bluuebunny/crossref_metadata_2025_split huggingface dataset:
Engineering oncogenic kinases to activate apoptotic machinery: a novel rewiring of Bcr-Abl and PKCβII (TECH1P.862)
Authors: Hoekstra David , Carlson Louise , Lee Kelvin 
Published: January 2023

Abstract
Oncogenic kinases are crucial for the development of several cancers and have become popular targets for small molecule based inhibitor therapy. However, despite the development of 2nd and 3rd generation inhibitors, resistance to this therapy remains a significant clinical obstacle. To overcome this resistance, we hypothesized that we could hijack oncogenic kinases to activate a rewired ‘pro-drug’, whose activation would lead to the death of the transformed cell. To test this, we selected Bcr-Abl as the oncogenic kinase, which is constitutively active and essential for chronic myeloid leukemia (CML) cell survival. For our pro-drug we selected protein kinase C, isoform βII (PKCβII) whose activation has been shown to increase apoptosis in CML cells. To test whether we could engineer PKCβII as a pro-drug to be activated by Bcr-Abl, we designed four engineered PKCβII constructs and found that were specifically and significantly activated by Bcr-Abl, with no effect on intrinsic Bcr-Abl signaling observed. Activation of the PKCβII pro-drug resulted in a significant increase in apoptosis and a significant decrease in proliferation in Bcr-Abl+ leukemic cells with no effect on Bcr-Abl- cells. These findings demonstrate that Bcr-Abl mediated activation of our PKCβII pro-drug is feasible, induces death in leukemic cells, and provides a novel mechanism by which oncogenic kinases can be re-wired to activate pro-death agents to potentially improve patient outcomes.
Discrimination of spectrally overlapping fluorochromes using Sony Corporations’s spectral analyzer (SP6800) (TECH1P.865)
Authors: Divekar Anagha , Lee MIchael , Kakuta Masaya , Nao Nitta , Motohiro Furuki , Yang Xifeng , Ransom John 
Published: January 2023

Abstract
A maximum of seven colors can be separated using traditional 2-laser flow cytometers and it is required to have the necessary band pass filters to separate these fluorochromes. In addition, these systems cannot differentiate between spectrally overlapping fluorophores such as PerCP and PerCP/Cy5.5; or FITC, PE and Alexa Fluor 514® (A514). SP6800, a novel spectral analyzer from Sony Corporation uses an innovative optics system that collects all emitted light and eliminates the need for band pass filters. We performed whole blood staining using a panel of 9 markers, which included significantly overlapping fluorochromes PerCP and PerCP/Cy5.5 and A514, PE and FITC and analyzed it on the SP6800. We were able clearly identify CD56+ (PerCP/Cy5.5), CD19+ (PerCP) and CD3+ (A514) cells. Further comparison of a 7-color panel of consisting of CD3 FITC, CD56 PE, CD19 PerCP, CD8 PE/Cy7, CD16 APC, CD4 Alexa Fluor 700® and CD62L APC/Cy7 on the BD LSRFortessa™ system and SP6800 gave similar results. Our data show that the SP6800 can perform all the analysis similar to a traditional cytometer with the added feature of separating spectrally overlapping fluorophores.
Optimizing adoptive cancer immunotherapy with γδ T cells through “pulse” zoledronate stimulation and IL-15 (TUM2P.886)
Authors: Nada Mohanad , Wang Hong , Workalemahu Grefachew , Morita Craig 
Published: January 2023

Abstract
Human γδ T cells expressing Vγ2Vδ2 TCRs monitor foreign- and self-prenyl pyrophosphate metabolites in isoprenoid biosynthesis to mediate immunity to microbes and cancer cells. Vγ2Vδ2 T cells have been used for adoptive cancer immunotherapy with some partial and complete remissions. Most clinical trials have used zoledronate to expand Vγ2Vδ2 T cells. Zoledronate inhibits farnesyl pyrophosphate synthase causing isopentenyl pyrophosphate to accumulate that then stimulates Vγ2Vδ2 T cells. Because constant zoledronate exposure is toxic, we hypothesized that a short "pulse" of zoledronate would reduce T cell toxicity because the monocytes that function as APC would preferentially take up zoledronate. We now find that "pulse" exposure of PBMC to zoledronate results in more uniform expansion of Vγ2Vδ2 T cells with higher purity and cell numbers along with better preservation of the early memory subsets that are the most effective for cancer immunotherapy. IL-15 was superior to IL-2 because it further preserved early (central) memory subsets. The Vγ2Vδ2 T cells also exhibited increased tumor cytotoxicity. Thus, “pulse” zoledronate exposure and substituting IL-15 for IL-2 maximizes the quantity and quality of expanded Vγ2Vδ2 T cells and represent improvements on the protocols being used in clinical trials. Moreover, because Vγ2Vδ2 T cells exhibit no autoimmunity or alloreactivity, they are potential recipients for the expression of chimeric antigen receptors.
Efficient identification of mutated cancer antigens recognized by T cells associated with durable tumor regressions (TUM2P.878)
Authors: Lu Yong-Chen , Yao Xin , Crystal Jessica , Li Yong , El-Gamil Mona , Gross Colin , Davis Lindy , Dudley Mark , Yang James , Samuels Yardena , Rosenberg Steven , Robbins Paul 
Published: January 2023

