Dose Methotrexate Research Articles

Outcomes in Routine Clinical Practice with Methotrexate-Temozolomide-Rituximab Induction and Modified Etoposide-Cytarabine Consolidation in Newly Diagnosed Primary CNS Lymphoma

Background: Recent studies have reported improved outcomes for patients with Primary CNS lymphoma (PCNSL) with addition of agents in induction (for e.g. cytarabine, thiotepa and rituximab in IELSG studies), intense consolidation (Autologous stem cell transplant (ASCT) as in IELSG 43) and intense non-myeloablative chemotherapy with etoposide/cytarabine (EA) as in CALGB 50202). Delivery of such an intense therapy is often difficult in clinical practice due to advanced age, comorbidities, poor performance status and resource constraints. We started using induction with methotrexate, temozolomide and rituximab (MTR) as per CALGB 50202 study however the intense non-myeloablative EA consolidation was not feasible. We used a strategy of reduced dose EA consolidation for 2 cycles with etoposide at 100 mg/m 2 over 2 hours and cytarabine 2 gm/m 2 twice daily for 2 or 3 days based on clinical judgement of the treating physician. Here we report outcomes with this approach in newly diagnosed PCNSL patients treated at our centre. Method: We did a retrospective analysis of patients with newly diagnosed PCNSL treated with MTR induction followed by modified EA consolidation. Kaplan-Meier was used to estimate progression free (PFS) and overall survival (OS). Variables influencing the outcomes with P less than 0.05 were considered significant. Results: Between January 2015 to December 2022, 72 patients were treated with above protocol with a median age of 46.5 years (range, 17 to 68 years) and 50 (69.4%) were males. Median ECOG performance score at presentation was 1 with 16 (22%) patients having ECOG performance score of 3 or more. IELSG scores were high and intermediate risk in 26 (36.1%) and 27 (37.5%) respectively while only 7 (9.7%) were in the low risk (data was not available for remaining 12 (17%) patients). MSKCC scores were available for all patients with 43 (59%) patients in class 1, 14 (19%) patients in class 2 and 15 (20%) patients in class 3. MTR induction with a dose of 8 gm/m2 could be delivered for all 4 cycles in 64 (88%) patients. Dose reduction was required in 14 (19.4%) patients; reasons were renal dysfunction in 8 (11%), infections in 3 (4%), liver dysfunction in 2 (3%). Nine patients (12.5%) had disease progression during induction. Total of 54 patients received consolidation with systemic therapy. Dose reduced EA consolidation was delivered in 46 (63%) patients while 8 patients received single agent high dose cytarabine consolidation and only 3 patients underwent ASCT. Seven patients received WBRT consolidation. EA consolidation was given for 2 days in 30 (65%) patients while 16 (35%) patients received 3 days of EA consolidation. Twenty six patients received maintenance lenalidomide. With a median follow up of 39.2 months (range, 2.1 to 91), median PFS is 46.9 months and median overall OS has not reached. Three years PFS & OS for all patients is 59.5% and 78% respectively ( Fig 1A &B). Median PFS and OS of patients receiving EA consolidation has not been reached with a 3-year PFS and OS of 77% and 79% respectively. MSKCC score was predictive of OS with 3 year OS in class 1, 2 and 3 of 89.5%, 68% and 58% respectively. Consolidation with split dose EA was associated with better OS and PFS as compared to single agent cytarabine alone. None of the 3 patients receiving ASCT have relapsed. Patients receiving full dose methotrexate had better OS compared to patients requiring dose reduction. Two versus 3 day EA consolidation had no impact on OS. MTR induction was well tolerated with grade 3 or more toxicity as follows: renal dysfunction in 8 (11%) patients, transaminitis in 4 and febrile neutropenia in 4 patients with no treatment related deaths. EA consolidation resulted in 18 episodes of febrile neutropenia with two deaths. Grade 3 or more hematologic toxicities were common with grade 3 or more anaemia, neutropenia and thrombocytopenia in 15 (32%), 40 (86%), 33 (71%) patients respectively. There were two deaths in the consolidation phase, one due to intracranial bleeding and another due to febrile neutropenia. Conclusion: MTR induction with modified EA consolidation is feasible and results in optimal outcomes in real world setting. Higher cumulative methotrexate dose and use of EA consolidation were associated with better OS. These outcomes compare favourably to CALGB 50202 study and represent a feasible option in resource limited setting.

