Exploring RAS mutation incidence and temporal heterogeneity in metastatic colorectal cancer patients - a single-institution experience utilising circulating tumour DNA.

Introduction: Colorectal cancer (CRC) remains a significant global health challenge, ranking among the leading causes of neoplastic mortality. Despite transformative therapeutic advances, a considerable proportion of patients are diagnosed with metastatic disease, and 15-30% of those initially presenting with early-stage CRC eventually experience recurrence. Comprehensive molecular testing, especially the evaluation of microsatellite instability and mutations in KRAS/NRAS or BRAF genes, is essential upon diagnosis of stage IV disease, guiding treatment decisions. Material and methods: This manuscript explores the mutational landscape of KRAS and NRAS in patients with CRC, employing digital polymerase chain reaction (PCR) BEAMing for the detection of mutations in liquid biopsy. Our study enrolled patients with histologically confirmed CRC and stage IV disease, focusing on identifying mutations in KRAS and NRAS genes during various stages of therapy. Results: Evaluating baseline, midline, and progression samples, we found that 66.6% maintained consistent mutational status post-disease progression, while 33.3% exhibited a shift in mutational status. The application of techniques with high sensitivity, such as BEAMing Digital PCR, is pivotal for accurate circulating tumour DNA (ctDNA) mutation detection. The study underscores the significance of continuous molecular monitoring in guiding therapeutic decisions for patients with metastatic CRC. Conclusions: Our findings contribute to our understanding of the evolving mutational landscape and the potential clinical implications of ctDNA ana- lysis in the era of personalised cancer medicine.

Development of saxitoxin biosynthesis gene sxtB-targeted qPCR assay for the quantification of toxic dinoflagellates Alexandrium catenella (group I) and A. pacificum (group IV) occurring in the Korean coast.

Toxic dinoflagellate Alexandrium can produce saxitoxins (STXs) and cause paralytic shellfish poisoning (PSP), and thus they are monitored for environmental safety management. Microscopic discrimination of dinoflagellates is difficult to distinguish between toxic and non-toxic species due to their similar morphology. Meanwhile, an alternative quantitative PCR (qPCR) assay is sensitive, rapid, and cost-effective for harmful species monitoring. Herein, we developed a novel qPCR assay to detect the STXs biosynthesis gene sxtB of Alexandrium catenella and A. pacificum, the leading cause of PSP outbreaks in Asian coasts and worldwide. The newly designed sxtB TaqMan probes target the species without any positive signal in other relative dinoflagellates. Deming regression analysis revealed that the sxtB copy number of A. catenella and A. pacificum was 3.6 and 4.1 copies per cell, respectively. During the blooming periods (April 13th-14th, 2020), only A. catenella cells were detected through the qPCR assay, ranging from 5.0 × 10 to 2.5 × 104 eq cells L-1. In addition, sxtB qPCR quantified more accurately compared to large subunit (LSU) rRNA targeting qPCR assay that overestimate cell density. Besides, the sensitivity of sxtB was higher compared to the microscope when the species were rarely present (5.0 × 102 cells L-1). These suggest that the sxtB qPCR assay can be applied to toxic Alexandrium monitoring in the Korean coast, even in the early stage of bloomings.

Genomic surveillance of Anopheles mosquitoes on the Bijagós Archipelago using custom targeted amplicon sequencing identifies mutations associated with insecticide resistance.

