Stacy Malaker

All Publications

• Canonic and Cross-reacrtive Binding of NK cell Inhibitory Recweptors to HLA-C Allotypes Is Dication by Peptides Bound to HLA-C FRONTIERS IN IMMUNOLOGY Sim, M. J., Malaker, S. A., Khan, A., Stowell, J. M., Shabanowitz, J., Peterson, M. E., Rajagopalan, S., Hunt, D. F., Altmann, D. M., Long, E. O., Boyton, R. J. 2017; 8

  Abstract

  Human natural killer (NK) cell activity is regulated by a family of killer cell immunoglobulin-like receptors (KIRs) that bind human leukocyte antigen (HLA) class I. Combinations of KIR and HLA genotypes are associated with disease, including susceptibility to viral infection and disorders of pregnancy. KIR2DL1 binds HLA-C alleles of group C2 (Lys(80)). KIR2DL2 and KIR2DL3 bind HLA-C alleles of group C1 (Asn(80)). However, this model cannot explain HLA-C allelic effects in disease or the impact of HLA-bound peptides. The goal of this study was to determine the extent to which the endogenous HLA-C peptide repertoire can influence the specific binding of inhibitory KIR to HLA-C allotypes.The impact of HLA-C bound peptide on inhibitory KIR binding was investigated taking advantage of the fact that HLA-C*05:01 (HLA-C group 2, C2) and HLA-C*08:02 (HLA-C group 1, C1) have identical sequences apart from the key KIR specificity determining epitope at residues 77 and 80. Endogenous peptides were eluted from HLA-C*05:01 and used to test the peptide dependence of KIR2DL1 and KIR2DL2/3 binding to HLA-C*05:01 and HLA-C*08:02 and subsequent impact on NK cell function. Specific binding of KIR2DL1 to the C2 allotype occurred with the majority of peptides tested. In contrast, KIR2DL2/3 binding to the C1 allotype occurred with only a subset of peptides. Cross-reactive binding of KIR2DL2/3 with the C2 allotype was restricted to even fewer peptides. Unexpectedly, two peptides promoted binding of the C2 allotype-specific KIR2DL1 to the C1 allotype. We showed that presentation of endogenous peptides or HIV Gag peptides by HLA-C can promote KIR cross-reactive binding.KIR2DL2/3 binding to C1 is more peptide selective than that of KIR2DL1 binding to C2, providing an explanation for KIR2DL3-C1 interactions appearing weaker than KIR2DL1-C2. In addition, cross-reactive binding of KIR is characterized by even higher peptide selectivity. We demonstrate a hierarchy of functional peptide selectivity of KIR-HLA-C interactions with relevance to NK cell biology and human disease associations. This selective peptide sequence-driven binding of KIR provides a potential mechanism for pathogen as well as self-peptide to modulate NK cell activation through altering levels of inhibition.

  View details for DOI 10.3389/fimmu.2017.00193

  View details for Web of Science ID 000396207100001

  View details for PubMedID 28352266

  View details for PubMedCentralID PMC5348643

• Peptide-binding motifs of two common equine class I MHC molecules in Thoroughbred horses. Immunogenetics Bergmann, T., Lindvall, M., Moore, E., Moore, E., Sidney, J., Miller, D., Tallmadge, R. L., Myers, P. T., Malaker, S. A., Shabanowitz, J., Osterrieder, N., Peters, B., Hunt, D. F., Antczak, D. F., Sette, A. 2017

  Abstract

  Quantitative peptide-binding motifs of MHC class I alleles provide a valuable tool to efficiently identify putative T cell epitopes. Detailed information on equine MHC class I alleles is still very limited, and to date, only a single equine MHC class I allele, Eqca-1*00101 (ELA-A3 haplotype), has been characterized. The present study extends the number of characterized ELA class I specificities in two additional haplotypes found commonly in the Thoroughbred breed. Accordingly, we here report quantitative binding motifs for the ELA-A2 allele Eqca-16*00101 and the ELA-A9 allele Eqca-1*00201. Utilizing analyses of endogenously bound and eluted ligands and the screening of positional scanning combinatorial libraries, detailed and quantitative peptide-binding motifs were derived for both alleles. Eqca-16*00101 preferentially binds peptides with aliphatic/hydrophobic residues in position 2 and at the C-terminus, and Eqca-1*00201 has a preference for peptides with arginine in position 2 and hydrophobic/aliphatic residues at the C-terminus. Interestingly, the Eqca-16*00101 motif resembles that of the human HLA A02-supertype, while the Eqca-1*00201 motif resembles that of the HLA B27-supertype and two macaque class I alleles. It is expected that the identified motifs will facilitate the selection of candidate epitopes for the study of immune responses in horses.