Early Post-Infusion Clearance of High-Dose Methotrexate Predicts Delayed Methotrexate Elimination and Acute Kidney Injury

Introduction High-dose methotrexate (HDMTX) is used for the treatment of hematologic malignancies but its use may be limited by delayed methotrexate elimination and acute kidney injury (AKI). Current guidelines for glucarpidase recommend administration between 48-60 hours following the start of a 24-hour HDMTX infusion based on methotrexate concentration at 36-, 42-, or 48-hours [Ramsey 2016]. However, this may be too late to prevent permanent toxicities in cases where significant AKI develops early during infusion. Recent evidence from pig models have shown that early methotrexate measurements can predict the development of AKI [Buddington 2023]. The aim of this study is to determine whether early elimination of methotrexate (soon after completion of the infusion) is a useful “early clearance biomarker” of delayed methotrexate elimination and AKI in humans. Methods Retrospective data from two sites in Spain that collected two or more early measurements of methotrexate levels from the end of HDMTX infusion were analyzed from a larger HDMTX registry. Methotrexate half-life was calculated employing exponential decay equations and the slope of a line connecting two out of three early time points; each combination was tested, and the best performing pair is reported. Early levels were tagged according to visual exploration and k-means cluster analysis (Figure 1). Delayed methotrexate elimination was defined as a concentration above 1 μmol/L after 42-hours from the start of infusion (micromolar criterion) or above 2 standard deviations (SD) from the population mean modeled using MTXPK.org at 42- or 48-hours from the start of infusion (SD criterion). Receiver operating characteristic (ROC) curves were fitted, with their respective area under the curve (AUC) and confidence intervals for delayed methotrexate elimination and AKI. Additionally, a logistic regression model was employed to predict AKI using a base model with covariates (age, sex, HDMTX dose, and first course) plus early methotrexate clearance. Results Data from 144 patients who received 453 HDMTX courses were included. Patient characteristics were as follows: 41% females, age ranged from 1.2 to 81.5 years at the time of HDMTX infusion; diagnoses included acute lymphoblastic leukemia, 79%; non-Hodgkin lymphoma, 21%. Median methotrexate dose was 4,856.9 mg/m 2 [IQR 2,871.7 - 4,985.0], with 88% of the courses infused over 24 hours and 12% over 4 hours. Delayed methotrexate elimination was found in 41% and 6.6% of the courses according to the micromolar criterion and SD criterion, respectively. AKI developed in 29.6% of courses. Median time for the first, second and third measurements were 0.25 [IQR 0.00 - 0.71], 6.32 [IQR 5.86 - 6.69] and 12 [IQR 11.68 - 12.50] hours, respectively. Half-life estimation between the second and third early time-points predicted delayed methotrexate elimination by the micromolar criterion with an ROC AUC of 0.81 [CI95 0.76 - 0.87] or SD criterion with an ROC AUC of 0.84 [CI95 0.68 - 1.00] (Fig. 2) and any grade AKI with an ROC AUC of 0.62 [CI95 0.55 - 0.69]. After adjusting for age, sex, HDMTX dose, and first course, half-life remained statistically significant in the logistic regression model, with an odds ratio of 1.4 [CI95 1.1 - 1.8] (p < 0.01). The addition of half-life to the base model improved the overall performance (ANOVA, p < 0.01). Conclusions The half-life of methotrexate elimination soon after completing the HDMTX infusion defines an early clearance biomarker that predicts delayed methotrexate elimination and AKI. It requires only the measurement of 2 early methotrexate levels soon after completing the infusion, and thus represents a practical and applicable method to identify patients at risk who may need pre-emptive interventions like increased hydration or glucarpidase.

Screening Cerebrospinal Fluid for Aggressive B-Cell Lymphoma: Significance of Clonal Populations Found with 8-Color Flow Cytometry

Background: Secondary central nervous system (CNS) lymphoma confers a poor prognosis, and prompt identification is critical for guiding management. Over the last decade, we have employed cerebrospinal fluid (CSF) analysis with 8-color flow cytometry (FCM) in screening patients deemed at risk of CNS relapse. However, there is little data to guide interpretation and management of positive results, and concurrent clinical and radiographic features of FCM positive (FCM+) patients, remain areas of uncertainty. Methods: We analyzed newly diagnosed (ND) and relapsed/refractory (R/R) diffuse large B cell lymphoma (DLBCL) and high grade B cell lymphoma (HGBCL) patients who underwent a screening lumbar puncture (LP) from 2011-2020. Clinical, imaging, and CSF findings (differential, cytology, FCM) were collected. Patients with prior CNS involvement or FCM showing CLL/low-grade B-NHL were excluded. “FCM-only” patients were defined as FCM+ but without any other evidence of CNS spread (negative cell count/cytology, MRI, and no symptoms), whereas others were deemed to have overt CNS disease. Results: There were 415 LPs among 361 patients (243 ND, 172 R/R). 21 (5%) patients were FCM+, 10 ND and 11 R/R patients (Table 1). Four had FCM-only involvement (2 ND, 2 R/R). Among these, the clonal population was 1.63% of WBC (range, 0.3-2.7%), and 3 had concurrent bone marrow (BM) or peripheral blood (PB) involvement. All 4 received intrathecal (IT) therapy. At median follow up of 23 months, 3 (75%) have died (2 from CNS disease progression, 1 unknown due to loss in follow up), and 1 remains alive in remission 44 months later following IT methotrexate (MTX) and CAR T-cell therapy. 17 had overt CNS disease (8 ND, 9 R/R). The clonal population was 9% (range, 0.2-98%). Eight (47%) had concurrent BM or PB involvement. Twelve (70%) received IV high dose MTX, and the remainder received only IT therapy. At median follow up of 42 months, 9 (53%) have relapsed (2 CNS only, 1 systemic, 6 both), and 9 have died. Median time to CNS relapse was 5 months. Conclusions: CSF 8 color FCM screening of DLBCL/HGBCL patients yields a low proportion of positive results, mostly in the setting of overt CNS disease. However, even FCM-only patients had poor outcomes (2/4 with CNS progression), suggesting that blood contamination does not explain CSF findings, and that these patients should be approached as secondary CNS involvement. Better pt selection for CSF screening is needed to reduce unnecessary LPs. Intensified or novel approaches for all FCM+ patients are needed.

