Erik Ames

All Publications

• Unexpectedly low tacrolimus concentrations attributed to inappropriately labeled water container from the instrument manufacturer. Clinical chemistry and laboratory medicine Ames, E., Bowen, R. A. 2022

  View details for DOI 10.1515/cclm-2022-0204

  View details for PubMedID 35405044

• Killers on the loose: Immunotherapeutic strategies to improve NK cell-based therapy for cancer treatment. International review of cell and molecular biology Dunai, C., Ames, E., Ochoa, M. C., Fernandez-Sendin, M., Melero, I., Simonetta, F., Baker, J., Alvarez, M. 2022; 370: 65-122

  Abstract

  Natural killer (NK) cells are innate lymphocytes that control tumor progression by not only directly killing cancer cells, but also by regulating other immune cells, helping to orchestrate a coordinated anti-tumor response. However, despite the tremendous potential that this cell type has, the clinical results obtained from diverse NK cell-based immunotherapeutic strategies have been, until recent years, rather modest. The intrinsic regulatory mechanisms that are involved in the control of their activation as well as the multiple mechanisms that tumor cells have developed to escape NK cell-mediated cytotoxicity likely account for the unsatisfactory clinical outcomes. The current approaches to improve long-term NK cell function are centered on modulating different molecules involved in both the activation and inhibition of NK cells, and the latest data seems to advocate for combining strategies that target multiple aspects of NK cell regulation. In this review, we summarize the different strategies (such as engineered NK cells, CAR-NK, NK cell immune engagers) that are currently being used to take advantage of this potent and complex immune cell.

  View details for DOI 10.1016/bs.ircmb.2022.04.001

  View details for PubMedID 35798507

• Polytypic T-cell prolymphocytic leukemia. Blood Ames, E., Silva, O. 2021; 137 (15): 2125

  View details for DOI 10.1182/blood.2021010694

  View details for PubMedID 33856441

• Bortezomib Augments Natural Killer Cell Targeting of Stem-Like Tumor Cells CANCERS Luna, J. I., Grossenbacher, S. K., Sturgill, I. R., Ames, E., Judge, S. J., Bouzid, L. A., Darrow, M. A., Murphy, W. J., Canter, R. J. 2019; 11 (1)

  Abstract

  Tumor cells harboring stem-like/cancer stem cell (CSC) properties have been identified and isolated from numerous hematological and solid malignancies. These stem-like tumor cells can persist following conventional cytoreductive therapies, such as chemotherapy and radiotherapy, thereby repopulating the tumor and seeding relapse and/or metastasis. We have previously shown that natural killer (NK) cells preferentially target stem-like tumor cells via non- major histocompatibility complex (MHC) restricted mechanisms. Here, we demonstrated that the proteasome inhibitor, bortezomib, augments NK cell targeting of stem cell-like tumor cells against multiple solid human tumor-derived cancer lines and primary tissue samples. Mechanistically, this was mediated by the upregulation of cell surface NK ligands MHC class I chain-related protein A and B (MICA and MICB) on aldehyde dehydrogenases (ALDH)-positive CSCs. The increased expression of MICA and MICB on CSC targets thereby enhanced NK cell mediated killing in vitro and ex vivo from both human primary tumor and patient-derived xenograft samples. In vivo, the combination of bortezomib and allogeneic NK cell adoptive transfer in immunodeficient mice led to increased elimination of CSCs as well as tumor growth delay of orthotopic glioblastoma tumors. Taken together, our data support the combination bortezomib and NK transfer as a strategy for both CSC targeting and potentially improved outcomes in clinical cancer patients.

  View details for DOI 10.3390/cancers11010085

  View details for Web of Science ID 000457233300008

  View details for PubMedID 30646520

  View details for PubMedCentralID PMC6356940

• Sclerosing mucoepidermoid carcinoma with eosinophilia: Cytologic characterization of a rare distinct entity in the thyroid DIAGNOSTIC CYTOPATHOLOGY Ames, E., Campbell, M. J., Afify, A., Krane, J. F., Huang, E. C. 2018; 46 (7): 632-635

  Abstract

  Sclerosing mucoepidermoid carcinoma with eosinophilia (SMECE) is an extremely rare thyroid carcinoma with limited cytologic descriptions in the literature. Here, we present a 52-year-old woman with a 3.9 cm thyroid nodule. Fine-needle aspiration smears consisted of a highly cellular specimen with tumor cells in isolated patterns and solid squamoid nests. Tumor cells had round to oval nuclei, prominent nucleoli, smooth nuclear contours, and moderate amounts of dense cytoplasm. In addition to the polymorphous population of lymphocytes, the background contained a striking abundance of eosinophils. The subsequent right thyroidectomy showed histologic features diagnostic for SMECE.

