bluuebunny/crossref_metadata_2025_split huggingface dataset:
Precision immunology for discovery and development of the Precision Vaccines Program (PVP)-037 small molecule series: imidazopyrimidine adjuvants identified via age-specific human in vitro modeling
Authors: Soni Dheeraj , Borriello Francesco , Scott David A , Ozonoff Al , Brightman Spencer , Smith Jennifer , Shamu Caroline , Ramirez Juan C , Baden Lindsey Robert , Cheng Wing Ki , van Haren Simon Daniel , Pettengill Matthew , Dowling David J , Levy Ofer 
Published: January 2023

Abstract
Infection is a significant cause of mortality and immunization is the most promising biomedical intervention to reduce this burden. However, multiple booster doses are often required to protect those with distinct immunity such as the young, elders, and immunocompromised individuals. Adjuvants are molecules that potentiate vaccine responses. There is a need for a larger toolbox of adjuvants, since Alum is the only adjuvant currently incorporated into licensed vaccines in the pediatric setting. We utilized a unique in vitro approach to discover new adjuvants, screening a ~10k small molecule library via a no-wash 384-well quantitative TNF immunoassay in peripheral blood mononuclear cells (PBMCs) from 3 adult healthy human donors. Preliminary hits were based on induced TNF production in at least two of the three donors screened. We identified a novel imidazopyrimidine (IMP) scaffold named PVP-037 which demonstrated efficacy, potency, and adjuvanticity. PVP-037 demonstrated broad age-independent immune-stimulatory ability in human PBMCs and dendritic cells, including key Th-polarizing cytokines such as IL-12, and significant enhancement murine humoral responses to recombinant hemagglutinin (rHA) proteins in the quadrivalent influenza. PVP-037 further demonstrated limited activity in several common cell lines (THP-1) and murine/porcine primary leukocytes. Structure activity relationship (SAR) studies identified the analog PVP-144-3 with greater in vitro efficacy across species (porcine, non-human primates) and murine in vivo adjuvanticity. Overall, we discovered an IMP adjuvant via high throughput screening of human mononuclear cells in vitro which correlated with enhanced influenza immunization in mice.
Corn based nanoparticle delivered inactivated influenza virus vaccine intranasally augments mucosal immune response in pigs
Authors: Gourapura Renukaradhya J , Feliciano-Ruiza Ninoshkaly , Renu Sankar , Lu Fangjia , Han Yi , Schrock Jennifer , Dhakal Santosh , Patil Veerupaxagouda , HogenEsch Harm 
Published: January 2023

Abstract
Swine influenza A virus (SwIV) causes respiratory tract infection in pigs. Available SwIV vaccines fail to provide cross-protective immunity in pigs. Nano-11 is an amphiphilic nanoparticle (70–80nm) obtained from sweet corn-derived phytoglycogen. Nano-11 carries high surface positive charge and thus facilitates easy preparation of nanoparticle based vaccine by electrostatic interaction with killed SwIAV antigen (KAg) or peptides (negative charge). Earlier we showed that Nano-11 bound killed SwIV H1N2 Ag (Nano-11+KAg) delivered intranasally in pigs induced mucosal antibody response, but the challenge heterologous H1N1 SwIV load was not substantially reduced in the airways. In this study, KAg or conserved ten IAV peptides co-adsorbed with adjuvant Poly(I:C) (negative charge) on Nano-11 [Nano-11+KAg/peptides+Poly(I:C)] was vaccinated to influenza-free pigs intranasally, twice, and challenged with a heterologous SwIV. We observed increased SIgA and IgG responses in the airways and enhanced proliferation of IFN-g+ gd T cells in PBMCs in Nano-11+KAg+Poly(I:C) vaccinates compared to control. In Nano-11+peptides+Poly(I:C) vaccinates noticed an increased proliferation of IFN-g+ gd T cells and IFN-g+ cytotoxic T cells in PBMCs compared to control. Commercial vaccine group induced higher IgG response in serum and proliferation of IFN-g+ T-helper/memory cells in PBMCs compared to control. However, reduction in challenge virus load in any of the vaccinated groups was not statistically significant. In conclusion, inclusion of Poly(I:C) in Nano-11 flu vaccine improved the T cell response, but further improvements in the vaccine formulation is required to take advantage of this easy to prepare particle based mucosal flu vaccine.
Trispecific antibody against HIV-1 demonstrate anti-viral activity in vivo and have potent Fc-dependent effector functions
Authors: Almasri Cassandra , Xu Ling , Dias Joana , Promsote Wanwisa , Wei Ronnie , Lord Dana M , Rao Ercole , Beninga Jochen , Yang Zhi-yong , Pegu Amarendra , Mascola John R. , Nabel Gary J. 
Published: January 2023

