Publications by authors named "Sagar Bapat"

12 Publications

  • Page 1 of 1

Surface Proteomics Reveals CD72 as a Target for -Evolved Nanobody-Based CAR-T Cells in -Rearranged B-ALL.

Cancer Discov 2021 Aug 16;11(8):2032-2049. Epub 2021 Mar 16.

Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California.

Alternative strategies are needed for patients with B-cell malignancy relapsing after CD19-targeted immunotherapy. Here, cell surface proteomics revealed CD72 as an optimal target for poor-prognosis /-rearranged (MLLr) B-cell acute lymphoblastic leukemia (B-ALL), which we further found to be expressed in other B-cell malignancies. Using a recently described, fully system, we selected synthetic CD72-specific nanobodies, incorporated them into chimeric antigen receptors (CAR), and demonstrated robust activity against B-cell malignancy models, including CD19 loss. Taking advantage of the role of CD72 in inhibiting B-cell receptor signaling, we found that SHIP1 inhibition increased CD72 surface density. We establish that CD72-nanobody CAR-T cells are a promising therapy for MLLr B-ALL. SIGNIFICANCE: Patients with MLLr B-ALL have poor prognoses despite recent immunotherapy advances. Here, surface proteomics identifies CD72 as being enriched on MLLr B-ALL but also widely expressed across B-cell cancers. We show that a recently described, fully nanobody platform generates binders highly active in CAR-T cells and demonstrate its broad applicability for immunotherapy development..
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http://dx.doi.org/10.1158/2159-8290.CD-20-0242DOI Listing
August 2021

A diagnostic host response biosignature for COVID-19 from RNA profiling of nasal swabs and blood.

Sci Adv 2021 02 3;7(6). Epub 2021 Feb 3.

Department of Laboratory Medicine, University of California, San Francisco, San Francisco, CA, USA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease-19 (COVID-19), has emerged as the cause of a global pandemic. We used RNA sequencing to analyze 286 nasopharyngeal (NP) swab and 53 whole-blood (WB) samples from 333 patients with COVID-19 and controls. Overall, a muted immune response was observed in COVID-19 relative to other infections (influenza, other seasonal coronaviruses, and bacterial sepsis), with paradoxical down-regulation of several key differentially expressed genes. Hospitalized patients and outpatients exhibited up-regulation of interferon-associated pathways, although heightened and more robust inflammatory responses were observed in hospitalized patients with more clinically severe illness. Two-layer machine learning-based host classifiers consisting of complete (>1000 genes), medium (<100), and small (<20) gene biomarker panels identified COVID-19 disease with 85.1-86.5% accuracy when benchmarked using an independent test set. SARS-CoV-2 infection has a distinct biosignature that differs between NP swabs and WB and can be leveraged for COVID-19 diagnosis.
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http://dx.doi.org/10.1126/sciadv.abe5984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7857687PMC
February 2021

Magnitude and Kinetics of Anti-Severe Acute Respiratory Syndrome Coronavirus 2 Antibody Responses and Their Relationship to Disease Severity.

Clin Infect Dis 2021 01;72(2):301-308

Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California, USA.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can be detected indirectly by measuring the host immune response. For some viruses, antibody concentrations correlate with host protection and viral neutralization, but in rare cases, antiviral antibodies can promote disease progression. Elucidation of the kinetics and magnitude of the SARS-CoV-2 antibody response is essential to understand the pathogenesis of coronavirus disease 2019 (COVID-19) and identify potential therapeutic targets.

Methods: Sera (n = 533) from patients with real-time polymerase chain reaction-confirmed COVID-19 (n = 94 with acute infections and n = 59 convalescent patients) were tested using a high-throughput quantitative immunoglobulin M (IgM) and immunoglobulin G (IgG) assay that detects antibodies to the spike protein receptor binding domain and nucleocapsid protein. Individual and serial samples covered the time of initial diagnosis, during the disease course, and following recovery. We evaluated antibody kinetics and correlation between magnitude of the response and disease severity.

Results: Patterns of SARS-CoV-2 antibody production varied considerably. Among 52 patients with 3 or more serial specimens, 44 (84.6%) and 42 (80.8%) had observed IgM and IgG seroconversion at a median of 8 and 10 days, respectively. Compared to those with milder disease, peak measurements were significantly higher for patients admitted to the intensive care unit for all time intervals between 6 and 20 days for IgM, and all intervals after 5 days for IgG.