BACKGROUND: Insecticide resistance is reducing the efficacy of vector control interventions, consequently threatening efforts to control vector-borne diseases, including malaria. Investigating the prevalence of molecular markers of resistance is a useful tool for monitoring the spread of insecticide resistance in disease vectors. The Bijagós Archipelago (Bijagós) in Guinea-Bissau is a region of stable malaria transmission where insecticide-treated nets are the mainstay for malaria control. However, the prevalence of molecular markers of insecticide resistance in malaria vectors is not well understood. METHODS: A total of 214 Anopheles mosquitoes were analysed from 13 islands across the Bijagós. These mosquitoes were collected using CDC light traps in November 2019, during the peak malaria transmission season. High-throughput multiplex amplicon sequencing was used to investigate the prevalence of 17 different molecular markers associated with insecticide resistance in four genes: vgsc, rdl, ace1 and gste2. RESULTS: Of the 17 screened mutations, four were identified in mosquitoes from the Bijagós: vgsc L995F (12.2%), N1570Y (6.2%) and A1746S (0.7%) and rdl A269G (1.1%). This study is the first to report the L995F knock-down resistance (kdr)-west allele in Anopheles melas on the Archipelago. An additional eight non-synonymous single-nucleotide polymorphisms were identified across the four genes which have not been described previously. The prevalences of the vgsc L995F and N1570Y mutations were higher on Bubaque Island than on the other islands in this study; Bubaque is the most populous island in the archipelago, with the greatest population mobility and connection to continental Guinea-Bissau. CONCLUSIONS: This study provides the first surveillance data for genetic markers present in malaria vectors from islands across the Bijagós Archipelago. Overall prevalence of insecticide resistance mutations was found to be low. However, the identification of the vgsc L995F and N1570Y mutations associated with pyrethroid resistance warrants further monitoring. This is particularly important as the mainstay of malaria control on the islands is the use of pyrethroid insecticide-treated nets.

[Detection of BCR-ABL Fusion Gene in Chronic Myeloid Leukemia by Novel Digital PCR].

OBJECTIVE: To establish a new digital polymerase chain reaction (dPCR) system for the detection of BCR-ABL fusion gene in patients with chronic myeloid leukemia (CML), and explore its analytical performance and clinical applicability in the detection of BCR-ABLp190/210/230. METHODS: A new dPCR system for detecting BCR-ABLp190/210/230 was successfully developed, and its sensitivity difference with qPCR and improvement of drug side effects in patients with CML during drug reduction or withdrawal were compared. RESULTS: Among 176 samples, qPCR and dPCR showed high consistency in the sensitivity of detecting BCR-ABL (82.39%), and the positive rate of dPCR was about 5 times higher that of qPCR (20.45% vs 3.98%). During follow-up, blood routine (25% vs 10%), kidney/liver/stomach (25% vs 20%) and cardiac function (10% vs 0) were significantly improved after drug reduction or withdrawal in patients with initial dPCR negative compared with before drug reduction or withdrawal. CONCLUSIONS: This new dPCR detection system can be applied to the detection of BCR-ABLp190/210/230. It has better consistency and higher positive detection rate than qPCR. Drug withdrawal or dose reduction guided by dPCR has a certain effect on improving drug side effects.

Exploration of treatment-free remission in CML, based on molecular monitoring.

BACKGROUND: Typical chronic myelogenous leukemia (CML) is a myeloproliferative neoplasm caused by t(9; 22)(q34; q11) translocation. This chromosomal translocation forms the BCR::ABL1 fusion gene. The tyrosine kinase encoded by the BCR::ABL1 is considered to be the main pathogenic diver. BCR::ABL1 is not only a therapeutic target, but also a monitoring target. Monitoring of BCR::ABL1 reveals the progression of the disease and guides the next treatment. Now for CML, the target of treatment has been focused on treatment-free remission (TFR). METHODS: We conducted a literature review of current developments of treatment-free remission and molecular monitoring methods. RESULTS: More effective and sensitive CML monitoring methods such as digital droplet PCR (ddPCR) and next generation sequencing (NGS) have further studied the measurable residual disease (MRD) and clonal heterogeneity, which provides strong support for the exploration of TFR. We discussed some of the factors that may be related to TFR outcomes at the molecular level, along with some monitoring strategies. CONCLUSION: Currently, predictive indicators for treatment-free remission outcomes and recurrence are lacking in clinical practice. In future, treatment-free remission research should focus on combining the clinical indicators with molecular monitoring and biological markers to personalize patient conditions and guide clinicians to develop individualized treatment plans, so that more patients with CML can achieve safer and stabler treatment-free remission.

