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Principal Investigator: Yasmine Belkaid
Organization: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
Fiscal Year: 2020
Award: $3,460,631
Funding agency: National Institute of Allergy and Infectious Diseases
In response to the SARS-CoV-2 pandemic we have made the technologies and platforms at CHI available for characterization of COVID-19 patient samples, as a priority ahead of our ongoing collaborative studies. By early April we had completed adaptation of our transcriptomic and flow cytometry platforms, and operating procedures, to begin analysing peripheral blood from Covid-19 patients. CHI began receiving samples from Northern Italy and CITE-Seq was performed for more than 100 samples in close collaboration with the Tsang lab. BCR sequencing and scATAC seq was performed for a subset of these, and proteomic analysis was obtained for all of the samples by outsourcing this work at SomaLogic. 27-color flow cytometry phenotyping was also performed by CHI to validate the novel CITE-seq platform. Further, CHIs automated pipeline was used to isolate high quality RNA from PAXgene samples of 200 Covid-19 samples, to be used for both RNA-Seq at CHI as well as distribution to the Goldbach-Mansky lab. Together these efforts have contributed to initial immune characterization of acutely infected Covid-19 patients. We hope these platforms can now be applied to new cohorts becoming available which are able to address questions such as variation in the outcomes of infection.
In addition to this COVID work CHI has continued to advance its previous studies that comprise 25 active collaborations across 8 Institutes and the Clinical Center. These are divided into 3 stages: initiation/sample collection (12 studies), assay (8 studies), and analysis/write up (5 studies). These all comprise multi-modal high-dimensional immune phenotyping of primary human samples. The current studies include characterizing immune changes in vaccine trials (malaria, HSV), clinical intervention trials (lupus, CHAPLE), and cohort studies of rare diseases (mitochondrial dysfunction). One of the new studies added is an exciting collaboration with Kevin Hall at NIDDK to characterize immune differences after periods of highly controlled diet changes. The monoclonal antibody therapy study of CHAPLE disease patients, with Mike Lenardo, is an example of how high dimensional phenotyping can lead to better understanding of patient pathology, and may inform use of a nascent therapy. The proteomic changes we observed give basic biology insight into, in this case, CD55 function and the mechanisms of complement inhibition, and this work is under review at Nature Immunology. Studies published this year included an integrated proteomic and cellular phenotyping analysis of maternal peripheral blood in longitudinally sampled human pregnancies, and characterization of changes in peripheral blood during treatment of healthy subjects with statins, and obese individuals undergoing a clinical trial of colchicine treatment.
To support CHIs mission we continue to develop wet-lab and computational infrastructure. For single cell sequencing technologies using the 10x platform, we have expanded the number of protein antigens measured in CITE-Seq, and developed scATAC-seq approaches with greater throughput and lower cost. CITE-Seq, in particular, led to a high-profile finding characterizing variation in baseline immune states that predicts immune responses (see section Scientific Advances). For flow cytometry, on a new Cytek Aurora we have validated a 27 color cytometry for broad immune phenotyping, as well as a 33 color T cell focused panel run after PMA stimulation to detect cells primed for production of characteristic cytokines or transcription factors. For flow cytometry analysis we are applying methods to enable conventionally gated populations to be recognized and interrogated using high dimensional parameters, to help overcome the difficulties of interpreting unbiased clustering of high dimensional data. For mass cytometry analysis we are finalising a package that uses high dimensional observations to discriminate only the subset of cells that respond in ex vivo stimulation assays, such as our mass cytometry assay that quantifies phosphorylation of 10 intracellular proteins, in addition to 20 cell phenotype markers, in response to 12 in vitro stimulation conditions.
CHI also provides fee-for-service access to assays not otherwise accessible to NIH researchers. The SomaLogic proteomic assay has been run for over 1000 samples, for 8 investigators, in the last year. This included human and murine samples from serum, plasma, cerebrospinal fluid, bronchioalveolar lavage fluid, and tissue homogenates. We continue to monitor alternative platforms and have run tests on O-Link that is offering increasingly high parameter antibody based analysis, although currently plan to transition our SomaLogic platform from the current 1.3k antigen panel to an analysis of 4.5k antigens.
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