Programme for Harmonization to the International Scale in Latin America for BCR-ABL1 quantification in CML patients: Findings and recommendations

The quantitation of BCR-ABL1 mRNA is mandatory for chronic myeloid leukemia (CML) patients, and RT-qPCR is the most extensively used method in testing laboratories worldwide. Nevertheless, substantial variation in RT-qPCR results makes inter-laboratory comparability hard. To facilitate inter-laboratory comparative assessment, an international scale (IS) for BCR-ABL1 was proposed.The laboratory-specific conversion factor (CF) to the IS can be derived from the World Health Organization (WHO) genetic reference panel; however, this material is limited to the manufacturers to produce and calibrate secondary reference reagents. Therefore, we developed secondary reference calibrators, as lyophilized cellular material, aligned to the IS. Our purpose was both to re-evaluate the CF in 18 previously harmonized laboratories and to propagate the IS to new laboratories.Results Our field trial including 30 laboratories across Latin America showed that, after correction of raw BCR-ABL1/ABL1 ratios using CF, the relative mean bias was significantly reduced. We also performed a follow-up of participating laboratories by annually revalidating the process; our results support the need for continuous revalidation of CFs. All participating laboratories also received a calibrator to determine the limit of quantification (LOQ); 90% of them could reproducibly detect BCR-ABL1, indicating that these laboratories can report a consistent deep molecular response. In addition, aiming to investigate the variability of BCR-ABL1 measurements across different RNA inputs, we calculated PCR efficiency for each individual assay by using different amounts of RNA.Conclusions In conclusion, for the first time in Latin America, we have successfully organized a harmonization platform for BCR-ABL1 measurement that could be of immediate clinical benefit for monitoring the molecular response of patients in low-resource regions.Fil: Ruiz, María Sol. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Sánchez, María Belén. Fundación Cáncer. Centro de Investigaciones Oncológicas; ArgentinaFil: Vera Contreras, Yuly Masiel. Fundación Cáncer. Centro de Investigaciones Oncológicas; ArgentinaFil: Agrielo, Evangelina. Laboratorio Especialidades Bioquímicas; ArgentinaFil: Alonso, Marta. Hospital Nacional Profesor Alejandro Posadas; ArgentinaFil: Altuna, María Eugenia. Clinica Roberto Raña; ArgentinaFil: Anchordoqui, María Sol. Argenomics; ArgentinaFil: Asinari, Mariana. Hospital Privado Centro Médico de Córdoba; ArgentinaFil: Bonetto, María Elisa. Hospital Rawson San Juan; ArgentinaFil: Camargo, Mauricio. Laboratorio Medellin; ColombiaFil: Giere, Isabel. Fundación Para Combatir la Leucemia; ArgentinaFil: González, Javier. Meyer Lab; ParaguayFil: Granda Alacote, Ana Cecilia. Laboratorios Medicos Lima; PerúFil: Guerra, Javier. Nanopharmacia Diagnóstica Mexico; MéxicoFil: Gutiérrez, Marina. Stamboulian; ArgentinaFil: Maldonado, Cecilia. Manlab; ArgentinaFil: Makiya, Ricard. No especifíca;Fil: Manrique, Gonzalo. Asociacion Española de Montevideo; UruguayFil: Monaco, María Eugenia. Laboratorio Tucuman; ArgentinaFil: Rozo, Juan Carlos. Unidad de Diagnóstico Hemato Oncológico; ColombiaFil: Santamaría Martín, Carlos Jose. Hospital de Niños San Jose de Costa Rica; Costa Rica. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Seravalle, Analía. Civic Rosario; ArgentinaFil: Zea, Olga. Laboratorio Medellin Colombia; ColombiaFil: Zubillaga, María Noel. Asociación Española de Socorros Mutuos; UruguayFil: Mordoh, José. Fundación Cáncer. Centro de Investigaciones Oncológicas; ArgentinaFil: Larripa, Irene Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Bianchini, Michele. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Technology as 'Applied Science': a Serious Misconception that Reinforces Distorted and Impoverished Views of Science

The current consideration of technology as 'applied science', this is to say, as something that comes 'after' science, justifies the lack of attention paid to technology in science education. In our paper we question this simplistic view of the science-technology relationship, historically rooted in the unequal appreciation of intellectual and manual work, and we try to show how the absence of the technological dimension in science education contributes to a na¿ ve and distorted view of science which deeply affects the necessary scientific and technological literacy of all citizens

In-depth characterization of NK cell markers from CML patients who discontinued tyrosine kinase inhibitor therapy