  View details for DOI 10.1002/dc.23914

  View details for Web of Science ID 000435082600015

  View details for PubMedID 29479842

• Immune targeting of cancer stem cells in gastrointestinal oncology JOURNAL OF GASTROINTESTINAL ONCOLOGY Canter, R. J., Grossenbacher, S. K., Ames, E., Murphy, W. J. 2016; 7: S1-S10

  Abstract

  The cancer stem cell (CSC) hypothesis postulates that a sub-population of quiescent cells exist within tumors which are resistant to conventional cytotoxic/anti-proliferative therapies. It is these CSCs which then seed tumor relapse, even in cases of apparent complete response to systemic therapy. Therefore, therapies, such as immunotherapy, which add a specific anti-CSC strategy to standard cytoreductive treatments may provide a promising new direction for future cancer therapies. CSCs are an attractive target for immune therapies since, unlike chemotherapy or radiotherapy, immune effector cells do not specifically require target cells to be proliferating in order to effectively kill them. Although recent advances have been made in the development of novel systemic and targeted therapies for advanced gastro-intestinal (GI) malignancies, there remains an unmet need for durable new therapies for these refractory malignancies. Novel immunotherapeutic strategies targeting CSCs are in pre-clinical and clinical development across the spectrum of the immune system, including strategies utilizing adaptive immune cell-based effectors, innate immune effectors, as well as vaccine approaches. Lastly, since important CSC functions are affected by the tumor microenvironment, targeting of both cellular (myeloid derived suppressor cells and tumor-associated macrophages) and sub-cellular (cytokines, chemokines, and PD1/PDL1) components of the tumor microenvironment is under investigation in the immune targeting of CSCs. These efforts are adding to the significant optimism about the potential utility of immunotherapy to overcome cancer resistance mechanisms and cure greater numbers of patients with advanced malignancy.

  View details for DOI 10.3978/j.issn.2078-6891.2015.066

  View details for Web of Science ID 000375061800001

  View details for PubMedID 27034806

  View details for PubMedCentralID PMC4783622

• NK Cells Preferentially Target Tumor Cells with a Cancer Stem Cell Phenotype JOURNAL OF IMMUNOLOGY Ames, E., Canter, R. J., Grossenbacher, S. K., Mac, S., Chen, M., Smith, R. C., Hagino, T., Perez-Cunningham, J., Sckisel, G. D., Urayama, S., Monjazeb, A. M., Fragoso, R. C., Sayers, T. J., Murphy, W. J. 2015; 195 (8): 4010-4019

  Abstract

  Increasing evidence supports the hypothesis that cancer stem cells (CSCs) are resistant to antiproliferative therapies, able to repopulate tumor bulk, and seed metastasis. NK cells are able to target stem cells as shown by their ability to reject allogeneic hematopoietic stem cells but not solid tissue grafts. Using multiple preclinical models, including NK coculture (autologous and allogeneic) with multiple human cancer cell lines and dissociated primary cancer specimens and NK transfer in NSG mice harboring orthotopic pancreatic cancer xenografts, we assessed CSC viability, CSC frequency, expression of death receptor ligands, and tumor burden. We demonstrate that activated NK cells are capable of preferentially killing CSCs identified by multiple CSC markers (CD24(+)/CD44(+), CD133(+), and aldehyde dehydrogenase(bright)) from a wide variety of human cancer cell lines in vitro and dissociated primary cancer specimens ex vivo. We observed comparable effector function of allogeneic and autologous NK cells. We also observed preferential upregulation of NK activation ligands MICA/B, Fas, and DR5 on CSCs. Blocking studies further implicated an NKG2D-dependent mechanism for NK killing of CSCs. Treatment of orthotopic human pancreatic cancer tumor-bearing NSG mice with activated NK cells led to significant reductions in both intratumoral CSCs and tumor burden. Taken together, these data from multiple preclinical models, including a strong reliance on primary human cancer specimens, provide compelling preclinical evidence that activated NK cells preferentially target cancer cells with a CSC phenotype, highlighting the translational potential of NK immunotherapy as part of a combined modality approach for refractory solid malignancies.

  View details for DOI 10.4049/jimmunol.1500447

  View details for Web of Science ID 000362968100053

  View details for PubMedID 26363055

  View details for PubMedCentralID PMC4781667

• Out-of-Sequence Signal 3 Paralyzes Primary CD4(+) T-Cell-Dependent Immunity IMMUNITY Sckisel, G. D., Bouchlaka, M. N., Monjazeb, A. M., Crittenden, M., Curti, B. D., Wilkins, D. C., Alderson, K. A., Sungur, C. M., Ames, E., Mirsoian, A., Reddy, A., Alexander, W., Soulika, A., Blazar, B. R., Longo, D. L., Wiltrout, R. H., Murphy, W. J. 2015; 43 (2): 240-250

