Publications by authors named "Dario Campana"

134 Publications

Phase I Trial of Expanded, Activated Autologous NK-cell Infusions with Trastuzumab in Patients with HER2-positive Cancers.

Clin Cancer Res 2020 Sep 10;26(17):4494-4502. Epub 2020 Jun 10.

Department of Paediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Purpose: Natural killer (NK) cells exert antibody-dependent cell cytotoxicity (ADCC). We infused expanded, activated autologous NK cells to potentiate trastuzumab-mediated ADCC in patients with HER2-positive malignancies.

Patients And Methods: In a phase I trial, patients with treatment-refractory HER2-positive solid tumors received trastuzumab, with or without bevacizumab, and autologous NK cells expanded by 10-day coculture with K562-mb15-41BBL cells. Primary objectives included safety and recommended phase II dose determination; secondary objectives included monitoring NK-cell activity and RECIST antitumor efficacy.

Results: In 60 cultures with cells from 31 subjects, median NK-cell expansion from peripheral blood was 340-fold (range, 91-603). NK cells expressed high levels of CD16, the mediator of ADCC, and exerted powerful killing of trastuzumab-targeted cells. In the 22 subjects enrolled in phase I dose escalation, trastuzumab plus NK cells were well tolerated; MTD was not reached. Phase IB ( = 9) included multiple cycles of NK cells (1 × 10/kg) and addition of bevacizumab. Although no objective response was observed, 6 of 19 subjects who received at least 1 × 10/kg NK cells at cycle 1 had stable disease for ≥6 months (median, 8.8 months; range 6.0-12.0). One patient, the only one with the high-affinity F158V CD16 variant, had a partial response. Peripheral blood NK cells progressively downregulated CD16 postinfusion; paired tumor biopsies showed increased NK cells, lymphocytic infiltrates, and apoptosis posttreatment.

Conclusions: NK-cell therapy in combination with trastuzumab was well tolerated, with target engagement and preliminary antitumor activity, supporting continued assessment of this approach in phase II trials.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0768DOI Listing
September 2020

Chimeric antigen receptor-T cells with cytokine neutralizing capacity.

Blood Adv 2020 04;4(7):1419-1431

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Infusion of T lymphocytes expressing chimeric antigen receptors (CARs) can produce extraordinary antitumor activity in patients with leukemia, lymphoma, and myeloma. The signaling mechanisms activating T cells and provoking tumor cell killing also trigger cytokine secretion and macrophage activation, leading to cytokine release syndrome (CRS). CRS is a serious side effect of CAR-T cells, and proinflammatory interleukin-6 (IL-6) is central to its pathogenesis. To endow T cells with anti-CRS activity, we designed a nonsignaling membrane-bound IL-6 receptor (mbaIL6) constituted by a single chain variable fragment derived from an anti-IL-6 antibody linked to a transmembrane anchoring peptide. We found that mbaIL6 expressed on the surface of T cells could rapidly remove IL-6 from the culture supernatant. IL-6 removal was proportional to the number of mbaIL6+ cells, increased with T-cell proliferation, and neutralized IL-6 signaling and function. A construct encoding for mbaIL6 and an anti-CD19-41BB-CD3ζ CAR allowed simultaneous expression of both receptors. T cells with mbaIL6 and CAR neutralized macrophage-derived IL-6 while exerting powerful antitumor activity. Cytotoxicity and proliferation were identical to those of cells expressing CAR alone in vitro, and CAR-T cells were effective in xenograft models regardless of mbaIL6 expression. Levels of human IL-6 in mice, however, were greatly reduced if T cells expressed both receptors instead of CAR alone. Thus, CAR-T cells with on-board capacity to extinguish IL-6 represent a new approach to prevent CRS and suppress its severity without affecting the antitumor potential of CAR-T cells.
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http://dx.doi.org/10.1182/bloodadvances.2019001287DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7160280PMC
April 2020

Reduced-dose intensity therapy for pediatric lymphoblastic leukemia: long-term results of the Recife RELLA05 pilot study.

Blood 2020 04;135(17):1458-1466

Department of Global Pediatric Medicine.

Treatment-related mortality is common among children with acute lymphoblastic leukemia (ALL) treated in poor-resource settings. We applied a simplified flow cytometric assay to identify patients with precursor B-cell ALL (B-ALL) at very low risk (VLR) of relapse and treated them with a reduced-intensity treatment plan (RELLA05). VLR criteria include favorable presenting features (age ≥ 1 and < 10 years), white blood cell count of <50 ×109/L, lack of extramedullary leukemia, and minimal residual disease level of <0.01% on remission induction day 19. Except for 2 doses of daunorubicin, treatment of patients with VLR B-ALL consisted of a combination of agents with relatively low myelotoxicity profiles, including corticosteroids, vincristine, L-asparaginase, methotrexate, and 6-mercaptopurine. Cyclophosphamide, systemic cytarabine, and central nervous system radiotherapy were not used. Of 454 patients with ALL treated at the Instituto de Medicina Integral Professor Fernando Figueira in Recife, Brazil, between December 2005 and June 2015, 101 were classified as having VLR B-ALL. There were no cases of death resulting from toxicity or treatment abandonment during remission induction. At a median follow-up of 6.6 years, there were 8 major adverse events: 6 relapses, 1 treatment-related death (from septicemia) during remission, and 1 secondary myeloid leukemia. The estimated 5-year event-free and overall survival rates were 92.0% ± 3.9% and 96.0% ± 2.8%, respectively. The 5-year cumulative risk of relapse was 4.24% ± 2.0%. The treatment was well tolerated. Episodes of neutropenia were of short duration. Patients with B-ALL selected by a combination of presenting features and degree of early response can be successfully treated with a mildly myelosuppressive chemotherapy regimen.
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http://dx.doi.org/10.1182/blood.2019004215DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7180080PMC
April 2020

NK cells for cancer immunotherapy.

Nat Rev Drug Discov 2020 03 6;19(3):200-218. Epub 2020 Jan 6.

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.