Conclusions: High-sensitivity assays with a robust dynamic range provide a comprehensive picture of host antibody response to SARS-CoV-2. IgM and IgG responses were significantly higher in patients with severe than mild disease. These differences may affect strategies for seroprevalence studies, therapeutics, and vaccine development.
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http://dx.doi.org/10.1093/cid/ciaa979DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7454426PMC
January 2021

SARS-CoV-2 seroprevalence and neutralizing activity in donor and patient blood.

Nat Commun 2020 09 17;11(1):4698. Epub 2020 Sep 17.

Creative Testing Solutions, Tempe, AZ, USA.

Given the limited availability of serological testing to date, the seroprevalence of SARS-CoV-2-specific antibodies in different populations has remained unclear. Here, we report very low SARS-CoV-2 seroprevalence in two San Francisco Bay Area populations. Seroreactivity was 0.26% in 387 hospitalized patients admitted for non-respiratory indications and 0.1% in 1,000 blood donors in early April 2020. We additionally describe the longitudinal dynamics of immunoglobulin-G (IgG), immunoglobulin-M (IgM), and in vitro neutralizing antibody titers in COVID-19 patients. The median time to seroconversion ranged from 10.3-11.0 days for these 3 assays. Neutralizing antibodies rose in tandem with immunoglobulin titers following symptom onset, and positive percent agreement between detection of IgG and neutralizing titers was >93%. These findings emphasize the importance of using highly accurate tests for surveillance studies in low-prevalence populations, and provide evidence that seroreactivity using SARS-CoV-2 anti-nucleocapsid protein IgG and anti-spike IgM assays are generally predictive of in vitro neutralizing capacity.
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http://dx.doi.org/10.1038/s41467-020-18468-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7499171PMC
September 2020

Evaluation of SARS-CoV-2 serology assays reveals a range of test performance.

Nat Biotechnol 2020 10 27;38(10):1174-1183. Epub 2020 Aug 27.

Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA, USA.

Appropriate use and interpretation of serological tests for assessments of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exposure, infection and potential immunity require accurate data on assay performance. We conducted a head-to-head evaluation of ten point-of-care-style lateral flow assays (LFAs) and two laboratory-based enzyme-linked immunosorbent assays to detect anti-SARS-CoV-2 IgM and IgG antibodies in 5-d time intervals from symptom onset and studied the specificity of each assay in pre-coronavirus disease 2019 specimens. The percent of seropositive individuals increased with time, peaking in the latest time interval tested (>20 d after symptom onset). Test specificity ranged from 84.3% to 100.0% and was predominantly affected by variability in IgM results. LFA specificity could be increased by considering weak bands as negative, but this decreased detection of antibodies (sensitivity) in a subset of SARS-CoV-2 real-time PCR-positive cases. Our results underline the importance of seropositivity threshold determination and reader training for reliable LFA deployment. Although there was no standout serological assay, four tests achieved more than 80% positivity at later time points tested and more than 95% specificity.
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http://dx.doi.org/10.1038/s41587-020-0659-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7740072PMC
October 2020

Test performance evaluation of SARS-CoV-2 serological assays.

medRxiv 2020 May 17. Epub 2020 May 17.

Department of Microbiology and Immunology, University of California, San Francisco, CA 94143, USA.

Background: Serological tests are crucial tools for assessments of SARS-CoV-2 exposure, infection and potential immunity. Their appropriate use and interpretation require accurate assay performance data.

Method: We conducted an evaluation of 10 lateral flow assays (LFAs) and two ELISAs to detect anti-SARS-CoV-2 antibodies. The specimen set comprised 128 plasma or serum samples from 79 symptomatic SARS-CoV-2 RT-PCR-positive individuals; 108 pre-COVID-19 negative controls; and 52 recent samples from individuals who underwent respiratory viral testing but were not diagnosed with Coronavirus Disease 2019 (COVID-19). Samples were blinded and LFA results were interpreted by two independent readers, using a standardized intensity scoring system.

Results: Among specimens from SARS-CoV-2 RT-PCR-positive individuals, the percent seropositive increased with time interval, peaking at 81.8-100.0% in samples taken >20 days after symptom onset. Test specificity ranged from 84.3-100.0% in pre-COVID-19 specimens. Specificity was higher when weak LFA bands were considered negative, but this decreased sensitivity. IgM detection was more variable than IgG, and detection was highest when IgM and IgG results were combined. Agreement between ELISAs and LFAs ranged from 75.7-94.8%. No consistent cross-reactivity was observed.