Abstract
Broadly neutralizing antibodies (bNAbs) against HIV-1 have been suggested as a complementary immunotherapy to current combination small molecule anti-retroviral therapies (cART) for treatment of HIV-1 infection. Monoclonal bNAbs when used as a monotherapy demonstrate high susceptibility for escape in HIV-1 infected individuals, thus a trispecific bNAb targeting three independent HIV-1 envelope determinants was developed to achieve more robust suppression. Upon treatment of viremic simian-human immunodeficiency virus (SHIV)-infected macaques with the trispecific bNAb, plasma viral suppression of up to 1000-fold was observed. To elucidate a mechanism for the observed plasma viremia control, in vitro assays using the trispecific bNAb were conducted to determine Fc-mediated effector functions. First, we determined that the trispecific bNAb bound to HIV-1 infected cell lines with an equal or higher affinity than its parental bNAbs alone. We then conducted antibody-dependent cell cytotoxicity (ADCC) and antibody-dependent cell phagocytosis (ADCP) assays and found that the trispecific bNAb had comparable or better activity compared to its parental bNAbs alone. Similarly, when we measured direct virolysis of HIV-1 virus using an antibody-dependent complement-mediated virus lysis (ADCML) assay, the trispecific bNAb had comparable or better activity compared to its parental bNAbs alone. These studies demonstrate that the trispecific bNAb shows anti-viral activity in vivo and has potent Fc-dependent effector functions that target both free virus and virally infected cells compared to monoclonal bNAbs alone. Thus, the trispecific bNAb is a promising single immunotherapeutic protein for treatment of HIV-1 infection.
Role of antibody in vaccine-mediated protection against pneumonic tularemia in the rabbit model
Authors: Reed Douglas S , Olsen Emily , Bowling Jennifer , Willett Katherine , Hazlett Karsten , Barry Eileen 
Published: January 2023

Abstract
Francisella tularensis, a gram negative coccobacillus, is the causative agent of tularemia, a severe zoonotic infection. When inhaled, as few as 15 cfu is sufficient to cause disease. There is concern that tularemia could be used as a biological weapon so vaccines are urgently needed. We have previously shown that the New Zealand White (NZW) rabbit is a good model of human disease; after exposure to small particle aerosols containing virulent F. tularensis at doses as low as 66 cfu, naive NZW rabbits develop fever within 3 days and succumb within 4–7 days. Disease pathology in the NZW rabbit strongly resembles what has been reported in humans. Recombinant derivatives of a virulent strain of F. tularensis are highly attenuated in NZW rabbits and protect against aerosol challenge with virulent F. tularensis. The level of protection is dependent on the strain used, the route of vaccination, and the number of vaccinations. Using a prime-boost small particle aerosol vaccination regimen, 73% percent of rabbits survived a robust (100 LD50) challenge when vaccinated with a strain that is deficient in the aromatic amino acid synthase pathway. Pre-challenge, post-vaccination plasma IgG and IgM titers against heat-killed F. tularensis correlate with protection. A single inoculation of plasma from hyper-immune rabbits extended the survival of naïve rabbits by one day. We will report our efforts to evaluate the role of antibody in protection of NZW rabbits against tularemia, including passive immunization of mice using purified immunoglobulin from hyper-immune rabbits and in vitro assays of antibody function.
In silico design of a universal carrier protein: A potential solution to the variable immunogenicity observed in past vaccines against drugs of abuse
Authors: Saylor Kyle Lucas , Donnan Ben , Zhang Chenming 
Published: January 2023