Low-Dose Methotrexate and Serious Adverse Events Among Older Adults With Chronic Kidney Disease

Low-dose methotrexate is used to treat rheumatoid arthritis and psoriasis. Due to its kidney elimination, better evidence is needed to inform its safety in adults with chronic kidney disease (CKD). To compare the 90-day risk of serious adverse events among adults with CKD who started low-dose methotrexate vs those who started hydroxychloroquine and to compare the risk of serious adverse events among adults with CKD starting 2 distinct doses of methotrexate vs those starting hydroxychloroquine. This retrospective, population-based, new-user cohort study was conducted in Ontario, Canada (2008-2021) using linked administrative health care data. Adults aged 66 years or older with CKD (defined as an estimated glomerular filtration rate [eGFR] <60 mL/min/1.73 m2 but not receiving dialysis) who started low-dose methotrexate (n = 2309) were matched 1:1 with those who started hydroxychloroquine. Low-dose methotrexate (5-35 mg/wk) vs hydroxychloroquine (200-400 mg/d). The primary outcome was a composite of serious adverse events: a hospital visit with myelosuppression, sepsis, pneumotoxic effects, or hepatotoxic effects within 90 days of starting the study drug. Prespecified subgroup analyses were conducted by eGFR category. Propensity score matching was used to balance comparison groups on indicators of baseline health. Risk ratios (RRs) were obtained using modified Poisson regression, and risk differences (RDs) using binomial regression. In a propensity score-matched cohort of 4618 adults with CKD (3192 [69%] women; median [IQR] age, 76 [71-82] years), the primary outcome was higher in patients who started low-dose methotrexate vs those who started hydroxychloroquine (82 of 2309 [3.55%] vs 40 of 2309 [1.73%]; RR, 2.05 (95% CI, 1.42-2.96); RD, 1.82% [95% CI, 0.91%-2.73%]). In subgroup analysis, the risks increased progressively at lower eGFR (eg, eGFR <45 mL/min/1.73 m2: RR, 2.79 [95% CI, 1.51-5.13]). In the secondary comparison with hydroxychloroquine, methotrexate users at 15 to 35 mg/wk had a higher risk of the primary outcome. In this cohort of 4618 older patients with CKD, the 90-day risk of serious adverse events was higher among those who started low-dose methotrexate than those who started hydroxychloroquine. If verified, these risks should be balanced against the benefits of low-dose methotrexate use.

La artritis reumatoide de inicio en el anciano recibe terapias menos agresivas que la artritis reumatoide de inicio en el adulto en una cohorte argentina

ObjectivesWhen rheumatoid arthritis (RA) starts after the age of 60 it is called elderly-onset rheumatoid arthritis (EORA) and when it starts earlier, young-onset rheumatoid arthritis (YORA). There are few Latin American studies that compared both groups. The objective of the study was to evaluate differences in the clinical characteristics, evolution and treatment among patients with RA with onset before or after 60years of age. Materials and methodsObservational study of patients with RA attended consecutively in four centers in Argentina. Sociodemographic data, comorbidities, clinical manifestations at diagnosis, presence of rheumatoid factor and/or anti-CCP (cyclic citrullinated peptide) and treatments received were collected. At the last visit, swollen and tender joints, assessment of disease activity by the patient and physician, the presence of radiographic erosions, and functional status using the HAQ-DI were recorded. ResultsFifty-one patients from each group were analyzed. The EORA group had a significantly higher proportion of smokers (58.8% vs. 35.3%, P=.029), cardiovascular history (54.9% vs. 21.6%, P=.001), abrupt onset (49% vs. 29.4%, P=.034) or with symptoms similar to PMR (19.6% vs. 0%, P=.001). Lower methotrexate doses were used in the EORA group: 19mg (15-25) vs. 21.9mg (20-25) (P=.0036) and more frequently did not receive bDMARDs or tsDMARDs. Discussion and conclusionsThe benefits of intensive treatment in patients with RA have been described. In this study, the use of DMARDs in the EORA group was less intensive, suggesting that advanced age constitutes a barrier in the therapeutic choice.

Pilot study of treatment of patients with deep infiltrative endometriosis with methotrexate carried in lipid nanoparticles.