  Abstract

  Primary T cell activation involves the integration of three distinct signals delivered in sequence: (1) antigen recognition, (2) costimulation, and (3) cytokine-mediated differentiation and expansion. Strong immunostimulatory events such as immunotherapy or infection induce profound cytokine release causing "bystander" T cell activation, thereby increasing the potential for autoreactivity and need for control. We show that during strong stimulation, a profound suppression of primary CD4(+) T-cell-mediated immune responses ensued and was observed across preclinical models and patients undergoing high-dose interleukin-2 (IL-2) therapy. This suppression targeted naive CD4(+) but not CD8(+) T cells and was mediated through transient suppressor of cytokine signaling-3 (SOCS3) inhibition of the STAT5b transcription factor signaling pathway. These events resulted in complete paralysis of primary CD4(+) T cell activation, affecting memory generation and induction of autoimmunity as well as impaired viral clearance. These data highlight the critical regulation of naive CD4(+) T cells during inflammatory conditions.

  View details for DOI 10.1016/j.immuni.2015.06.023

  View details for Web of Science ID 000360101200008

  View details for PubMedID 26231116

  View details for PubMedCentralID PMC4770886

• Enhanced targeting of stem-like solid tumor cells with radiation and natural killer cells ONCOIMMUNOLOGY Ames, E., Canter, R. J., Grossenbacher, S. K., Mac, S., Smith, R. C., Monjazeb, A. M., Chen, M., Murphy, W. J. 2015; 4 (9): e1036212

  Abstract

  Natural killer (NK) cells are innate lymphocytes postulated to mediate resistance against primary haematopoietic but not solid tumor malignancies. Cancer stem cells (CSCs) are a small subset of malignant cells with stem-like properties which are resistant to chemo- and radiotherapies and are able to repopulate a tumor after cytoreductive treatments. We observed increased frequencies of stem-like tumor cells after irradiation, with increased expression of stress ligands on surviving stem-like cells. Ex vivo NK cells activated by low dose IL2 in vitro and IL15 in vivo displayed an increased ability to target solid tumor stem-like cells both in vitro and in vivo after irradiation. Mechanistically, both upregulation of stress-related ligands on the stem-like cells as well as debulking of non-stem populations contributed to these effects as determined by data from cell lines, primary tumor samples, and most relevant patient derived specimens. In addition, pretreatment of tumor-bearing mice with local radiation prior to NK transfer resulted in significantly longer survival indicating that radiation therapy in conjunction with NK cell adoptive immunotherapy targeting stem-like cancer cells may offer a promising novel radio-immunotherapy approach in the clinic.

  View details for DOI 10.1080/2162402X.2015.1036212

  View details for Web of Science ID 000360239800016

  View details for PubMedID 26405602

  View details for PubMedCentralID PMC4570100

• Anti-proliferative but not anti-angiogenic tyrosine kinase inhibitors enrich for cancer stem cells in soft tissue sarcoma BMC CANCER Canter, R. J., Ames, E., Mac, S., Grossenbacher, S. K., Chen, M., Li, C., Borys, D., Smith, R. C., Tellez, J., Sayers, T. J., Monjazeb, A. M., Murphy, W. J. 2014; 14: 756

  Abstract

  Increasing studies implicate cancer stem cells (CSCs) as the source of resistance and relapse following conventional cytotoxic therapies. Few studies have examined the response of CSCs to targeted therapies, such as tyrosine kinase inhibitors (TKIs). We hypothesized that TKIs would have differential effects on CSC populations depending on their mechanism of action (anti-proliferative vs. anti-angiogenic).We exposed human sarcoma cell lines to sorafenib, regorafenib, and pazopanib and assessed cell viability and expression of CSC markers (ALDH, CD24, CD44, and CD133). We evaluated survival and CSC phenotype in mice harboring sarcoma metastases after TKI therapy. We exposed dissociated primary sarcoma tumors to sorafenib, regorafenib, and pazopanib, and we used tissue microarray (TMA) and primary sarcoma samples to evaluate the frequency and intensity of CSC markers after neoadjuvant therapy with sorafenib and pazopanib. Parametric and non-parametric statistical analyses were performed as appropriate.After functionally validating the CSC phenotype of ALDHbright sarcoma cells, we observed that sorafenib and regorafenib were cytotoxic to sarcoma cell lines (P < 0.05), with a corresponding 1.4 - 2.8 fold increase in ALDHbright cells from baseline (P < 0.05). In contrast, we observed negligible effects on viability and CSC sub-populations with pazopanib. At low doses, there was progressive CSC enrichment in vitro after longer term exposure to sorafenib although the anti-proliferative effects were attenuated. In vivo, sorafenib improved median survival by 11 days (P < 0.05), but enriched ALDHbright cells 2.5 - 2.8 fold (P < 0.05). Analysis of primary human sarcoma samples revealed direct cytotoxicity following exposure to sorafenib and regorafenib with a corresponding increase in ALDHbright cells (P < 0.05). Again, negligible effects from pazopanib were observed. TMA analysis of archived specimens from sarcoma patients treated with sorafenib demonstrated significant enrichment for ALDHbright cells in the post-treatment resection specimen (P < 0.05), whereas clinical specimens obtained longitudinally from a patient treated with pazopanib showed no enrichment for ALDHbright cells (P > 0.05).Anti-proliferative TKIs appear to enrich for sarcoma CSCs while anti-angiogenic TKIs do not. The rational selection of targeted therapies for sarcoma patients may benefit from an awareness of the differential impact of TKIs on CSC populations.