Abstract
Cancer immunotherapy with adoptive transfer of tumor infiltrating lymphocytes (TILs) represents an effective treatment for patients with metastatic melanoma, with the objective regressions in up to 72% of patients in three sequential pilot clinical trials. However, the antigen targets recognized by effective TILs remain largely unclear. Melanoma patients 2359 and 2591 both experienced durable complete regressions of metastases ongoing beyond five years following adoptive TIL transfer. A novel approach was developed to identify mutated T-cell antigens by screening a tandem minigene library, which comprised non-synonymous mutation sequences identified by whole-exome sequencing of autologous tumors. The screening of tandem minigene libraries resulted in the identification of mutated kinesin family member 2C (KIF2C) antigen as a target of TIL 2359, and mutated DNA polymerase alpha subunit B (POLA2) antigen as a target of TIL 2591. Both KIF2C and POLA2 have been found to play important roles in cell proliferation. These findings suggest that the minigene screening approach may facilitate the antigen repertoire analysis of tumor reactive T cells, and lead to the development of novel adoptive cell therapies based on the in vitro stimulation of T cells that recognize candidate mutated antigens derived from genes essential for the carcinogenesis.
Chronic alcohol consumption decreases the survival and compromises the antitumor immune response of mice bearing estrogen receptor positive E0771 breast cancer (TUM2P.898)
Authors: Zhang Hui , Zhang Faya , Zhu Zhaohui , Meadows Gary 
Published: January 2023

Abstract
Epidemiological data convincingly indicate that chronic alcohol consumption increases the risk of breast cancer, especially estrogen receptor positive (ER+) breast cancer in women. The effect of alcohol on antitumor immunity and survival remains to be elucidated. Herein, we implanted ER+ E0771 breast cancer cells into the mammary pad of female C57BL/6 mice consuming 20% w/v alcohol for three months and examined the effect of chronic alcohol consumption on tumor growth, dynamic changes in the antitumor immune response and host survival. Alcohol did not affect primary tumor growth or the body weight of tumor-bearing mice; however, it decreased survival. Alcohol did not alter the percentage of MDSC, Treg, CD4+ T or CD8+ T cells in the spleen and blood, but increased theCD4/CD8 ratio. B cells decreased during tumor growth, and iNKT cells increased in the blood of alcohol-consuming mice. In non-tumor-bearing mice, alcohol increases IFN-γ-producing CD8+ T cells and NK cells. Four weeks after E0771 inoculation, the percentage of IFN-γ-producing CD8+ T cells and NK cells were lower in alcohol-consuming mice compared to water-drinking mice. Alcohol consumption also decreased survival of mice immunized with an E0771 cell lysate and boosted with αGalCer compared to water-drinking counterparts. Collectively, chronic alcohol consumption compromises antitumor immunity and decreases the survival of mice bearing ER+ breast cancer.
Breast cancer diagnosis by high throughput sequencing of T cell receptors from peripheral blood samples (TUM2P.905)
Authors: Wang Chunlin , Alleyne Jessica , Yang Qunying , Byrne-Steele Miranda , Ng Enders , Kwong Ava , Shin Vivian , Han Jian 
Published: January 2023