Natural killer (NK) cells can swiftly kill multiple adjacent cells if these show surface markers associated with oncogenic transformation. This property, which is unique among immune cells, and their capacity to enhance antibody and T cell responses support a role for NK cells as anticancer agents. Although tumours may develop several mechanisms to resist attacks from endogenous NK cells, ex vivo activation, expansion and genetic modification of NK cells can greatly increase their antitumour activity and equip them to overcome resistance. Some of these methods have been translated into clinical-grade platforms and support clinical trials of NK cell infusions in patients with haematological malignancies or solid tumours, which have yielded encouraging results so far. The next generation of NK cell products will be engineered to enhance activating signals and proliferation, suppress inhibitory signals and promote their homing to tumours. These modifications promise to significantly increase their clinical activity. Finally, there is emerging evidence of increased NK cell-mediated tumour cell killing in the context of molecularly targeted therapies. These observations, in addition to the capacity of NK cells to magnify immune responses, suggest that NK cells are poised to become key components of multipronged therapeutic strategies for cancer.
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http://dx.doi.org/10.1038/s41573-019-0052-1DOI Listing
March 2020

Engineering of Natural Killer Cells for Clinical Application.

Methods Mol Biol 2020 ;2097:91-105

Departments of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.

The clinical success of chimeric antigen receptor-directed T cells in leukemia and lymphoma has boosted the interest in cellular therapy of cancer. It has been known for nearly half a century that a subset of lymphocytes called natural killer (NK) cells can recognize and kill cancer cells, but their clinical potential as therapeutics has not yet been fully explored. Progress in methods to expand and genetically modify human NK cells has resulted in technologies that allow the production of large numbers of highly potent cells with specific anticancer activity. Here, we describe clinically applicable protocols for NK cell engineering, including expansion of NK cells and genetic modification using electroporation of messenger RNA.
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http://dx.doi.org/10.1007/978-1-0716-0203-4_6DOI Listing
January 2021

Specific stimulation of T lymphocytes with erythropoietin for adoptive immunotherapy.

Blood 2020 02;135(9):668-679

Department of Paediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

In adoptive T-cell immunotherapy of cancer, expansion and persistence of effector cells is a key determinant of response. We tested whether T lymphocytes could be rendered sensitive to erythropoietin (Epo) through ectopic expression of its wild-type receptor or a truncated form (EpoRm), which augments Epo signaling in erythrocyte progenitors. Both receptors could be expressed in human T lymphocytes; Epo ligation induced STAT5 phosphorylation, which was abrogated by nontoxic concentrations of the JAK1/2 inhibitor ruxolitinib. EpoRm had higher expression and triggered more potent stimulation than its wild-type counterpart, including superior T-cell survival and proliferation. Using a bicistronic vector, we expressed EpoRm together with an anti-CD19-41BB-CD3ζ chimeric antigen receptor (CAR), while maintaining the functions of each receptor. In the presence of Epo, EpoRm-CAR T cells had greater ex vivo expansion than CAR T cells and killed CD19+ leukemic cells more effectively in long-term cultures. In immunodeficient mice, physiologic levels of murine Epo were sufficient to preferentially expand EpoRm-CAR T cells, yielding a significantly higher antileukemic activity. Thus, outfitting adoptive T cells with EpoRm should yield greater effector-to-target ratios with a smaller number of infused cells; Epo or ruxolitinib administration could be used to adjust their levels postinfusion, maximizing antitumor activity and minimizing toxicity.
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http://dx.doi.org/10.1182/blood.2019001645DOI Listing
February 2020

Improved CNS Control of Childhood Acute Lymphoblastic Leukemia Without Cranial Irradiation: St Jude Total Therapy Study 16.

J Clin Oncol 2019 12 28;37(35):3377-3391. Epub 2019 Oct 28.

St Jude Children's Research Hospital, Memphis, TN.

Purpose: Despite contemporary treatment, up to 10% of children with acute lymphoblastic leukemia still experience relapse. We evaluated whether a higher dosage of PEG-asparaginase and early intensification of triple intrathecal therapy would improve systemic and CNS control.

Patients And Methods: Between 2007 and 2017, 598 consecutive patients age 0 to 18 years received risk-directed chemotherapy without prophylactic cranial irradiation in the St Jude Total Therapy Study 16. Patients were randomly assigned to receive PEG-asparaginase 3,500 U/m versus the conventional 2,500 U/m. Patients presenting features that were associated with increased risk of CNS relapse received two extra doses of intrathecal therapy during the first 2 weeks of remission induction.

Results: The 5-year event-free survival and overall survival rates for the 598 patients were 88.2% (95% CI, 84.9% to 91.5%) and 94.1% (95% CI, 91.7% to 96.5%), respectively. Cumulative risk of any-isolated or combined-CNS relapse was 1.5% (95% CI, 0.5% to 2.5%). Higher doses of PEG-asparaginase did not affect treatment outcome. T-cell phenotype was the only independent risk factor for any CNS relapse (hazard ratio, 5.15; 95% CI, 1.3 to 20.6; = . 021). Among 359 patients with features that were associated with increased risk for CNS relapse, the 5-year rate of any CNS relapse was significantly lower than that among 248 patients with the same features treated in the previous Total Therapy Study 15 (1.8% [95% CI, 0.4% to 3.3%] 5.7% [95% CI, 2.8% to 8.6%]; = .008). There were no significant differences in the cumulative risk of seizure or infection during induction between patients who did or did not receive the two extra doses of intrathecal treatment.

Conclusion: Higher doses of PEG-asparaginase failed to improve outcome, but additional intrathecal therapy during early induction seemed to contribute to improved CNS control without excessive toxicity for high-risk patients.
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http://dx.doi.org/10.1200/JCO.19.01692DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7351342PMC
December 2019

Pathologic Features of Down Syndrome Myelodysplastic Syndrome and Acute Myeloid Leukemia: A Report From the Children's Oncology Group Protocol AAML0431.

Arch Pathol Lab Med 2020 04 20;144(4):466-472. Epub 2019 Aug 20.