Abstract
Vaccines against drugs of abuse (VADAs) show promise as a possible therapeutic for the alleviation of addiction due to their ability to modulate intake behavior via the altering of drug pharmacokinetics. In the past, however, these vaccines have failed to show clinical efficacy due to extensive variability in the magnitude of antibody response between individual vaccinees. Correspondingly, the future success of VADAs hinges directly on whether or not researchers can find ways to engineer more broadly effective vaccines. Differences in response must, to some extent, be the result of variable capacities for each individual’s immune system to process and/or present antigens. As such, we speculate that future VADAs will need to target the three key stages of vaccine processing, recognition, processing, and presentation, in order to achieve clinical effectiveness. Here, we describe a project aiming to engineer a broadly effective carrier protein based on this concept. Using haplotype frequency data provided by Be the Match, HLA-DQ and -DR haplotypes covering 99% of the population were selected for analysis. Next, the most common carrier proteins used in conjugate vaccine formulations were analyzed for MHC II epitope content using these haplotypes and NetMHCIIPan 3.2 prediction software. Finally, a modified version of the prediction output was used to excise the most likely MHC II epitopes. The highest-ranked epitopes were then stitched together using linkers designed with chemical conjugation sites and cathepsin cleavage sequences and subsequently tethered to cholera toxin subunit B. These results present the successful design of an in silico derived, broadly effective carrier protein to be used in future VADA formulations.
Cell-based cancer immunotherapy using chimeric antigen receptor (CAR)-engineered natural killer (NK) cells surface-modified with drug-loaded nanoparticles targeting adenosine receptors
Authors: Cinay Gunce Ezgi 
Published: January 2023

Abstract
The immunosuppressive tumor microenvironment (TME) plays a key role in failed eradication of solid tumors and tumor recurrence upon cancer treatment, as resident immune cells lose their cancer surveillance abilities. One of the underlying mechanisms responsible for the loss of adoptively transferred immune cell function in the TME is the A2a adenosine receptor (A2aR) inhibitory pathway. Encapsulating SCH-58261 (SCH), a potent antagonists of A2aR, in crosslinked multilamellar liposomal vesicles (cMLVs) was shown to efficiently deliver the drug to adoptively transferred cells in tumors. We genetically modified human NK92 cells to express anti-mesothelin CAR (ameso.CAR.NK) to be used in this study.
We hypothesized that the efficacy of NK cell adoptive transfer therapies could be enhanced if CAR.NK cells were used as active carriers to direct SCH-loaded cMLVs to the immunosuppressive TME to block the A2aR pathway. We first covalently attached cMLVs to NK cell surface. We next verified that cMLV conjugation did not have adverse effects on NK cell viability and migration. In addition, surface conjugation of cMLVs did not reduce IFN-γ secretion and cancer cell killing ability of NK cells, suggesting similar levels of cognate antigen-specific activation of ameso.CAR.NK cells by mesothelin-overexpressing ovarian cancer cells.
In the future, we will test the in vitro function of irradiated CAR.NK cells, given the requirement of irradiating NK92 cell lines prior to clinical use. Next, we will test our platform in an ovarian cancer xenograft mouse model and examine the effects of our therapy on tumor growth. Finally, we will investigate the potential of this therapy to reverse immune suppression on the TME-resident tumor-infiltrating NK cells.
The length of the intracellular domain conditions the antitumor efficacy of CAR-T cells
Authors: Ramello M Cecilia , Roselli Emiliano , Abate-Daga Daniel 
Published: January 2023

Abstract
Among the variables affecting CAR-T cell function, evidence shows that CAR architecture is a common factor influencing clinical outcomes. Previously, we demonstrated that a 2nd generation (gen.) CAR targeting the prostate stem cell antigen (PSCA) was more efficient than a 3rd gen. counterpart, in a preclinical model of pancreatic adenocarcinoma. This superior anti-tumor capacity was associated with greater tonic signaling. To further characterize this phenomenon, we compared the antitumor activity and phenotype of two 2nd gen. CAR-T cells (CAR-Ts) that differed in the length of the intracellular domain (ICD) by only 24 amino acids: PSCA-8t28Z vs. PSCA-8t(+24)28Z. Upon adoptive transfer into tumor-bearing mice, PSCA-8t28Z CAR-Ts displayed significantly greater antitumor efficacy. This was not due to differences in CAR surface expression, or in trafficking or persistence of T cells, since CAR-T numbers in spleen and tumor were comparable among groups. However, it was associated with significantly higher phosphorylation of CD3ζ in PSCA-8t28Z CAR-Ts in spleen, suggesting spontaneous CAR-signaling activity. Unsupervised analysis revealed that proliferation was similar between CAR-T and GFP T cells in spleen. In contrast, tumor-infiltrating (TI) PSCA-8t28Z CAR-Ts exhibited a cluster of Ki-67hi cells that was absent in the other groups. Moreover, TI PSCA-8t28Z CAR-Ts showed the highest expression of inhibitory receptors, suggesting antigen-induced CAR signaling within the tumor. Together, our results show that shorter ICDs are associated with higher tonic signaling and higher T cell activation after antigen recognition. These data highlights the importance of CAR-ICD length for optimal CAR-T function in solid tumors.
A commensal Clostridium species restricts systemic alphavirus dissemination through a type I interferon signaling axis
Authors: Winkler Emma S , Thackray Larissa B , Hykes Barry L , Handley Scott A , Droit Lindsay , Andhey Prabhakar , Mack Matthias , Shrihari Swathi , Swain Amanda , Cella Marina , Colonna Marco , Artyomov Maxim , Stappenbeck Thaddeus S , Diamond Michael S 
Published: January 2023