IntroductionTreatment-free remission (TFR) in patients with chronic myeloid leukemia in chronic phase is considered a safe option if suitable molecular monitoring is available. However, the question arises as to which factors can contribute to the maintenance of TFR, and immunologic surveillance of the remaining leukemic cells is believed to be one of them. Argentina Stop Trial is an open-label, single-arm, multicenter trial assessing TFR after tyrosine kinase inhibitors interruption, that after more than 4 years showed a successful TFR rate of 63%.MethodsIn this context, we set up an immunological study by flow cytometry in order to analyze specific NK cell subsets from peripheral blood patient samples both at the time of discontinuation as well as during the subsequent months.ResultsAt the time of discontinuation, patients show a mature NK cell phenotype, probably associated to TKI treatment. However, 3 months after discontinuation, significant changes in several NK cell receptors occurred. Patients with a higher proportion of CD56dim NK and PD-1+ NK cells showed better chances of survival. More interestingly, non-relapsing patients also presented a subpopulation of NK cells with features associated with the expansion after cytomegalovirus infection (expression of CD57+NKG2C+), and higher proportion of NKp30 and NKp46 natural cytotoxicity receptors, which resulted in greater degranulation and associated with better survival (p<0.0001).DiscussionThis NK cell subset could have a protective role in patients who do not relapse, thus further characterization could be useful for patients in sustained deep molecular response

Planned Pregnancy in a Chronic Myeloid Leukemia Patient in Molecular Remission

Excellent response rates and a good quality of life have been observed since the introduction of tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML) treatment. Consequently, some challenges began to appear in CML women in child-bearing age wishing to become pregnant. Currently, many women around the world are in stable major/complete molecular response MMR/CMR (MMR: <0.1% BCR-ABL/ABL and CMR: undetectable BCR-ABL mRNA by RQ-PCR transcript levels on the international scale). The condition of stable MMR/CMR is linked to a long-term virtual absence of progression to the accelerated and blastic phase and to the possibility of stopping the TKI treatment with the maintenance of a condition of CMR in a proportion of cases. Imatinib teratogenic and prescribing information prohibits the use of it during pregnancy. We describe the case of a 36-year-old female patient with CML in chronic phase who stopped imatinib after 2 years in major molecular response (MMR) to plan a pregnancy. Molecular monitoring by RQ-PCR was performed quarterly. She achieved a safe pregnancy and delivery maintaining an optimal molecular response throughout the pregnancy. Isolated literature reports have been described, but no formal advice has been described at present time

GSTM1 and GSTP1, but not GSTT1 genetic polymorphisms are associated with chronic myeloid leukemia risk and treatment response

Background: Chronic myeloid leukemia (CML) is associated to the BCR-ABL1 oncogene and can successfully be treated with tyrosine kinase inhibitors (TKIs). However, it remains still under investigation which molecular factors may influence CML risk or varying responses to TKIs. The aim of this study was to assess the role of Glutathione-S- transferases (GSTs) genetic polymorphisms in CML susceptibility and TKI clinical outcome.Materials: Deletion polymorphisms in GSTM1 and GSTT1 genes and the single nucleotide polymorphism in GSTP1 c.319A>G (rs1695; p.105Ile>Val) were genotyped by PCR methods in 141 CML treated patients and 141 sex- and age-matched healthy individualsResults: Individual analysis of each GST gene showed no association with CML risk. A trend toward significance (p=0.07) for a recessive model was found for GSTP1 (OR: 2.04; IC: 0.94-4.4). However, the combined analysis showed that GSTM1-null/GSTP1- GG as well as GSTT1-null/GSTP1- GG were associated with CML development (p= 0.03; OR: 3.54 IC: 1.2-14.57; p= 0.05; OR: 12.65; IC: 1.17-21.5). The relationship with treatment outcome showed that the presence of GSTM1 gene was significantly linked with an inferior rate of major molecular response (p=0.048) and poor event free-survival (EFS)(p=0.02).Furthermore, a group of patients with GSTP1-GG genotype were significantly associated with reduced EFS comparing to those carrying other GSTP1 genotypes (p=0.049). GSTP1-GG genotypes had short time to treatment failure in a group of patients unresponsive to TKIs comparing to other GSTP1 genotypes (p=0.03).Conclusions: This study highlights the significance of GSTs polymorphisms on CML susceptibility and response to TKIs in the Argentinean population.Fil: Weich, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Ferri, Cristian Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Moiraghi, Beatriz. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos ; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; ArgentinaFil: Bengio, Raquel. Academia Nacional de Medicina de Buenos Aires. Instituto de Investigaciones Hematológicas ; ArgentinaFil: Giere, Isabel. Fundaleu; ArgentinaFil: Pavlovsky, Carolina. Fundaleu; ArgentinaFil: Larripa, Irene Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Fundia, Ariela Freya. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentin

