Effect of BCR::ABL1 transcript type and droplet digital polymerase chain reaction on successful treatment-free remission in chronic myeloid leukemia patients who discontinued tyrosine kinase inhibitor.

Background: Droplet digital polymerase chain reaction (ddPCR) is an exact method of measurement. Objectives: We conducted this study to identify the prognostic factors for successful treatment-free remission in patients with chronic-phase chronic myeloid leukemia who discontinued tyrosine kinase inhibitors (TKIs). We also aimed to validate ddPCR for predicting molecular relapse. Design: This is a prospective, multicenter study. Methods: We enrolled patients treated with TKIs for at least 3 years with a confirmed sustained deep molecular response (DMR) for at least 1 year. TKI was re-administered in patients who experienced the loss of major molecular response (MMR). Results: A total of 66 patients from five institutions in South Korea were enrolled. During a median follow-up period of 16.5 months, 29/66 (43.9%) patients experienced molecular relapse; the probability of molecular relapse-free survival (RFS) at 6 or 12 months after TKI discontinuation was 65.6% or 57.8%, respectively, with most molecular relapses occurring within the first 7 months. All patients who lost MMR were re-treated with TKI, and all re-achieved MMR at a median of 2.8 months. E14a2 transcript type (p = 0.005) and longer DMR duration (⩾48 months) prior to TKI discontinuation (p = 0.002) were associated with prolonged molecular RFS and with sustained DMR. Patients with both e13a2 transcript type and detectable BCR::ABL1 (⩾MR5.0) by ddPCR at the time of TKI discontinuation showed shorter duration of molecular RFS (p = 0.015). Conclusion: Our data suggest that transcript type and BCR::ABL1 transcript levels on ddPCR should be taken into consideration when deciding whether to discontinue TKI therapy.

The association of genetic alterations with response rate in newly diagnosed chronic myeloid leukemia patients.

Genetic differences may be associated with the response to tyrosine kinase inhibitor (TKI) in patients with chronic myeloid leukemia (CML). In this study, we identified genetic alterations between rapid and slow responders (BCR/ABL1 International Scale at 6 months: ≤0.1 % vs. > 0.1 %) of TKI treatment in chronic phase CML patients. Our analyses involved single nucleotide polymorphism (SNP), a Genome Wide Association Study and a Network-wide Association Study (NetWAS). Seventy-two patients from 16 institutions were enrolled and treated with a TKI, nilotinib. Gene Set Analysis identified genetic alterations in pathways related to the differentiation, proliferation, and activity of various innate immune cells. The NetWAS analysis found that genes associated with natural killer (NK) cells (PTPRCAP, BLNK, HCK, ARHGEF11, GPR183, TRPV2, SHKBP1, CD2) showed significant differences between rapid and slow responders of nilotinib. However, we found no significantly different genetic alterations according to the response in the SNP analysis. In conclusion, we found that rapidity of response to TKI was associated with pathway-associated genetic alterations in immune cells, particularly with respect to NK cell activity. These results suggested that the innate immune system at initial diagnosis had an important role in treatment response in patients with CML.

IDH1/2 mutations in acute myeloid leukemia.

The mutational and epigenetic landscape of acute myeloid leukemia (AML) has become increasingly well understood in recent years, informing on biological targets for precision medicine. Among the most notable findings was the recognition of mutational hot-spots in the isocitrate dehydrogenase (IDH) genes. In this review, we provide an overview on the IDH1/2 mutation landscape in Korean AML patients, and compare it with available public data. We also discuss the role of IDH1/2 mutations as biomarkers and drug targets. Taken together, occurrence of IDH1/2 mutations is becoming increasingly important in AML treatment, thus requiring thorough examination and follow-up throughout the clinical course of the disease.

Ultra-deep sequencing mutation analysis of the BCR/ABL1 kinase domain in newly diagnosed chronic myeloid leukemia patients.