Previously, lipid nanoparticles (LDE) injected in women with endometriosis were shown to concentrate in the lesions. Here, the safety and feasibility of LDE carrying methotrexate (MTX) to treat deep infiltrating endometriosis was tested. Prospective pilot study. Perola Byington Hospital Reference for Women's Health. Eleven volunteers (aged 30-47years, BMI 26.15 ± 6.50kg/m2) with endometriosis with visual analog scale pelvic pain scores (VAS) > 7 and rectosigmoid lesions were enrolled in the study. Three patients were treated with LDE-MTX at single intravenous 25mg/m2 dose of MTX and eight patients with two 25mg/m2 doses with 1-week interval. Clinical complaints, blood count, and biochemistry were analyzed before treatment and on days 90, 120, and 180 after LDE-MTX administration. Endometriotic lesions were evaluated by pelvic and transvaginal ultrasound (TVUS) before treatment and on days 30 and 180 after LDE-MTX administration. No clinical complaints related with LDE-MTX treatment were reported by the patients, and no hematologic, renal, or hepatic toxicities were observed in the laboratorial exams. FSH, LH, TSH, free T4, anti-Müllerian hormone, and prolactin levels were also within normal ranges during the observation period. Scores for deep dyspareunia (p < 0.001), chronic pelvic pain (p = 0.008), and dyschezia (p = 0.025) were improved over the 180-day observation period. There was a non-significant trend for reduction of VAS scores for dysmenorrhea. Bowel lesions by TVUS were unchanged. No clear differences between the two dose levels in therapeutic responses were observed. Results support the safety and feasibility of using LDE-MTX in women with deep infiltrating endometriosis as a novel and promising therapy for the disease. More prolonged treatment schemes should be tested in future placebo-controlled studies aiming to establish the usefulness of this novel nanomedicine approach.

Tooth Abnormalities and Their Age-Dependent Occurrence in Leukemia Survivors.

The multidrug nature of anticancer treatment and different treatment protocols used in the studies are likely to be a major limitation in establishing real risk factors determining the occurrence of dental abnormalities. The authors aimed to establish a relationship between the duration and the dose of chemotherapy and the number of tooth adverse effects in the group receiving the same treatment. Of the 40 anticancer therapy recipients who attended the outpatient dental clinic, 7 leukemia survivors receiving the treatment according to the ALL IC-BFM 2002 protocol were selected. The study group consisted of four females and three males aged 92 to 207 months at the time of dental examination and 29 to 91 months at leukemia diagnosis. As a result of the clinical and radiological examination, dental abnormalities such as agenesis, tooth size reduction, root abnormalities, and taurodontia were identified, and the medical records of all survivors were reviewed in terms of drugs administered, their doses, and treatment schedules. No correlation was observed between the treatment duration of an intensive therapy, the entire therapy, and the number of tooth abnormalities. No relationship was also found between the number of dental abnormalities and the cumulative dose of vincristine, L-asparaginase, methotrexate, cyclophosphamide, cytarabine, and 6-mercaptopurine. The age at the onset of antineoplastic therapy is likely to be the strongest risk factor for toxic injury during tooth development.

Consolidation with Ponatinib Plus Sequential Blinatumomab and Chemotherapy after Low Intensity Dasatinib-Based Induction in Adults with Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia: Outcomes from a Single Institution

