No abstract

Infusions of natural killer (NK) cells are an emerging tool for cancer immunotherapy. The development of clinically applicable methods to produce large numbers of fully functional NK cells is a critical step to maximize the potential of this approach. We determined the capacity of the leukemia cell line K562 modified to express a membrane-bound form of interleukin (IL)-15 and 41BB ligand (K562-mb15-41BBL) to generate human NK cells with enhanced cytotoxicity. Sevenday coculture with irradiated K562-mb15-41BBL induced a median 21.6-fold expansion of CD56 + CD3 -NK cells from peripheral blood (range, 5.1-to 86.6-fold; n = 50), which was considerably superior to that produced by stimulation with IL-2, IL-12, IL-15, and/or IL-21 and caused no proliferation of CD3 + lymphocytes. Similar expansions could also be obtained from the peripheral blood of patients with acute leukemia undergoing therapy (n = 11). Comparisons of the gene expression profiles of the expanded NK cells and their unstimulated or IL-2-stimulated counterparts showed marked differences. The expanded NK cells were significantly more potent than unstimulated or IL-2-stimulated NK cells against acute myeloid leukemia cells in vitro. They could be detected for >1 month when injected into immunodeficient mice and could eradicate leukemia in murine models of acute myeloid leukemia. We therefore adapted the K562-mb15-41BBL stimulation method to large-scale clinical-grade conditions, generating large numbers of highly cytotoxic NK cells. The results that we report here provide rationale and practical platform for clinical testing of expanded and activated NK cells for cell therapy of cancer. [Cancer Res 2009;69(9):4010-7]

A B S T R A C T PurposeMethotrexate plasma concentration is related to its clinical effects. Our aim was to identify the genetic basis of interindividual variability in methotrexate pharmacokinetics in children with newly diagnosed acute lymphoblastic leukemia (ALL). Patients and MethodsWe performed a genome-wide analysis of 500,568 germline single-nucleotide polymorphisms (SNPs) to identify how inheritance affects methotrexate plasma disposition among 434 children with ALL who received 3,014 courses of methotrexate at 2 to 5 g/m 2 . SNPs were validated in an independent cohort of 206 patients. ResultsAdjusting for age, race, sex, and methotrexate regimen, the most significant associations were with SNPs in the organic anion transporter polypeptide, SLCO1B1. Two SNPs in SLCO1B1, rs11045879 (P ϭ 1.7 ϫ 10 Ϫ10 ) and rs4149081 (P ϭ 1.7 ϫ 10 Ϫ9 ), were in linkage disequilibrium (LD) with each other (r 2 ϭ 1) and with a functional polymorphism in SLCO1B1, T521C (rs4149056; r 2 Ͼ 0.84). rs11045879 and rs4149081 were validated in an independent cohort of 206 patients (P ϭ .018 and P ϭ .017), as were other SLCO1B1 SNPs residing in different LD blocks. SNPs in SLCO1B1 were also associated with GI toxicity (odds ratio, 15.3 to 16.4; P ϭ .03 to .004). ConclusionA genome-wide interrogation identified inherited variations in a plausible, yet heretofore lowpriority candidate gene, SLCO1B1, as important determinants of methotrexate's pharmacokinetics and clinical effects.