bluuebunny/crossref_metadata_2025_split huggingface dataset:
Immunization protocols towards shorter development time for monoclonal antibodies (130.44)
Authors: Hawksworth David , Brophy Susan 
Published: January 2023

Abstract
In the field of hybridoma development a wide variety of immunization regimens have been employed to develop fusion candidate animals in the shortest time possible. In studies presented here, we sought to discern the most immediate path to the highest affinity immune response. In the first study, we immunized two strains of mice using two distinct immunization regimens - 20 ug every six weeks or 5 ug bi-weekly, for 2 different proteins antigens. Results showed that, while additional boosts resulted in an improvement in average affinity, there was no difference in affinity between dosing groups at the end of the regimens. More importantly, it was discovered that some of the highest affinity animals had reached their maximum response without the need for an additional boost. Finally, for these two protein targets, BALB/c mice developed a higher affinity immune response than A/J mice. In a second study, mice were immunized three times with 20 ug protein using either a four or six week rest period between boosts. Results here showed no significant difference in relative affinity between mice rested 4 vs. 6 weeks between boosts. Overall, our results suggest that high affinity animals can be identified earlier in the boosting regimen and using shorter rest periods, thus allowing for the development of hybridoma cell lines in a shorter amount of time. Additionally, the more laborious bi-weekly dosing regimen showed no advantage over one with fewer boosts.
CD99 signal regulates the proliferation of multiple myeloma cell lines, RPMI8226. (131.10)
Authors: Gil Minchan , Park Seo-Jeong , Park Young Soo , Lee Hyangsin , Kim Hye-Mi , Park Chan-Sik 
Published: January 2023

Abstract
CD99 is a 32-kDa cell surface protein that is variously expressed on hematopoietic cells and hematopoietic-derived cancer cells. CD99 is highly expressed in some multiple myeloma cells including RPMI8226. In this study, we show that engagement of CD99 by agonistic anti-CD99 monoclonal antibody (mAb), YG32, inhibited proliferation of RPMI8226 multiple myeloma cells. However, YG32 mAb did not induce the cell death of RPMI 8226 cells. We carried out the microarray analysis to investigate the expression of genes involved in growth retardation by CD99 engagement. In microarray data, RPMI8226 cells engaged by anti-CD99 mAb had reduced expression of cyclin D1 and D2. Expression of CDKN2A gene, which codes cell cycle inhibitors p16INK4A and p19ARF, were induced by anti-CD99 mAb. These results suggest that CD99 signal regulates the proliferation of multiple myeloma cells by modulating proliferative and anti-prolifearative genes. The study was supported by National Research Foundation (NRF) grant of the Korean government A (R13-2008-023-01003)
Long-term cytokine therapy with IL-15/IL-15Rα complexes leads to impaired NK cell function (131.11)
Authors: Turley Shannon , Rubinstein Mark , Goldrath Ananda , Elpek Kutlu 
Published: January 2023

Abstract
Immunotherapies targeting effector lymphocytes are promising approaches for the treatment of cancer and infectious diseases. One such approach employs IL-15/IL-15Rα complexes which are considerably more potent than IL-15 in expanding cytotoxic lymphocytes including NK and CD8 T cells. Recent studies demonstrated that short-term treatment with IL-15/IL-15Rα complexes is highly effective in driving a transient but substantial reduction in tumor burden via immune-mediated mechanisms whereas continuous cytokine therapy improves survival only modestly and fails to control tumor outgrowth. Given the limited efficacy of long-term therapy, we sought to investigate the impact of continuous in vivo stimulation with IL-15/IL-15Rα complexes on NK and CD8 T cells in tumor-free mice. Long-term treatment with IL-15/IL-15Rα complexes triggered massive expansion of NK and CD8 T cells. Strikingly, however, NK cells exhibited marked defects in activation, IFNγ production, degranulation, and cytotoxicity whereas CD8 T cells showed enhanced functional capacity. Additionally, continuous treatment generated a suppressive environment for transferred naïve NK and CD8 T cells and resulted in accumulation of CD11b+Gr1+ cells. The mechanistic basis for NK cell dysfunction upon chronic stimualation with IL-15/IL-15Ra complexes will be discussed. Our results have important clinical implications for the design of immunotherapies and vaccines using multiple doses of immunostimulatory agents.
Abrogation of SHP-1 in tumor-specific effector T cells during adoptive immunotherapy of disseminated leukemia (131.15)
Authors: Stromnes Ingunn , Fowler Carla , Blattman Joseph , Tan Xiaoxia , Greenberg Philip 
Published: January 2023