Introduction: Dasatinib plus prednisone is a standard induction approach for adults with Philadelphia Chromosome positive (Ph+) ALL known to result in high remission rates with minimal toxicity, however optimal post-remission therapy has not been determined. Given the excellent outcomes observed in recent clinical studies using ponatinib in adult Ph+ ALL, we explored a strategy of switching the TKI to ponatinib and consolidating with sequential blinatumomab (blina) and chemotherapy after dasatinib + prednisone induction. In contrast to recent studies of blina + TKI wherein patients have received up to 3-5 cycles of blina, we limited the number of blina cycles to 2 to minimize cost and logistical burden. We also administered systemic chemotherapy after blina with the goal of decreasing risk of extramedullary relapse, which has been observed in studies of blina-based “chemotherapy-free” regimens. Methods: Since June 2021, all adult patients with Ph+ ALL (n=14) at our institution were switched to ponatinib (30mg/d then 15mg/d once complete molecular remission [CMR] achieved) after dasatinib + prednisone induction. Concurrent with TKI, patients could also receive blina for one or two cycles (at discretion of treating clinician), followed by up to four cycles of consolidative chemotherapy alternating between high-dose methotrexate (1.5g/m 2) and intermediate-dose cytarabine (1g/m 2 x 6 doses). Patients not proceeding to stem cell transplant continue ponatinib monotherapy for up to 5 years, and 12 doses of intrathecal methotrexate 15mg are given throughout the total course of treatment. Prior to June 2021, our institutional approach was to recommend allogeneic SCT in CR1 for all transplant candidates immediately after dasatinib + prednisone induction. We reviewed charts of patients with newly diagnosed Ph+ ALL at University of Colorado Hospital from 2014-2023 to compare outcomes of patients treated before 06/2021 (“dasatinib/SCT” cohort) and after 06/2021 (“ponatinib” cohort). Results: From 6/2021-02/2023, 14 patients (median age 54) with newly diagnosed Ph+ ALL were switched to ponatinib after dasatinib + prednisone induction. After induction, 12 patients received blina (median 1 cycle; range 1-2) and consolidation chemotherapy (median 4 cycles; range 3-4). Blina could not be obtained for 2 patients who instead received chemotherapy only for 6 cycles. 3 patients (21%) underwent allogeneic SCT in CR1 due to persistent detectable bcr/abl transcripts by qPCR, and all 3 continued ponatinib as maintenance after SCT. With a median follow-up of 15 months (range: 4-24 months) no relapses have occurred and all patients remain alive at last follow-up. No patients have discontinued ponatinib due to intolerance. Chemotherapy plus ponatinib was well-tolerated after blina, with all except for one patient having received all intended cycles (one patient stopped chemotherapy after 3 cycles due to excessive hematologic toxicity). Between 11/2014-03/2021, 36 patients (median age 50) with Ph+ ALL were treated with dasatinib + prednisone induction, after which 31 (86%) underwent allogeneic SCT in CR1. All non-transplant candidates continued TKI alone. With a median follow-up of 37 months (range: 4-102 months), 11 relapses have occurred, 9 of which occurred after SCT. Among 13 patients who died during follow-up, 7 deaths were due to relapsed/refractory ALL and 6 were due to treatment-related mortality. Comparing outcomes between the two cohorts, relapse-free survival is improved in the more recent (ponatinib) cohort compared to the earlier (dasatinib/SCT) cohort (p=0.044, log-rank test) (Figure 1) despite a lower rate of allogeneic SCT in CR1 (21% vs 86%). No difference in overall survival (p=0.12) has been observed (not shown). Conclusions: For adults with Ph+ ALL, switching to ponatinib after dasatinib-based induction and consolidating with blina and chemotherapy is a safe and potentially effective strategy. The optimal number of blina cycles given post-remission is unclear, though we observed favorable outcomes in patients receiving a median of only 1 cycle, and maximum of 2 cycles. Favorable outcomes despite the low rate of allogeneic SCT (21%) seen in our cohort suggest that SCT may be safely avoided for some patients who receive ponatinib plus blina and chemotherapy-based consolidation.

Association between High Antimetabolite Dose Intensity during Maintenance, Toxicities and Relapse Risk in Children with Acute Lymphoblastic Leukemia (ALL): COG-AALL03N1 Report

Introduction: Current COG protocols for childhood ALL recommend increasing 6-mercaptopurine (6MP) and methotrexate (MTX) dose intensity (DI=prescribed therapy dose/ planned protocol dose) to &amp;gt;100% during maintenance for patients exhibiting inadequate myelosuppression (surrogate for systemic exposure to 6MP/MTX). Whether increasing DI &amp;gt;100% (high DI) to achieve myelosuppression is associated with increased toxicity is not clear. Additionally, whether high DI influences relapse risk, and the direction of that association, remains unknown. Nonadherence to 6MP results in inadequate myelosuppression and prompts clinicians to increase the prescribed dose. Nonadherence to 6MP is also associated with relapse (Bhatia, JAMA Onc 2015). Prior literature examining the association between high DI and relapse risk did not consider adherence to chemotherapy. Here, we examined whether high 6MP/MTX DI affects toxicity or relapse, taking 6MP adherence into consideration. Methods: We used data collected by COG-AALL03N1 (primary aim: 6MP adherence and risk of relapse) to examine the association between high DI and (i) hematologic toxicity, and (ii) relapse risk. Inclusion criteria: (i) age &amp;lt;21y at ALL diagnosis, (ii) receiving maintenance therapy in first remission and &amp;gt;6mo since starting maintenance, (iii) wild-type thiopurine methyltransferase ( TPMT) or nucleoside diphosphate-linked motif 15 (NUDT15) and (iv) objectively measured 6MP adherence for 6mo per patient (using MEMS-TrackCaps). 6MPDI and MTXDI were calculated monthly for 6mo. Updates on vital status, relapse or second neoplasm were collected every 6mo-1y from participating sites for up to 10y. Exposure: ‘high DI’ phenotype: (i) 6MPDI or MTXDI ≥110% at any time during the first 4 study months, or (ii) 6MPDI or MTXDI 100% to 110% at study enrollment and ≥25% increase over the first 4 study months. All other patients were categorized as having ‘normal DI’. Outcomes: (i) hematologic toxicity (absolute neutrophil count &amp;lt;500 cells/µL, platelet count &amp;lt;50,000 cells/mm 3, or treatment interruption due to low ANC/platelets) during the subsequent 2 study months (months 5 and 6); and (ii) ALL relapse any time after month 4. We used logistic regression to examine the association between high DI and hematologic toxicity, adjusting for age at study entry, sex, race/ethnicity. The association between DI and relapse was estimated using proportional sub-distribution hazard regression models, adjusting for age at study entry, blast cytogenetics, time from start of maintenance, and treatment interruptions. Analyses were stratified by 6MP adherence (adherers: median adherence ≥85%; non-adherers: median adherence &amp;lt;85%). Adherence was included in both models to account for residual confounding. Results: Characteristics of the 410 eligible participants are summarized in the Table; the cohort was followed for a median 7.9y (range, 0.2-13). There were no significant differences between patients with normal (71.5%) vs high (28.5%) DI. Patients with high DI had lower median 6MP adherence (94.6% vs 97.0%, P=0.02). Hematologic DLT: As shown in the Figure, the proportion of patients with hematologic toxicities was significantly higher among adherers with high DI compared with adherers with normal DI (38.1% vs 18.5%, P=0.0003). In contrast, the proportion of patients with hematologic toxicities was not significantly different by DI status among non-adherers (18.5% vs 10.5%, P=0.4). Multivariable analysis revealed that high DI was associated with 2.9-fold greater odds of hematologic toxicity among adherers (95%CI=1.6-5.1, P&amp;lt;0.001) and a non-significant 2.1-fold greater odds of hematologic toxicity among non-adherers (95%CI=0.4-10.1, P=0.4). Relapse risk: The adjusted hazard of relapse among adherent patients exposed to high DI was 2.2-fold greater (95%CI=1.0-5.1, P=0.06) than adherent patients exposed to normal DI after adjusting for covariates. In contrast, the hazard of relapse among non-adherent patients exposed to high DI (vs. normal DI) was not elevated (HR=0.9, 95%CI=0.2-3.8, P=0.8). Conclusions: High antimetabolite dose intensity in children with ALL who adhere to oral 6MP during maintenance therapy is associated with higher odds of hematologic toxicities as well as increased hazard of relapse. These findings inform clinicians about the issues related to dose escalation beyond protocol doses, especially among adherent patients.