A review on characterization of BCR - ABL transcript variants for molecular monitoring of chronic myeloid leukemia phenotypes.

Chronic myeloid leukemia (CML) is a clonal myeloproliferative growth of human pluripotent stem cells which is estimated to occur at a rate of 1/100000 populations every year worldwide. A characteristic feature of this disease is the presence of the Philadelphia chromosome genotype, which results from the reciprocal translocation between human chromosomes 9 and 22. Two types of major genotypes are involved, which consequently result in two major types of expressed fusion mRNA transcripts: b3a2 and b2a2, i.e. major breakpoint segments (happening after exon 13 & after exon 14) of the BCR gene on chromosome 22 fuze with the ABL1 gene breakpoint (happening after exon 2) on chromosome 9, forming two genotypes coding for two transcripts: b3a2 (e14a2) and b2a2 (e13a2). The protein 'p210 BCR-ABL1', a protein which characteristically exhibits a high tyrosine kinase activity which is followed by the activation of various cellular processes that lead to increased cellular proliferation and cancer, is coded by both major BCR - ABL1 mRNA transcripts. Recent developments in the treatment of CML through molecular monitoring of the disease have managed to reduce patient morbidity and mortality. Advanced molecular techniques are aimed at detecting BCR-ABL1 transcript levels to monitor treatment response. Transcript typing is necessary to detect minimal residual disease and to achieve molecular response by helping to provide selective therapy based on the type of transcript identified, as transcript type is correlated with the disease course.The purpose of this review is to discuss: the role of the BCR-ABL1 fusion gene in the pathogenesis of CML; the role of BCR-ABL1 transcript characterization in the molecular monitoring of CML therapy; the association of BCR - ABL1 transcript types with different CML phenotypes, molecular responses, and treatment responses; and the laboratory techniques employed to detect and characterize BCR - ABL1 transcripts.

Epidemiology and Resistance Phenotypes of Carbapenem-Resistant Klebsiella pneumoniae in Corfu General Hospital (2019-2022): A Comprehensive Time Series Analysis of Resistance Gene Dynamics.

Antimicrobial resistance is a significant global health challenge, with Klebsiella pneumoniae being one of the most common antibiotic-resistant pathogens. This study provides an in-depth analysis of the prevalence and resistance patterns of antibiotic-resistant Klebsiella pneumoniae in the General Hospital of Corfu, Greece, between 2019 and 2022, with the aim of understanding the potential impact of the COVID-19 pandemic on the epidemiology of this bacterium. Utilizing a retrospective epidemiological approach, this study analyzed 212 isolates obtained from the hospital's Microbiology Department. These isolates were subjected to genotypic and phenotypic identification, with resistance genes (bla-KPC, bla-NDM, bla-VIM, bla-OXA-48, and mcr-1) and antibiotic resistance patterns as the primary focus. The results revealed a significant shift in resistance gene prevalence, with a notable increase in bla-KPC from 16.67% in 2021 to 58.46% in 2022, and a decrease in bla-NDM from 81.48% in 2021 to 38.46% in 2022. In terms of antibiotic resistance patterns, there was a consistent increase in resistance to amikacin and a significant decrease in resistance to ceftazidime/avibactam. These findings underscore the dynamic nature of carbapenem-resistant Klebsiella pneumoniae (CRKP) resistance and highlight the need for ongoing surveillance and adaptive therapeutic strategies in the face of evolving resistance patterns.

Recent Development on Sensing Strategies for Small Molecules Detections.