Ultra-deep sequencing detects low-frequency genetic mutations with high sensitivity. We used this approach to prospectively examine mutations in the BCR/ABL1 tyrosine kinase from patients with newly diagnosed, chronic-phase chronic myeloid leukemia (CML) treated with the tyrosine kinase inhibitor nilotinib. Between May 2013 and November 2014, 50 patients from 18 institutions were enrolled in the study. We screened 103 somatic mutations and found that mutations in the P-loop domain were the most frequent (173/454 mutations in the P-loop) and noted the presence of the V299 L mutation (dasatinib-resistant/nilotinib-sensitive) in 98 % of patients (49/50). No patients had Y253H, E255 V, or F359 V/C/I mutations, which would recommend dasatinib rather than nilotinib treatment. The S417Y mutation was associated with lower achievement of a major molecular response (MMR) at 6 months, and the V371A mutation was associated with reduced MMR and MR4.5 durations (MMR for 2 years: 100 % for no mutation vs. 75 % for mutation, P=0.039; MR4.5 for 15 months: 94.1 % vs. 25 %, P=0.002). Patients with known nilotinib-resistant mutations had lower rates of MR4.5 achievement. In conclusion, ultra-deep sequencing is a sensitive method for genetic-based treatment decisions. Based on the results of these mutational analyses, nilotinib treatment is a promising option for Korean patients with CML.

Use of Droplet Digital Polymerase Chain Reaction for Detecting Minimal Residual Disease: A Prospective Multi-Institutional Study.

BACKGROUND/AIM: Droplet digital polymerase chain reaction (ddPCR) is an exact method of measuring nucleic acids. The aim of this prospective study was to evaluate minimal residual disease (MRD) using ddPCR in chronic myeloid leukemia (CML) patients. PATIENTS AND METHODS: Between May 2013 and November 2014, CML patients treated with nilotinib were enrolled in our study. BCR/ABL1 transcripts levels were evaluated using ddPCR at the first time of complete molecular response (CMR). We enrolled 15 patients from 7 Institutions. The treatment period and median follow-up period were 45 months and 47 months, respectively. RESULTS: Patients with a high level of BCR/ABL1 transcript had a greater tendency to lose the CMR during the follow-up period (p=0.095). In addition, patients with a low level of BCR/ABL1 transcript showed a longer duration of CMR compared to those with a high level (p=0.032). CONCLUSION: We found that ddPCR is a sensitive method for detecting MRD and that MRD could affect the duration of the treatment response.

A phase 4 study of nilotinib in Korean patients with Philadelphia chromosome-positive chronic myeloid leukemia in chronic phase: ENESTKorea.

Although nilotinib has improved efficacy compared to imatinib, suboptimal response and intolerable adverse events (AEs) limit its effectiveness in many patients with chronic myeloid leukemia in chronic phase (CML-CP). We investigated the 2-year efficacy and safety of nilotinib and their relationships with plasma nilotinib concentrations (PNCs). In this open-label, multi-institutional phase 4 study, 110 Philadelphia chromosome-positive CML-CP patients were treated with nilotinib at a starting dose of 300 mg twice daily. Molecular responses (MRs) and AEs were monitored for up to 24 months. The 24-month cumulative MR4.5 rate was evaluated as the primary endpoint. Plasma samples were collected from 94 patients to determine PNCs, and the per-patient mean was used to categorize them into four mean PNC (MPNC) groups. Cumulative MR rates and safety were compared between groups. With a median follow-up of 22.2 months, the 24-month cumulative MR4.5 rate was 56.2% (95% confidence interval, 44.0%-8.3%), and the median time to MR4.5 was 23.3 months. There were no significant differences in the cumulative rates of major molecular response, MR4 , and MR4.5 between MPNC groups. One patient died due to acute viral hepatitis, and two developed hematological or cytogenetic relapse, while no progression to accelerated or blast phase was observed. Safety results were consistent with previous studies with no new safety signal identified. Across the MPNC groups, there was no significant linear trend in the frequency of AEs. Nilotinib is highly effective for the treatment of CML-CP with manageable AEs. The measurement of PNC has no predictive value for patient outcomes and is thus not found to be clinically useful. This study is registered with clinicaltrials.gov, Number NCT03332511.