Clinical Features and Therapeutic Evaluation of Central Nervous System Involvement in Pediatric Anaplastic Large Cell Lymphoma

Objective：To analyze the clinical features of central nervous system involvement (CNS3) in pediatric anaplastic large cell lymphoma (ALCL) and evaluate the efficacy of CNCL-ALCL-2017 protocol for the treatment of CNS3. Methods：The clinical data of 215 pediatric ALCL patients ≤18 years old enrolled in the Chinese Children's Cancer Group (CNCL) between April 2017 and March 2023 were collected sequentially. All enrolled patients staging and CNS layering were according to the IPNHLSS staging system and treated with CNCL-ALCL-2017 protocol (modified BFM-ALCL99 protocol). The clinical features at diagnosis and treatment outcomes of CNS3 patients were analyzed. All patients were followed up until June 30, 2023. Data were collected using a unified information platform. Associations between patient characteristics were analyzed with Chi-squared or Fisher's exact tests. Eventfree survival (EFS) and overall survival (OS) curves were estimated according to the KaplanMeier method and compared by Logrank test. Furthermore, statistical analysis was performed using SPSS 22.0 software. P&amp;lt;0.05 was considered statistically significant. Results: Among the 215 ALCL pediatric patients, 17 (7.9%) had CNS3, including 13 males and 4 females with median age of 8.0 (range 1.0-14.0) years. In this patients, brain parenchyma and spinal cord involvement were found in 58.8% (10/17) patients on imaging, with space-occupying lesions in 7 cases and abnormal signals on MRI in 4 cases. Concurrent space-occupying lesions and abnormal signals on MRI was detected in 1 case. Cerebrospinal fluid (CSF) positivity was detected in 35.3% (6/17) patients, including 2 cases with positive ALK gene and 5 cases with positive flow cytometry, with 1 case positive for both. Cranial nerve deficits such as facial nerve palsy, visual or hearing impairment were observed in 23.5% (4/17) patients. Concurrent CSF, imaging and cranial nerve abnormalities were present in 5.9% (1/17), while two of this were present in 29.4% (5/17). Isolated CSF positivity without other abnormalities was detected in 23.5% (4/17). Pathological subtypes included common type (76.5%, 13/17), histiocyte-rich variant (5.9%, 1/17), small cell variant (5.9%, 1/17), others (11.8%, 2/17). Immunohistochemistry showed ALK + in 94.1% (16/17) and CD3 + in 76.5% (13/17). Concurrent ALK gene positivity in both peripheral blood and bone marrow was detected in 64.7% (11/17), with 58.8% (10/17) positive in both samples. The median follow-up of all patients was 35.4 months (range 0.5-74.9). All CNS3 patients had stage IV disease and were treated with regimen D (with 5 g/m 2 methotrexate). The 3-year OS was 94.3%, 98.2% and 93.3% for CNS1 (144, 67.0%), CNS2 (54, 25.1%) and CNS3 (17, 7.9%) patients, respectively; the 3-year EFS was 85.6%, 90.3% and 86.7%, respectively. No significant differences were found among the three groups. The 3-year OS and EFS for the whole cohort were 95.1% and 84.7%, respectively, also with no significant differences from the CNS3 group. Of the CNS3 patients, 1 was lost to follow-up after the first course, 1 died during treatment, and the interim CR rate was 75.0% (12/16). CSF conversion was achieved in 3/6 (50%) CSF-positive patients. 6.25% (1/16) patient had CSF ALK gene re-positivity during maintenance and persistent positivity despite additional ALK inhibitor alectinib and intrathecal therapy. In the CNS1 group, 2 cases were lost to follow-up and 4 cases died. The relapse rate was 11.4% (16/140), and the rate of central nervous system relapse was 2.1% (3/140). Additionally, the rate of CNS1 relapse was slightly higher but not statistically significantly different for CNS2. Conclusions: CNS involvement is relatively rare in pediatric ALCL. Detection rate of CNS3 can be improved by CSF flow cytometry and gene screening. The prognosis of CNS3 was significantly improved with CNCL-ALCL-2017 protocol, which may be related to the increased dose of methotrexate in the protocol. The short follow-up time and the small number of cases in this group may affect the results. Furthermore, CNS relapse rate of CNS2 was not markedly higher compared to CNS1. Keywords: Anaplastic large cell lymphoma; Pediatric; Central nervous system; Clinical features; Prognosis