Sensors play a critical role in the detection and monitoring of various substances present in our environment, providing us with valuable information about the world around us. Within the field of sensor development, one area that holds particular importance is the detection of small molecules. Small molecules encompass a wide range of organic or inorganic compounds with low molecular weight, typically below 900 Daltons including gases, volatile organic compounds, solvents, pesticides, drugs, biomarkers, toxins, and pollutants. The accurate and efficient detection of these small molecules has attracted significant interest from the scientific community due to its relevance in diverse fields such as environmental pollutants monitoring, medical diagnostics, industrial optimization, healthcare remedies, food safety, ecosystems, and aquatic and terrestrial life preservation. To meet the demand for precise and efficient monitoring of small molecules, this summary aims to provide an overview of recent advancements in sensing and quantification strategies for various organic small molecules including Hydrazine, Glucose, Morpholine, Ethanol amine, Nitrosamine, Oxygen, Nitro-aromatics, Phospholipids, Carbohydrates, Antibiotics, Pesticides, Drugs, Adenosine Triphosphate, Aromatic Amine, Glutathione, Hydrogen Peroxide, Acetone, Methyl Parathion, and Thiophenol. The focus is on understanding the receptor sensing mechanism, along with the electrical, optical, and electrochemical response. Additionally, the variations in UV-visible spectral properties of the ligands upon treatment with the receptor, fluorescence and absorption titration analysis for limit of detection (LOD) determination, and bioimaging analysis are discussed wherever applicable. It is anticipated that the information gathered from this literature survey will be helpful for the perusal of innovation regarding sensing strategies.

Effectiveness of digital care platform CMyLife for patients with chronic myeloid leukemia: results of a patient-preference trial.

BACKGROUND: Two most important factors determining treatment success in chronic myeloid leukemia (CML) are adequate medication compliance and molecular monitoring albeit still being suboptimal. The CMyLife platform is an eHealth innovation, co-created with and for CML patients, aiming to improve their care, leading to an increased quality of life and the opportunity of hospital-free care. OBJECTIVE: To explore the effectiveness of CMyLife in terms of information provision, patient empowerment, medication compliance, molecular monitoring, and quality of life. METHODS: Effectiveness of CMyLife was explored using a patient-preference trial. Upon completion of the baseline questionnaire, participants actively used (intervention group) or did not actively use (questionnaire group) the CMyLife platform for at least 6 months, after which they completed the post-intervention questionnaire. Scores between the intervention group and the questionnaire group were compared with regard to the within-subject change between baseline and post-measurement using Generalized Estimating Equation models. RESULTS: At baseline, 33 patients were enrolled in the questionnaire group and 75 in the intervention group. Online health information knowledge improved significantly when actively using CMyLife and patients felt more empowered. No significant improvements were found regarding medication compliance and molecular monitoring, which were already outstanding. Self-reported effectiveness showed that patients experienced that using CMyLife improved their medication compliance and helped them to oversee their molecular monitoring. Patients using CMyLife reported more symptoms but were better able to manage these. CONCLUSIONS: Since hospital-free care has shown to be feasible in time of the COVID-19 pandemic, eHealth-based innovations such as CMyLife could be a solution to maintain the quality of care and make current oncological health care services more sustainable. TRIAL REGISTRATION: ClinicalTrials.gov NCT04595955 , 22/10/2020.

Landscape of BCL2 Resistance Mutations in a Real-World Cohort of Patients with Relapsed/Refractory Chronic Lymphocytic Leukemia Treated with Venetoclax.

The oral, highly selective Bcl2 inhibitor venetoclax has substantially improved the therapeutic landscape of chronic lymphocytic leukemia (CLL). Despite the remarkable response rates in patients with relapsed/refractory (R/R) disease, acquired resistance is the leading cause of treatment failure, with somatic BCL2 mutations being the predominant genetic drivers underpinning venetoclax resistance. To assess the correlation between disease progression and the most common BCL2 mutations G101V and D103Y, sensitive (10-4) screening for the most common BCL2 mutations G101V and D103Y was performed in 67 R/R CLL patients during venetoclax single-agent or venetoclax-rituximab combination therapy. With a median follow-up time of 23 months, BCL2 G101V and D103Y were detected in 10.4% (7/67) and 11.9% (8/67) of the cases, respectively, with four patients harboring both resistance mutations. Ten out of eleven patients carrying BCL2 G101V and/or D103Y experienced relapse during the follow-up period, representing 43.5% of the cases (10/23) showing clinical signs of disease progression. All BCL2 G101V or D103Y variants were detected in patients receiving venetoclax as a continuous single-agent treatment while these mutations were not observed during or after fixed-duration venetoclax therapy. Targeted ultra-deep sequencing of BCL2 uncovered three additional variants in four patient samples obtained at relapse, suggesting convergent evolution and implying a cooperating role of BCL2 mutations in driving venetoclax resistance. This cohort is the largest R/R CLL patient population reported to date in which BCL2 resistance mutations were investigated. Our study demonstrates the feasibility and clinical value of sensitive screening for BCL2 resistance mutations in R/R CLL.