  View details for DOI 10.1186/1471-2407-14-756

  View details for Web of Science ID 000343258800001

  View details for PubMedID 25301268

  View details for PubMedCentralID PMC4200119

• Regulating ES or Induced Pluripotent Stem Cells by Innate Lymphoid Cells Reply TRANSPLANTATION Ames, E., Murphy, W. J. 2014; 98 (5): E39-E40

  View details for DOI 10.1097/TP.0000000000000345

  View details for Web of Science ID 000341854800004

  View details for PubMedID 25171530

  View details for PubMedCentralID PMC4343530

• Treatment of chronic graft-versus-host disease with bortezomib BLOOD Pai, C., Chen, M., Mirsoian, A., Grossenbacher, S. K., Tellez, J., Ames, E., Sun, K., Jagdeo, J., Blazar, B. R., Murphy, W. J., Abedi, M. 2014; 124 (10): 1677-1688

  Abstract

  Chronic graft-versus-host disease (cGVHD) following allogeneic hematopoietic stem cell transplantation (HSCT) has emerged as a predominant complication following HSCT and has a distinct etiology. We and others have previously demonstrated that bortezomib, a proteasome inhibitor, can prevent but not treat acute GVHD in mice. To assess the effects of bortezomib on cGVHD, a mouse minor histocompatibility antigen-mismatched strain combination was used to mimic clinical cGVHD sclerodermatous pathogenesis and phenotype. Treatment of ongoing cGVHD with bortezomib ameliorated cutaneous lesions, which were also associated with a reduction in total numbers of germinal center B cells and lower B-cell activating factor gene expression levels in cutaneous tissues. Importantly, lymphoma-bearing mice receiving allogeneic HSCT with bortezomib preserved graft-versus-tumor (GVT) effects. Based on these animal studies, we initiated an intrapatient dose escalation clinical trial in patients with extensive steroid-intolerant, dependent, or resistant cGVHD. Marked clinical improvement was observed in patients, which was also associated with reductions of peripheral B cells and minimal toxicity. These results indicate that bortezomib can be of significant use in the treatment of cGVHD and may also allow for maintenance of GVT. This trial was registered at www.clinicaltrials.gov as #NCT01672229.

  View details for DOI 10.1182/blood-2014-02-554279

  View details for Web of Science ID 000342762300023

  View details for PubMedID 25009225

  View details for PubMedCentralID PMC4155274

• Natural Killer Cell Subsets Differentially Reject Embryonic Stem Cells Based on Licensing TRANSPLANTATION Perez-Cunningham, J., Ames, E., Smith, R. C., Peter, A. K., Nolta, J. A., Murphy, W. J. 2014; 97 (10): 992-998

  Abstract

  Embryonic stem cells (ESC) and induced pluripotent stem cells provide great promise to the future of medicine. Because immune rejection represents a major obstacle to the success of all stem cell-based therapies, many recent studies have sought to determine the key immune mediators involved in ESC rejection. The role of natural killer (NK) cells and specifically the role of NK cell licensing is not well understood in ESC rejection.Mouse or human ESCs were subjected to cytotoxicity assays involving their respective species-matched activated NK cells. Mouse ESCs were then transplanted to allogeneic recipients after depletion of NK cell subsets in the host. ESC engraftment was analyzed by bioluminescent imaging.Depletion of all NK cells in vivo resulted in the greatest amount of ESC engraftment, confirming a role for NK cells in ESC rejection. Importantly, depletion of the Ly49C/I or Ly49G2 NK cell subsets resulted in differential ESC engraftment and rejection. This indicates that NK cell rejection of allogeneic ESC is highly differential based on the presence of licensed NK cells. Blocking NKG2D in vitro resulted in less killing of mESC by allogeneic NK cells, indicating NKG2D is a likely mechanism for NK-mediated killing of mESC.In this study, we show that expression of inhibitory Ly49s correlates with the ability of NK cells to kill murine ESC in an NKG2D-dependent manner. This further suggests that the rejection of similar stem cell transplants in humans will be dependent upon the presence of licensed NK cells.