Conclusion: Our evaluation showed heterogeneous assay performance. Reader training is key to reliable LFA performance, and can be tailored for survey goals. Informed use of serology will require evaluations covering the full spectrum of SARS-CoV-2 infections, from asymptomatic and mild infection to severe disease, and later convalescence. Well-designed studies to elucidate the mechanisms and serological correlates of protective immunity will be crucial to guide rational clinical and public health policies.
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http://dx.doi.org/10.1101/2020.04.25.20074856DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7273265PMC
May 2020

SARS-CoV-2 seroprevalence and neutralizing activity in donor and patient blood from the San Francisco Bay Area.

medRxiv 2020 May 25. Epub 2020 May 25.

Creative Testing Solutions, Tempe, AZ, USA.

We report very low SARS-CoV-2 seroprevalence in two San Francisco Bay Area populations. Seropositivity was 0.26% in 387 hospitalized patients admitted for non-respiratory indications and 0.1% in 1,000 blood donors. We additionally describe the longitudinal dynamics of immunoglobulin-G, immunoglobulin-M, and in vitro neutralizing antibody titers in COVID-19 patients. Neutralizing antibodies rise in tandem with immunoglobulin levels following symptom onset, exhibiting median time to seroconversion within one day of each other, and there is >93% positive percent agreement between detection of immunoglobulin-G and neutralizing titers.
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http://dx.doi.org/10.1101/2020.05.19.20107482DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7273245PMC
May 2020

Characterization of Immune Cells from Adipose Tissue.

Curr Protoc Immunol 2019 09;126(1):e86

Nomis Foundation Laboratories of Immunobiology and Microbial Pathogenesis, The Salk Institute for Biological Studies, La Jolla, California.

Adipose tissue (AT) serves a crucial role in maintaining organismal metabolic homeostasis. Studies have demonstrated that AT is populated with a diverse array of immune cells that coordinate and regulate AT function. This adipo-immune system is highly dynamic, reflecting the physiologic state of the organism (e.g., obese, lean, aged, or young) as well as the constant physiologic remodeling of AT associated with the daily rhythms of fasting and feeding. Many of the adaptive and maladaptive functional changes of AT are regulated by changes in the quantity and quality of distinct sets of AT-resident immune cells. Here we present protocols to assess the dynamic state of the immune system within AT by constructing censuses of adipose-resident immune cells (macrophages, dendritic cells, neutrophils, eosinophils, NK cells, innate lymphocytes, T cells, and B cells, etc.) based on flow cytometry, which we term adipo-immune profiles (AIPs). Constructing AIPs can be an integral part of assessment for AT health and function. This article describes the protocols to generate such AIPs. © 2019 by John Wiley & Sons, Inc.
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http://dx.doi.org/10.1002/cpim.86DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6814145PMC
September 2019

The nuclear receptor REV-ERBα modulates Th17 cell-mediated autoimmune disease.

Proc Natl Acad Sci U S A 2019 09 27;116(37):18528-18536. Epub 2019 Aug 27.

NOMIS Center for Immunobiology and Microbial Pathogenesis, The Salk Institute for Biological Studies, La Jolla, CA 92037;

T helper 17 (Th17) cells produce interleukin-17 (IL-17) cytokines and drive inflammatory responses in autoimmune diseases such as multiple sclerosis. The differentiation of Th17 cells is dependent on the retinoic acid receptor-related orphan nuclear receptor RORγt. Here, we identify REV-ERBα (encoded by ), a member of the nuclear hormone receptor family, as a transcriptional repressor that antagonizes RORγt function in Th17 cells. REV-ERBα binds to ROR response elements (RORE) in Th17 cells and inhibits the expression of RORγt-dependent genes including and Furthermore, elevated REV-ERBα expression or treatment with a synthetic REV-ERB agonist significantly delays the onset and impedes the progression of experimental autoimmune encephalomyelitis (EAE). These results suggest that modulating REV-ERBα activity may be used to manipulate Th17 cells in autoimmune diseases.
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http://dx.doi.org/10.1073/pnas.1907563116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6744854PMC
September 2019

Electronic cigarette inhalation alters innate immunity and airway cytokines while increasing the virulence of colonizing bacteria.

