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Abacavir-reactive memory T cells are present in drug naïve individuals.

Lucas A, Lucas M, Strhyn A, Keane NM, McKinnon E, Pavlos R, Moran EM, Meyer-Pannwitt V, Gaudieri S, D'Orsogna L, Kalams S, Ostrov DA, Buus S, Peters B, Mallal S, Phillips E - PLoS ONE (2015)

Bottom Line: Fifty-five percent of individuals with HLA-B*57:01 exposed to the antiretroviral drug abacavir develop a hypersensitivity reaction (HSR) that has been attributed to naïve T-cell responses to neo-antigen generated by the drug.Abacavir reactive CD8+ T-cell responses were detected in vitro in one hundred percent of abacavir unexposed HLA-B*57:01 positive healthy donors.Abacavir-specific CD8+ T cells from such donors can be expanded from sorted memory, and sorted naïve, CD8+ T cells without need for autologous CD4+ T cells.

View Article: PubMed Central - PubMed

Affiliation: Institute for Immunology and Infectious Diseases, Murdoch University, Perth, Australia.

ABSTRACT

Background: Fifty-five percent of individuals with HLA-B*57:01 exposed to the antiretroviral drug abacavir develop a hypersensitivity reaction (HSR) that has been attributed to naïve T-cell responses to neo-antigen generated by the drug. Immunologically confirmed abacavir HSR can manifest clinically in less than 48 hours following first exposure suggesting that, at least in some cases, abacavir HSR is due to re-stimulation of a pre-existing memory T-cell population rather than priming of a high frequency naïve T-cell population.

Methods: To determine whether a pre-existing abacavir reactive memory T-cell population contributes to early abacavir HSR symptoms, we studied the abacavir specific naïve or memory T-cell response using HLA-B*57:01 positive HSR patients or healthy controls using ELISpot assay, intra-cellular cytokine staining and tetramer labelling.

Results: Abacavir reactive CD8+ T-cell responses were detected in vitro in one hundred percent of abacavir unexposed HLA-B*57:01 positive healthy donors. Abacavir-specific CD8+ T cells from such donors can be expanded from sorted memory, and sorted naïve, CD8+ T cells without need for autologous CD4+ T cells.

Conclusions: We propose that these pre-existing abacavir-reactive memory CD8+ T-cell responses must have been primed by earlier exposure to another foreign antigen and that these T cells cross-react with an abacavir-HLA-B*57:01-endogenous peptide ligand complex, in keeping with the model of heterologous immunity proposed in transplant rejection.

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Assessment of abacavir responsive memory or naïve T cells.A. Sorting gates used to collect total CD4+ and CD8+ memory phenotype (green polygon) and naïve phenotype T cells (blue rectangle) or B. CD8+ memory or naïve phenotype T cells only. C. Protocol used for the selection, priming and re-stimulation of sorted naïve and memory T cells from cryopreserved HLA-B*57:01 positive donor PBMC. D. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype T cells derived from sorting strategy A. Cultures were re-stimulated with APCs treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57), respectively. E. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype CD8+ T cells derived from sort strategy B. Cultures were re-stimulated with APCs as described above.
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pone.0117160.g004: Assessment of abacavir responsive memory or naïve T cells.A. Sorting gates used to collect total CD4+ and CD8+ memory phenotype (green polygon) and naïve phenotype T cells (blue rectangle) or B. CD8+ memory or naïve phenotype T cells only. C. Protocol used for the selection, priming and re-stimulation of sorted naïve and memory T cells from cryopreserved HLA-B*57:01 positive donor PBMC. D. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype T cells derived from sorting strategy A. Cultures were re-stimulated with APCs treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57), respectively. E. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype CD8+ T cells derived from sort strategy B. Cultures were re-stimulated with APCs as described above.

Mentions: We have previously demonstrated that robust abacavir treatment specific CD8+ T-cell responses can be reproducibly induced in in vitro 10 day cultures of PBMC from drug naïve HLA B*57:01 positive donors (2). We have used this culture protocol to examine the contributions of pre-existing memory T cells, and/or naïve T cells in the establishment of abacavir associated immunity, using abacavir-unexposed HLA-B*57:01 positive and negative PBMC from healthy donors, which included three of the four donors examined in Fig. 3. We examined the abacavir specific response of unsorted donor PBMC or sorted memory (Fig. 4A: Sort gate shaded green), or sorted naïve (Fig. 4A: Sort gate shaded blue) T cells or sorted CD8+ memory (Fig. 4B: Sort gate shaded green) or sorted CD8+ naïve T cells Fig. 4B: Sort gate shaded blue). Cultured cells were stimulated with irradiated C1R.B57 APC treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57) and cultured for 10 days. At that time aliquots of the cultures were re-stimulated with similarly treated APC to detect IFN-γ production from abacavir responsive CD8+ T cells as shown in Fig. 4C. Representative responses for memory and naïve cultures derived from a CD4+ and CD8+ T-cell sort (Fig. 4D) or a CD8+ T-cell sort (Fig. 4E) are shown.


Abacavir-reactive memory T cells are present in drug naïve individuals.