Abstract
Adoptive therapy is a promising modality for treating patients with cancer. Currently, there are many obstacles to success, such as the low avidity of self/tumor-reactive T cells and the failure of transferred T cells to persist and to function in a tolerizing tumor environment. To overcome some of these obstacles, we asked if abrogating the Src homology region 2 domain-containing tyrosine phosphatase -1 (SHP-1), a negative regulator of lymphocyte activation, in tumor-reactive effector T cells would influence efficacy of adoptive therapy. A conditional knockout of SHP-1 in mature, tumor-specific transgenic T cells (TCRgag) was developed to analyze cell intrinsic effects of complete or partial ablation of SHP-1 on effector CD8 T cell function, persistence and efficacy in adoptive therapy of disseminated leukemia. In vitro-expanded TCRgag SHP-1-/- cells exhibited greater therapeutic efficacy than TCRgag SHP-1+/+ or SHP-1+/- cells, due to enhanced T cell expansion and survival in vivo, independent of providing exogenous IL-2. The increased expansion and cytokine production of SHP-1-/- effector cells was most pronounced in settings of lymphopenia. The role of SHP-1 in regulating signals received during lymphopenia-induced expansion and on the activation threshold of effector T cells will be discussed. Targeting SHP-1 in human T cells may provide a means to increase in vivo expansion following transfer and avidity for tumor targets during adoptive therapy of cancer.
Combination of CTLA-4 blockade with anti-CD40/IL-2 immunotherapy augments the CD8+ T cell response in resting and tumor-bearing mice (131.17)
Authors: Sckisel Gail , Nzigira Ngabo , Hurwitz Andrew , Blazar Bruce , Murphy William 
Published: January 2023

Abstract
Immunotherapy is a promising new option for the treatment of disseminated cancers. We have demonstrated that combined interleukin-2 (IL-2) and anti-CD40 (αCD40) therapy results in synergistic anti-tumor effects. While this strategy has shown success, it does not work well on less immunogenic tumors and can be extremely toxic. We sought to determine if removal of negative regulators such as cytotoxic T lymphocyte antigen-4 (CTLA-4) could result in augmentation of CD8+ T cell effects. We assessed total CD8+ T cell numbers and lytic ability after combination therapy in naïve and tumor-bearing mice. Here we present evidence that when the therapies are combined, there are increases in CD8+ T cells both systemically and intratumorally as well as a prolonged CD8+ memory expansion. We also saw increases in the lytic ability of CD8+ T cells against tumor cells as measured by chromium-51 release assay after combination with CTLA-4 blockade. We hypothesize that the amplification of T cell responses due to CTLA-4 blockade will enhance the anti-tumor effects of immunostimulatory therapies. It may also be possible to augment anti-tumor responses such that less immunotherapy will be needed to achieve equivalent anti-tumor responses.
SA-4-1BBL serves as an effective immunostimulatory component of HPV-16 E7 protein based therapeutic vaccine in a mouse model of cervical cancer (131.21)
Authors: Sharma Rajesh , Srivastava Abhishek , Yolcu Esma , Schabowsky Rich-Henry , Shirwan Haval 
Published: January 2023

Abstract
Cervical cancer is a major health problem in women worldwide. Current prophylactic vaccines are ineffective in therapeutic settings. Therefore, there is an acute need for the development of therapeutic vaccines. The HPV E7 oncoprotein is consistently expressed in cervical cancer, required for the maintenance of the transformed phenotype, and is not subjected to antigenic drift. However, the low antigenicity of this protein may require potent adjuvants for therapeutic efficacy. We have recently generated a novel form of the 4-1BBL costimulatory molecule chimeric with core streptavidin (SA-4-1BBL) and demonstrated its pleiotropic effects in soluble from on various cells of the immune system. We herein tested the utility of SA-4-1BBL as the immunomodulatory component of HPV-16 E7 recombinant protein based therapeutic vaccine in the E7 expressing TC-1 cervical cancer mouse model as a prelude to clinical trials. A single subcutaneous vaccination was effective in eradicating established TC-1 tumors in ~70% of mice. The therapeutic efficacy of the vaccine was associated with robust primary and memory CD4+ and CD8+ T cell responses, incresed CD8+ T cells and decreased T reg cells in tumor, enhanced NK cell killing, and generation of IFN-γ and IL-2 responses. Taken together, these data demonstrate the utility of SA-4-1BBL as a new class of immunomodulator for the development of therapeutic vaccines against cancer and chronic infections.
TNFRSF25 agonists as potent cancer vaccine adjuvants (131.25)
Authors: Schreiber Taylor , Podack Eckhard 
Published: January 2023

