Search

Molecular testing foriJAK2/imutations is part of the standard diagnostic workup for patients with suspected polycythemia vera. We sought to characterize evolving practice patterns in the investigation of erythrocytosis and the prevalence of secondary causes, including use of medications such as sodium-glucose cotransporter-2 (SGLT2) inhibitors, among patients who underwent molecular testing.We reviewed charts of all consecutive patients investigated for erythrocytosis (hemoglobingt; 160 g/L for women,gt; 165 g/L for men) withiJAK2/itesting between 2015 and 2021 at London Health Sciences Centre, a tertiary referral centre in Ontario, Canada, to assess changes in rates ofiJAK2/imutation positivity, average hemoglobin levels and the prevalence of secondary causes of erythrocytosis.A total of 891 patients with erythrocytosis underwentiJAK2/imutation testing with an increase in number of tests (particularly from 2017 to 2018), a decrease in the rate ofiJAK2/ipositivity and similar average hemoglobin levels over the study period. We observed a high proportion of patients with secondary causes of erythrocytosis, ranging from 59% to 74% over the study period, including medications associated with erythrocytosis, namely testosterone (6%-11%) and SGLT2 inhibitors (2%-19%). Stopping SGLT2 inhibitors was associated with a significant decrease in hemoglobin levels (mean -14.7 g/L, 95% confidence interval -18.9 to -10.5 g/L) compared with continuation.Use of SGLT2 inhibitors may be a common and underrecognized secondary cause of elevated hemoglobin levels in patients investigated for erythrocytosis. Our findings underscore the importance of a detailed medical history to support judicious use of molecular testing, in adherence with the current guideline on the investigation of erythrocytosis.

Erythrocytosis, most often measured as an increase in hemoglobin and/or hematocrit, is a common reason for referral to internal medicine and hematology clinics and a rational approach is required to effectively identify patients with polycythemia vera while avoiding over-investigation.We aimed to develop and validate a simple rule to predict JAK2 mutation positivity based on complete blood count parameters to aid in the diagnostic approach to patients referred for elevated hemoglobin.Internal medicine and hematology clinics at an academic tertiary referral center.The JAK2 Prediction Cohort (JAKPOT), a large retrospective cohort (n = 901) of patients evaluated by internal medicine and hematology specialists for elevated hemoglobin.JAK2 mutation analysis was performed in all patients and clinical and laboratory variables were collected. Patients were randomly divided into derivation and validation cohorts. A prediction rule was developed using data from the derivation cohort and tested in the validation cohort.The JAKPOT prediction rule included three variables: (i) red blood cell count6.45×10In patients with elevated hemoglobin, the use of a simple prediction rule helps to accurately identify patients with a low likelihood of having a JAK2 mutation, potentially limiting costly over-investigation in this common referral population.

Background: Since the identification of JAK2 mutations in polycythemia vera (PV) in 2005 (Kralovics et al., NEJM 2005), molecular testing of JAK2 in patients with erythrocytosis has become part of routine clinical practice. We hypothesized that changes in the World Health Organization (WHO) diagnostic criteria for PV in 2016, which lowered the hemoglobin threshold to >165 g/L for men and >160 g/L for women, may have resulted in increased molecular testing. This study examines changing patterns of utilization of molecular diagnostics in patients referred for erythrocytosis at a tertiary care center. Methods: We examined all patients with erythrocytosis who underwent JAK2 testing, which included testing for JAK2 V617F with PCR between 2015 and 2017, and JAK2 V617F and exon 12 mutations with Next-Generation Sequencing (NGS) between 2018 and 2020 at London Health Sciences Centre in Ontario, Canada. We performed a retrospective chart review to extract laboratory and clinical data, including information on medical comorbidities and medications, with a focus on known secondary causes of erythrocytosis. Results: A total of 668 patients with erythrocytosis underwent JAK2 testing at our institution between August 1, 2015 and December 31, 2020. There was an overall increase in testing over the five-year study period, with a decline in the positive detection rate: 8/29 (28%) in 2015, 15/94 (16%) in 2016, 15/100 (15%) in 2017, 19/136 (14%) in 2018, 17/162 (10%) in 2019, and 14/147 (10%) in 2020 (Figure 1). The average hemoglobin levels in patients with erythrocytosis who underwent testing remained similar across all years (range 170-173 g/L for women, 179-181 g/L for men). In our cohort, there was a high proportion of