J Mol Med (Berl) 2016 06 25;94(6):667-79. Epub 2016 Jan 25.

Pulmonary and Critical Care Section, VA San Diego Healthcare System, 3350 La Jolla Village Dr, MC 111J, San Diego, CA, 92161, USA.

Unlabelled: Electronic (e)-cigarette use is rapidly rising, with 20 % of Americans ages 25-44 now using these drug delivery devices. E-cigarette users expose their airways, cells of host defense, and colonizing bacteria to e-cigarette vapor (EV). Here, we report that exposure of human epithelial cells at the air-liquid interface to fresh EV (vaped from an e-cigarette device) resulted in dose-dependent cell death. After exposure to EV, cells of host defense-epithelial cells, alveolar macrophages, and neutrophils-had reduced antimicrobial activity against Staphylococcus aureus (SA). Mouse inhalation of EV for 1 h daily for 4 weeks led to alterations in inflammatory markers within the airways and elevation of an acute phase reactant in serum. Upon exposure to e-cigarette vapor extract (EVE), airway colonizer SA had increased biofilm formation, adherence and invasion of epithelial cells, resistance to human antimicrobial peptide LL-37, and up-regulation of virulence genes. EVE-exposed SA were more virulent in a mouse model of pneumonia. These data suggest that e-cigarettes may be toxic to airway cells, suppress host defenses, and promote inflammation over time, while also promoting virulence of colonizing bacteria.

Key Message: Acute exposure to e-cigarette vapor (EV) is cytotoxic to airway cells in vitro. Acute exposure to EV decreases macrophage and neutrophil antimicrobial function. Inhalation of EV alters immunomodulating cytokines in the airways of mice. Inhalation of EV leads to increased markers of inflammation in BAL and serum. Staphylococcus aureus become more virulent when exposed to EV.
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http://dx.doi.org/10.1007/s00109-016-1378-3DOI Listing
June 2016

Depletion of fat-resident Treg cells prevents age-associated insulin resistance.

Nature 2015 Dec 18;528(7580):137-41. Epub 2015 Nov 18.

Immunobiology and Microbial Pathogenesis Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA.

Age-associated insulin resistance (IR) and obesity-associated IR are two physiologically distinct forms of adult-onset diabetes. While macrophage-driven inflammation is a core driver of obesity-associated IR, the underlying mechanisms of the obesity-independent yet highly prevalent age-associated IR are largely unexplored. Here we show, using comparative adipo-immune profiling in mice, that fat-resident regulatory T cells, termed fTreg cells, accumulate in adipose tissue as a function of age, but not obesity. Supporting the existence of two distinct mechanisms underlying IR, mice deficient in fTreg cells are protected against age-associated IR, yet remain susceptible to obesity-associated IR and metabolic disease. By contrast, selective depletion of fTreg cells via anti-ST2 antibody treatment increases adipose tissue insulin sensitivity. These findings establish that distinct immune cell populations within adipose tissue underlie ageing- and obesity-associated IR, and implicate fTreg cells as adipo-immune drivers and potential therapeutic targets in the treatment of age-associated IR.
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http://dx.doi.org/10.1038/nature16151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4670283PMC
December 2015

Innate immunity. Dermal adipocytes protect against invasive Staphylococcus aureus skin infection.

Science 2015 Jan;347(6217):67-71

Division of Dermatology, University of California, San Diego (UCSD), La Jolla, CA 92093, USA.

Adipocytes have been suggested to be immunologically active, but their role in host defense is unclear. We observed rapid proliferation of preadipocytes and expansion of the dermal fat layer after infection of the skin by Staphylococcus aureus. Impaired adipogenesis resulted in increased infection as seen in Zfp423(nur12) mice or in mice given inhibitors of peroxisome proliferator-activated receptor γ. This host defense function was mediated through the production of cathelicidin antimicrobial peptide from adipocytes because cathelicidin expression was decreased by inhibition of adipogenesis, and adipocytes from Camp(-/-) mice lost the capacity to inhibit bacterial growth. Together, these findings show that the production of an antimicrobial peptide by adipocytes is an important element for protection against S. aureus infection of the skin.
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http://dx.doi.org/10.1126/science.1260972DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4318537PMC
January 2015
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