Detection of BCR-ABL Fusion Gene in Chronic Myeloid Leukemia by Novel Digital PCR / 中国实验血液学杂志

OBJECTIVE@#To establish a new digital polymerase chain reaction (dPCR) system for the detection of BCR-ABL fusion gene in patients with chronic myeloid leukemia (CML), and explore its analytical performance and clinical applicability in the detection of BCR-ABLp190/210/230.@*METHODS@#A new dPCR system for detecting BCR-ABLp190/210/230 was successfully developed, and its sensitivity difference with qPCR and improvement of drug side effects in patients with CML during drug reduction or withdrawal were compared.@*RESULTS@#Among 176 samples, qPCR and dPCR showed high consistency in the sensitivity of detecting BCR-ABL (82.39%), and the positive rate of dPCR was about 5 times higher that of qPCR (20.45% vs 3.98%). During follow-up, blood routine (25% vs 10%), kidney/liver/stomach (25% vs 20%) and cardiac function (10% vs 0) were significantly improved after drug reduction or withdrawal in patients with initial dPCR negative compared with before drug reduction or withdrawal.@*CONCLUSIONS@#This new dPCR detection system can be applied to the detection of BCR-ABLp190/210/230. It has better consistency and higher positive detection rate than qPCR. Drug withdrawal or dose reduction guided by dPCR has a certain effect on improving drug side effects.

Potential and limitations for monitoring of pesticide biodegradation at trace concentrations in water and soil.

Pesticides application on agricultural fields results in pesticides being released into the environment, reaching soil, surface water and groundwater. Pesticides fate and transformation in the environment depend on environmental conditions as well as physical, chemical and biological degradation processes. Monitoring pesticides biodegradation in the environment is challenging, considering that traditional indicators, such as changes in pesticides concentration or identification of pesticide metabolites, are not suitable for many pesticides in anaerobic environments. Furthermore, those indicators cannot distinguish between biotic and abiotic pesticide degradation processes. For that reason, the use of molecular tools is important to monitor pesticide biodegradation-related genes or microorganisms in the environment. The development of targeted molecular (e.g., qPCR) tools, although laborious, allowed biodegradation monitoring by targeting the presence and expression of known catabolic genes of popular pesticides. Explorative molecular tools (i.e., metagenomics & metatranscriptomics), while requiring extensive data analysis, proved to have potential for screening the biodegradation potential and activity of more than one compound at the time. The application of molecular tools developed in laboratory and validated under controlled environments, face challenges when applied in the field due to the heterogeneity in pesticides distribution as well as natural environmental differences. However, for monitoring pesticides biodegradation in the field, the use of molecular tools combined with metadata is an important tool for understanding fate and transformation of the different pesticides present in the environment.

Impact of the COVID-19 pandemic on the regular follow-up and outcomes of patients with chronic myeloid leukemia in chronic-phase.

Introduction: COVID-19 immediately became a major consideration in the management of chronic myeloid leukemia (CML). The influence of such considerations on viral transmission rates and leukemic control remain to be explored. We conducted this study to identify these alterations and to investigate their clinical consequences. Methods: This was a cross-sectional study, performed at a single institution on CML patients who were interviewed with a survey. We compared variables concerning new attitudes in the pandemic era between the 12-month periods before and after the pandemic onset. Outcome data were attained from the hospital archives. Findings: The number of patients receiving regular outpatient care for CML in chronic phase was 210, 91% had achieved at least major molecular responses. We assessed survival, progression, number of clinical visits of all, performed the survey on 89% and evaluated molecular responses on 86.6% of these patients. The frequency of clinical and molecular monitoring was significantly reduced during the pandemic deviating significantly from the guidelines. Frequency of death, progression, loss of molecular response was not significantly increased during the pandemic era despite a few cases where the delay in assessment possibly played a role in the unfavorable outcomes. There were no COVID related deaths or disabilities. Conclusion: The case-based untoward events would have probably been better managed with a more efficient communication web between patients, hematologists, and the laboratory. Therefore, it seems reasonable to consider whether such communicative paths are functional before giving up on the set schedule of CML management at times of uncertainty.