Severe Methotrexate Toxicity Following a Capizzi Cycle in an Obese Adolescent With Acute Lymphoblastic Leukemia and Hepatic Steatosis.

Methotrexate is a major component of pediatric leukemia treatment. While toxicities are common after high-dose methotrexate, escalating dose methotrexate (Capizzi methotrexate) is typically well-tolerated. We report an adolescent Hispanic female with pre-B acute lymphoblastic leukemia, preexisting obesity and hepatic steatosis who developed severe multiorgan failure following an escalating dose of methotrexate with delayed methotrexate excretion of 11 days. We identified one similar report in an obese adult; however, this case is the first to our knowledge involving a pediatric patient. With the rising incidence of obesity and associated comorbidities among children and adolescents with leukemia, attention to potential risks for this population is warranted.

Ruxolitinib Promotes Clinical Responses in Large Granular Lymphocytic Leukemia Via Suppression of JAK/STAT-Dependent Inflammatory Cascades

Background: Large granular lymphocytic leukemias (LGL) are T or NK-cell lymphoproliferative disorders that are characterized by myelosuppression and frequent mutations in STAT3. Commonly used agents include low doses of methotrexate or cyclophosphamide. Among 5 patients (pts) with LGL treated on our phase II study of ruxolitinib in T-cell lymphoma (Moskowitz, et al. Blood 2021), 2 achieved partial response (PR) and 2 achieved stable disease, suggesting oncogenic dependence on JAK/STAT pathway activity in LGL. Methods: To further explore the efficacy and mechanism of JAK1/2 blockade in LGL, we expanded our phase II study to include 23 pts with relapsed/refractory LGL. This sample size was chosen to allow for narrowing of the 95% confidence interval to 54-87% (Wilson score method) if similar efficacy was observed. Pts received ruxolitinib 20mg PO BID continuously. Responses were assessed after cycles 2, 5, and every 3 cycles thereafter using the response assessment from the ECOG-5998 study for LGL. Furthermore, stable disease (SD) was defined as improvement in cytopenia(s) by 1 grade (without meeting criteria for PR) for a minimum of 6 months. Overall response rate (ORR) included complete response (CR) plus PR; clinical benefit rate (CBR) included ORR plus SD. In parallel, we performed single-cell RNA sequencing (scRNA-seq) on paired C1D1 and C1D8 peripheral blood mononuclear cells (PBMCs) from 7 patients (5 PR, 1 SD, and 1 non-responder) and plasma proteomic profiling on 5 patients (3 PR, 1 SD, and 1 POD). Results: Among 23 pts, median age was 58 (31-86), 13 (57%) were male, 19 (83%) were white, 1 (4%) was black. Median number of prior therapies was 2 (range 0-5). 12/22 (54%) had STAT3 mutations (of whom 1 also had a JAK1 mutation); 1 had a JAK2 mutation, 1 pt was missing sequencing. Baseline cytopenias included anemia (n=14, 61%), thrombocytopenia (n=12, 52%), and/or neutropenia (n=16, 70%). Among 20 pts evaluable for response (3 too early), there were 5 (25%) CRs, 6 (30%) PRs, and 4 (20%) SD. ORR and CBR were 55% and 75% respectively. Median event free survival (EFS) was not reached and was 68% (95% CI: 50-93) at 14 months ( Fig. 1A). STAT3 mutation status predicted for improved EFS (14-month EFS 100% vs 40%, p=0.007). After a median duration of treatment of 13.1 months, there were no unexpected toxicities. Severe adverse events included febrile neutropenia (n=3, 13%) and herpes zoster (n=1, 4%). To understand the mechanism by which ruxolitinib improved hematologic parameters in LGL, we performed unbiased proteomic analysis of peripheral blood plasma samples using the OLink 96 Inflammation and Immune Response Platforms. Ruxolitinib led to a decrease in several soluble mediators known to be driven by interferon-driven JAK/STAT activation, including CXCL9, CXCL10, and PD-L1, as well as TNF ( Fig. 1B). Several of these factors have been shown to suppress hematopoietic stem cell function, suggesting a JAK/STAT-dependent paracrine mechanism by which LGL leads to myelosuppression. ScRNA-seq of PBMCs identified LGL cells based on expression of cytotoxic genes, including NKG7, CST7, CCL5, GZMB and PRF1. Analysis of cell type proportions within the peripheral blood showed that patients achieving a PR to Ruxolitinib showed a decrease in the proportion of circulating myeloid cells. Gene set enrichment analysis (GSEA) of LGL cells revealed substantial downregulation of both JAK/STAT signaling and TNF signaling in LGL cells in PR but not SD patients, whereas persistently elevated TNF signaling was seen not only in LGL cells but also circulating myeloid and naïve T-cells in SD patients, suggesting that LGL cells might promote bone marrow suppression via paracrine effects on the bystander host immune system. Network analysis of LGL cells and non-malignant peripheral blood immune cells is ongoing. Conclusions: Ruxolitinib induced durable responses in pts with LGL and therefore may represent an option for pts requiring treatment. STAT3 mutation status was predictive of EFS and responses were associated with JAK/STAT pathway down-regulation, demonstrating on-target effects. Integrated immune profiling suggests that ruxolitinib may interrupt JAK/STAT-dependent paracrine inflammatory signals that promote bone marrow suppression. An ongoing phase II study of ruxolitinib in LGL will aid in clarifying optimal dosing and confirming both efficacy and mechanism of action (NCT05592015).

