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.
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.


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 Vbeta-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 IL2Rgamma(null) mice.


CONCLUSIONS
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.
Distler
Eva
E
Department of Medicine III-Hematology and Oncology, Johannes Gutenberg-University of Mainz, Mainz, Germany.
Wölfel
Catherine
C
Köhler
Sylvia
S
Nonn
Marion
M
Kaus
Nina
N
Schnürer
Elke
E
Meyer
Ralf G
RG
Wehler
Thomas C
TC
Huber
Christoph
C
Wölfel
Thomas
T
Hartwig
Udo F
UF
Herr
Wolfgang
W
D000483:Alleles; D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000166:cytology* / Q000378:metabolism; D002478:Cells, Cultured; D002999:Clone Cells; D003601:Cytotoxicity Tests, Immunologic; D005805:Genes, MHC Class I; D006801:Humans; D053631:Interleukin Receptor Common gamma Subunit / Q000235:genetics*; D019041:L-Selectin / Q000096:biosynthesis*; D015470:Leukemia, Myeloid, Acute / Q000097:blood / Q000276:immunology* / Q000517:prevention & control*; D051379:Mice; D016688:Mice, Inbred NOD; D018345:Mice, Knockout; D016513:Mice, SCID; D009368:Neoplasm Transplantation; D012015:Reference Standards; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology / Q000276:immunology / Q000637:transplantation*
D053631:Interleukin Receptor Common gamma Subunit; D019041:L-Selectin