  View details for DOI 10.1097/TP.0000000000000063

  View details for Web of Science ID 000336745900009

  View details for PubMedID 24704665

• Advantages and clinical applications of natural killer cells in cancer immunotherapy CANCER IMMUNOLOGY IMMUNOTHERAPY Ames, E., Murphy, W. J. 2014; 63 (1): 21-28

  Abstract

  The past decade has witnessed a burgeoning of research and further insight into the biology and clinical applications of natural killer (NK) cells. Once thought to be simple innate cells important only as cytotoxic effector cells, our understanding of NK cells has grown to include memory-like responses, the guidance of adaptive responses, tissue repair, and a delicate paradigm for how NK cells become activated now termed "licensing" or "arming." Although these cells were initially discovered and named for their spontaneous ability to kill tumor cells, manipulating NK cells in therapeutic settings has proved difficult and complex in part due to our emerging understanding of their biology. Therapies involving NK cells may either activate endogenous NK cells or involve transfers of exogenous cells by hematopoietic stem cell transplantation or adoptive cell therapy. Here, we review the basic biology of NK cells, highlighting characteristics which make NK cells particularly useful in cancer therapies. We also explore current treatment strategies that have been used for cancer as well as discuss potential future directions for the field.

  View details for DOI 10.1007/s00262-013-1469-8

  View details for Web of Science ID 000329227700003

  View details for PubMedID 23989217

  View details for PubMedCentralID PMC3880590

• Contrasting Effects of Anti-Ly49A Due to MHC Class I cis Binding on NK Cell-Mediated Allogeneic Bone Marrow Cell Resistance JOURNAL OF IMMUNOLOGY Alvarez, M., Sungur, C. M., Ames, E., Anderson, S. K., Pomeroy, C., Murphy, W. J. 2013; 191 (2): 688-698

  Abstract

  NK subsets have activating and inhibitory receptors that bind MHC-I. Ly49A is a mouse inhibitory receptor that binds with high affinity to H2(d) in both a cis- and trans-manner. Ly49A cis-associations limit trans-interactions with H2(d)-expressing targets as well as mAb binding. We demonstrate that cis-interactions affect mAb effector functions. In vivo administration of anti-Ly49A depleted NK cells in H2(b) but not H2(d) mice. Despite lack of depletion, in vivo treatment with anti-Ly49A reduced NK killing capabilities and inhibited activation, partially due to its agonistic effect. These data explain the previously described in vivo effects on bone marrow allograft rejection observed with anti-Ly49A treatment in H2(d)-haplotype mice. However, prior treatment of mice with poly(I:C) or mouse CMV infection resulted in increased Ly49A expression and Ly49A(+) NK cell depletion in H2(d) mice. These data indicate that, although Ly49 mAbs can exert similar in vivo effects in mice with different MHC haplotypes, these effects are mediated via different mechanisms of action correlating with Ly49A expression levels and can be altered within the same strain contingent on stimuli. This illustrates the marked diversity of mAb effector functions due to the regulation of the level of expression of target Ags and responses by stimulatory incidents such as infection.

  View details for DOI 10.4049/jimmunol.1300202

  View details for Web of Science ID 000321260100021

  View details for PubMedID 23752612

  View details for PubMedCentralID PMC3733663

• Murine natural killer cell licensing and regulation by T regulatory cells in viral responses PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Sungur, C. M., Tang-Feldman, Y. J., Ames, E., Alvarez, M., Chen, M., Longo, D. L., Pomeroy, C., Murphy, W. J. 2013; 110 (18): 7401-7406

  Abstract

  Natural killer (NK) cells show differential functionality based on their capability of binding to self-MHC consistent with licensing. Here we show in vivo confirmation of the physiologic effects of licensing with differential effects of NK subsets on anti-murine cytomegalovirus (anti-MCMV) responses after syngeneic hematopoietic stem cell transplantation (HSCT) or regulatory T-cell (Treg) depletion. After HSCT, depletion of licensed NK cells led to far greater viral loads in target organs early after infection compared with nondepleted and unlicensed depleted mice. There was a preferential expansion of licensed, C-type lectin-like activating receptor Ly49H+ NK cells with increased IFNγ production after infection in nondepleted mice post-HSCT and after Treg depletion. Adoptive transfer of licensed NK subsets into immunodeficient hosts provided significantly greater MCMV resistance compared with transfer of total NK populations or unlicensed subsets. In non-HSCT mice, only concurrent depletion of Tregs or TGF-β neutralization resulted in detection of NK licensing effects. This suggests that licensed NK cells are the initial and rapidly responding population of NK cells to MCMV infection, but are highly regulated by Tregs and TGF-β.