From the Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee (Drs Mast, Mosse, Jones, and Head); the Division of Hematology/Oncology, Children's Hospital of Michigan, Wayne State University, Detroit (Dr Taub); the Department of Biostatistics, University of Southern California, Monrovia (Dr Alonzo and Mr Wang); the Division of Hematology/Oncology/Bone Marrow Transplantation, Children's Mercy Hospital, Kansas City, Missouri (Dr Gamis); the Pathology and Laboratory Medicine Service, VA Tennessee Valley Healthcare System, Nashville (Dr Mosse); the Department of Pediatrics, University of New Mexico, Albuquerque (Dr Mathew); the Division of Hematology-Oncology, IWK Health Centre, Halifax, Nova Scotia, Canada (Dr Berman); the Departments of Oncology (Dr Campana and Ms Coustan-Smith) and Pathology (Dr Raimondi), St. Jude Children's Research Hospital, Memphis, Tennessee; the Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, The National University Cancer Institute, NUH Medical Centre, Singapore (Dr Campana and Ms Coustan-Smith); the Department of Laboratory Medicine and Pathology, University of Minnesota Medical School, Minneapolis (Dr Hirsch); the Department of Pediatrics, University of Toronto, Toronto, Ontario, Canada (Dr Hitzler); and the Division of Hematology/Oncology, The Hospital for Sick Children Developmental and Stem Cell Biology, The Hospital for Sick Children Research Institute, Toronto, Ontario, Canada (Dr Hitzler). Dr Mast now has a joint appointment at the Pathology and Laboratory Medicine Service, VA Tennessee Valley Healthcare System, Nashville. Dr Mathew is currently at the Department of Pediatrics, Presbyterian Health Services, Albuquerque, New Mexico. Dr Berman is currently at the Department of Pediatrics, Dalhousie University, Halifax, Nova Scotia, Canada. Dr Jones is currently at Pathgroup Labs, Nashville, Tennessee. Dr Campana is no longer at the Department of Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee. Ms Coustan-Smith is no longer at the Department of Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee.

Context.—: Detailed diagnostic features of acute myeloid leukemia in Down syndrome are lacking, leading to potential misdiagnoses as standard acute myeloid leukemia occurring in patients with Down syndrome.

Objective.—: To evaluate diagnostic features of acute myeloid leukemia and myelodysplastic syndrome in patients with Down syndrome.

Design.—: Diagnostic bone marrow samples from 163 patients enrolled in the Children's Oncology Group study AAML0431 were evaluated by using central morphologic review and institutional immunophenotyping. Results were compared to overall survival, event-free survival, mutation status, cytogenetics, and minimal residual disease results.

Results.—: Sixty myelodysplastic syndrome and 103 acute myeloid leukemia samples were reviewed. Both had distinctive features compared to those of patients without Down syndrome. They showed megakaryocytic and erythroid but little myeloid dysplasia, and marked megakaryocytic hyperplasia with unusual megakaryocyte morphology. In acute myeloid leukemia cases, megakaryoblastic differentiation of blasts was most common (54 of 103, 52%); other cases showed erythroblastic (11 of 103, 11%), mixed erythroid/megakaryoblastic (20 of 103, 19%), or no differentiation (10 of 103, 10%). Myelodysplastic syndrome and acute myeloid leukemia cases had similar event-free survival and overall survival. Leukemic subgroups showed interesting, but not statistically significant, trends for survival and minimal residual disease. Cases with institutional diagnoses of French American British M1-5 morphology showed typical features of Down syndrome disease, with survival approaching that of other cases.

Conclusions.—: Myelodysplastic syndrome and acute myeloid leukemia in Down syndrome display features that allow discrimination from standard cases of disease. These distinctions are important for treatment decisions, and for understanding disease pathogenesis. We propose specific diagnostic criteria for Down syndrome-related subtypes of acute myeloid leukemia and myelodysplastic syndrome.
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http://dx.doi.org/10.5858/arpa.2018-0526-OADOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7031019PMC
April 2020

Blocking expression of inhibitory receptor NKG2A overcomes tumor resistance to NK cells.

J Clin Invest 2019 05 12;129(5):2094-2106. Epub 2019 Mar 12.

Department of Pediatrics and National University Cancer Institute Singapore, National University of Singapore, Singapore.

A key mechanism of tumor resistance to immune cells is mediated by expression of peptide-loaded HLA-E in tumor cells, which suppresses natural killer (NK) cell activity via ligation of the NK inhibitory receptor CD94/NKG2A. Gene expression data from approximately 10,000 tumor samples showed widespread HLAE expression, with levels correlating with those of KLRC1 (NKG2A) and KLRD1 (CD94). To bypass HLA-E inhibition, we developed a way to generate highly functional NK cells lacking NKG2A. Constructs containing a single-chain variable fragment derived from an anti-NKG2A antibody were linked to endoplasmic reticulum-retention domains. After retroviral transduction in human peripheral blood NK cells, these NKG2A Protein Expression Blockers (PEBLs) abrogated NKG2A expression. The resulting NKG2Anull NK cells had higher cytotoxicity against HLA-E-expressing tumor cells. Transduction of anti-NKG2A PEBL produced more potent cytotoxicity than interference with an anti-NKG2A antibody and prevented de novo NKG2A expression, without affecting NK cell proliferation. In immunodeficient mice, NKG2Anull NK cells were significantly more powerful than NKG2A+ NK cells against HLA-E-expressing tumors. Thus, NKG2A downregulation evades the HLA-E cancer immune-checkpoint, and increases the anti-tumor activity of NK cell infusions. Because this strategy is easily adaptable to current protocols for clinical-grade immune cell processing, its clinical testing is feasible and warranted.
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http://dx.doi.org/10.1172/JCI123955DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486333PMC
May 2019

No evidence that G6PD deficiency affects the efficacy or safety of daunorubicin in acute lymphoblastic leukemia induction therapy.

Pediatr Blood Cancer 2019 06 7;66(6):e27681. Epub 2019 Mar 7.

Department of Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee.

Background/objectives: Anthracyclines are used in induction therapy of pediatric acute lymphoblastic leukemia (ALL) and are known to generate oxidative stress; whether this translates into enhanced antileukemic activity or hemolytic effects in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency is unknown.

Design/methods: Among 726 pediatric patients with newly diagnosed ALL treated at St. Jude Children's Research Hospital, 22 had deficient G6PD activity. We compared the prevalence of positive minimal residual disease (MRD) ≥1% at Day 15/Day 19 of induction or ≥0.01% at Day 42/Day 46 (end of induction) and the number of red blood cell (RBC) transfusions after daunorubicin in induction between patients with or without G6PD deficiency, adjusting for ALL risk group, treatment protocol, age, and gender.

Results: There was no difference in Day 15/19 (P = 1) or end of induction MRD (P = 0.76) nor in the number of RBC transfusions (P = 0.73); the lack of association with MRD was confirmed in a dataset of 1192 newly diagnosed male patients enrolled in a Children's Oncology Group trial (P = 0.78).

Conclusion: We found no evidence that G6PD deficiency affects daunorubicin activity during induction treatment for ALL.
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http://dx.doi.org/10.1002/pbc.27681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6518412PMC
June 2019

Reverse-engineering flow-cytometry gating strategies for phenotypic labelling and high-performance cell sorting.

Bioinformatics 2019 01;35(2):301-308

Singapore Immunology Network, Agency for Science Technology and Research, Singapore.