Abstract
The role of the gut microbiota in shaping antiviral immune responses at non-intestinal sites has been described in multiple models of viral pathogenesis. Although chikungunya virus (CHIKV), an emerging alphavirus, is a substantial global health threat, the impact of the microbiota in modulating immune responses to alphavirus infection is entirely unexplored. Here, we show that CHIKV infection of oral antibiotic-treated or germ-free mice resulted in increased viremia within one day of infection, and this enhanced viral dissemination. Microbiota depletion resulted in greater CHIKV infection of circulating monocytes, which was linked to dampened systemic type I interferon (IFN) responses. Differences in CHIKV tropism and replication following microbiota depletion depended on MyD88 signaling in plasmacytoid dendritic cells, which contributed to systemic type I IFN production within hours of infection. Colonization of antibiotic-treated mice with a single bacterial microbiota-derived Clostridium species resulted in MyD88-dependent type I IFN responses that restricted CHIKV infection in blood. We propose that select bacterial symbionts impact antiviral immunity to alphaviruses within hours of infection through a type I IFN signaling axis and by virtue of effects on viremia may determine disease severity and affect vector transmission and epidemic spread.
HLA-B recycling and trafficking pathways in primary human monocytes
Authors: Olson Eli C , Raghavan Malini 
Published: January 2023

Abstract
Human leukocyte antigen class I (HLA-I) molecules are a heterotrimeric protein complex responsible for presenting peptide antigens to CD8+ T cells. Typically, peptides are loaded onto the HLA-I complex in the ER and trafficked to the cell surface through the secretory pathway. Efficient peptide loading in the ER involves specific chaperone interactions to stabilize open conformers. While peptide loading is a tightly regulated process, HLA-B allotypes vary in their assembly and expression characteristics. We have recently found that some HLA-B allotypes such as B*35:01 are expressed at high levels in monocytes when compared with other allotypes such as B*08:01. Monocytes contained high levels of intracellular HLA-I, which is accumulated in a non-ER AP-1+ compartment. We hypothesized that monocytes possess a secondary mode of assembly involving recycling through the endo-lysosomal pathway. We find that HLA-B accumulates in various endo-lysosomal compartments. Additionally, endosomal recycling appears to be important for maintaining surface expression. Certain allotypes exhibit greater cell-surface rescue when lysosomal acidification is inhibited, consistent with a model of differential dependence on ER assembly factors. Taken together, these results indicate that endosomal pathways play a role in maintaining constitutive HLA-B surface expression, and that the inhibition of recycling pathways results in a loss of surface HLA-B. Some allotypes are more susceptible to degradative trafficking due to their chaperone-dependent assembly mechanisms, leaving them more vulnerable to peptide dissociation in the endosomes and trafficking to lysosomes.
Isoform repertoire of Blood mononuclear cells from SLE patients
Authors: Carmen Silvina del , Singh Prashant , Nath Pulak , Zhao Yuqi , Huynh Richie , Chung Cheng-Han , Rossi Robert , Pascual Virginia , Banchereau Jacques 
Published: January 2023