The Need to Pair Molecular Monitoring Devices with Molecular Imaging to Personalize Health.

By enabling the non-invasive monitoring and quantification of biomolecular processes, molecular imaging has dramatically improved our understanding of disease. In recent years, non-invasive access to the molecular drivers of health versus disease has emboldened the goal of precision health, which draws on concepts borrowed from process monitoring in engineering, wherein hundreds of sensors can be employed to develop a model which can be used to preventatively detect and diagnose problems. In translating this monitoring regime from inanimate machines to human beings, precision health posits that continual and on-the-spot monitoring are the next frontiers in molecular medicine. Early biomarker detection and clinical intervention improves individual outcomes and reduces the societal cost of treating chronic and late-stage diseases. However, in current clinical settings, methods of disease diagnoses and monitoring are typically intermittent, based on imprecise risk factors, or self-administered, making optimization of individual patient outcomes an ongoing challenge. Low-cost molecular monitoring devices capable of on-the-spot biomarker analysis at high frequencies, and even continuously, could alter this paradigm of therapy and disease prevention. When these devices are coupled with molecular imaging, they could work together to enable a complete picture of pathogenesis. To meet this need, an active area of research is the development of sensors capable of point-of-care diagnostic monitoring with an emphasis on clinical utility. However, a myriad of challenges must be met, foremost, an integration of the highly specialized molecular tools developed to understand and monitor the molecular causes of disease with clinically accessible techniques. Functioning on the principle of probe-analyte interactions yielding a transducible signal, probes enabling sensing and imaging significantly overlap in design considerations and targeting moieties, however differing in signal interpretation and readout. Integrating molecular sensors with molecular imaging can provide improved data on the personal biomarkers governing disease progression, furthering our understanding of pathogenesis, and providing a positive feedback loop toward identifying additional biomarkers and therapeutics. Coupling molecular imaging with molecular monitoring devices into the clinical paradigm is a key step toward achieving precision health.

Molecular Monitoring of BCR-ABL Fusion Transcripts in Patients with Chronic Myeloid Leukemia During Treatment Using the Endpoint Fluorescence Method.

OBJECTIVE: The purpose of the study was to compare results and evaluate the agreement between the endpoint fluorescence (EPF) method and quantitative real-time polymerase chain reaction (QPCR) during molecular monitoring of patients with chronic myeloid leukemia (CML) receiving treatment. MATERIALS AND METHODS: The study was conducted at Molecular Lab of Riphah International University, Islamabad, Pakistan, from January 2017 to December 2018. A total of 150 blood specimens from 30 patients with CML were analyzed at regular intervals during therapy. The detection/quantification of transcript mRNA was done simultaneously using QPCR and the EPF method. RESULTS: Out of a total of 150 RNA specimens analyzed, 117 (78%) specimens were positive, whereas 33 (22%) were negative for the transcript using both methods at various stages of treatment. Strong linear negative correlations between the cycle threshold and relative fluorescence unit values were observed with P <.0001 at 0, 3, 6, 9, and 12 months of treatment. No significant difference (P >.05) between the means of the BCR-ABL percentage was observed in either method at all stages of treatment. The bias between the 2 methods was calculated as 0.069 ± 3.50, and 95% limits of agreement were 6.92% to -6.79%. CONCLUSION: We found that EPF is s simple method to detect/quantify BCR-ABL mRNA expression during treatment with comparable results to QPCR.

