Enhanced POC assay for TB in HIV-infected children based on the ultrasensitive detection of the urinary form of the lipoarabinomannan antigen
基于尿形式阿拉伯脂甘露聚糖抗原的超灵敏检测,增强 HIV 感染儿童结核病的 POC 检测
基本信息
- 批准号:10611413
- 负责人:
- 金额:$ 74.73万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2020
- 资助国家:美国
- 起止时间:2020-04-14 至 2025-03-31
- 项目状态:未结题
- 来源:
- 关键词:5 year oldAffinityAliquotAntibodiesAntigenic DiversityAntigensAreaAvidityB-LymphocytesBindingBiological AssayBiological MarkersCD4 Positive T LymphocytesCarbohydratesCharacteristicsChildChildhoodCollaborationsCollectionDetectionDevicesDiagnosisDiagnosticEngineeringEpitopesFreezingFutureGenerationsGlycolipidsGoalsHIVHIV InfectionsHIV SeronegativityHIV-1HIV/TBHumanImmuneImmunologicsIndustrializationInfectionKenyaLateralLeadLibrariesLinkMannosidesMemory B-LymphocyteMethodsMonoclonal AntibodiesMycobacterium tuberculosisNaturePatient SelectionPatientsPediatric cohortPolysaccharidesPopulationProductionPropertyRandomizedReagentSamplingSerumSourceSouth AfricanSpecificityStandardizationStructureSurfaceTestingTuberculosisTuberculosis diagnosisUrineVariantVertebral columnWorkXyloseYeastsco-infectioncohortdetection assaydetection platformdiagnosis standardefficacy testinghuman monoclonal antibodiesimmunoreactivityimprovedlateral flow assaylipoarabinomannanmanmanufacturemortalitymycobacterialnovelpatient screeningpoint of carescreeningsugarurinary
项目摘要
Abstract
TB remains a major source of mortality in young children in endemic areas, particularly in areas where there is
a high rate of HIV infection. These children are difficult to diagnose with existing assays, and it is believed that
many of the >230,000 children who died because of TB in 2017 could have been saved with better diagnostics.
A highly promising biomarker for diagnosing TB is lipoarabinomannan (LAM), a major mycobacterial surface
glyolipid that accumulates at different concentrations in the great majority of actively infected TB patients. Our
lab has recently made major contributions towards understanding the diversity of the antigenic properties of
LAM and the complexity of the humoral immune rsponse against this antigen. One surprising discovery
resulting from our work is that the urinary form of TB LAM (uLAM) is antigenically distinct from the bacterially
associated form (ManLAM), and we have identified novel combinations of monoclonal antibodies that allow the
sensitive and specific detection of uLAM. This has recently led to the production of a new lateral flow assay
(the Fujifilm SILVAMP TB LAM assay) that has a significantly increased sensitivity for uLAM over that of the
commercial assay. However, despite this improvement, the sensitivity and robustness of this assay is still not
sufficient to meet the WHO requirements for an optimal assay. This proposal will build on our previous work to
further improve the affinities and specificities of our existing reagents, and identify new reagents with increased
sensitivity and accuracy for uLAM. To better understand the nature of uLAM and provide standardized samples
for future assays we will solate large volumes (~ 2L) of urine from positive patients with a range of uLAM levels,
and freeze away multiple aliquots of these samples for future use. A selected set of these urines will be used to
purify high concentrations of uLAM from multiple patients, and the structural and immunological properties of
these antigens will be compared. We have engineered forms of existing capture and detection reagents that
have improved affinities/avidities for ManLAM, and the sensitivity of these reagents will be compared to those
of existing reagents and lateral flow assays against several cohorts of pediatric urine samples that meet the
requirements of this RFA (<5 years old, >20% HIV+). We will screen memory B cell populations from selected
patients to identify new mabs with high affinities and/or specificities for uLAM, and select yeast display libraries
generated by randomization of the key CDR regions of our lead detection reagent for variants with higher
affinities and specificites than the parental antibody. The binding properties of these new mAbs will be
characterized, and their efficacy in detecting uLAM in the pediatric samples will be tested. Antibody
combinations with demonstrably better sensitivities than existing reagents will be transferred to our commercial
collaborators at Fujifilm and other companies for incorporation and testing in their platforms. The overall goals
of this study are to develop an improved LAM detection assay that meets or exceeds the Target Product Profile
(TPP) proposed by the WHO for high priority biomarker-based, non-sputum-based, POC tests for TB detection.