Role of induction and consolidation chemotherapy in elderly acute myeloid leukemia patients.

The present study sought to elucidate the role of induction and consolidation therapy in elderly patients. We retrospectively collected data of 477 patients who were aged over 60 years at the time of acute myeloid leukemia (AML) diagnosis. The median overall survival (OS) was 339 days in the induction group (n = 266) and 86 days in the best supportive care group (n = 211) (P < 0.001). In the induction group, the complete remission (CR) rate was 58.3 %, and treatment-related death was 15.4 %. Successful induction was related to good performance [Eastern Cooperative Oncology Group (ECOG <2)] [hazard ratio (HR) 3.215; P = 0.002]. Mortality correlated with failure to achieve CR (HR 4.059; P < 0.001) and poor performance status (ECOG >2) (HR 2.731; P = 0.035). In CR patients, poor karyotype and absence of consolidation (HR 2.313; P = 0.003) correlated with mortality. More than one cycle of consolidation was associated with better OS (P < 0.001). Lack of salvage therapy was associated with mortality in patients who did not achieve CR (HR 3.223; P = 0.005). Intensive induction in patients with good performance and >1 cycle of consolidation after CR may be the best strategy for improving OS in elderly AML patients.

Multicenter study evaluating the impact of hypomethylating agents as bridging therapy to hematopoietic stem cell transplantation in myelodysplastic syndromes.

Allogeneic hematopoietic stem cell transplantation (alloSCT) is currently the only curative treatment modality for myelodysplastic syndromes (MDS). The treatment paradigm for MDS has changed in recent years with the introduction of hypomethylating agents (HMAs). The present retrospective multicenter study was designed to assess the effects of pre-transplant HMA on transplant outcome and determine which patients would benefit most from this therapy. A total of 109 patients who received alloSCT at one of five institutions between 2007 and 2010 were enrolled in this study regardless of pre-transplant HMA therapy. 81 of the 109 patients enrolled were treated with HMA prior to alloSCT. 28 patients received alloSCT without HMA bridging. The distributions of WHO classification groups and IPSS scores were similar between the two groups (P = 0.752 and P = 0.265, respectively). Pre-transplant HMA did not affect OS (P = 0.244), and there were no differences in response to HMA therapy within the HMA-treated group. The cumulative incidence of NRM was not significantly different between the two groups (P = 0.500). However, for patients with a high blast count (>5 % of bone marrow at the time of diagnosis), pre-transplant HMA therapy had a NRM benefit (83.3 vs. 48.6 %, P = 0.014).

Analysis of cariogenic bacteria in saliva of cancer patients.

This study examined salivary flow and salivary pH and the prevalence and levels of cariogenic bacteria in the saliva of oncological patients and healthy controls. Quantitative real-time polymerase chain reaction was used to assess the levels of microbes including Streptococcus mutans, Streptococcus sobrinus, Lactobacillus salivarius, and Lactobacillus acidophilus in the saliva of 41 patients with a solid tumor (SO), 30 patients with a hematologic malignancy (HE), and 40 healthy controls. Salivary flow and pH were lower in oncological patients than in controls. The frequencies of all four cariogenic bacteria were highest in the SO group. S. mutans and L. salivarius were the most commonly detected in all three study groups. Mean numbers of S. sobrinus and L. salivarius in the SO group were significantly higher than in controls (p<0.05). There were no significant differences between patients and controls with respect to mean numbers of S. mutans and L. acidophilus in saliva. However, the proportions of S. mutans, S. sobrinus, and L. salivarius versus total bacteria in the SO group were significantly higher than in controls. Within patients, both mean numbers and the proportions of S. mutans and S. sobrinus were significantly different (p<0.05). In summary, significant differences were found in salivary pH values and the levels of S. mutans, S. sobrinus, and L. salivarius between SO patients and healthy controls.