Abstract
Signaling through TNFRSF25 by its natural ligand, TNFSF15, or agonistic antibodies enhances cytokine production and effector function by antigen-activated T cells. Paradoxically, we recently discovered that TNFRSF25 signaling induces rapid and specific proliferation of CD4+FoxP3+ regulatory T cells (Treg) in vivo in the absence of exogenous antigen. When TNFRSF25 agonistic antibody, clone 4C12, was used in combination with heat shock protein (gp96-Ig) based vaccination, 4C12 treatment led to synergistic antigen-specific CTL expansion when given together with the vaccine, which occurred simultaneously with Treg expansion in vivo. The frequency of CTL and Treg reached 65% of total CD8+ cells and 45% of total CD4+ cells, respectively, within the same tissue following combined administration of 4C12 and gp96-Ig. TNFR25 driven synergism with gp96-Ig vaccination was dependent upon mTOR signaling. In a therapeutic setting, combined administration of 4C12 and gp96-Ig enhanced rejection of established EG7, LLC and 3-MCA induced sarcoma tumors. These data indicate that TNFRSF25 agonists enhance vaccine-induced anti-tumor immunity which correlates with increased magnitude and duration of tumor-specific CTL expansion, systemically. These findings illustrate that TNFRSF25 signaling may enhance either inflammatory or regulatory immunity depending on the temporal availability of its ligand and the inflammatory milieu.
Targeted delivery of interferon alpha via fusion to anti-CD20 results in potent antitumor activity against B-cell lymphoma (131.30)
Authors: Xuan Caiyun , Steward Kristopher , Timmerman John , Morrison Sherie 
Published: January 2023

Abstract
The anti-CD20 antibody rituximab has substantially improved outcomes in patients with B-cell non-Hodgkin lymphomas. However, many patients are not cured by rituximab-based therapies, and overcoming de novo or acquired rituximab resistance remains an important challenge to successful treatment of B-cell malignancies. Interferon alpha (IFNα) has potent immunostimulatory properties and anti-proliferative effects against some B-cell cancers, but its clinical utility is limited by systemic toxicity. To improve the efficacy of CD20-targeted therapy, we constructed fusion proteins consisting of anti-CD20 and murine or human IFNα. Fusion proteins had reduced IFNα activity in vitro compared with native IFNα, but CD20 targeting permitted efficient anti-proliferative and pro-apoptotic effects against an aggressive rituximab-insensitive human CD20+ murine lymphoma (38C13-huCD20) and a human B-cell lymphoma (Daudi). In vivo efficacy was demonstrated against established 38C13-huCD20 grown in syngeneic immunocompetent mice and large, established Daudi xenografts grown in nude mice. Optimal tumor eradication required CD20 targeting, with 87% of mice cured of rituximab-insensitive tumors. Gene knockdown studies revealed that tumor eradication required expression of type I IFN receptors on the tumor cell surface. Targeting type I IFNs to sites of B-cell lymphoma by fusion to anti-CD20 antibodies represents a potentially useful strategy for treatment of B-cell malignancies.
MUC1-specific Th1 immunotherapy induce differential levels and proportions of "inducible" and "natural" CD4 TReg cell subpopulations that results in enhanced tumor immunity and increased ovarian cancer patient survival (131.38)
Authors: Dobrzanski Mark , Rewers-Felkins Kathleen , Samad Khaliquzzaman , Robinson William , Wright Stephen 
Published: January 2023