Abstract
T cells play vital roles in the identification and elimination of transformed cells. It might be possible to diagnose cancer by reading imprints on the immune system made by cancer cells. High-throughput immune repertoire sequencing allows the depiction of T cell repertoire details at a nucleotide resolution and enables us to search for cancer-specific biomarkers. RNA was isolated from whole blood in breast cancer and matched controls and the CDR3 region of the TCR beta chain amplified and sequenced by Illumina next-generation sequencing platform. Initial studies showed around 200 breast cancer-specific CDR3 sequences. Two thirds of those CDR3 sequences show a pattern of convergent evolution, i.e., the same CDR3 peptide sequence is coded by different sequences. To leverage those disease-specific CDR3’s for diagnostics, we developed a novel metric called immune repertoire sharing index (SI) to measure the similarity between a repertoire and the disease library. With SI, we were able to predict breast cancer cases with both the sensitivity and the specificity around 90%. The diagnostics process requires only 10ml peripheral blood. We hypothesized that the immune repertoire sequencing and sharing index method could be generalized to other diseases. An ‘ultimate diagnostics’ may be emerged where taking one sample, with one procedure; hundreds of diseases can be diagnosed by evaluating immune repertoires.
An inflammatory T cell signature predicts obesity-associated type 2 diabetes (HUM3P.262)
Authors: Ip Blanche , Cilfone Nicholas , Zhu Min , Kuchibhatla Ramya , Azer Michel , McDonnell Marie , Apovian Caroline , Lauffenburger Douglas , Nikolajczyk Barbara 
Published: January 2023

Abstract
T cell inflammation plays pivotal roles in obesity-associated type 2 diabetes (T2D). Multiple T cell subsets, including Th1, Th2, Th17 and regulatory T cells are critical for the development of insulin resistance in animal models of T2D. However, the relative importance of T cell subsets in human disease pathogenesis remains a critical gap in knowledge with important implications for defining the most potent inflammatory mediators of T2D. To identify T cell subsets that disproportionately contribute to T2D-associated inflammation, we broadly quantified T cell cytokine production by circulating or adipose-associated immune cells from obese subjects clinically defined as non-T2D, pre-T2D or T2D. Cytokines traditionally associated with specific T cell subsets, including IL-2 (Th1), IL-4/10/13 (Th2) and IL-17A/F (Th17), were produced at highest amounts by T2D samples. We combined cytokine outcomes with multivariate mathematical approaches to show that T cell cytokine profiles predicted T2D status with &gt;85% accuracy. Furthermore, Th17 signature cytokines were more powerful than either Th1 or Th2 cytokines for differentiation of non-T2D from T2D samples. We conclude that Th17 cells are an underappreciated source of T cell cytokines that support inflammation and predict human T2D. Study outcomes unify contradictory claims of the importance of single cytokines and justify including Th17 cells as druggable sources of inflammation in human T2D.
MSPrecise: a new diagnostic test for relapsing-remitting multiple sclerosis by cerebrospinal fluid B cell sequencing (HUM3P.267)
Authors: Rounds William , Salinas Edward , Wilks Tom , Levin Mikhail , Ligocki Ann , Ionete Carolina , Pardo-Villamizar Carlos , Greenberg Benjamin , Bigwood Douglas , Eastman Eric , Cowell Lindsay , Monson Nancy 
Published: January 2023

Abstract
Patients with relapsing-remitting multiple sclerosis (RRMS), or who will convert to RRMS, have cerebrospinal fluid (CSF) B cells that accumulate specific replacement mutations in their rearranged VH4 gene segments as identified by single-cell Sanger sequencing. To harness this observation in a clinical setting, we have developed a diagnostic test to extract and score this genetic information from bulk CSF lymphocytes by next-generation DNA sequencing (NGS). Our current aim is to determine the accuracy of this test on a cohort of patients with varying times of RRMS onset and disease modifying therapies, as well as on a diverse cohort of patients with other neurological diseases (OND) that generally mimic the presentation of MS. Our initial testing on 39 patients has shown an accuracy of 84% in distinguishing RRMS from OND patients. As we continue to expand the size of the cohorts being tested, we will also focus on identifying differences between sub-cohorts of patients at different stages of RRMS or with particular OND diagnoses. With a growing need for a faster and more accurate diagnosis of RRMS, MSPrecise may be a useful addition to the existing diagnostic toolkit that clinicians use to achieve this goal.
B lymphocytes limit the severity of emphysema caused by N. brasiliensis infection in mice (INC4P.332)
Authors: Chen Fei , Wu Wenhui , Millman Ariel , Palma Mark , Gause William 
Published: January 2023

