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Development of a TCR beta repertoire assay for profiling liquid biopsies from NSCLC donors

Cancer Drug Resistance 2020年第3期3卷 563-571页

作者： Leisa P.Jackson Benjamin A.Tjoa Hestia Mellert gary A.pestano Development Biodesix Inc.BoulderCO 80301USA. Cellero LLCBothellWashingtonWA 98021USA.

Aim:The aim of this study was to demonstrate the utility of T-Cell receptor beta(TCRβ)sequencing as a robust method for assessing T-cell repertoire changes in donors with non-small cell lung cancer(NSCLC).We further demonstrated the use of the assay by monitoring repertoire modulation in a defined model antigen system,cytomegalovirus(CMV).Methods:Peripheral blood mononuclear cells from four healthy donors were challenged with a 1-week exposure to whole-cell lysate from CMV-infected cells or CMVpp65495-503 peptide(NLVPMVATV).T-cell repertoire perturbations were assessed using the Oncomine TCR Beta-SR Assay and Ion GeneStudio S5 Plus Sequencer.A pp65 tetramer flow cytometry assay was used as an orthogonal method to assess clonal expansion of a subset of CMV-specific *** evaluation of the assay in peripheral blood lymphocytes from NSCLC donors,five whole blood specimens were evaluated using the same sequencing ***:The TCR beta assay identified 6,683-61,936 unique clones from 1-2 million reads per sample,and an average of 80%of the total reads were usable for TCR *** the NSCLC donors,TCR convergence and clonality values were consistent with published results and ranged 0.016-0.033 for convergence and 0.09-0.48 for *** the CMV study,TCR sequencing detected the expansion of a common family of clones in all 4 samples in response to antigen *** expansion corresponded to an increase in pp65 tetramer staining by flow *** TCR convergence scores ranged 0.009-0.041 and increased 5-fold in one sample as a result of pp65 antigen ***:The results of this study demonstrated the utility of profiling of the TCRβrepertoire in a model system and in donors with ***,we demonstrated the correlation between RNA-seq methods and protein-tetramer analysis using flow *** techniques represent an emerging solution that could complement other liquid and tissue diagnostic assays in the cl

关键词： TCRβ liquid biopsy Ion Torrent immunotherapy

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Longitudinal monitoring for the emergence of epidermal growth factor C797S resistance mutations in non-small cell lung cancer using blood-based droplet digital PCR

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Cancer Drug Resistance 2019年第3期2卷 912-916页

作者： Mark Bowling Hyder Arastu Victoria Edwards Leisa Jackson Scott Thurston Jordan Reese Hestia Mellert gary pestano Paul Walker East Carolina University Mail Stop 628Internal Medicine-Hematology/OncologyGreenvilleNC 27834USA Biodesix Inc.2970 Wilderness PlaceBoulderCO 80301USA.

Validation of assays for the C797S mutation as a biomarker for osimertinib resistance is promising in guiding treatment decision-making for multidrug resistant non-small cell lung cancer. A newly developed droplet digital PCR (ddPCR) assay was used to retrospectively evaluate the emergence of the C797S mutation in six remnant plasma samples in this case report. It was found that the detected emergence of C797S clearly correlated with clinical signs of treatment resistance. Had these data been available to aid treatment selection in real time, there would have been hope for recaptured disease response and control instead of treatment cessation. The results of this study show that highly sensitive ddPCR methods can be used for the monitoring of emergent epidermal growth factor somatic variant mutations in circulation.

关键词： Epidermal growth factor non-small cell lung cancer C797S drug resistance

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利用HLA-DRβ1亚型同源多肽筛选类风湿关节炎相关抗原的研究

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中国免疫学杂志 2002年第3期18卷 152-154页

作者：栗占国肖利群韩蕾 gary pestano 北京大学人民医院风湿免疫科北京100034 美国哈佛大学医学院Dana Farber研究所免疫实验室

目的 :筛选与类风湿关节炎易感基因HLA DRβ1亚型同源多肽结合的抗原性蛋白。方法 :利用HLA DRβ1亚型同源多肽亲和层析柱结合类风湿关节炎患者B细胞可溶性蛋白 ,并以pH梯度洗脱获取HLA DRβ1同源多肽结合蛋白。结果 :类风湿关节炎易... 详细信息

目的 :筛选与类风湿关节炎易感基因HLA DRβ1亚型同源多肽结合的抗原性蛋白。方法 :利用HLA DRβ1亚型同源多肽亲和层析柱结合类风湿关节炎患者B细胞可溶性蛋白 ,并以pH梯度洗脱获取HLA DRβ1同源多肽结合蛋白。结果 :类风湿关节炎易感基因HLA DRβ1亚型同源多肽可高亲和力结合II型胶原、热休克蛋白 90和组蛋白等 5种抗原蛋白。结论 :II型胶原及热休克蛋白 90可能通过与HLA DRβ1分子形成二聚体复合物参与类风湿关节炎的发病。

关键词：类风湿关节炎 HLA-DRβ1 亚型结合抗原同源多肽 PH梯度洗脱 Ⅱ型胶原热休克蛋白

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