Clinical networking results in continuous improvement of the outcome of patients with acute promyelocytic leukemia.

ABSTRACT: The introduction of all-trans retinoic acid combined with anthracyclines has significantly improved the outcomes for patients diagnosed with acute promyelocytic leukemia (APL), and this strategy remains the standard of care in countries in which arsenic trioxide is not affordable. However, data from national registries and real-world databases indicate that low- and middle-income countries (LMIC) still face disappointing results, mainly because of high induction mortality and suboptimal management of complications. The American Society of Hematology established the International Consortium on Acute Leukemias (ICAL) to address this challenge through international clinical networking. Here, we present the findings from the International Consortium on Acute Promyelocytic Leukemia study involving 806 patients with APL recruited from 2005 to 2020 in Brazil, Chile, Paraguay, Peru, and Uruguay. The induction mortality rate has notably decreased to 14.6% compared with the pre-ICAL rate of 32%. Multivariable logistic regression analysis revealed as factors associated with induction death: age of ≥40 years, Eastern Cooperative Oncology Group performance status score of 3, high-risk status based on the Programa Español de Tratamiento en Hematologia/Gruppo Italiano Malattie EMatologiche dell'Adulto classification, albumin level of ≤3.5 g/dL, bcr3 PML/RARA isoform, the interval between presenting symptoms to diagnosis exceeding 48 hours, and the occurrence of central nervous system and pulmonary bleeding. With a median follow-up of 53 months, the estimated 4-year overall survival rate is 81%, the 4-year disease-free survival rate is 80%, and the 4-year cumulative incidence of relapse rate is 15%. These results parallel those observed in studies conducted in high-income countries, highlighting the long-term effectiveness of developing clinical networks to improve clinical care and infrastructure in LMIC.

Reduced SLIT2 is Associated with Increased Cell Proliferation and Arsenic Trioxide Resistance in Acute Promyelocytic Leukemia.

The SLIT-ROBO axis plays an important role in normal stem-cell biology, with possible repercussions on cancer stem cell emergence. Although the Promyelocytic Leukemia (PML) protein can regulate SLIT2 expression in the central nervous system, little is known about SLIT2 in acute promyelocytic leukemia. Hence, we aimed to investigate the levels of SLIT2 in acute promyelocytic leukemia (APL) and assess its biological activity in vitro and in vivo. Our analysis indicated that blasts with SLIT2high transcript levels were associated with cell cycle arrest, while SLIT2low APL blasts displayed a more stem-cell like phenotype. In a retrospective analysis using a cohort of patients treated with all-trans retinoic acid (ATRA) and anthracyclines, high SLIT2 expression was correlated with reduced leukocyte count (p = 0.024), and independently associated with improved overall survival (hazard ratio: 0.94; 95% confidence interval: 0.92-0.97; p < 0.001). Functionally, SLIT2-knockdown in primary APL blasts and cell lines led to increased cell proliferation and resistance to arsenic trioxide induced apoptosis. Finally, in vivo transplant of Slit2-silenced primary APL blasts promoted increased leukocyte count (p = 0.001) and decreased overall survival (p = 0.002) compared with the control. In summary, our data highlight the tumor suppressive function of SLIT2 in APL and its deteriorating effects on disease progression when downregulated.

NTAL is associated with treatment outcome, cell proliferation and differentiation in acute promyelocytic leukemia.

Non-T cell activation linker (NTAL) is a lipid raft-membrane protein expressed by normal and leukemic cells and involved in cell signaling. In acute promyelocytic leukemia (APL), NTAL depletion from lipid rafts decreases cell viability through regulation of the Akt/PI3K pathway. The role of NTAL in APL cell processes, and its association with clinical outcome, has not, however, been established. Here, we show that reduced levels of NTAL were associated with increased all-trans retinoic acid (ATRA)-induced differentiation, generation of reactive oxygen species, and mitochondrial dysfunction. Additionally, NTAL-knockdown (NTAL-KD) in APL cell lines led to activation of Ras, inhibition of Akt/mTOR pathways, and increased expression of autophagy markers, leading to an increased apoptosis rate following arsenic trioxide treatment. Furthermore, NTAL-KD in NB4 cells decreased the tumor burden in (NOD scid gamma) NSG mice, suggesting its implication in tumor growth. A retrospective analysis of NTAL expression in a cohort of patients treated with ATRA and anthracyclines, revealed that NTAL overexpression was associated with a high leukocyte count (P = 0.007) and was independently associated with shorter overall survival (Hazard Ratio: 3.6; 95% Confidence Interval: 1.17-11.28; P = 0.026). Taken together, our data highlights the importance of NTAL in APL cell survival and response to treatment.