Abstract
The intestinal nematode parasite, Nippostrongylus brasiliensis (Nb), transiently resides in the lung causing lung damage, resulting in emphysema. However, the mechanisms contributing to this chronic lung pathology remain not well-defined. Here we show that emphysema develops as early as 7 days after Nb inoculation, as measured by alveolar mean linear chord length and is associated with increased neutrophil elastase (NE) and macrophage matrix metalloproteinase 12 (Mmp12) gene expression in the lung. Transfer of sorted Nb-primed neutrophils or macrophages to naïve recipients results in emphysema, indicating that either of these populations can trigger lung emphysema in naïve mice. Parasite infection also results in more severe emphysema in B cell deficient Jh-/- mice compared to wild-type mice or FcRγ-/-mice and is associated with higher numbers of neutrophils and macrophages, as well as increased expression of NE and Mmp12 in the lung. Transfer of B lymphocytes from wild-type mice to the Jh-/- mice can restore control of emphysema. These data suggest that Nb-primed neutrophils and macrophages contribute to lung emphysema. Furthermore, parasite-induced B cells exert regulatory effects that can markedly reduce the severity of emphysema.
FcRγ-chain, Grb2 and SHP-1 are invovled in mannose receptor (CD206)-mediated signaling in relation to tuberculosis (INC4P.329)
Authors: Rajaram Murugesan , Ni Bin , Arnett Eusondia , Abul Azad , Schlesinger Larry 
Published: January 2023

Abstract
The mannose receptor (MR, CD206) is highly expressed on human macrophages. It is a major pattern recognition receptor for pathogens, mediates phagocytosis and endocytosis, and regulates inflammatory signaling pathways. We have shown that the MR mediates Mycobacterium tuberculosis (M.tb) phagocytosis by human macrophages which leads to up-regulation of PPARγ expression and a delay in phagosome-lysosome (P-L) fusion. However, the associated receptor-specific downstream signaling molecules are unknown. We assayed for MR-associated proteins by MALDI-TOF and Western blot using a pull down assay with macrophage lysates and phosphorylated and un-phosphorylated MR cytoplasmic tail peptides. We have identified that FcRg-chain interacts with the MR and this interaction is necessary for MR surface exposure on human monocyte-derived macrophages (MDMs). Furthermore, M.tb infection of MDMs leads to phosphorylation of MR tyrosine residues and recruitment of Grb2 which initiates the Rac/Pak/Cdc-42 signaling cascade required for phagocytosis, and activation of MAP kinases and the PI3k/Akt pathway. MR activation also recruits and activates SHP-1. SHP-1 blockade increases M.tb P-L fusion and inflammatory cytokines. In conclusion, we have identified key MR signaling molecules and regulatory pathways which initiate phagocytosis and other immune-related functions in human macrophages.
Identification and characterization of phosphodiesterases that specifically degrade 3’3’-cyclic GMP-AMP (INM2P.358)
Authors: Jiang Zhengfan 
Published: January 2023

Abstract
Cyclic di-nucleotides act as intracellular second messengers, modulating a variety of cellular activities including innate immune activation. Whereas phosphodiesterases (PDEs) hydrolyzing c-di-GMP and c-di-AMP have been identified, no PDEs for cGAMPs have been reported. Here we identified the first three cGAMP specific PDEs in V. cholerae (herein designated V-cGAP1/2/3 respectively). V-cGAPs belong to HD-GYP domain containing proteins, and specifically break 3’3’-cGAMP, but not other forms of cGAMP. 3’3’-cGAMP was linearized to 5’-pApG by all three PDEs, and was further hydrolyzed to 5’-ApG by V-cGAP1, which functions as both a phosphodiesterase and a 5’-nucleotidase. In vivo experiments demonstrated that V-cGAPs play non-redundant roles in cGAMP degradation. The high specificity of V-cGAPs on 3’3’-cGAMP suggests the existence of specified PDEs for other cGAMPs, including 2’3’-cGAMP in mammalian cells; and the absolute requirement of GYP motif for 3’3’-cGAMP degradation indicates that HD-domain containing PDEs in eukaryotes are probably not able to hydrolyze cGAMPs. The fact that all V-cGAPs attack 3’3’-cGAMP on one specific phosphodiester bond suggests that PDEs for other cGAMPs would utilize the similar strategy. These results will provide valuable information to search for and characterize the mammalian 2’3’-cGAMP-specific PDEs in future studies.
Despite KLRG1 expression, highly cytotoxic ThEO/TcEO polarized T cells efficiently establish immunologic memory (LYM5P.662)
Authors: Bartkowiak Todd , Ai Midan , Ager Casey , Curran Michael 
Published: January 2023