摘要
结核病仍然是流行地区幼儿死亡的主要来源,特别是在有
艾滋病毒感染率很高。这些儿童很难用现有的检测方法进行诊断,而且据信
2017年死于结核病的23万名儿童中,如果有更好的诊断方法,许多人本可以获救。
诊断结核病的一个很有前途的生物标志物是脂阿拉伯甘露聚糖(LAM),它是一种主要的分枝杆菌表面
糖脂在绝大多数活跃感染的结核病患者体内以不同浓度积聚。我们的
最近,实验室在了解猪瘟病毒抗原性的多样性方面做出了重大贡献。
LAM和针对该抗原的体液免疫反应的复杂性。一个令人惊讶的发现
我们工作的结果是尿液形式的TB LAM(ULAM)在抗原性上与细菌的不同
相关形式(ManLAM),我们已经确定了新的单抗组合,使
对ULAM的敏感和特异性检测。最近,这导致了一种新的侧向流动分析的产生
(Fujifilm SILVAMP TB LAM试验)对ULAM的敏感性显著高于ULAM
商业化化验。然而,尽管有了这一改进,该检测的灵敏度和稳健性仍然不是
足以满足世卫组织对最佳化验的要求。这项提议将建立在我们先前工作的基础上,以
进一步提高我们现有试剂的亲和力和特异性,并确定增加了
ULAM的敏感度和准确性。为了更好地了解ULAM的性质并提供标准化样本
在未来的检测中,我们将从ULAM水平范围内的阳性患者中提取大量(~2L)尿液,
并将这些样品中的多份冷冻起来以备将来使用。一组精选的小便器将被用于
从多个患者中纯化高浓度的ULAM,并对其结构和免疫学性质进行研究
将对这些抗原进行比较。我们设计了多种形式的现有捕获和检测试剂
已经改善了ManLAM的亲和力/亲和力,这些试剂的敏感性将与那些
对符合条件的几组儿童尿样进行现有试剂和侧向流动分析
要求此RFA(<;5岁,>;20%HIV+)。我们将筛选出记忆B细胞群体
患者识别与ULAM具有高亲和力和/或特异性的新单抗,并选择酵母展示文库
由我们的铅检测试剂的关键CDR区域随机生成,适用于具有更高
亲和力和特异度高于亲本抗体。这些新单抗的结合属性将是
将测试它们在儿科样本中检测ULAM的有效性。抗体
敏感性明显好于现有试剂的组合将转移到我们的商业
富士胶片和其他公司的合作者在他们的平台上进行整合和测试。总体目标
这项研究的目的是开发一种改进的LAM检测方法,以满足或超过目标产品配置文件
(TPP)由世界卫生组织提出,用于高优先级的基于生物标记物的、非基于痰的POC检测结核病。
项目成果
期刊论文数量(0)
专著数量(0)
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会议论文数量(0)
专利数量(0)
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ABRAHAM PINTER其他文献
ABRAHAM PINTER的其他文献
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{{ truncateString('ABRAHAM PINTER', 18)}}的其他基金
Development of a highly-sensitive urine test for tuberculosis (TB) that detects diverse forms of urinary TB lipoarabinomannan (uLAM)
开发一种高灵敏度的结核病尿液检测方法,可检测多种形式的尿液结核菌脂阿拉伯甘露聚糖 (uLAM)
- 批准号:
10667871 - 财政年份:2022
- 资助金额:
$ 74.73万 - 项目类别:
Complementary diagnostic biomarkers of sputum culture-negative TB [R21]
痰培养阴性结核病的补充诊断生物标志物 [R21]
- 批准号:
10557869 - 财政年份:2022
- 资助金额:
$ 74.73万 - 项目类别:
Complementary diagnostic biomarkers of sputum culture-negative TB [R21]
痰培养阴性结核病的补充诊断生物标志物 [R21]
- 批准号:
10433028 - 财政年份:2022
- 资助金额:
$ 74.73万 - 项目类别:
Enhanced POC assay for TB in HIV-infected children based on the ultrasensitive detection of the urinary form of the lipoarabinomannan antigen
基于尿形式阿拉伯脂甘露聚糖抗原的超灵敏检测,增强 HIV 感染儿童结核病的 POC 检测
- 批准号:
10675836 - 财政年份:2020
- 资助金额:
$ 74.73万 - 项目类别:
Enhanced POC assay for TB in HIV-infected children based on the ultrasensitive detection of the urinary form of the lipoarabinomannan antigen
基于尿形式阿拉伯脂甘露聚糖抗原的超灵敏检测,增强 HIV 感染儿童结核病的 POC 检测
- 批准号:
10378761 - 财政年份:2020
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Novel epitopes that mediate broad neutralization of clade B and C HIV-1 isolates
介导 B 型和 C 型 HIV-1 分离株广泛中和的新表位
- 批准号:
8701676 - 财政年份:2013
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Optimizing protective vaccine targets in the V1/V2 domain of HIV-1 gp120
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8501371 - 财政年份:2012
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Optimizing protective vaccine targets in the V1/V2 domain of HIV-1 gp120
优化 HIV-1 gp120 V1/V2 结构域中的保护性疫苗靶标
- 批准号:
8410364 - 财政年份:2012
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Strategies for Eliciting bnAbs against Conserved HIV-1 Quaternary Epitopes
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8035414 - 财政年份:2010
- 资助金额:
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Characterization of sequence specificities and structures of QNEs
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- 批准号:
7904630 - 财政年份:2010
- 资助金额:
$ 74.73万 - 项目类别:
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