  View details for DOI 10.1073/pnas.1218767110

  View details for Web of Science ID 000318682300063

  View details for PubMedID 23589894

  View details for PubMedCentralID PMC3645578

• IFN-gamma Receptor-Deficient Donor T Cells Mediate Protection from Graft-versus-Host Disease and Preserve Graft-versus-Tumor Responses after Allogeneic Bone Marrow Transplantation JOURNAL OF IMMUNOLOGY Sun, K., Hsiao, H., Li, M., Ames, E., Bouchlaka, M., Welniak, L. A., Hagino, T., Jagdeo, J., Pai, C., Chen, M., Blazar, B. R., Abedi, M., Murphy, W. J. 2012; 189 (4): 2033-2042

  Abstract

  Graft-versus-host disease (GVHD) is a major complication of allogeneic bone marrow transplantation. It has been previously reported that lung GVHD severity directly correlates with the expansion of donor Th17 cells in the absence of IFN-γ. However, the consequence of Th17-associated lung GVHD in the presence of IFN-γ has not been well characterized. In the current study, T cells from IFN-γ receptor knockout (IFN-γR(-/-)) mice, capable of producing IFN-γ but unable to signal in response to IFN-γ, have been used to elucidate further the role of IFN-γ in GVHD. We found the transfer of donor T cells from either IFN-γR(-/-) or IFN-γ knockout (IFN-γ(-/-)) mice resulted in significant increases in donor Th17 cells in the lung. Marked increases in IL-4-producing Th2 cells infiltrating the lungs were also observed in the mice of donor IFN-γR(-/-) T cells. Notably, despite the presence of these cells, these mice did not show the severe immune-mediated histopathological lung injury observed in mice receiving donor IFN-γ(-/-) T cells. Increases in lung GVHD did occur in mice with donor IFN-γR(-/-) T cells when treated in vivo with anti-IFN-γ demonstrating that the cytokine has a protective role on host tissues in GVHD. A survival benefit from acute GVHD was also observed using donor cells from IFN-γR(-/-) T cells compared with control donors. Importantly, tumor-bearing mice receiving IFN-γR(-/-) T cells versus wild-type donor T cells displayed similar graft-versus-tumor (GVT) effects. These results demonstrate the critical role of IFN-γ on host tissues and cell effector functions in GVHD/GVT.

  View details for DOI 10.4049/jimmunol.1102853

  View details for Web of Science ID 000307216000057

  View details for PubMedID 22778394

  View details for PubMedCentralID PMC3509544

• Delineation of antigen-specific and antigen-nonspecific CD8(+) memory T-cell responses after cytokine-based cancer immunotherapy BLOOD Tietze, J. K., Wilkins, D. C., Sckisel, G. D., Bouchlaka, M. N., Alderson, K. L., Weiss, J. M., Ames, E., Bruhn, K. W., Craft, N., Wiltrout, R. H., Longo, D. L., Lanier, L. L., Blazar, B. R., Redelman, D., Murphy, W. J. 2012; 119 (13): 3073-3083

  Abstract

  Memory T cells exhibit tremendous antigen specificity within the immune system and accumulate with age. Our studies reveal an antigen-independent expansion of memory, but not naive, CD8(+) T cells after several immunotherapeutic regimens for cancer resulting in a distinctive phenotype. Signaling through T-cell receptors (TCRs) or CD3 in both mouse and human memory CD8(+) T cells markedly up-regulated programmed death-1 (PD-1) and CD25 (IL-2 receptor α chain), and led to antigen-specific tumor cell killing. In contrast, exposure to cytokine alone in vitro or with immunotherapy in vivo did not up-regulate these markers but resulted in expanded memory CD8(+) T cells expressing NKG2D, granzyme B, and possessing broadly lytic capabilities. Blockade of NKG2D in mice also resulted in significantly diminished antitumor effects after immunotherapy. Treatment of TCR-transgenic mice bearing nonantigen expressing tumors with immunotherapy still resulted in significant antitumor effects. Human melanoma tissue biopsies obtained from patients after topically applied immunodulatory treatment resulted in increased numbers of these CD8(+) CD25(-) cells within the tumor site. These findings demonstrate that memory CD8(+) T cells can express differential phenotypes indicative of adaptive or innate effectors based on the nature of the stimuli in a process conserved across species.

  View details for DOI 10.1182/blood-2011-07-369736

  View details for Web of Science ID 000302141200022

  View details for PubMedID 22251483

  View details for PubMedCentralID PMC3321869

• The Triterpenoid CDDO-Me Promotes Hematopoietic Progenitor Expansion and Myelopoiesis in Mice BIOLOGY OF BLOOD AND MARROW TRANSPLANTATION Ames, E., Harouna, S., Meyer, C., Welniak, L. A., Murphy, W. J. 2012; 18 (3): 396-405

  Abstract

  The synthetic triterpenoid CDDO-Me has been shown to directly inhibit the growth of myeloid leukemias and lends itself to a wide array of therapeutic indications, including inflammatory conditions, because of its inhibition of NF-κB. We have previously demonstrated protection from acute graft-versus-host disease after CDDO-Me administration in an allogeneic bone marrow transplantation model. In the current study, we observed that CDDO-Me promoted myelopoiesis in both naive and transplanted mice. This effect was dose dependent, as high doses of CDDO-Me inhibited myeloid growth in vitro. All lineages (granulocyte macrophage colony-forming unit, BFU-E) were promoted by CDDO-Me. We then compared the effects with granulocyte colony-stimulating factor, a known inducer of myeloid expansion and mobilization from the bone marrow. Whereas both drugs induced terminal myeloid expansion in the spleen, peripheral blood, and bone marrow, granulocyte colony-stimulating factor only induced granulocyte macrophage colony-forming unit precursors in the spleen, while CDDO-Me increased these precursors in the spleen and bone marrow. After sublethal total-body irradiation, mice pretreated with CDDO-Me further displayed an accelerated recovery of myeloid progenitors and total nucleated cells in the spleen. A similar expansion of myeloid and myeloid progenitors was noted with CDDO-Me treatment after syngeneic bone marrow transplantation. Combined, these data suggest that CDDO-Me may be of use posttransplantation to accelerate myeloid recovery in addition to the prevention of graft-versus-host disease.