Abstract
Targeting immune inhibitory (CTLA-4, PD-1) or stimulatory (OX-40, 4-1BB) co-receptors has proven a potent anti-cancer therapeutic strategy, eliciting robust anti-tumor T cell responses and compelling pre-clinical and clinical data in a variety of cancers. We have previously shown in pre-clinical models that 4-1BB agonist antibodies engender strong anti-tumor T cell responses, imparting enhanced survival benefit in addition to evoking durable tumor regression. This impressive anti-tumor effect is due, in part, to the unique ability of 4-1BB co-stimulation to differentiate a novel subset of T cells that acquires a highly cytotoxic effector profile typically only seen in NK cells. This subset-dubbed ThEO (CD4+) or TcEO (CD8+)-is driven by the T-box transcription factor Eomesodermin, and is further characterized by expression of the inhibitory receptor KLRG1. We show here that, despite their strong effector function and expression of several co-inhibitory receptors (PD-1, TIM-3, KLRG1), the ThEO/TcEO phenotype establishes immunological memory and recalls with high frequency upon secondary tumor challenge with potent anti-tumor effector function. Further, these cells manifest characteristics of central memory (Tcm), effector memory (Tem), as well as stem cell memory (Tscm), and reside in multiple memory niches. These findings provide mechanistic insight into the process of memory formation and give evidence for the capacity of ThEO/TcEO cells to prevent tumor recurrence.
CD4+ CD44v.low cells are peripheral CD4+ T cell precursors that are distinct from recent thymic emigrants and stem cell-like memory cells (LYM6P.713)
Authors: Narsale Aditi , Zhao Chunfang , Marrero Idania , Davies Joanna 
Published: January 2023

Abstract
CD4+ CD44v.low cells are peripheral precursor T cells that inhibit lymphopenia by generating a large CD4+ T cell pool containing balanced numbers of naïve, memory, and regulatory Foxp3+ cells with a diverse TCR repertoire. They can also inhibit cachexia, a profound wasting syndrome that is associated with lymphopenia and immune dysregulation. Recent thymic emigrants (RTE) and stem cell-like memory T cells (TSCM) can also replenish a T cell pool. In this study we formally test whether CD44v.low cells are the same population as RTE and TSCM. Our data show that, in contrast to RTE, CD44v.low cells express high levels of CD45RB and low levels of CD24. Moreover, CD44v.low cells isolated from mice devoid of RTE retain their capacity to repopulate lymphopenic mice with naïve and memory cells and Foxp3+ Tregs. In addition, CD44v.low cells do not express IL-2Rb, Sca-1, and CXCR3, the phenotypic hallmarks of TSCM. Further characterization of the cells generated by CD44v.low cells shows that they secrete significantly more Th2-type cytokines IL-4 and IL-6, significantly less Th17/Th22-type cytokines IL-17A and IL-22, but equivalent Th1-type cytokines IFN-g and TNF-a compared to cells generated by their naïve cell counterparts. Overall, these data demonstrate that CD44v.low cells are neither RTE nor TSCM. Moreover, their capacity to promote a balance cytokine profile might explain their ability to inhibit cachexia.
Human pDCs express the C-type Lectin receptor Dectin-1 and uptake and kill Aspergillus fumigatus spores in vitro (MPF4P.734)
Authors: Maldonado Samuel , Dai Jihong , Singh Sukhwinder , Mwangi David , Rivera Amariliz , Fitzgerald-Bocarsly Patricia 
Published: January 2023

Abstract
Plasmacytoid dendritic cells (pDCs) are the body’s most potent producers of type-1 IFNs, and are key responders to viruses like HIV, serving as an important link between the innate and adaptive immune responses. Although they are mainly anti-viral, evidence suggests pDCs also play an integral part in anti-fungal immunity. We demonstrated that human pDCs express the C-type Lectin receptor (CLR), Dectin-1, and that it is further upregulated after stimulation with HSV, Influenza, and the synthetic DNA ODN, CpG-A. In contrast, we did not find expression of Dectin-2, another CLR, on pDCs. Dectin-1 recognizes β-linked glucans, a component of the cell wall of opportunistic fungi like Aspergillus, Pneumocystis, and Candida, and is involved in immune functions including the uptake of fungi, induction of cytokine production, and direction of T-helper cell differentiation. Using Fluorescent Aspergillus Reporter (FLARE)-labelled A. fumigatus spores, which allow us to detect and differentiate between live and dead spores using flow cytometry, we showed that human pDCs can phagocytose and kill Aspergillus spores in vitro within a six-hour time frame. We hypothesize that Dectin-1 is the receptor responsible for this uptake. Although pDCs are classically considered IFN producers, stimulation with spores did not result in IFN-α production by pDC. Results from this ongoing study will shed further light on the role of human pDCs in anti-fungal immunity.