patients with known or suspected secondary causes of erythrocytosis who underwent molecular testing. Between 2018 and 2020, 324/445 (73%) of patients who underwent molecular testing had either chronic obstructive pulmonary disease, obstructive sleep apnea, other hypoxic lung disease, smoking history, erythropoietin-secreting tumor, or potential drug-induced erythrocytosis. Specifically, we observed an increase in proportion of patients who underwent molecular testing on sodium-glucose cotransporter-2 (SGLT-2) inhibitors, a known secondary cause of erythrocytosis, with 15/136 (11%) in 2018, 17/162 (10%) in 2019, and 25/147 (17%) in 2020. In contrast, the proportion of patients on testosterone was relatively constant at 15/136 (11%) in 2018, 11/162 (6.8%) in 2019, and 11/147 (7.5%) in 2020. Conclusion: This study revealed that a high proportion of patients with known or suspected secondary causes of erythrocytosis underwent JAK2 testing, resulting in increase in molecular testing over time and a decline in positive detection rate. In particular, we observed a number of patients on SGLT-2 inhibitors who had investigation, suggesting that this class of medications may be an underrecognized cause of drug-induced erythrocytosis (Chin-Yee et al., CMAJ 2020). Our findings underscore the importance of careful medical history and medication review to support more judicious use of molecular testing. Similarity in average hemoglobin levels across the five-year study period suggests that other factors, such as increased availability of 'routine' molecular testing, rather than changes in the WHO diagnostic criteria may explain increases in JAK2 testing. Our study indicates a need to develop an effective clinical prediction rule for JAK2 positivity to better risk stratify patients with suspected PV based on clinical and laboratory parameters to optimize utilization of molecular diagnostics. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Background: The widespread availability of molecular testing for JAK2 mutations has facilitated the diagnosis of polycythemia vera (PV) but also raises the concern of test overutilization in patients referred for elevated hemoglobin. At our institution, we have observed increased molecular testing in these patients with declining rates of JAK2 mutation positivity, suggesting that a prediction rule could be useful to guide such testing. In this study, we report the derivation and validation of a simple rule using complete blood count (CBC) parameters to predict the likelihood of having a JAK2 mutation in patients referred for elevated hemoglobin. Methods: We examined all patients with elevated hemoglobin (≥160 g/L for women, or ≥165 g/L for men), who underwent JAK2 mutation testing using the Next-Generation Sequencing (NGS)-based Oncomine Myeloid Research Assay (ThermoFisher Scientific, MA, USA), between 2018 and 2021 at the London Health Sciences Centre in Ontario, Canada. We extracted data including age and sex as well as CBC parameters at the time of testing, including hemoglobin, hematocrit, erythrocytes, leukocytes, neutrophils, platelets and mean corpuscular volume. All CBCs were performed on a Sysmex XN Analyzer (Sysmex Corporation, Japan). In the derivation cohort, JAK2-positive and -negative groups were compared using Student's t-tests or c 2 tests, as appropriate. We dichotomized potentially significant continuous variables at an optimal cut-off point using receiving operating characteristic curves. Potentially significant predictors were evaluated using multiple variable stepwise logistic regression analysis with JAK2 positivity as the dependent variable. The model was evaluated using Hosmer-Lemeshow tests and pseudo-R2 measures. A dichotomous score was derived based on the presence or absence of significant variables and subsequently evaluated and internally validated using logistic regression and c 2 tests using non-parametric bootstrapping with 1000 samples. The model was subsequently validated in the second cohort. Results: The derivation cohort included 308 patients tested between January 9, 2018 and December 19, 2019, and the validation cohort included 223 patients tested between January 7, 2020 and May 12, 2021. The characteristics of both cohorts are shown in Table 1. The final model included platelets above the upper quintile (308 × 10 9/L) and erythrocytes above the upper quartile (6.17 × 10 12/L) and a score of one was assigned to patients with either of these characteristics. The odds ratio for JAK2 positivity in patients with a score of 1 was 14.6 (95% CI 5.5-38.8) compared to those with a score of 0. The model had a sensitivity of 87.8% and a negative predictive value of 97.4% in the derivation cohort, and of 100% for both in the validation cohort. The percentage of JAK2 positive patients in patients with a score of 1 was 28%. The percent of false negatives was 2.6% (95% CI 