  View details for DOI 10.1016/j.bbmt.2011.11.013

  View details for Web of Science ID 000301204800008

  View details for PubMedID 22100978

  View details for PubMedCentralID PMC3502000

• Mouse NK cell-mediated rejection of bone marrow allografts exhibits patterns consistent with Ly49 subset licensing BLOOD Sun, K., Alvarez, M., Ames, E., Barao, I., Chen, M., Longo, D. L., Redelman, D., Murphy, W. J. 2012; 119 (6): 1590-1598

  Abstract

  Natural killer (NK) cells can mediate the rejection of bone marrow allografts and exist as subsets based on expression of inhibitory/activating receptors that can bind MHC. In vitro data have shown that NK subsets bearing Ly49 receptors for self-MHC class I have intrinsically higher effector function, supporting the hypothesis that NK cells undergo a host MHC-dependent functional education. These subsets also play a role in bone marrow cell (BMC) allograft rejection. Thus far, little in vivo evidence for this preferential licensing across mouse strains with different MHC haplotypes has been shown. We assessed the intrinsic response potential of the different Ly49(+) subsets in BMC rejection by using β2-microglobulin deficient (β2m(-/-)) mice as donors. Using congenic and allogeneic mice as recipients and depleting the different Ly49 subsets, we found that NK subsets bearing Ly49s, which bind "self-MHC" were found to be the dominant subset responsible for β2m(-/-) BMC rejection. This provides in vivo evidence for host MHC class I-dependent functional education. Interestingly, all H2(d) strain mice regardless of background were able to resist significantly greater amounts of β2m(-/-), but not wild-type BMC than H2(b) mice, providing evidence that the rheostat hypothesis regarding Ly49 affinities for MHC and NK-cell function impacts BMC rejection capability.

  View details for DOI 10.1182/blood-2011-08-374314

  View details for Web of Science ID 000300420900036

  View details for PubMedID 22184406

  View details for PubMedCentralID PMC3286220

• Mouse Ly49G2(+) NK cells dominate early responses during both immune reconstitution and activation independently of MHC BLOOD Barao, I., Alvarez, M., Ames, E., Orr, M. T., Stefanski, H. E., Blazar, B. R., Lanier, L. L., Anderson, S. K., Redelman, D., Murphy, W. J. 2011; 117 (26): 7032-7041

  Abstract

  Natural killer (NK) cell subsets can be defined by the differential expression of inhibitory receptors for MHC class I molecules. Early after congenic HSCT, we found that Ly49G2(high) single-positive NK cells repopulated, displayed an activated phenotype, and were highly cytolytic. Over time, this subset was replaced with NK cells with a normal pattern of Ly49 expression. Treatment of mice with IL-2 also resulted in the rapid expansion of these Ly49G2(high) single-positive NK cells. Only the Ly49g (Klra7) Pro1 transcript was highly induced in both HSCT- and IL-2-treated recipients. MHC-independent expansion of the Ly49G2(+) subset was also observed after Listeria monocytogenes or mouse cytomegalovirus infection. Our data indicate that during reconstitution after HSCT and various activation stimuli, Ly49G2(+) NK cells represent the "first-responder" NK cells, which occur independently of NK-cell licensing via Ly49-MHC interactions. These data suggest that the inhibitory Ly49G2 receptor represents an activation marker on mouse NK cells under various conditions.

  View details for DOI 10.1182/blood-2010-11-316653

  View details for Web of Science ID 000292244000011

  View details for PubMedID 21498673

  View details for PubMedCentralID PMC3143551

• Sensitization of human breast cancer cells to natural killer cell-mediated cytotoxicity by proteasome inhibition CLINICAL AND EXPERIMENTAL IMMUNOLOGY Ames, E., Hallett, W. D., Murphy, W. J. 2009; 155 (3): 504-513

