.11.1637 ◽

2000 ◽

Vol 192 (11) ◽

pp. 1637-1644 ◽

Cited By ~ 322

Author(s):

Cassian Yee ◽

John A. Thompson ◽

Patrick Roche ◽

David R. Byrd ◽

Peter P. Lee ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cd8 T Cells ◽

Specific Immunotherapy ◽

Tumor Antigens ◽

Normal Tissues ◽

Cell Clones ◽

Expression Evidence ◽

Tumor Biopsies

Current strategies for the immunotherapy of melanoma include augmentation of the immune response to tumor antigens represented by melanosomal proteins such as tyrosinase, gp100, and MART-1. The possibility that intentional targeting of tumor antigens representing normal proteins can result in autoimmune toxicity has been postulated but never demonstrated previously in humans. In this study, we describe a patient with metastatic melanoma who developed inflammatory lesions circumscribing pigmented areas of skin after an infusion of MART-1–specific CD8+ T cell clones. Analysis of the infiltrating lymphocytes in skin and tumor biopsies using T cell–specific peptide–major histocompatibility complex tetramers demonstrated a localized predominance of MART-1–specific CD8+ T cells (>28% of all CD8 T cells) that was identical to the infused clones (as confirmed by sequencing of the complementarity-determining region 3). In contrast to skin biopsies obtained from the patient before T cell infusion, postinfusion biopsies demonstrated loss of MART-1 expression, evidence of melanocyte damage, and the complete absence of melanocytes in affected regions of the skin. This study provides, for the first time, direct evidence in humans that antigen-specific immunotherapy can target not only antigen-positive tumor cells in vivo but also normal tissues expressing the shared tumor antigen.

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A Detailed Analysis of Parameters Supporting the Engraftment and Growth of Chronic Lymphocytic Leukemia Cells in Immune-Deficient Mice

Frontiers in Immunology ◽

10.3389/fimmu.2021.627020 ◽

2021 ◽

Vol 12 ◽

Author(s):

Piers E. M. Patten ◽

Gerardo Ferrer ◽

Shih-Shih Chen ◽

Jonathan E. Kolitz ◽

Kanti R. Rai ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Intraperitoneal Administration ◽

Human Condition ◽

Lymphocytic Leukemia ◽

Lymphoid Tissues ◽

Quiescent State

Patient-derived xenograft models of chronic lymphocytic leukemia (CLL) can be created using highly immunodeficient animals, allowing analysis of primary tumor cells in an in vivo setting. However, unlike many other tumors, CLL B lymphocytes do not reproducibly grow in xenografts without manipulation, proliferating only when there is concomitant expansion of T cells. Here we show that in vitro pre-activation of CLL-derived T lymphocytes allows for a reliable and robust system for primary CLL cell growth within a fully autologous system that uses small numbers of cells and does not require pre-conditioning. In this system, growth of normal T and leukemic B cells follows four distinct temporal phases, each with characteristic blood and tissue findings. Phase 1 constitutes a period during which resting CLL B cells predominate, with cells aggregating at perivascular areas most often in the spleen. In Phase 2, T cells expand and provide T-cell help to promote B-cell division and expansion. Growth of CLL B and T cells persists in Phase 3, although some leukemic B cells undergo differentiation to more mature B-lineage cells (plasmablasts and plasma cells). By Phase 4, CLL B cells are for the most part lost with only T cells remaining. The required B-T cell interactions are not dependent on other human hematopoietic cells nor on murine macrophages or follicular dendritic cells, which appear to be relatively excluded from the perivascular lymphoid aggregates. Notably, the growth kinetics and degree of anatomic localization of CLL B and T cells is significantly influenced by intravenous versus intraperitoneal administration. Importantly, B cells delivered intraperitoneally either remain within the peritoneal cavity in a quiescent state, despite the presence of dividing T cells, or migrate to lymphoid tissues where they actively divide; this dichotomy mimics the human condition in that cells in primary lymphoid tissues and the blood are predominately resting, whereas those in secondary lymphoid tissues proliferate. Finally, the utility of this approach is illustrated by documenting the effects of a bispecific antibody reactive with B and T cells. Collectively, this model represents a powerful tool to evaluate CLL biology and novel therapeutics in vivo.

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