Evaluation of high-throughput genomic assays for the Fc gamma receptor locus

Chantal E. Hargreaves; Chisako Iriyama; Matthew J.J. Rose-Zerilli; Sietse Q. Nagelkerke; Khiyam Hussain; Rosalind Ganderton; Charlotte Lee; Lee R. Machado; Edward J. Hollox; Helen Parker; Kate V. Latham; Taco W. Kuijpers; Kathleen N. Potter; Sarah E. Coupland; Andrew Davies; Michael Stackpole; Melanie Oates; Andrew R. Pettitt; Martin J. Glennie; Mark S. Cragg; Jonathan C. Strefford

doi:10.1371/journal.pone.0142379

Evaluation of high-throughput genomic assays for the Fc gamma receptor locus

Chantal E. Hargreaves, Chisako Iriyama, Matthew J.J. Rose-Zerilli, Sietse Q. Nagelkerke, Khiyam Hussain, Rosalind Ganderton, Charlotte Lee, Lee R. Machado, Edward J. Hollox, Helen Parker, Kate V. Latham, Taco W. Kuijpers, Kathleen N. Potter, Sarah E. Coupland, Andrew Davies, Michael Stackpole, Melanie Oates, Andrew R. Pettitt, Martin J. Glennie, Mark S. CraggJonathan C. Strefford

Research output: Contribution to journal › Article › peer-review

12 Citations (Scopus)

Abstract

Cancer immunotherapy has been revolutionised by the use monoclonal antibodies (mAb) that function through their interaction with Fc gamma receptors (FcγRs). The low-affinity FcγR genes are highly homologous, map to a complex locus at 1p23 and harbour single nucleotide polymorphisms (SNPs) and copy number variation (CNV) that can impact on receptor function and response to therapeutic mAbs. This complexity can hinder accurate characterisation of the locus. We therefore evaluated and optimised a suite of assays for the genomic analysis of the FcγR locus amenable to peripheral blood mononuclear cells and formalin-fixed paraffin-embedded (FFPE) material that can be employed in a highthroughput manner. Assessment of TaqMan genotyping for FCGR2A-131H/R, FCGR3A-158F/V and FCGR2B-232I/T SNPs demonstrated the need for additional methods to discriminate genotypes for the FCGR3A-158F/V and FCGR2B-232I/T SNPs due to sequence homology and CNV in the region. A multiplex ligation-dependent probe amplification assay provided high quality SNP and CNV data in PBMC cases, but there was greater data variability in FFPE material in a manner that was predicted by the BIOMED-2 multiplex PCR protocol. In conclusion, we have evaluated a suite of assays for the genomic analysis of the FcγR locus that are scalable for application in large clinical trials of mAb therapy. These assays will ultimately help establish the importance of FcγR genetics in predicting response to antibody therapeutics.

Original language	English
Article number	e0142379
Journal	PloS one
Volume	10
Issue number	11
DOIs	https://doi.org/10.1371/journal.pone.0142379
Publication status	Published - 06-11-2015
Externally published	Yes

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Hargreaves, C. E., Iriyama, C., Rose-Zerilli, M. J. J., Nagelkerke, S. Q., Hussain, K., Ganderton, R., Lee, C., Machado, L. R., Hollox, E. J., Parker, H., Latham, K. V., Kuijpers, T. W., Potter, K. N., Coupland, S. E., Davies, A., Stackpole, M., Oates, M., Pettitt, A. R., Glennie, M. J., ... Strefford, J. C. (2015). Evaluation of high-throughput genomic assays for the Fc gamma receptor locus. PloS one, 10(11), [e0142379]. https://doi.org/10.1371/journal.pone.0142379

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title = "Evaluation of high-throughput genomic assays for the Fc gamma receptor locus",

abstract = "Cancer immunotherapy has been revolutionised by the use monoclonal antibodies (mAb) that function through their interaction with Fc gamma receptors (FcγRs). The low-affinity FcγR genes are highly homologous, map to a complex locus at 1p23 and harbour single nucleotide polymorphisms (SNPs) and copy number variation (CNV) that can impact on receptor function and response to therapeutic mAbs. This complexity can hinder accurate characterisation of the locus. We therefore evaluated and optimised a suite of assays for the genomic analysis of the FcγR locus amenable to peripheral blood mononuclear cells and formalin-fixed paraffin-embedded (FFPE) material that can be employed in a highthroughput manner. Assessment of TaqMan genotyping for FCGR2A-131H/R, FCGR3A-158F/V and FCGR2B-232I/T SNPs demonstrated the need for additional methods to discriminate genotypes for the FCGR3A-158F/V and FCGR2B-232I/T SNPs due to sequence homology and CNV in the region. A multiplex ligation-dependent probe amplification assay provided high quality SNP and CNV data in PBMC cases, but there was greater data variability in FFPE material in a manner that was predicted by the BIOMED-2 multiplex PCR protocol. In conclusion, we have evaluated a suite of assays for the genomic analysis of the FcγR locus that are scalable for application in large clinical trials of mAb therapy. These assays will ultimately help establish the importance of FcγR genetics in predicting response to antibody therapeutics.",

author = "Hargreaves, {Chantal E.} and Chisako Iriyama and Rose-Zerilli, {Matthew J.J.} and Nagelkerke, {Sietse Q.} and Khiyam Hussain and Rosalind Ganderton and Charlotte Lee and Machado, {Lee R.} and Hollox, {Edward J.} and Helen Parker and Latham, {Kate V.} and Kuijpers, {Taco W.} and Potter, {Kathleen N.} and Coupland, {Sarah E.} and Andrew Davies and Michael Stackpole and Melanie Oates and Pettitt, {Andrew R.} and Glennie, {Martin J.} and Cragg, {Mark S.} and Strefford, {Jonathan C.}",

note = "Publisher Copyright: {\textcopyright} 2015 Hargreaves et al.This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.",

year = "2015",

month = nov,

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doi = "10.1371/journal.pone.0142379",