Motivation: Recent flow and mass cytometers generate datasets of dimensions 20 to 40 and a million single cells. From these, many tools facilitate the discovery of new cell populations associated with diseases or physiology. These new cell populations require the identification of new gating strategies, but gating strategies become exponentially more difficult to optimize when dimensionality increases. To facilitate this step, we developed Hypergate, an algorithm which given a cell population of interest identifies a gating strategy optimized for high yield and purity.

Results: Hypergate achieves higher yield and purity than human experts, Support Vector Machines and Random-Forests on public datasets. We use it to revisit some established gating strategies for the identification of innate lymphoid cells, which identifies concise and efficient strategies that allow gating these cells with fewer parameters but higher yield and purity than the current standards. For phenotypic description, Hypergate's outputs are consistent with fields' knowledge and sparser than those from a competing method.

Availability And Implementation: Hypergate is implemented in R and available on CRAN. The source code is published at http://github.com/ebecht/hypergate under an Open Source Initiative-compliant licence.

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/bty491DOI Listing
January 2019

A novel λ integrase-mediated seamless vector transgenesis platform for therapeutic protein expression.

Nucleic Acids Res 2018 09;46(16):e99

School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551.

Advances in stem cell engineering, gene therapy and molecular medicine often involve genome engineering at a cellular level. However, functionally large or multi transgene cassette insertion into the human genome still remains a challenge. Current practices such as random transgene integration or targeted endonuclease-based genome editing are suboptimal and might pose safety concerns. Taking this into consideration, we previously developed a transgenesis tool derived from phage λ integrase (Int) that precisely recombines large plasmid DNA into an endogenous sequence found in human Long INterspersed Elements-1 (LINE-1). Despite this advancement, biosafety concerns associated with bacterial components of plasmids, enhanced uptake and efficient transgene expression remained problematic. We therefore further improved and herein report a more superior Int-based transgenesis tool. This novel Int platform allows efficient and easy derivation of sufficient amounts of seamless supercoiled transgene vectors from conventional plasmids via intramolecular recombination as well as subsequent intermolecular site-specific genome integration into LINE-1. Furthermore, we identified certain LINE-1 as preferred insertion sites for Int-mediated seamless vector transgenesis, and showed that targeted anti-CD19 chimeric antigen receptor gene integration achieves high-level sustained transgene expression in human embryonic stem cell clones for potential downstream therapeutic applications.
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http://dx.doi.org/10.1093/nar/gky500DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6144826PMC
September 2018

Universal monitoring of minimal residual disease in acute myeloid leukemia.

JCI Insight 2018 05 3;3(9). Epub 2018 May 3.

Department of Pediatrics, National University of Singapore, Singapore.

Background: Optimal management of acute myeloid leukemia (AML) requires monitoring of treatment response, but minimal residual disease (MRD) may escape detection. We sought to identify distinctive features of AML cells for universal MRD monitoring.

Methods: We compared genome-wide gene expression of AML cells from 157 patients with that of normal myeloblasts. Markers encoded by aberrantly expressed genes, including some previously associated with leukemia stem cells, were studied by flow cytometry in 240 patients with AML and in nonleukemic myeloblasts from 63 bone marrow samples.

Results: Twenty-two (CD9, CD18, CD25, CD32, CD44, CD47, CD52, CD54, CD59, CD64, CD68, CD86, CD93, CD96, CD97, CD99, CD123, CD200, CD300a/c, CD366, CD371, and CX3CR1) markers were aberrantly expressed in AML. Leukemia-associated profiles defined by these markers extended to immature CD34+CD38- AML cells; expression remained stable during treatment. The markers yielded MRD measurements matching those of standard methods in 208 samples from 52 patients undergoing chemotherapy and revealed otherwise undetectable MRD. They allowed MRD monitoring in 129 consecutive patients, yielding prognostically significant results. Using a machine-learning algorithm to reduce high-dimensional data sets to 2-dimensional data, the markers allowed a clear visualization of MRD and could detect 1 leukemic cell among more than 100,000 normal cells.

Conclusion: The markers uncovered in this study allow universal and sensitive monitoring of MRD in AML. In combination with contemporary analytical tools, the markers improve the discrimination between leukemic and normal cells, thus facilitating data interpretation and, hence, the reliability of MRD results.

Funding: National Cancer Institute (CA60419 and CA21765); American Lebanese Syrian Associated Charities; National Medical Research Council of Singapore (1299/2011); Viva Foundation for Children with Cancer, Children's Cancer Foundation, Tote Board & Turf Club, and Lee Foundation of Singapore.
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http://dx.doi.org/10.1172/jci.insight.98561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6012500PMC
May 2018

A novel method to generate T-cell receptor-deficient chimeric antigen receptor T cells.

Blood Adv 2018 03;2(5):517-528

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Practical methods are needed to increase the applicability and efficacy of chimeric antigen receptor (CAR) T-cell therapies. Using donor-derived CAR-T cells is attractive, but expression of endogenous T-cell receptors (TCRs) carries the risk for graft-versus-host-disease (GVHD). To remove surface TCRαβ, we combined an antibody-derived single-chain variable fragment specific for CD3ε with 21 different amino acid sequences predicted to retain it intracellularly. After transduction in T cells, several of these protein expression blockers (PEBLs) colocalized intracellularly with CD3ε, blocking surface CD3 and TCRαβ expression. In 25 experiments, median TCRαβ expression in T lymphocytes was reduced from 95.7% to 25.0%; CD3/TCRαβ cell depletion yielded virtually pure TCRαβ-negative T cells. Anti-CD3ε PEBLs abrogated TCRαβ-mediated signaling, without affecting immunophenotype or proliferation. In anti-CD3ε PEBL-T cells, expression of an anti-CD19-41BB-CD3ζ CAR induced cytokine secretion, long-term proliferation, and CD19 leukemia cell killing, at rates meeting or exceeding those of CAR-T cells with normal CD3/TCRαβ expression. In immunodeficient mice, anti-CD3ε PEBL-T cells had markedly reduced GVHD potential; when transduced with anti-CD19 CAR, these T cells killed engrafted leukemic cells. PEBL blockade of surface CD3/TCRαβ expression is an effective tool to prepare allogeneic CAR-T cells. Combined PEBL and CAR expression can be achieved in a single-step procedure, is easily adaptable to current cell manufacturing protocols, and can be used to target other T-cell molecules to further enhance CAR-T-cell therapies.
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http://dx.doi.org/10.1182/bloodadvances.2017012823DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5851418PMC
March 2018

Blockade of CD7 expression in T cells for effective chimeric antigen receptor targeting of T-cell malignancies.

