6533b833fe1ef96bd129c17a

RESEARCH PRODUCT

Acute myeloid leukemia (AML)-reactive cytotoxic T lymphocyte clones rapidly expanded from CD8(+) CD62L((high)+) T cells of healthy donors prevent AML engraftment in NOD/SCID IL2Rgamma(null) mice.

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Objective Current in vitro techniques for isolating leukemia-reactive cytotoxic T lymphocytes (CTLs) from healthy donors are of relatively low efficiency and yield responder populations with unknown biological significance. This study aimed at the development of a more reliable approach, allowing generation and expansion of acute myeloid leukemia (AML)-reactive CTLs using primary in vitro stimulation. Materials and Methods We established allogeneic mini-mixed lymphocyte-leukemia cultures (mini-MLLCs) by stimulating donor CD8 + T cells with human leukocyte antigen (HLA) class I–matched AML blasts in microtiter plates. Before culture, CD8 + T cells were separated into CD62L (high)+ and CD62L (low)+/neg subsets enriched for naive/central memory and effector memory cells, respectively. Results In eight different related and unrelated donor/AML pairs, numerous CTL populations were isolated that specifically lysed myeloid leukemias in association with various HLA-A, -B, or -C alleles. These CTLs expressed T-cell receptors of single Vβ-chain families, indicating their clonal origin. The majority of CTL clones were obtained from mini-MLLCs initiated with CD62L (high)+ cells. Using antigen-specific stimulation, multiple CTL populations were amplified to 10 8 –10 10 cells within 6 to 8 weeks. Three of four representative CTL clones were capable of completely preventing engraftment of human primary AML blasts in nonobese diabetic/severe combined immune deficient IL2Rγ null mice. Conclusion The mini-MLLC approach allows the efficient in vitro expansion of AML-reactive CTL clones from CD8 + CD62L (high)+ precursors of healthy donors. These CTLs can inhibit leukemia engraftment in immunodeficient mice, suggesting their potential biological relevance.