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Hotspot Hunter: a computational system for large-scale screening and selection of candidate immunological hotspots in pathogen proteomes. G.L. Zhang, A.M. Khan , K.N. Srinivasan, A.T. Heiny, K.X. Lee, C.K. Kwoh, J.T. August and V. Brusic. Outline. Background & Motivations
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Hotspot Hunter: a computational system for large-scale screening and selection of candidate immunological hotspots in pathogen proteomes G.L. Zhang, A.M. Khan, K.N. Srinivasan, A.T. Heiny, K.X. Lee, C.K. Kwoh, J.T. August and V. Brusic
Outline Background & Motivations System – Hotspot Hunter Discussion
Identification of T-cell epitopes for the study of vaccines and immunotherapies Peptide TCR HLA http://immuneweb.xxmc.edu.cn/Lymphoid%20System.files/UntiPCT8.jpeg
P1 Promiscuous epitopes P2 P3 P4 One supertype H1 H2 H3 H4 T-cell epitope clusters (hotspots) for the development of epitope-based vaccines • Promiscuous T-cell epitopes relevant to large proportion of the human population • Presence of clusters of promiscuous T-cell epitopes (hotspots) in antigens
Mapping hotspots experimentally is a challenging task • Large size of pathogen proteomes (sequence length versus sequence number) • Low natural prevalence of T-cell epitopes (~1-5%) for a given HLA molecule • High cost of peptide synthesis • Limited access to human PBMC • Time-consumingexperimental assays Peptide TCR HLA
Existing prediction systems are not suitable for large-scale study of hotspots in pathogen proteomes • Limitations of existing promiscuous epitope prediction systems • Single protein sequence per submission • Do not predict for hotspots • Impractical for large-scale systematic study of hotspots in large proteomes
Outline • Background & Motivations • System – Hotspot Hunter • Discussion
Hotspot Hunter • Screen and select of hotspots specific to four common HLA supertypes • HLA class I A2, A3, B7 cover ~ 88% of human population • HLA class II DR cover ~100% of human population
Hotspot Hunter Implementation • Predictive Engines ANN and SVM methods • Predictions results integrated using soft computing principles • 10-fold cross-validation results showed that the system is of high accuracy
Hotspot Hunter can reliably identify real hotspots • HLA-DR supertype specific hotspots for HCV Core protein sequence Experimental verified 30-51 118-173 95-108 FP 30-47 FN 130-147 Hotspot Hunter predictions
Hotspot Hunter Functions • Single sequence query • Multiple sequence query • Target selection • Selection of common hotspot across more than one HLA supertype
Outline Background & Motivations System – Hotspot Hunter Discussion
Hotspot Hunter is a new generation computational tool aiding in epitope-based vaccine design • Allows prediction of immunological hotspots • Combines the strengths of the ANN and SVM robust prediction performance • Multiple sequence query suitable for large-scale study • Provides a utility for selecting candidate hotspots and experimental targets
Application of Hotspot Hunter • Our system can be customized and integrated into specialized databases • Tumor Antigen Database: http://research.i2r.a-star.edu.sg/Templar/DB/cancer_antigen/ • CandiVF - Candida albicans Virulence Factor Database Tongchusak et al., (2005) Int J Pep Res Ther.
Acknowledgment • Funding Agency • NIH, USA
Validation using experimental binders • Human pappilomavirus type 16 proteins E6 (Kast et al.,1994) • E6 hot-spot regions • HLA-A2 E6 7-34 (7-15, 18-26, and 26-34) • HLA-A2 E6 52-60 (single peptide) • HLA-A3 E6 33-67 (33-41, 42-50, and 59-67) • E6 75-101 (75-83, 89-97, and 93-101) • E6 125-151 (125-133 and 143-151) E6 HLA-A2 HLA-A3