TP53 codon 72 polymorphism predicts chronic myeloid leukemia susceptibility and treatment outcome

BCR-ABL1 gene is a key molecular marker of chronic myeloid leukemia (CML), but it is still unclear which molecular factors may influence CML risk or lead to variable responses to tyrosine kinase inhibitors (TKIs). The aim of this study was to investigate the impact of TP53 c.213 G > C(Arg72Pro; rs1042522) polymorphism on CML risk and its correlation with clinical outcome. Peripheral blood samples from 141 treated CML patients and 141 sex- and age-matched healthy individuals were genotyped by PCR-RFLP. Standard genetic models for disease penetrance were evaluated by logistic regression analysis and Kaplan-Meier method was performed to estimate survival curves. Our study suggests that TP53 c.213 G > C polymorphism may be involved in CML development considering a recessive model (p = 0.01; OR: 0.19; CI: 0.06–0.68). In addition, a non-homogenous distribution was found for this polymorphism in males and patients youngers than 50 years (p = 0.02). According to clinical response, TP53-GG genotype was associated with higher levels of BCR-ABL1 transcripts (p = 0.04) and shorter event free survival (p = 0.04). Moreover, a trend toward significance was found for failure free survival (p = 0.06) and time to imatinib failure (p = 0.08). In conclusion, our data suggest that a; TP53 c.213 G > C may be a potential biomarker of CML susceptibility and clinical outcome.Fil: Weich, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Ferri, Cristian Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Moiraghi, Elena Beatriz. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos "Ramos Mejía"; ArgentinaFil: Bengió, Raquel. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Giere, Isabel. Fundaleu; ArgentinaFil: Pavlovsky, Carolina. Fundaleu; ArgentinaFil: Larripa, Irene Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Fundia, Ariela Freya. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentin

Elevated plasma levels of IL-6 and MCP-1 selectively identify CML patients who better sustain molecular remission after TKI withdrawal

Abstract Treatment-free remission (TFR) in chronic myeloid leukemia (CML) is safe under adequate molecular monitoring, but questions remain regarding which factors may be considered predictive for TFR. Argentina Stop Trial (AST) is a multicenter TFR trial showing that 65% of patients sustain molecular remission, and the prior time in deep molecular response (DMR) was associated with successful TFR. Luminex technology was used to characterize cytokines in plasma samples. Using machine learning algorithms, MCP-1 and IL-6 were identified as novel biomarkers and MCP-1low/IL-6low patients showed eightfold higher risk of relapse. These findings support the feasibility of TFR for patients in DMR and MCP-1/IL-6 plasma levels are strong predictive biomarkers

DataSheet_1_In-depth characterization of NK cell markers from CML patients who discontinued tyrosine kinase inhibitor therapy.docx

IntroductionTreatment-free remission (TFR) in patients with chronic myeloid leukemia in chronic phase is considered a safe option if suitable molecular monitoring is available. However, the question arises as to which factors can contribute to the maintenance of TFR, and immunologic surveillance of the remaining leukemic cells is believed to be one of them. Argentina Stop Trial is an open-label, single-arm, multicenter trial assessing TFR after tyrosine kinase inhibitors interruption, that after more than 4 years showed a successful TFR rate of 63%.MethodsIn this context, we set up an immunological study by flow cytometry in order to analyze specific NK cell subsets from peripheral blood patient samples both at the time of discontinuation as well as during the subsequent months.ResultsAt the time of discontinuation, patients show a mature NK cell phenotype, probably associated to TKI treatment. However, 3 months after discontinuation, significant changes in several NK cell receptors occurred. Patients with a higher proportion of CD56dim NK and PD-1+ NK cells showed better chances of survival. More interestingly, non-relapsing patients also presented a subpopulation of NK cells with features associated with the expansion after cytomegalovirus infection (expression of CD57+NKG2C+), and higher proportion of NKp30 and NKp46 natural cytotoxicity receptors, which resulted in greater degranulation and associated with better survival (pDiscussionThis NK cell subset could have a protective role in patients who do not relapse, thus further characterization could be useful for patients in sustained deep molecular response.</p