Analysis of the competitiveness between a non-aflatoxigenic and an aflatoxigenic Aspergillus flavus strain on maize kernels by droplet digital PCR.

Non-aflatoxigenic Aspergillus flavus strains are used as a biocontrol system on maize fields to decrease the aflatoxin biosynthesis of aflatoxigenic A. flavus strains. A. flavus strain AF36 was the first commercially available biocontrol strain and is authorized for use on maize fields by the US Environmental Protection Agency, e.g., in Texas and Arizona. A droplet digital PCR (ddPCR) assay was developed to analyze the mechanisms of competition and interaction of aflatoxigenic and non-aflatoxigenic A. flavus strains. This assay enables the parallel identification and quantification of the biocontrol strain A. flavus AF36 and the aflatoxigenic A. flavus strain MRI19. To test the assay, spores of both strains were mixed in varying ratios and were incubated on maize-based agar or maize kernels for up to 20 days. Genomic equivalent ratios (genome copy numbers) of both strains were determined by ddPCR at certain times after incubation and were compared to the spore ratios used for inoculation. The aflatoxin biosynthesis was also measured. In general, A. flavus MRI19 had higher competitiveness in the tested habitats compared to the non-aflatoxigenic strain, as indicated by higher final genomic equivalent ratios of this strain compared to the spore ratios used for inoculation. Nevertheless, A. flavus AF36 effectively controlled aflatoxin biosynthesis of A. flavus MRI19, as a clear aflatoxin inhibition was already seen by the inoculation of 10% spores of the biocontrol strain mixed with 90% spores of the aflatoxigenic strain compared to samples inoculated with only spores of the aflatoxigenic A. flavus MRI19.

Genomic and clinical findings in myeloid neoplasms with PDGFRB rearrangement.

Platelet-derived growth factor receptor B (PDGFRB) gene rearrangements define a unique subgroup of myeloid and lymphoid neoplasms frequently associated with eosinophilia and characterized by high sensitivity to tyrosine kinase inhibition. To date, various PDGFRB/5q32 rearrangements, involving at least 40 fusion partners, have been reported. However, information on genomic and clinical features accompanying rearrangements of PDGFRB is still scarce. Here, we characterized a series of 14 cases with a myeloid neoplasm using cytogenetic, single nucleotide polymorphism array, and next-generation sequencing. We identified nine PDGFRB translocation partners, including the KAZN gene at 1p36.21 as a novel partner in a previously undescribed t(1;5)(p36;q33) chromosome change. In all cases, the PDGFRB recombination was the sole cytogenetic abnormality underlying the phenotype. Acquired somatic variants were mainly found in clinically aggressive diseases and involved epigenetic genes (TET2, DNMT3A, ASXL1), transcription factors (RUNX1 and CEBPA), and signaling modulators (HRAS). By using both cytogenetic and nested PCR monitoring to evaluate response to imatinib, we found that, in non-AML cases, a low dosage (100-200 mg) is sufficient to induce and maintain longstanding hematological, cytogenetic, and molecular remissions.

Outcome of acute myeloid leukaemia in Nigeria: clinician's perspective.

The outcome of acute myeloid leukaemia (AML) has remained a major concern even in developed countries. In resource poor countries, it is envisaged that the outcome will be far worse because of late presentations, lack of appropriate diagnostic facilities and supportive care. However, data to validate this is lacking and many of these countries lack an effective cancer registry. This study determined the clinician's perspective of the outcome of care of AML patients in Nigeria and their attitudes to the care of these patients. Structured self-administered questionnaire was used to assess the clinician's perception of outcomes of care, contributory factors and attitude to care of AML patients. Ninety-eight percent of clinicians reported that the outcome of care was suboptimal; 73.3% and 90.6% of the clinicians reported having less than 31% of AML patients surviving induction and post-induction therapies, respectively. Sixty-six-point one percent (66.1%), 50% and 62.7% of the clinicians have never used immunophenotyping, cytogenetic or molecular studies, respectively, in the management of AML patients under their care. Access to blood components other than Red cells was low; 23.3% had access to apheresis platelets and 55% to fresh frozen plasma. Forty-six percent of clinicians will either give half dose of chemotherapy or offer only supportive care. This reported early death rate is three times higher than that reported in developed countries with only 9% likely to survive the first year of induction compared to about 32.9% in Ontario. Approximately 28 units of pooled or apheresis derived platelet may be required in course of therapy but just 10% of clinicians have access to platelet apheresis. Lack of diagnostic facilities, blood components and clinicians' attitudes are contributing factors to the extremely poor outcomes of patients with AML in Nigeria.