Abstract
Adoptive T cell therapy is a viable treatment for cancer patients and optimally requires participation of CD4 T cells. Here, we assessed the therapeutic effects of adoptively transferred autologous MUC1 peptide-stimulated CD4+ effector cells in patients with ovarian cancer. Using MUC1 peptide and IL-2 for ex vivo CD4+/Th1 effector cell generation, we show that 3 monthly treatment cycles of T cell restimulation and intraperitoneal re-infusion modulated T cell mediated immune responses that correlated with diminished serum CA125 tumor marker levels and enhanced patient survival. One patient remains disease free, another patient survived long-term for nearly 18 months with recurrent disease and two patients expired within 3-5 months following final infusion. Long term survivors showed elevated levels of systemic CD3/CD4/CD25+ and CD3/CD4/CD25- T cells when compared to that of pre-treatment levels and similarly treated short-term survivors. Such cells co-expressed different levels of "Inducible" Tr1 (CD4/CD25-/FoxP3-/IL-10+) and "Natural" (CD4/CD25+/FoxP3+) TReg cell ratios that further correlated with disease-free survival. These patients showed elevated levels of Th1 cells and MUC1-specific T cells expressing chemokine receptors associated with T cell differentiation/memory. This suggests that MUC1-stimulated CD4 T cells induce differential levels and proportions of systemic TReg subpopulations that influence long term tumor immunity in such patients following treatment.
Evaluation of cellular therapies utilizing a fluorine-based magnetic resonance imaging (MRI) tracer agent (131.40)
Authors: Helfer Brooke , Balducci Anthony , Mailliard Robbie , Wesa Amy , Ahrens Eric 
Published: January 2023

Abstract
Natural killer cells (NK), T cells and dendritic cells (DC) are increasingly being used in the clinic to adjust patient immune function in response to cancer. One of the current limitations of such therapies is the inability to track the therapeutic cells once they are administered to a patient. Here we utilize a novel, non-radioactive fluorine-based (19F) MRI tracer agent to label clinically relevant cells ex-vivo and to non-invasively detect the transferred cells in vivo over time. We demonstrate that human NK, T cells and DC (immature and mature) can be effectively labeled without effect on their health, phenotype or function. Using a murine xenograft model as a pre-clinical “proof of principle” assay, we visualized and tracked the migration of human DC in-vivo. Following injection, the 19F labeled cells were detected and anatomically located by MRI. Mature DC but not immature DC were observed to migrate into the region of draining lymph nodes 18h post injection by MRI. Isolation and flow cytometric analysis of cells from draining lymph nodes confirmed the presence of human DC as well as the efficacy of this method. The application of this tracer agent across several cell types, along with non-invasive highly specific detection of 19F labeled cells by MRI, demonstrates the potential use of 19F in translational and clinical applications of cellular therapies. Ultimately, this technology may expedite a therapy’s transition from bench to bedside.
The role of Actin Binding Protein One (Abp1) in B cell development, activation, and B cell tolerance. (132.23)
Authors: Seeley Margaret , Onabajo Dr. Olusegun , Song Dr. Wenxia 
Published: January 2023

Abstract
The B cell receptor (BCR) serves as both signal-transducer and antigen-transporter. Optimal activation of B cells depends on coordinated BCR signaling, antigen processing, and actin cytoskeleton rearrangement events. Actin-binding protein 1 (Abp1/HIP-55/SH3P7) is a unique adaptor protein that has been shown to directly interact with components of signal transduction, endocytosis, and actin cytoskeleton apparatuses through its proline-rich, SH3, and actin binding domains. This suggests a role for Abp1 in mediating cross-talk among the three cell apparatuses during BCR activation. Here we study the role of Abp1 in B cell development, activation, and B cell tolerance. We discovered major defects in B cell development in Abp1 knockout mice, exhibited as reduced numbers of pre- and immature B cells in the bone marrow, and transitional B cells in the spleen. Furthermore, Abp1 knockout mice exhibit splenomegaly and have increased levels of anti-nuclear antibodies. At the cellular level, Abp1 is recruited to BCR surface signalosomes in response to antigenic stimulation. In Abp1 knockout mice, BCR-induced protein tyrosine phosphorylation of splenic B cells is elevated and phosphorylation of the MAP kinase JNK reduced compared to wild type B cells. These results suggest that Abp1 can regulate B cell development, B cell activation, and B cell tolerance via modulation of BCR signaling. This work supported by a NIH grant (AI059617) for W.S.
IL-2 signals promote the differentiation of effector CD8+ T cells (132.26)
Authors: Mitchell Diana , Williams Matthew 
Published: January 2023