Blood Adv 2017 Nov 21;1(25):2348-2360. Epub 2017 Nov 21.

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Effective immunotherapies for T-cell malignancies are lacking. We devised a novel approach based on chimeric antigen receptor (CAR)-redirected T lymphocytes. We selected CD7 as a target because of its consistent expression in T-cell acute lymphoblastic leukemia (T-ALL), including the most aggressive subtype, early T-cell precursor (ETP)-ALL. In 49 diagnostic T-ALL samples (including 14 ETP-ALL samples), median CD7 expression was >99%; CD7 expression remained high at relapse (n = 14), and during chemotherapy (n = 54). We targeted CD7 with a second-generation CAR (anti-CD7-41BB-CD3ζ), but CAR expression in T lymphocytes caused fratricide due to the presence of CD7 in the T cells themselves. To downregulate CD7 and control fratricide, we applied a new method (protein expression blocker [PEBL]), based on an anti-CD7 single-chain variable fragment coupled with an intracellular retention domain. Transduction of anti-CD7 PEBL resulted in virtually instantaneous abrogation of surface CD7 expression in all transduced T cells; 2.0% ± 1.7% were CD7 vs 98.1% ± 1.5% of mock-transduced T cells (n = 5; < .0001). PEBL expression did not impair T-cell proliferation, interferon-γ and tumor necrosis factor-α secretion, or cytotoxicity, and eliminated CAR-mediated fratricide. PEBL-CAR T cells were highly cytotoxic against CD7 leukemic cells in vitro and were consistently more potent than CD7 T cells spared by fratricide. They also showed strong anti-leukemic activity in cell line- and patient-derived T-ALL xenografts. The strategy described in this study fits well with existing clinical-grade cell manufacturing processes and can be rapidly implemented for the treatment of patients with high-risk T-cell malignancies.
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http://dx.doi.org/10.1182/bloodadvances.2017009928DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5729624PMC
November 2017

Minimal residual disease in pediatric ALL.

Oncotarget 2017 Oct 13;8(45):78251-78252. Epub 2017 Sep 13.

Ching-Hon Pui: Departments of Oncology and Pathology, St. Jude Children's Research Hospital, Memphis, TN, USA.

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http://dx.doi.org/10.18632/oncotarget.20856DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5667954PMC
October 2017

Improved outcomes for myeloid leukemia of Down syndrome: a report from the Children's Oncology Group AAML0431 trial.

Blood 2017 06 7;129(25):3304-3313. Epub 2017 Apr 7.

Division of Hematology/Oncology, Children's Mercy Hospitals and Clinics, Kansas City, MO.

Patients with myeloid leukemia of Down syndrome (ML-DS) have favorable event-free survival (EFS), but experience significant treatment-related morbidity and mortality. ML-DS blast cells ex vivo have increased sensitivity to cytarabine (araC) and daunorubicin, suggesting that optimizing drug dosing may improve outcomes while reducing toxicity. The Children's Oncology Group (COG) AAML0431 trial consisted of 4 cycles of induction and 2 cycles of intensification therapy based on the treatment schema of the previous COG A2971 trial with several modifications. High-dose araC (HD-araC) was used in the second induction cycle instead of the intensification cycle, and 1 of 4 daunorubicin-containing induction cycles was eliminated. For 204 eligible patients, 5-year EFS was 89.9% and overall survival (OS) was 93.0%. The 5-year OS for 17 patients with refractory/relapsed leukemia was 34.3%. We determined the clinical significance of minimal residual disease (MRD) levels as measured by flow cytometry on day 28 of induction I. MRD measurements, available for 146 of the 204 patients, were highly predictive of treatment outcome; 5-year disease-free survival for MRD-negative patients (n = 125) was 92.7% vs 76.2% for MRD-positive patients (n = 21) (log-rank = .011). Our results indicated that earlier use of HD-araC led to better EFS and OS in AAML0431 than in past COG studies. A 25% reduction in the cumulative daunorubicin dose did not impact outcome. MRD, identified as a new prognostic factor for ML-DS patients, can be used for risk stratification in future clinical trials. This trial was registered at www.clinicaltrials.gov as #NCT00369317.
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http://dx.doi.org/10.1182/blood-2017-01-764324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5482102PMC
June 2017

Minimal residual disease-guided therapy in childhood acute lymphoblastic leukemia.

Blood 2017 04 6;129(14):1913-1918. Epub 2017 Feb 6.

Departments of Oncology and Pathology, St. Jude Children's Research Hospital, Memphis, TN; and.

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http://dx.doi.org/10.1182/blood-2016-12-725804DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5383866PMC
April 2017

Concordance of two approaches in monitoring of minimal residual disease in B-precursor acute lymphoblastic leukemia: Fusion transcripts and leukemia-associated immunophenotypes.

J Formos Med Assoc 2017 Oct 4;116(10):774-781. Epub 2017 Jan 4.

Division of Hematology-Oncology, Department of Internal Medicine, Chang Gung Memorial Hospital at Linkou, Taoyuan, Taiwan; College of Medicine, Chang Gung University, Taoyuan, Taiwan. Electronic address:

Background/purpose: Real-time quantitative polymerase chain reaction (RQ-PCR) for fusion transcripts and flow cytometry for leukemia-specific markers are widely used for minimal residual disease (MRD) detection in acute lymphoblastic leukemia, but the relation between the results of either method is unclear.

Methods: Mononucleated cells from 108 bone marrow samples collected from 55 B-precursor acute lymphoblastic leukemia patients (30 with t(12;21)/ETV6-RUNX1, 16 with t(9;22)/BCR-ABL1 and nine with t(1;19)/TCF3-PBX1) were examined in tandem by RQ-PCR and six-color flow cytometry.

Results: MRD results were concordant in 91 of the 108 paired samples (84.2%; K=0.690); 49 samples were MRD-negative while 42 were MRD-positive by both methods, with < 1 log difference in positive MRD estimates in 39 samples (92.9%). Of the 17 discordant samples, 16 were MRD-positive by RQ-PCR but MRD-negative by flow cytometry; the opposite was true in one sample. Kappa value/concordance was 0.690/85.0% (n = 60) for ETV6-RUNX1, 0.842/93.3% (n = 15) for TCF3-PBX1, and 0.535/78.8% (n = 33) for BCR-ABL1. Specific immunophenotypic abnormalities were more prevalent in each genetic subgroup, such as CD38 underexpression, CD58 overexpression, and CD34 overexpression in ETV6-RUNX1, TCF3-PBX1, and BCR-ABL1, respectively.