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Hargreaves, CE, Iriyama, C, Rose-Zerilli, MJJ, Nagelkerke, SQ, Hussain, K, Ganderton, R, Lee, C, Machado, LR, Hollox, EJ, Parker, H, Latham, KV, Kuijpers, TW, Potter, KN, Coupland, SE, Davies, A, Stackpole, M, Oates, M, Pettitt, AR, Glennie, MJ, Cragg, MS & Strefford, JC 2015, 'Evaluation of high-throughput genomic assays for the Fc gamma receptor locus', PloS one, vol. 10, no. 11, e0142379. https://doi.org/10.1371/journal.pone.0142379

TY - JOUR

T1 - Evaluation of high-throughput genomic assays for the Fc gamma receptor locus

AU - Hargreaves, Chantal E.

AU - Iriyama, Chisako

AU - Rose-Zerilli, Matthew J.J.

AU - Nagelkerke, Sietse Q.

AU - Hussain, Khiyam

AU - Ganderton, Rosalind

AU - Lee, Charlotte

AU - Machado, Lee R.

AU - Hollox, Edward J.

AU - Parker, Helen

AU - Latham, Kate V.

AU - Kuijpers, Taco W.

AU - Potter, Kathleen N.

AU - Coupland, Sarah E.

AU - Davies, Andrew

AU - Stackpole, Michael

AU - Oates, Melanie

AU - Pettitt, Andrew R.

AU - Glennie, Martin J.

AU - Cragg, Mark S.

AU - Strefford, Jonathan C.

N1 - Publisher Copyright: © 2015 Hargreaves et al.This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

PY - 2015/11/6

Y1 - 2015/11/6

N2 - Cancer immunotherapy has been revolutionised by the use monoclonal antibodies (mAb) that function through their interaction with Fc gamma receptors (FcγRs). The low-affinity FcγR genes are highly homologous, map to a complex locus at 1p23 and harbour single nucleotide polymorphisms (SNPs) and copy number variation (CNV) that can impact on receptor function and response to therapeutic mAbs. This complexity can hinder accurate characterisation of the locus. We therefore evaluated and optimised a suite of assays for the genomic analysis of the FcγR locus amenable to peripheral blood mononuclear cells and formalin-fixed paraffin-embedded (FFPE) material that can be employed in a highthroughput manner. Assessment of TaqMan genotyping for FCGR2A-131H/R, FCGR3A-158F/V and FCGR2B-232I/T SNPs demonstrated the need for additional methods to discriminate genotypes for the FCGR3A-158F/V and FCGR2B-232I/T SNPs due to sequence homology and CNV in the region. A multiplex ligation-dependent probe amplification assay provided high quality SNP and CNV data in PBMC cases, but there was greater data variability in FFPE material in a manner that was predicted by the BIOMED-2 multiplex PCR protocol. In conclusion, we have evaluated a suite of assays for the genomic analysis of the FcγR locus that are scalable for application in large clinical trials of mAb therapy. These assays will ultimately help establish the importance of FcγR genetics in predicting response to antibody therapeutics.

AB - Cancer immunotherapy has been revolutionised by the use monoclonal antibodies (mAb) that function through their interaction with Fc gamma receptors (FcγRs). The low-affinity FcγR genes are highly homologous, map to a complex locus at 1p23 and harbour single nucleotide polymorphisms (SNPs) and copy number variation (CNV) that can impact on receptor function and response to therapeutic mAbs. This complexity can hinder accurate characterisation of the locus. We therefore evaluated and optimised a suite of assays for the genomic analysis of the FcγR locus amenable to peripheral blood mononuclear cells and formalin-fixed paraffin-embedded (FFPE) material that can be employed in a highthroughput manner. Assessment of TaqMan genotyping for FCGR2A-131H/R, FCGR3A-158F/V and FCGR2B-232I/T SNPs demonstrated the need for additional methods to discriminate genotypes for the FCGR3A-158F/V and FCGR2B-232I/T SNPs due to sequence homology and CNV in the region. A multiplex ligation-dependent probe amplification assay provided high quality SNP and CNV data in PBMC cases, but there was greater data variability in FFPE material in a manner that was predicted by the BIOMED-2 multiplex PCR protocol. In conclusion, we have evaluated a suite of assays for the genomic analysis of the FcγR locus that are scalable for application in large clinical trials of mAb therapy. These assays will ultimately help establish the importance of FcγR genetics in predicting response to antibody therapeutics.

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Evaluation of high-throughput genomic assays for the Fc gamma receptor locus

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