Abstract
CD8+ T cells that do not receive IL-2 signals during the primary immune response to acute infection form stable memory populations but are unable to mount successful recall responses. In this study, we found a specific defect in the differentiation and persistence of effector and effector memory CD8+ T cells in the absence of IL-2 signals during the response to lymphocytic choriomeningitis virus (LCMV) infection. Although expression levels of the effector-associated transcription factors T-bet, Eomes, and Blimp-1 did not differ significantly, unbiased analysis of gene expression by WT or IL-2Rα-deficient CTLs revealed that optimal expression of CTL effector molecules, including Granzymes and Perforin, was IL-2-dependent. The end result of CD8+ T cell activation in the absence of IL-2 was therefore decreased effector CTL activity. The defect in effector differentiation was even more profound for secondary CTL responses and in models of localized tissue-specific infection. CD8+ T cells that homed to the lung following influenza infection failed to express Granzyme B. We conclude that IL-2 provides a differentiation signal to responding CD8+ T cells that enhances access to an effector gene expression program during the primary response to infection. Furthermore, our findings suggest that IL-2 permanently imparts a gene expression imprint to CD8+ memory T cells that enables them to re-access the CTL effector program upon subsequent encounter with antigen.
Characterization of intracellular signaling profiles induced by antigen receptor activation in murine B cell subsets (132.30)
Authors: Lyubchenko Taras , Liubchenko Ganna , Holers V. Michael 
Published: January 2023

Abstract
Protein phosphorylation and Ca2+ mobilization are among major signaling parameters characterizing the activation status of B lymphocytes. This study identifies quantitative differences between intracellular signaling profiles in subsets of follicular, marginal zone, transitional T1, T2, T3 murine B cells. Phosphorylation levels of signaling proteins and Ca2+ responses upon BCR engagement with anti-IgM were measured by multicolor flow cytometry. The results revealed previously uncharacterized diversity of signaling profiles in different B cells subsets that correlated with their developmental stage and previous antigen exposure. At least three important trends were identified: 1) B cells at earlier developmental stages had higher activation threshold, baseline Ca2+ and phosphorylation activity, and Ca2+ dynamics that in some instances did not correlate with sIgM expression levels; 2) phosphorylation levels within particular B cell subsets were heterogeneous which suggests the presence of various cell groups with different signaling properties within each subset; 3) analysis of phosphorylation profiles in individual signaling molecules revealed molecule-specific nonlinear characteristics of dose-response curves that are indicative of quantitative signaling differences between B cell subsets and subset-specific balance of BCR signaling cascades. These preliminary findings provide rationale for investigating roles of different B cell subsets in alterations of immune tolerance.
Activation of human naïve CD8 T cells by artificial antigen presenting cells (132.31)
Authors: Blair David , Dustin Michael 
Published: January 2023

Abstract
The precise mechanisms by which an activated T cell differentiates to a memory cell remain unclear. Recent evidence indicates that asymmetric cell division (ACD), an evolutionarily conserved process whereby multiple cell fates are derived from a common progenitor cell, may play an important role in the differentiation of T cells. However, these studies were conducted in mouse models and it is unknown whether parallel mechanisms occur in human T cells. To better understand these mechanisms, we employed the use of an established artificial antigen presenting cell (aAPC) system for the activation of human naïve CD8 T cells. Our efforts have focused on the early events following T cell activation up to the first division of T cells. We investigated atypical protein kinase C zeta (PKCζ), a protein involved in ACD, and its role in human CD8 T cell differentiation upon engagement of the T cell receptor. Anti-CD3 presentation to T cells by the aAPCs resulted in the partitioning of PKCζ distal to the immunological synapse in a temporal manner. This polarization was first observed at 12 hours following activation and was maintained for at least up to 42 hours of stimulation. However, in the absence of anti-CD3 stimulation by the aAPC, polarization of PKCζ did not occur in the T cells. These data indicate that partitioning of polarization complex proteins occurs in human T cells and may be important in the differentiation of CD8 T cell subsets following activation.