Conclusion: In most follow-up samples, MRD estimates by two methods are in agreement, especially in patients with TCF3-PBX1.
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http://dx.doi.org/10.1016/j.jfma.2016.12.002DOI Listing
October 2017

Expanded and armed natural killer cells for cancer treatment.

Cytotherapy 2016 11 3;18(11):1422-1434. Epub 2016 Aug 3.

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore. Electronic address:

The capacity of natural killer (NK) cells to recognize and kill transformed cells suggests that their infusion could be used to treat cancer. It is difficult to obtain large numbers of NK cells ex vivo by exposure to cytokines alone but the addition of stimulatory cells to the cultures can induce NK cell proliferation and long-term expansion. Some of these methods have been validated for clinical-grade application and support clinical trials testing feasibility and safety of NK cell administration. Early data indicate that ex vivo expansion of NK cells from healthy donors or from patients with cancer is robust, allowing multiple infusions from a single apheresis. NK cells can transiently expand in vivo after infusion. Allogeneic NK cells are not direct effectors of graft-versus-host disease but this may occur if donor NK cells are infused after allogeneic hematopoietic stem cell transplant, which may activate T cell alloreactivity. NK cells can be directed with antibodies, or engineered using either transient modification by electroporation of mRNA or prolonged gene expression by viral transduction. Thus, expanded NK cells can be armed with activating receptors that enhance their natural anti-tumor capacity or with chimeric antigen receptors that can redirect them towards specific tumor targets. They can also be induced to express cytokines that promote their autonomous growth, further supporting their in vivo expansion. With the implementation of these approaches, expanded and armed NK cells should ultimately become a powerful component of immunotherapy of cancer.
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http://dx.doi.org/10.1016/j.jcyt.2016.06.013DOI Listing
November 2016

Human NK cells maintain licensing status and are subject to killer immunoglobulin-like receptor (KIR) and KIR-ligand inhibition following ex vivo expansion.

Cancer Immunol Immunother 2016 09 8;65(9):1047-59. Epub 2016 Jul 8.

Department of Human Oncology, University of Wisconsin, 4159 WIMR Bldg. UW-Madison Campus, 1111 Highland Avenue, Madison, WI, 53705, USA.

Infusion of allogeneic NK cells is a potential immunotherapy for both hematopoietic malignancies and solid tumors. Interactions between killer immunoglobulin-like receptors (KIR) on human NK cells and KIR-ligands on tumor cells influence the magnitude of NK function. To obtain sufficient numbers of activated NK cells for infusion, one potent method uses cells from the K562 human erythroleukemia line that have been transfected to express activating 41BB ligand (41BBL) and membrane-bound interleukin 15 (mbIL15). The functional importance of KIRs on ex vivo expanded NK cells has not been studied in detail. We found that after a 12-day co-culture with K562-mbIL15-41BBL cells, expanded NK cells maintained inhibition specificity and prior in vivo licensing status determined by KIR/KIR-ligand interactions. Addition of an anti-CD20 antibody (rituximab) induced NK-mediated antibody-dependent cellular cytotoxicity and augmented killing of CD20+ target cells. However, partial inhibition induced by KIR/KIR-ligand interactions persisted. Finally, we found that extended co-cultures of NK cells with stimulatory cells transduced to express various KIR-ligands modified both the inhibitory and activating KIR repertoires of the expanded NK cell product. These studies demonstrate that the licensing interactions known to occur during NK ontogeny also influence NK cell function following NK expansion ex vivo with HLA-null stimulatory cells.
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http://dx.doi.org/10.1007/s00262-016-1864-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5477646PMC
September 2016

Expanded and Activated Natural Killer Cells for Immunotherapy of Hepatocellular Carcinoma.

Cancer Immunol Res 2016 07 13;4(7):574-81. Epub 2016 May 13.

Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Viral infection of the liver is a major risk factor for hepatocellular carcinoma (HCC). Natural killer (NK) cells recognize virally infected and oncogenically transformed cells, suggesting a therapeutic role for NK-cell infusions in HCC. Using the K562-mb15-41BBL cell line as a stimulus, we obtained large numbers of activated NK cells from the peripheral blood of healthy donors. Expanded NK cells exerted remarkably high cytotoxicity against HCC cell lines, which was generally much higher than that of unstimulated or IL2-activated NK cells. In immunodeficient NOD/scid IL2RGnull mice engrafted with Hep3B, treatment with expanded NK cells markedly reduced tumor growth and improved overall survival. HCC cells exposed for 48 hours to 5 μmol/L of sorafenib, a kinase inhibitor currently used for HCC treatment, remained highly sensitive to expanded NK cells. HCC cell reductions of 39.2% to 53.8% caused by sorafenib in three cell lines further increased to 80.5% to 87.6% after 4 hours of culture with NK cells at a 1:1 effector-to-target ratio. NK-cell cytotoxicity persisted even in the presence of sorafenib. We found that NKG2D, an NK-cell-activating receptor, was an important mediator of anti-HCC activity. We therefore enhanced its signaling capacity with a chimeric NKG2D-CD3ζ-DAP10 receptor. This considerably increased the anti-HCC cytotoxicity of expanded NK cells in vitro and in immunodeficient mice. The NK expansion and activation method applied in this study has been adapted to clinical-grade conditions. Hence, these results warrant clinical testing of expanded NK-cell infusions in patients with HCC, possibly after genetic modification with NKG2D-CD3ζ-DAP10. Cancer Immunol Res; 4(7); 574-81. ©2016 AACR.
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http://dx.doi.org/10.1158/2326-6066.CIR-15-0229DOI Listing
July 2016

A novel anti-GD2/4-1BB chimeric antigen receptor triggers neuroblastoma cell killing.

Oncotarget 2015 Sep;6(28):24884-94

Department of Medical and Surgical Sciences for Children & Adults, Division of Oncology, University-Hospital of Modena and Reggio Emilia, Modena, Italy.