Abstract
Systemic Lupus Erythematosus (SLE) is characterized by numerous immunological alterations, including increased expression of immune activators, such as IFNa, along with alterations in cell composition, such as expansion of plasma cells that produce autoantibodies.
Alternative Splicing (AS) is a biological process that regulates post-transcriptional alterations of the RNA. Pathogenic AS events have been associated to several diseases. Furthermore, the description of SLE autoantibodies that target components of the spliceosome such as anti-Sm/RNP, anti-Ro/La, and anti-dsDNA has led us to hypothesize that SLE is associated to abnormal AS events.
RNA-seq uses standard Short-Read technologies which involve RNA or cDNA fragmentation for the generation of 150 bp sequencing libraries. Since most mammalian mRNAs are 1–2 kb long, using RNA-seq to identify novel alternatively spliced isoforms is extremely challenging. However, emerging Long-Read sequencing technologies allow full-length mRNA sequencing, de novo transcriptome assembly, and isoform profiling. Our lab has built a computational pipeline that combines Long-Read sequencing with RNA-seq data to identify and quantify novel transcripts in peripheral blood mononuclear cells (PBMCs) from SLE patients compared to healthy controls. From our computational analysis, we have observed that SLE patients display differential exon-inclusion and exclusion events in transcripts coding for immune activating and regulatory proteins, respectively. We conclude that SLE might be associated with pathogenic AS events that might place the immune balance under double jeopardy, through activation of immune activating receptors and inhibition of immune inhibiting receptors.
Characterization of new circular RNA products from the Pax-5 gene in B-cells
Authors: Robichaud Gilles A. , Hannay Brandon , Martin Danick , Veilleux Vanessa , Finn Nicholas 
Published: January 2023

Abstract
Circular RNAs (circRNAs) represent a new class of RNAs which are highly expressed and conserved in mammalian cells. CircRNAs regulate many biological processes and their aberrant expression is often associated with disease progression. We have identified and characterized circRNAs generated from the Pax-5 gene, an essential transcription factor for B-cell lineage commitment and also, a potent oncogene. Given the pivotal role of Pax-5 gene products in B-cell development and function, we set out to profile Pax-5 circular RNA (circPax5) expression and elucidate their role in B-cell biology. We identify and map five major circPax5 transcript isoforms which are found in B-cell lines and primary B-cells isolated from peripheral blood mononuclear cells (PBMCs) of healthy donors. These circPax5 variants consists of circular single-stranded RNA sequences corresponding to: i) exons 2–3; ii) exons 2–4; iii) exons 2–5; iv) exons 2–6; and, v) exons 2–8 of the human Pax-5 gene. Using TaqMan assays designed to target each unique circPax5 junction region, we found that circPax5 expression profiles vary with B-cell developmental stages. Furthermore, circPax5 exons 2–5 and 2–6 are overexpressed in patients suffering from chronic lymphocytic leukemia (CLL). Functionally, we observe that circPax5 products modulate B-cell processes and interact by sponging of specific micro-RNAs. Altogether, we identify new genetic products (i.e. circRNAs) generated from a prominent immune-regulating gene (Pax-5). The functional elucidation of circPax5 products will potentially bring insight into the function and signaling of B-cell processes and disease.
Missense mutation in non-recombinase domain of RAG1 results in B-cell developmental defect
Authors: Bodansky Aaron L , Burn Thomas , Behrens Edward M 
Published: January 2023

Abstract

Objective
To examine how a single amino acid change to a region of RAG1 that is not related to VDJ recombination affects B-cell development.

Methods
We studied B-cells within a Rag1 P326G missense mutant mouse (Rag1PG) utilizing flow cytometric analysis to phenotype B-cell development and peripheral populations, as well as biochemical studies examining the B-cell Receptor (BCR) molecular complex and signaling function. We also created a mixed bone marrow chimera to determine which alterations were cell-intrinsic.

Results
We find that overall there is an ~50% decrease in total B-cells in the Rag1PG mouse compared to wildtype (WT) due to a Pro-B to Pre-B-cell developmental block. Peripheral B cell cytopenias are in the Follicular Zone B-cell compartment, with sparing of the Marginal Zone B-cells. There is decreased Syk expression in Rag1PG—however no differences in BCR signaling strength were appreciated in Rag1PG. In the mixed bone marrow chimera, we find a competitive advantage in the WT developing B-cells, but peripheral B cell populations survive equally well.

Conclusion
Despite the Rag1PG protein being a single amino acid change to a region of RAG1 not involved in recombination, we nevertheless find a developmental defect consistent with leaky SCID. Given that there is no evidence of altered BCR signaling, and the developmental block occurs at the stage in B-cell development in which VDJ recombination is first required, these data suggest a recombination defect. The mutation is in a ubiquitin ligase domain, and it is possible this alters currently unappreciated roles of RAG1, such as histone degradation to allow RAG1 access to DNA. Our findings illustrate how mutations in the non-recombinase domain of RAG1 may result in leaky SCID.