Acute promyelocytic leukemia: evaluation of diagnostic tests from 2000 to 2018 in a public hospital / Leucemia promielocítica aguda: avaliação dos testes diagnósticos entre o período de 2000 e 2018 em um hospital público

ABSTRACT Introduction: Acute promyelocytic leukemia (APL) is caused by t(15;17)(q24;q21) translocation, which product is the fusion oncoprotein PML-RARa (promyelocytic leukemia-retinoic acid receptor alpha). The morphology of leukemic promyelocytes is usually characteristic, with the presence of faggot cells and coarse cytoplasmic granulations; immunophenotype is characteristic in most cases. However, definitive laboratory diagnosis should be performed by detecting t(15;17) or by PML-RARa fusion protein. Objectives: To compare cytomorphology, flow cytometry, and classical cytogenetic of bone marrow samples from patients with APL, treated at the Complexo Hospital de Clínicas da Universidade Federal do Paraná (CHC-UFPR), as well as describe the possible discrepancies between the methodologies. Method: Retrospective analysis of APL cases treated at the CHC-UFPR from January 2000 to July 2018. Results: Eighty-eight patients (42 man/ 46 woman; mean age: 34 years), 42.1% of them presented a high-risk prognosis. Flow cytometry was performed in 83 cases (94.3%); karyotype was performed in 79 cases (89.7%), but translocation t(15;17) was confirmed in only 53 cases (60.2%). From the 28 patients with a non-conclusive karyotype; fourteen (15.9%) of them presented the PML-RARa transcript in the molecular analysis. In total, 35 patients (39 8%) performed research of the PML-RARa gene by molecular biology. Only 45 patients (51.1%) presented concordant diagnosis among the three technical exams (morphology, flow cytometry and cytogenetics). Overall survival was 67% at 4.8 years, with 29 deaths. Conclusion: Genetic confirmation was observed in 76.1% of samples, 60.2% by conventional cytogenetics and 15.9% by molecular biology. There was a disagreement between the methodologies, and a low sensibility of the conventional cytogenetics, demonstrating the importance of performing molecular techniques for diagnostic confirmation.

RESUMEN Introducción: La leucemia promielocítica aguda (LPA) es causada por la translocación t(15;17)(q24;q21), cuyo producto es la oncoproteína de fusión PML-RARa (proteína de la leucemia promielocítica-receptor alfa de ácido retinoico). La morfología de los promielocitos leucémicos suele ser típica, con presencia de células faggot (células en haces) y gruesasgranulaciones citoplásmicas; el inmunofenotipo es característico en la mayor parte de los casos. No obstante, el diagnóstico final de laboratorio debe ser hecho por la detección de la t(15;17) o por la oncoproteína PML-RARa. Objetivos: Comparar la citomorfología, la citometría de flujo y la citogenética clásica de muestras de médula ósea de pacientes con LPA asistidos en el Complexo Hospital de Clínicas da Universidade Federal do Paraná (CHC-UFPR), así como describir lasposibles discrepancias entre los métodos. Método: Análisis retrospectivo de los casos de LPA asistidos en el CHC-UFPR entre enero de 2000 y julio de 2018. Resultados: De los 88 pacientes (42 hombres y 46 mujeres; edad promedio: 34 anos), 42,1% presentaron pronóstico de alto riesgo. Citometría de flujo se realizó en 83 casos (94,3%); cariotipo en 79 casos (89,7%),pero la translocación se confirmó en sólo53 (60,2%) casos. Entre los28 pacientes con cariotipo no concluyente, 14 (15,9%) presentaron el transcripto PML-RARa. En total, 35 pacientes (39,8%) realizaron la pesquisa del gen PML-RARa por biología molecular. Cuarenta y cinco pacientes (51,1%) tuvieron diagnóstico acorde entre los métodos (morfología, citometría de flujo y citogenética). La supervivencia global fue de 67% en 4,8 anos, con 29 muertes. Conclusión: Hubo confirmación genética en 76,1% de las muestras, siendo 60,2% por citogenética y 15,9%por biología celular. Hubo desacuerdo entre los métodos y baja sensibilidad de la citogenética convencional, lo que demuestra la importancia de la realización de técnicas moleculares para confirmación diagnóstica.