Chimeric antigen receptor (CAR)-expressing T cells are a promising therapeutic option for patients with cancer. We developed a new CAR directed against the disialoganglioside GD2, a surface molecule expressed in neuroblastoma and in other neuroectoderm-derived neoplasms. The anti-GD2 single-chain variable fragment (scFv) derived from a murine antibody of IgM class was linked, via a human CD8α hinge-transmembrane domain, to the signaling domains of the costimulatory molecules 4-1BB (CD137) and CD3-ζ. The receptor was expressed in T lymphocytes by retroviral transduction and anti-tumor activities were assessed by targeting GD2-positive neuroblastoma cells using in vitro cytotoxicity assays and a xenograft model. Transduced T cells expressed high levels of anti-GD2 CAR and exerted a robust and specific anti-tumor activity in 4- and 48-hour cultures with neuroblastoma cells. Cytotoxicity was associated with the release of pro-apoptotic molecules such as TRAIL and IFN-γ. These results were confirmed in a xenograft model, where anti-GD2 CAR T cells infiltrating tumors and persisting into blood circulation induced massive apoptosis of neuroblastoma cells and completely abrogated tumor growth. This anti-GD2 CAR represents a powerful new tool to redirect T cells against GD2. The preclinical results of this study warrant clinical testing of this approach in neuroblastoma and other GD2-positive malignancies.
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http://dx.doi.org/10.18632/oncotarget.4670DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4694800PMC
September 2015

Clinical utility of sequential minimal residual disease measurements in the context of risk-based therapy in childhood acute lymphoblastic leukaemia: a prospective study.

Lancet Oncol 2015 Apr 20;16(4):465-74. Epub 2015 Mar 20.

Department of Oncology, St Jude Children's Research Hospital, Memphis, TN, USA; Department of Pediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Background: The level of minimal residual disease during remission induction is the most important prognostic indicator in patients with acute lymphoblastic leukaemia (ALL). We aimed to establish the clinical significance of minimal residual disease in a prospective trial that used sequential minimal residual disease measurements to guide treatment decisions.

Methods: Between June 7, 2000, and Oct 24, 2007, 498 assessable patients with newly diagnosed ALL were enrolled in a clinical trial at St Jude Children's Research Hospital. We provisionally classified the risk of relapse as low, standard, or high according to patients' baseline clinical and laboratory features. Final risk assignment to establish treatment intensity was based mainly on minimal residual disease levels measured on days 19 and 46 of remission induction, and on week 7 of maintenance treatment. Additional measurements of minimal residual disease were made on weeks 17, 48, and 120 (end of treatment). The primary aim was to establish the association between event-free survival and patients' minimal residual disease levels during remission induction and sequentially post-remission. This trial was registered at ClinicalTrials.gov, number NCT00137111.

Findings: Irrespective of the provisional risk classification, 10-year event-free survival was significantly worse for patients with 1% or greater minimal residual disease levels on day 19 compared with patients with lower minimal residual disease levels (69·2%, 95% CI 49·6-82·4, n=36 vs 95·5%, 91·7-97·5, n=244; p<0·001 for the provisional low-risk group and 65·1%, 50·7-76·2, n=56 vs 82·9%, 75·6-88·2, n=142; p=0·01 for the provisional standard-risk group). 12 patients with provisional low-risk ALL and 1% or higher minimal residual disease levels on day 19 but negative minimal residual disease (<0·01%) on day 46 were treated for standard-risk ALL and had a 10-year event-free survival of 88·9% (43·3-98·4). For the 280 provisional low-risk patients, a minimal residual disease level of less than 1% on day 19 predicted a better outcome, irrespective of the minimal residual disease level on day 46. Of provisional standard-risk patients with minimal residual disease of less than 1% on day 19, the 15 with persistent minimal residual disease on day 46 seemed to have an inferior 10-year event-free survival compared with the 126 with negative minimal residual disease (72·7%, 42·5-88·8 vs 84·0%, 76·3-89·4; p=0·06) after receiving the same post-remission treatment for standard-risk ALL. Of patients attaining negative minimal residual disease status after remission induction, minimal residual disease re-emerged in four of 382 studied on week 7, one of 448 at week 17, and one of 437 at week 48; all but one of these six patients died despite additional treatment. By contrast, relapse occurred in only two of the 11 patients who had decreasing minimal residual disease levels between the end of induction and week 7 of maintenance therapy and were treated with chemotherapy alone.

Interpretation: Minimal residual disease levels during remission induction treatment have important prognostic and therapeutic implications even in the context of minimal residual disease-guided treatment. Sequential minimal residual disease monitoring after remission induction is warranted for patients with detectable minimal residual disease.

Funding: National Institutes of Health and American Lebanese Syrian Associated Charities.
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http://dx.doi.org/10.1016/S1470-2045(15)70082-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4612585PMC
April 2015

Ex vivo-expanded natural killer cells demonstrate robust proliferation in vivo in high-risk relapsed multiple myeloma patients.

J Immunother 2015 Jan;38(1):24-36

*Myeloma Institute for Research and Therapy §Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR †Center for Cell and Gene Therapy, The Methodist Hospital, Texas Children's Hospital, Houston, TX ‡Department of Paediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore ∥Department of Pediatrics, Baylor College of Medicine, Houston, TX.

Highly activated/expanded natural killer (NK) cells can be generated by stimulation with the human leukocyte antigen-deficient cell line K562, genetically modified to express 41BB-ligand and membrane-bound interleukin (IL)15. We tested the safety, persistence, and activity of expanded NK cells generated from myeloma patients (auto-NK) or haploidentical family donors (allo-NK) in heavily pretreated patients with high-risk relapsing myeloma. The preparative regimen comprised bortezomib only or bortezomib and immunosuppression with cyclophosphamide, dexamethasone, and fludarabine. NK cells were shipped overnight either cryopreserved or fresh. In 8 patients, up to 1×10⁸ NK cells/kg were infused on day 0 and followed by daily administrations of IL2. Significant in vivo expansion was observed only in the 5 patients receiving fresh products, peaking at or near day 7, with the highest NK-cell counts in 2 subjects who received cells produced in a high concentration of IL2 (500 U/mL). Seven days after infusion, donor NK cells comprised >90% of circulating leukocytes in fresh allo-NK cell recipients, and cytolytic activity against allogeneic myeloma targets was retained in vitro. Among the 7 evaluable patients, there were no serious adverse events that could be related to NK-cell infusion. One patient had a partial response and in another the tempo of disease progression decreased; neither patient required further therapy for 6 months. In the 5 remaining patients, disease progression was not affected by NK-cell infusion. In conclusion, infusion of large numbers of expanded NK cells was feasible and safe; infusing fresh cells was critical to their expansion in vivo.
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http://dx.doi.org/10.1097/CJI.0000000000000059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4352951PMC
January 2015

Prognostic factors in children with acute myeloid leukaemia and excellent response to remission induction therapy.