Predictive value of pretreatment risk group and baseline LDH levels in MDS patients receiving azacitidine treatment.

This study analyzed the outcomes of myelodysplastic syndrome patients treated with azacitidine. Between August 2006 and June 2008, a total of 126 patients were treated with azacitidine at a dose of 75 mg/m(2)/day subcutaneously for 7 days, which was repeated every 28 days. The median age of the patients was 64 (range, 20-82) years. Forty-three patients (33.4%) were classified as intermediate-2 and high risk according to International Prognostic Scoring System (IPSS), while 61 patients (47.3%) were classified as high and very high risk according to WHO Prognostic Scoring System (WPSS). Sixty patients (47.6%) exhibited a response at the median of 3 (range, 1-5) cycles. A complete response was observed in 21 patients (16.7%), a partial response in six patients (4.8%), and total hematologic improvement in 61 patients (48.4%). For the IPSS risk group, the median survival for the patients with intermediate-1 was 20.0 months, for intermediate-2 was 14.9 months, and for high risk was 6.3 months (p = 0.008). For the WPSS risk group, the median survival duration was 21.3 months for the very low and low risk patients, 16.5 months for intermediate risk patients, and 14.9 months for the high and very high risk patients (p = 0.003). The patients with higher than normal lactate dehydrogenase (LDH) levels at the time of diagnosis showed a poor survival (p = 0.003). The median survival duration for the patients with high LDH levels was 13.9 months, while that for the patients with normal LDH levels was 20.6 months. The multivariate analyses revealed that high LDH levels [hazard ratio (HR) 4.384, p < 0.001] and high and very high WPSS risk group (HR 3.855, p = 0.014) were significantly associated with a worse survival, whereas a response to azacitidine was identified as a good prognostic factor for survival (HR 0.224, p = 0.019). In conclusion, while the pretreatment risk group and initial LDH levels were both confirmed as important prognostic factors to predict the outcomes for patients treated with azacitidine, more effective therapies are still needed to prevent disease progression.

Differential involvement of protein kinase C in human promyelocytic leukemia cell differentiation enhanced by artemisinin.

Artemisinin, a sesquiterpene lactone endoperoxide that exists in several medicinal plants, is a well-known anti-malarial agent. In this report, we investigated the effect of artemisinin on cellular differentiation in the human promyelocytic leukemia HL-60 cell culture system. Artemisinin markedly increased the degree of HL-60 leukemia cell differentiation when simultaneously combined with low doses of 1 alpha,25-dihydoxyvitamin D(3) [1,25-(OH)(2)D(3)] or all-trans retinoic acid (all-trans RA). Artemisinin by itself had very weak effects on the differentiation of HL-60 cells. Cytofluorometric analysis and cell morphologic studies indicated that artemisinin potentiated 1,25-(OH)(2)D(3)-induced cell differentiation predominantly into monocytes and all-trans RA-induced cell differentiation into granulocytes, respectively. Extracellular-regulated kinase (ERK) inhibitors markedly inhibited HL-60 cell differentiation induced by artemisinin in combination with 1,25-(OH)(2)D(3) or all-trans RA, whereas phosphatidylinositol 3-kinase (PI3-K) inhibitors did not. Particularly, protein kinase C (PKC) inhibitors inhibited HL-60 cell differentiation induced by artemisinin in combination with 1,25-(OH)(2)D(3) but not with all-trans RA. Artemisinin enhanced PKC activity and protein level of PKC beta I isoform in only 1,25-(OH)(2)D(3)-treated HL-60 cells. Taken together, these results indicate that artemisinin strongly enhanced 1,25-(OH)(2)D(3)- and all-trans RA-induced cell differentiation in which PKC is differentially involved in arteminisin-mediated enhancement of leukemia cell differentiation.