1.1-6.0) and 0 (95% CI 0-2.8) in the derivation and validation cohorts, respectively. The use of this rule to guide molecular testing would have resulted in approximately 60% fewer tests. Conclusion: We developed and validated a simple rule to predict the likelihood of JAK2 mutation positivity in patients with a hemoglobin of 160 or higher, based on CBC parameters with a high negative predictive value (Figure 1). If implemented, this prediction rule could result in a significant reduction in molecular testing avoiding 60% or approximately 100 tests per year at our institution. This approach would be particularly beneficial for broader health system management of hematological malignancies, facilitating the reallocation of resources to emerging higher-yield molecular diagnostic investigation (Kawata et al., BJH 2021). Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Background: JAK2 V617F and exon 12 mutations are the characteristic driver mutations in polycythemia vera (PV), identified in more than 95% of patients. In addition, other genetic mutations have previously been described in JAK2-positive PV that appear to have prognostic significance (Tefferi et al., Blood 2016). The incidence of other driver mutations in unselected patients referred for elevated hemoglobin is less well studied. This study aims to characterize the genetic mutational landscape in a real-world population of patients referred for elevated hemoglobin using a targeted Next-Generation Sequencing (NGS)-based assay. Methods: We reviewed all patients referred for elevated hemoglobin levels (>160 g/L in females or >165 g/L in males) between 2018 and 2020 to hematology clinics at London Health Sciences Centre in Southwestern Ontario, Canada who underwent testing for genetic variants using the NGS-based Oncomine Myeloid Research Assay (ThermoFisher Scientific, MA, USA). This assay targets 40 key genes with diagnostic and prognostic implications in several myeloid malignancies (17 full genes and 23 genes with clinically relevant "hotspot" regions) and a panel of 29 fusion driver genes (>600 fusion partners). Patient demographics, laboratory data and final diagnosis were extracted from the electronic medical record. For all patients with genetic mutations, clinical diagnosis was confirmed by three independent reviewers. Results: A total of 529 patients underwent genetic testing for elevated hemoglobin levels: 389 (73.5%) were males (mean age 58; range 18-95) and 140 (26.5%) were female (mean age 60; range 24-85). JAK2 mutations were detected in 10.9% (58/529) of patients and a diagnosis of PV was confirmed. The majority of JAK2-mutated PV patients (n=57) were positive for JAK2 V617F, while one patient had an exon 12 mutation. Additional single myeloid mutations were detected in 34.5% (20/58) of JAK2-positive patients and involved the following genes: TET2 (11; 19%), DNMT3A (2; 3.4%), ASXL1 (2; 3.4%), SRSF2 (2; 3.4%), BCOR (1; 1.7%), TP53 (1; 1.7%) and ZRSR2 (1; 1.7%) (Figure 1A). JAK2 mutations were not detected in 89.0% (471/529) of our cohort. A diagnosis of PV was confirmed in 2 JAK2-negative patients based on clinical features and myeloid mutations were detected in both: SRSF2 and TET2 gene mutations in 1 patient and SRSF2, IDH2, ASXL1 gene mutations in the other patient. Three JAK2-negative patients tested positive for the BCR-ABL fusion and were diagnosed with chronic myeloid leukemia. The remaining 466 JAK2-negative patients were diagnosed with secondary erythrocytosis and myeloid mutations were found in 6% (28/466) of these cases. Mutations were detected in DNMT3A (12; 2.6%), TET2 (5; 1.1%), ASXL1 (5; 1.1%), TP53 (2; 0.4%), NF1 (2; 0.4%), KIT (1; 0.2%), U2AF1 (1; 0.2%) (Figure 1B). All patients with JAK2-negative secondary erythrocytosis had only one myeloid gene mutation detected. Conclusion: Additional myeloid mutations other than JAK2 mutations are frequently identified in patients referred for erythrocytosis, with the highest frequencies observed in the TET2, DNMT3A and ASXL1 genes. The spectrum of myeloid mutations and overall incidence in JAK2-negative patients with secondary erythrocytosis is similar to the reported incidence of Clonal Hematopoiesis of Indeterminate Potential (CHIP) (Jaiswal et al., NEJM 2014), and suggests that these may represent incidental age-related mutations. By contrast, among the JAK2-positive patients, 34.5% had at least one additional myeloid mutation supporting a pathogenic role in these patients with myeloproliferative neoplasms. While concomitant myeloid mutations in patients with PV are well-described, further research is required to elucidate the significance of variants identified in JAK2-negative patients classified as secondary erythrocytosis in order to determine whether these mutations contribute to clinical phenotype or represent background CHIP. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.