Br J Haematol 2015 Jan 28;168(1):94-101. Epub 2014 Aug 28.

Department of Oncology, St. Jude Children's Research Hospital, Memphis, TN, USA.

Minimal residual disease (MRD) is a strong prognostic factor in children and adolescents with acute myeloid leukaemia (AML) but nearly one-quarter of patients who achieve MRD-negative status still relapse. The adverse prognostic factors among MRD-negative patients remain unknown. We analysed the AML02 study cohort to identify demographic and genetic prognostic factors. Among the presenting features, certain 11q23 abnormalities, such as t(6;11) and t(10;11), acute megakaryoblastic leukaemia without the t(1;22), and age ≥10 years were associated with inferior outcome in patients who had MRD-negative status after either remission induction I or II. By contrast, those with rearrangement of CBF genes had superior outcome. Our study identifies patient populations for whom close post-remission MRD monitoring to detect and treat emerging relapse and adjustment in treatment intensity might be indicated.
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http://dx.doi.org/10.1111/bjh.13107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4262553PMC
January 2015

Outcomes of children with BCR-ABL1–like acute lymphoblastic leukemia treated with risk-directed therapy based on the levels of minimal residual disease.

J Clin Oncol 2014 Sep;32(27):3012-20

Purpose: BCR-ABL1–like acute lymphoblastic leukemia (ALL) is a recently identified B-cell ALL (B-ALL)subtype with poor outcome that exhibits a gene expression profile similar to BCR-ABL1-positive ALL but lacks the BCR-ABL1 fusion protein. We examined the outcome of children with BCR-ABL1–like ALL treated with risk-directed therapy based on minimal residual disease (MRD) levels during remission induction.

Patients And Methods: Among 422 patients with B-ALL enrolled onto the Total Therapy XV study between 2000 and 2007, 344 had adequate samples for gene expression profiling. Next-generation sequencing and/or analysis of genes known to be altered in B-ALL were performed in patients with BCR-ABL1–likeALL who had available material. Outcome was compared between patients with and those without BCR-ABL1–like ALL.

Results: Forty (11.6%) of the 344 patients had BCR-ABL1–like ALL. They were significantly more likely to be male, have Down syndrome, and have higher MRD levels on day 19 and at the end of induction than did other patients with B-ALL. Among 25 patients comprehensively studied for genetic abnormalities, 11 harbored a genomic rearrangement of CRLF2, six had fusion transcripts responsive to ABL tyrosine kinase inhibitors or JAK inhibitors, and seven had mutations involving the Ras signaling pathway. There were no significant differences in event-free survival (90.0% +/- 4.7% [SE] v. 88.4% +/- .9% at 5 years; P = .41or in overall survival (92.5% +/- 4.2% v. 95.1% +/- 1.3% at 5 years; P = .41) between patients with and without BCR-ABL1–like ALL.

Conclusion: Patients who have BCR-ABL1–like ALL with poor initial treatment response can be salvaged with MRD-based risk-directed therapy and may benefit from identification of kinase-activating lesions for targeted therapies.
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http://dx.doi.org/10.1200/JCO.2014.55.4105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4162497PMC
September 2014

Autonomous growth and increased cytotoxicity of natural killer cells expressing membrane-bound interleukin-15.

Blood 2014 Aug 8;124(7):1081-8. Epub 2014 Jul 8.

Department of Pediatrics, National University of Singapore, Singapore;

Natural killer (NK) cell survival and, hence, cytotoxicity requires cytokine support. We determined whether expression of interleukin-15 (IL-15) in a nonsecretory, membrane-bound form could sustain NK cell growth. We linked the human IL15 gene to that encoding CD8α transmembrane domain (mbIL15). After retroviral transduction, human NK cells expressed mbIL15 on the cell surface; IL-15 secretion was negligible. Survival of mbIL15-NK cells without interleukin-2 (IL-2) after 7-day culture was vastly superior to that of mock-transduced NK cells (P < .001, n = 15) and of NK cells expressing nonmembrane-bound IL-15 (P = .025, n = 9); viable mbIL15-NK cells were detectable for up to 2 months. In immunodeficient mice, mbIL15-NK cells expanded without IL-2 and were detectable in all tissues examined (except brain) in much higher numbers than mock-transduced NK cells (P < .001). Expansion further increased with IL-2. The primary mechanism of mbIL15 stimulation was autocrine; it activated IL-15 signaling and antiapoptotic signaling. NK cells expressing mbIL15 had higher cytotoxicity against leukemia, lymphoma, and solid tumor cells in vitro and against leukemia and sarcoma cells in xenograft models. Thus, mbIL15 confers independent growth to NK cells and enhances their antitumor capacity. Infusion of mbIL15-NK cells would allow NK cell therapy without the potential adverse effects of cytokine administration.
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http://dx.doi.org/10.1182/blood-2014-02-556837DOI Listing
August 2014

Definition of cure in childhood acute myeloid leukemia.

Cancer 2014 Aug 2;120(16):2490-6. Epub 2014 May 2.

Department of Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee; Department of Pediatrics, College of Medicine, University of Tennessee Health Science Center, Memphis, Tennessee.

Background: A better understanding of when cure can be declared in childhood acute myeloid leukemia (AML) would reduce anxiety and improve quality of life of AML survivors. The authors determined the likelihood that patients with AML would maintain long-term remission after the completion of therapy.

Methods: The cumulative risk of relapse, the time to relapse, event-free survival, and overall survival were analyzed for 604 patients with AML who were enrolled in 7 successive clinical trials divided into 3 treatment eras (1976-1991, 1991-1997, and 2002-2008).

Results: The median time to relapse did not change over time (0.93 years vs 0.76 years vs 0.8 years, respectively, for each consecutive era; P = .22), but the risk of relapse decreased significantly (5-year cumulative incidence of relapse: 52.6% ± 3.1% vs 31.5% ± 3.9% vs 22% ± 3%, respectively, for each consecutive era; P < .001). Among patients who were in remission 4 years from diagnosis, the probabilities of relapse were 1.7%, 2.9%, and 0.9%, respectively, for each consecutive era. In the most recent era, all but 1 of 44 relapses occurred within 4 years of diagnosis.

Conclusions: Children with AML who receive treatment with contemporary therapy and remain in remission 4 years from diagnosis probably are cured. Although late relapses and late deaths from other causes are rare, long-term follow-up of survivors is necessary for the timely management of late adverse effects.
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http://dx.doi.org/10.1002/cncr.28742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4282842PMC
August 2014