  View details for DOI 10.1007/s00251-017-0978-6

  View details for PubMedID 28315936

• Identification and Characterization of Complex Glycosylated Peptides Presented by the MHC Class II Processing Pathway in Melanoma JOURNAL OF PROTEOME RESEARCH Malaker, S. A., Ferracane, M. J., Depontieu, F. R., Zarling, A. L., Shabanowitz, J., Bai, D. L., Topalian, S. L., Engelhard, V. H., Hunt, D. F. 2017; 16 (1): 228-237

  View details for DOI 10.1021/acs.jproteome.6b00496

  View details for Web of Science ID 000391782100021

• Shared peptide binding of HLA Class I and II alleles associate with cutaneous nevirapine hypersensitivity and identify novel risk alleles. Scientific reports Pavlos, R., McKinnon, E. J., Ostrov, D. A., Peters, B., Buus, S., Koelle, D., Chopra, A., Schutte, R., Rive, C., Redwood, A., Restrepo, S., Bracey, A., Kaever, T., Myers, P., Speers, E., Malaker, S. A., Shabanowitz, J., Jing, Y., Gaudieri, S., Hunt, D. F., Carrington, M., Haas, D. W., Mallal, S., Phillips, E. J. 2017; 7 (1): 8653

  Abstract

  Genes of the human leukocyte antigen (HLA) system encode cell-surface proteins involved in regulation of immune responses, and the way drugs interact with the HLA peptide binding groove is important in the immunopathogenesis of T-cell mediated drug hypersensitivity syndromes. Nevirapine (NVP), is an HIV-1 antiretroviral with treatment-limiting hypersensitivity reactions (HSRs) associated with multiple class I and II HLA alleles. Here we utilize a novel analytical approach to explore these multi-allelic associations by systematically examining HLA molecules for similarities in peptide binding specificities and binding pocket structure. We demonstrate that primary predisposition to cutaneous NVP HSR, seen across ancestral groups, can be attributed to a cluster of HLA-C alleles sharing a common binding groove F pocket with HLA-C*04:01. An independent association with a group of class II alleles which share the HLA-DRB1-P4 pocket is also observed. In contrast, NVP HSR protection is afforded by a cluster of HLA-B alleles defined by a characteristic peptide binding groove B pocket. The results suggest drug-specific interactions within the antigen binding cleft can be shared across HLA molecules with similar binding pockets. We thereby provide an explanation for multiple HLA associations with cutaneous NVP HSR and advance insight into its pathogenic mechanisms.

  View details for DOI 10.1038/s41598-017-08876-0

  View details for PubMedID 28819312

  View details for PubMedCentralID PMC5561238

• Identification of glycopeptides as post-translationally modified neoantigens in leukemia. Cancer immunology research Malaker, S. A., Penny, S. A., Steadman, L. G., Myers, P. T., Loke, J., Raghavan, M., Bai, D. L., Shabanowitz, J., Hunt, D., Cobbold, M. 2017

  Abstract

  Leukemias are highly immunogenic but have a low mutational load, providing few mutated peptide targets. Thus, the identification of alternative neoantigens is a pressing need. Here, we identify 36 MHC class I-associated peptide antigens with O-linked β-N-acetylglucosamine (O-GlcNAc) modifications as candidate neoantigens, using three experimental approaches. Thirteen of these peptides were also detected with disaccharide units on the same residues and two contain either mono- and/or di-methylated arginine residues. A subset were linked with key cancer pathways, and these peptides were shared across all of the leukemia patient samples tested (5/5). Seven of the O-GlcNAc peptides were synthesized and five (71%) were shown to be associated with multifunctional memory T-cell responses in healthy donors. An O-GlcNAc-specific T-cell line specifically killed autologous cells pulsed with the modified peptide, but not the equivalent unmodified peptide. Therefore, these post-translationally modified neoantigens provide logical targets for cancer immunotherapy.

  View details for DOI 10.1158/2326-6066.CIR-16-0280

  View details for PubMedID 28314751

• MHC Class I-Associated Phosphopeptides Are the Targets of Memory-like Immunity in Leukemia SCIENCE TRANSLATIONAL MEDICINE Cobbold, M., De la Pena, H., Norris, A., Polefrone, J. M., Qian, J., English, A. M., Cummings, K. L., Penny, S., Turner, J. E., Cottine, J., Abelin, J. G., Malaker, S. A., Zarling, A. L., Huang, H., Goodyear, O., Freeman, S. D., Shabanowitz, J., Pratt, G., Craddock, C., Williams, M. E., Hunt, D. F., Engelhard, V. H. 2013; 5 (203)

  Abstract

  Deregulation of signaling pathways is a hallmark of malignant transformation. Signaling-associated phosphoproteins can be degraded to generate cancer-specific phosphopeptides that are presented by major histocompatibility complex (MHC) class I and II molecules and recognized by T cells; however, the contribution of these phosphoprotein-specific T cells to immune surveillance is unclear. We identified 95 phosphopeptides presented on the surface of primary hematological tumors and normal tissues, including 61 that were tumor-specific. Phosphopeptides were more prevalent on more aggressive and malignant samples. CD8(+) T cell lines specific for these phosphopeptides recognized and killed both leukemia cell lines and human leukocyte antigen-matched primary leukemia cells ex vivo. Notably, healthy individuals showed robust CD8(+) T cell responses against many of these phosphopeptides within the circulating memory compartment. This immunity was significantly reduced or absent in some leukemia patients. This reduction correlated with clinical outcome; however, immunity was restored after allogeneic stem cell transplantation. These results suggest that phosphopeptides may be targets of cancer immune surveillance in humans, and point to their importance for development of vaccine-based and T cell adoptive transfer immunotherapies.

  View details for DOI 10.1126/scitranslmed.3006061

  View details for Web of Science ID 000324589900004

  View details for PubMedID 24048523