RESUMO Introdução: A leucemia promielocítica aguda (LPA) écausada pela translocação t(15;17)(q24;q21), cujo produto éa oncoproteína de fusão PML-RARa (leucemia promielocítica-receptor alfa do ácido retinoico). A morfologia dos promielócitos leucêmicos é habitualmente característica, com presença de faggot cells (células em maços ou feixes) e granulações citoplasmáticas grosseiras; o imunofenótipo é característico na maioria dos casos. Porém, o diagnóstico laboratorial definitivo deve ser feito pela detecção da t(15;17) ou pela oncoproteína PML-RARa. Objetivos: Comparar a citomorfologia, a citometria de fluxo e a citogenética clássica de amostras de medula óssea de pacientes com LPA atendidos no Complexo Hospital de Clínicas da Universidade Federal do Paraná (CHC-UFPR), bem como descrever as possíveis discrepâncias entre as metodologias. Método: Análise retrospectiva dos casos de LPA atendidos no CHC-UFPR entre janeiro de 2000 e julho de 2018. Resultados: Dos 88 pacientes (42 homens e 46 mulheres; média de idade: 34 anos), 42,1% apresentaram prognóstico de alto risco. A citometria de fluxo foi realizada em 83 casos (94,3%); o cariótipo, em 79 casos (89,7%), mas a translocação foi confirmada em apenas 53 (60,2%) casos. Dos 28 pacientes com cariótipo não conclusivo, 14 (15,9%) tinham a presença do transcrito PML-RARa. No total, 35 pacientes (39,8%) realizaram a pesquisa do gene PML-RARa por biologia molecular. Quarenta e cinco pacientes (51,1%) obtiveram diagnóstico concordante entre as metodologias (morfologia, citometria de fluxo e citogenética). A sobrevida global foi de 67% em 4,8 anos;com 29 óbitos. Conclusão: A confirmação genética foi realizada em 76,1% das amostras, sendo 60,2%por citogenética e 15,9% por biologia molecular. Houve discordância entre as metodologias e baixa sensibilidade da citogenética convencional, o que demonstra a importância da realização de técnicas moleculares para confirmação diagnóstica.

Combining gene mutation with gene expression analysis improves outcome prediction in acute promyelocytic leukemia.

By combining the analysis of mutations with aberrant expression of genes previously related to poorer prognosis in both acute promyelocytic leukemia (APL) and acute myeloid leukemia, we arrived at an integrative score in APL (ISAPL) and demonstrated its relationship with clinical outcomes of patients treated with all-trans retinoic acid (ATRA) in combination with anthracycline-based chemotherapy. Based on fms-like tyrosine kinase-3-internal tandem duplication mutational status; the ΔNp73/TAp73 expression ratio; and ID1, BAALC, ERG, and KMT2E gene expression levels, we modeled ISAPL in 159 patients (median ISAPL score, 3; range, 0-10). ISAPL modeling identified 2 distinct groups of patients, with significant differences in early mortality (P < .001), remission (P = .004), overall survival (P < .001), cumulative incidence of relapse (P = .028), disease-free survival (P = .03), and event-free survival (P < .001). These data were internally validated by using a bootstrap resampling procedure. At least for patients treated with ATRA and anthracycline-based chemotherapy, ISAPL modeling may identify those who need to be treated differently to maximize their chances for a cure.