  Abstract

  The proteasome inhibitor, bortezomib, has direct anti-tumour effects and has been demonstrated to sensitize tumour cells to tumour necrosis factor-related apoptosis-inducing ligand-mediated apoptosis. Natural killer (NK) cells are effective mediators of anti-tumour responses, both through cytotoxic granule killing and apoptosis-inducing pathways. We therefore investigated if bortezomib sensitized human breast cancer cells to killing by the human NK cell line, NK-92. Bortezomib was unable to sensitize MDA-231 breast cancer cells to NK cell-mediated killing in short-term in vitro assays. However, bortezomib did cause these cells to up-regulate apoptosis-related mRNA as well as death receptors on the cell surface. In a long-term in vitro tumour outgrowth assay that allows NK cells to use their full repertoire of killing pathways, bortezomib sensitized three breast cancer cell lines to NK cell-mediated killing, which led to greater anti-tumour effects than either treatment alone. We then used a xenogeneic mouse model in which CB-17 SCID mice were injected with human breast cancer cells. This model displayed the effectiveness of NK-92 cells, but the addition of bortezomib did not increase the survival further or reduce the number of lung metastases in tumour-bearing mice. However, while bortezomib was highly cytotoxic to NK-92 cells in vitro, bortezomib treatment in vivo did not decrease NK-92 function, suggesting that through alternative dosing or timing of bortezomib, greater efficacy may occur from combined therapy. These data demonstrate that combined treatment of human breast cancer with bortezomib and NK cells has the potential to generate superior anti-tumour responses than either therapy alone.

  View details for DOI 10.1111/j.1365-2249.2008.03818.x

  View details for Web of Science ID 000262950100016

  View details for PubMedID 19220837

  View details for PubMedCentralID PMC2669527

• Combination therapy using IL-2 and anti-CD25 results in augmented natural killer cell-mediated antitumor responses BIOLOGY OF BLOOD AND MARROW TRANSPLANTATION Hallett, W. H., Ames, E., Alvarez, M., Barao, I., Taylor, P. A., Blazar, B. R., Murphy, W. J. 2008; 14 (10): 1088-1099

  Abstract

  Interleukin (IL)-2 has been extensively examined to promote clinical T and natural killer (NK) cell responses. Regulatory T cells (Tregs) have been shown to regulate many aspects of the immune system, including NK cell-mediated responses. We have demonstrated that in vivo administration of IL-2 led to activation and expansion of both NK cells and immunosuppressive Tregs. Therefore, we attempted to augment NK cell antitumor effects by concurrently depleting Tregs using anti-CD25. Increased NK cell activation by IL-2 was found to be correlated with an increase in classical, short-term NK cell in vitro killing assays regardless of the depletion of Tregs. But when splenocytes of the treated mice were used in long-term tumor outgrowth experiments, we observed that prior depletion of Tregs from IL-2 administration led to improved antitumor effects compared with either treatment alone. Importantly, these in vitro data are correlated with subsequent in vivo survival of leukemia-bearing mice, in which co-treatment of IL-2 with anti-CD25 led to significantly improved survival compared with mice treated with either IL-2 alone or with Treg depletion. Prior depletion of NK1.1(+) cells, but not of CD8(+) cells, completely abrogated all antitumor effects mediated by IL-2 and anti-CD25 combination therapy. These findings demonstrate that superior NK cell-mediated antileukemic effects can be achieved with IL-2 administration and concurrent depletion of CD25(+) cells.

  View details for DOI 10.1016/j.bbmt.2008.08.001

  View details for Web of Science ID 000259669100002

  View details for PubMedID 18804038

  View details for PubMedCentralID PMC2735407

• Sensitization of tumor cells to NK cell-mediated killing by proteasome inhibition JOURNAL OF IMMUNOLOGY Hallett, W. D., Ames, E., Motarjemi, M., Barao, I., Shanker, A., Tamang, D. L., Sayers, T. J., Hudig, D., Murphy, W. J. 2008; 180 (1): 163-170

  Abstract

  Bortezomib is a proteasome inhibitor that has direct antitumor effects. We and others have previously demonstrated that bortezomib could also sensitize tumor cells to killing via the death ligand, TRAIL. NK cells represent a potent antitumor effector cell. Therefore, we investigated whether bortezomib could sensitize tumor cells to NK cell-mediated killing. Preincubation of tumor cells with bortezomib had no effect on short-term NK cell killing or purified granule killing assays. Using a 24-h lysis assay, increases in tumor killing was only observed using perforin-deficient NK cells, and this increased killing was found to be dependent on both TRAIL and FasL, correlating with an increase in tumor Fas and DR5 expression. Long-term tumor outgrowth assays allowed for the detection of this increased tumor killing by activated NK cells following bortezomib treatment of the tumor. In a tumor purging assay, in which tumor:bone marrow cell mixtures were placed into lethally irradiated mice, only treatment of these mixtures with a combination of NK cells with bortezomib resulted in significant tumor-free survival of the recipients. These results demonstrate that bortezomib treatment can sensitize tumor cells to cellular effector pathways. These results suggest that the combination of proteasome inhibition with immune therapy may result in increased antitumor efficacy.

  View details for DOI 10.4049/jimmunol.180.1.163

  View details for Web of Science ID 000252162700022

  View details for PubMedID 18097016