Lucas A, Lucas M, Strhyn A, Keane NM, McKinnon E, Pavlos R, Moran EM, Meyer-Pannwitt V, Gaudieri S, D'Orsogna L, Kalams S, Ostrov DA, Buus S, Peters B, Mallal S, Phillips E - PLoS ONE (2015)

Assessment of abacavir responsive memory or naïve T cells.A. Sorting gates used to collect total CD4+ and CD8+ memory phenotype (green polygon) and naïve phenotype T cells (blue rectangle) or B. CD8+ memory or naïve phenotype T cells only. C. Protocol used for the selection, priming and re-stimulation of sorted naïve and memory T cells from cryopreserved HLA-B*57:01 positive donor PBMC. D. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype T cells derived from sorting strategy A. Cultures were re-stimulated with APCs treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57), respectively. E. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype CD8+ T cells derived from sort strategy B. Cultures were re-stimulated with APCs as described above.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4326126&req=5

pone.0117160.g004: Assessment of abacavir responsive memory or naïve T cells.A. Sorting gates used to collect total CD4+ and CD8+ memory phenotype (green polygon) and naïve phenotype T cells (blue rectangle) or B. CD8+ memory or naïve phenotype T cells only. C. Protocol used for the selection, priming and re-stimulation of sorted naïve and memory T cells from cryopreserved HLA-B*57:01 positive donor PBMC. D. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype T cells derived from sorting strategy A. Cultures were re-stimulated with APCs treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57), respectively. E. Representative plots of in-vitro cultures of HLA-B*57:01 positive memory (left most pairs of plots) or naïve (right most pairs of plots) phenotype CD8+ T cells derived from sort strategy B. Cultures were re-stimulated with APCs as described above.
Mentions: We have previously demonstrated that robust abacavir treatment specific CD8+ T-cell responses can be reproducibly induced in in vitro 10 day cultures of PBMC from drug naïve HLA B*57:01 positive donors (2). We have used this culture protocol to examine the contributions of pre-existing memory T cells, and/or naïve T cells in the establishment of abacavir associated immunity, using abacavir-unexposed HLA-B*57:01 positive and negative PBMC from healthy donors, which included three of the four donors examined in Fig. 3. We examined the abacavir specific response of unsorted donor PBMC or sorted memory (Fig. 4A: Sort gate shaded green), or sorted naïve (Fig. 4A: Sort gate shaded blue) T cells or sorted CD8+ memory (Fig. 4B: Sort gate shaded green) or sorted CD8+ naïve T cells Fig. 4B: Sort gate shaded blue). Cultured cells were stimulated with irradiated C1R.B57 APC treated with abacavir (C1R.B57.ABC) or untreated (C1R.B57) and cultured for 10 days. At that time aliquots of the cultures were re-stimulated with similarly treated APC to detect IFN-γ production from abacavir responsive CD8+ T cells as shown in Fig. 4C. Representative responses for memory and naïve cultures derived from a CD4+ and CD8+ T-cell sort (Fig. 4D) or a CD8+ T-cell sort (Fig. 4E) are shown.

Bottom Line: Fifty-five percent of individuals with HLA-B*57:01 exposed to the antiretroviral drug abacavir develop a hypersensitivity reaction (HSR) that has been attributed to naïve T-cell responses to neo-antigen generated by the drug.Abacavir reactive CD8+ T-cell responses were detected in vitro in one hundred percent of abacavir unexposed HLA-B*57:01 positive healthy donors.Abacavir-specific CD8+ T cells from such donors can be expanded from sorted memory, and sorted naïve, CD8+ T cells without need for autologous CD4+ T cells.

View Article: PubMed Central - PubMed

Affiliation: Institute for Immunology and Infectious Diseases, Murdoch University, Perth, Australia.

ABSTRACT

Background: Fifty-five percent of individuals with HLA-B*57:01 exposed to the antiretroviral drug abacavir develop a hypersensitivity reaction (HSR) that has been attributed to naïve T-cell responses to neo-antigen generated by the drug. Immunologically confirmed abacavir HSR can manifest clinically in less than 48 hours following first exposure suggesting that, at least in some cases, abacavir HSR is due to re-stimulation of a pre-existing memory T-cell population rather than priming of a high frequency naïve T-cell population.

Methods: To determine whether a pre-existing abacavir reactive memory T-cell population contributes to early abacavir HSR symptoms, we studied the abacavir specific naïve or memory T-cell response using HLA-B*57:01 positive HSR patients or healthy controls using ELISpot assay, intra-cellular cytokine staining and tetramer labelling.

Results: Abacavir reactive CD8+ T-cell responses were detected in vitro in one hundred percent of abacavir unexposed HLA-B*57:01 positive healthy donors. Abacavir-specific CD8+ T cells from such donors can be expanded from sorted memory, and sorted naïve, CD8+ T cells without need for autologous CD4+ T cells.

Conclusions: We propose that these pre-existing abacavir-reactive memory CD8+ T-cell responses must have been primed by earlier exposure to another foreign antigen and that these T cells cross-react with an abacavir-HLA-B*57:01-endogenous peptide ligand complex, in keeping with the model of heterologous immunity proposed in transplant rejection.

Show MeSH
Related in: MedlinePlus