Screening and monitoring of the BTKC481S mutation in a real-world cohort of patients with relapsed/refractory chronic lymphocytic leukaemia during ibrutinib therapy.

The Bruton's tyrosine kinase (BTK) inhibitor ibrutinib has revolutionised the therapeutic landscape of chronic lymphocytic leukaemia (CLL). Acquired mutations emerging at position C481 in the BTK tyrosine kinase domain are the predominant genetic alterations associated with secondary ibrutinib resistance. To assess the correlation between disease progression, and the emergence and temporal dynamics of the most common resistance mutation BTKC481S , sensitive (10-4 ) time-resolved screening was performed in 83 relapsed/refractory CLL patients during single-agent ibrutinib treatment. With a median follow-up time of 40 months, BTKC481S was detected in 48·2% (40/83) of the patients, with 80·0% (32/40) of them showing disease progression during the examined period. In these 32 cases, representing 72·7% (32/44) of all patients experiencing relapse, emergence of the BTKC481S mutation preceded the symptoms of clinical relapse with a median of nine months. Subsequent Bcl-2 inhibition therapy applied in 28/32 patients harbouring BTKC481S and progressing on ibrutinib conferred clinical and molecular remission across the patients. Our study demonstrates the clinical value of sensitive BTKC481S monitoring with the largest longitudinally analysed real-world patient cohort reported to date and validates the feasibility of an early prediction of relapse in the majority of ibrutinib-treated relapsed/refractory CLL patients experiencing disease progression.

Assessment of individual molecular response in chronic myeloid leukemia patients with atypical BCR-ABL1 fusion transcripts: recommendations by the EUTOS cooperative network.

PURPOSE: Approximately 1-2% of chronic myeloid leukemia (CML) patients harbor atypical BCR-ABL1 transcripts that cannot be monitored by real-time quantitative PCR (RT-qPCR) using standard methodologies. Within the European Treatment and Outcome Study (EUTOS) for CML we established and validated robust RT-qPCR methods for these patients. METHODS: BCR-ABL1 transcripts were amplified and sequenced to characterize the underlying fusion. Residual disease monitoring was carried out by RT-qPCR with specific primers and probes using serial dilutions of appropriate BCR-ABL1 and GUSB plasmid DNA calibrators. Results were expressed as log reduction of the BCR-ABL1/GUSB ratio relative to the patient-specific baseline value and evaluated as an individual molecular response (IMR). RESULTS: In total, 330 blood samples (2-34 per patient, median 8) from 33 CML patients (19 male, median age 62 years) were analyzed. Patients expressed seven different atypical BCR-ABL1 transcripts (e1a2, n = 6; e6a2, n = 1; e8a2, n = 2; e13a3, n = 4; e14a3, n = 6; e13a3/e14a3, n = 2; e19a2, n = 12). Most patients (61%) responded well to TKI therapy and achieved an IMR of at least one log reduction 3 months after diagnosis. Four patients relapsed with a significant increase of BCR-ABL1/GUSB ratios. CONCLUSIONS: Characterization of atypical BCR-ABL1 transcripts is essential for adequate patient monitoring and to avoid false-negative results. The results cannot be expressed on the International Scale (IS) and thus the common molecular milestones and guidelines for treatment are difficult to apply. We, therefore, suggest reporting IMR levels in these cases as a time-dependent log reduction of BCR-ABL1 transcript levels compared to baseline prior to therapy.