Abstract 8 — Risk of Liver Injury in Psoriasis and Rheumatoid Arthritis is Associated with Metabolic Risk Factors Rather than Methotrexate and Leflunomide

Objectives To assess the associations of hepatic injury and fibrosis in patients with psoriasis (PsO) and rheumatoid arthritis (RA). Methods A total of 318 patients attending dermatology and/or rheumatology out-patient clinics in Queen Mary Hospital, with clinical diagnosis of PsO, psoriatic arthritis (PsA) or RA were consecutively recruited from July 2020 to August 2022. Demographic data, comorbidities, and medications were recorded. Body mass index (BMI) was measured. Serum biomarkers for hepatic injury including autotaxin, matrix metalloproteinase (MMP) 3, MMP 8, and MMP 9 were determined. Transient elastography (TE) was performed to determine liver stiffness measure (LSM) and controlled attenuation parameter (CAP). Regression models were built to determine the association between methotrexate and leflunomide cumulative dosage, serum biomarkers and TE scores. Results A total of 67 (20.4%) patients had LSM&gt;7.1 kPa. Multivariate linear regression showed methotrexate cumulative dosage was associated with autotaxin level (B=4.77, 95% CI 1.39; 8.15, p=0.001). There was no association between methotrexate cumulative dosage and MMP 3, MMP 8, MMP 9, LSM, CAP. There was also no association between leflunomide cumulative dosage and autotaxin, MMP 3, MMP 8, MMP 9, LSM, CAP. BMI (B=6.18, 95% CI 4.95; 7.41, p&lt;0.001), hypertension (B=18.98, 95% CI 4.15; 31.82, p=0.01), hyperlipidaemia (B=20.21, 95% CI 3.50; 36.92, p=0.02) were associated with CAP. Conclusion Methotrexate and leflunomide have minimal hepatic risk in patient with PsO and RA. Metabolic risk factors had strong associations with hepatic injury.

Medical management/Non-surgical of ectopic pregnancy: Case report and review of literature

Ectopic pregnancy is a cause of maternal morbidity and mortality. Management options are surgical, medical and expectant management. Medical management also known as the non-surgical approach is not widely practiced in our environment despite its several advantages over the traditional open salpingectomy including its less invasiveness, reduced cost and absence of anaesthetic risk, surgical risk and need for hospital admission. It however requires early presentation and diagnosis before tubal rupture, a well motivated patient and fulfilment of strict criteria in well selected patients. Aim. To present this rare case report of the medical/non-surgical management of unruptured ectopic pregnancy and a review of the literature. Case report: Mrs AW, Para 3+0 (2 alive) who presented with lower abdominal pains and abnormal scanty vaginal bleeding following 6 weeks of amenorrhoea. Her physical examination revealed a stable cardiovascular state. There were no significant abdominal findings. Vaginal examination revealed positive cervical motion tenderness. A diagnosis unruptured ectopic pregnancy was made. Pelvic ultrasound scan revealed a cystic left adnexial mass measuring 1.94 cm by 1.7 cm with thickened, well circumscribed margins and it contained an echogenic mass (fetal pole) with cardiac pulsation (145/min). The crown-rump length was 0.76cm which corresponded to 6 weeks plus 5 days of gestation. Her serum β hCG was 3625.8miu/ml. She expressed her aversion for a surgical intervention having had 3 previous surgeries. She gave an informed consent for medical/non-surgical management with intramuscular Methotrexate. She subsequently received 3 doses of weekly 100mg intramuscular methotrexate. Each dose of Methotrexate was preceded by re-evaluation of liver/renal function test, full blood count and serum β hCG. By the 4th week of treatment the success of her treatment was confirmed by the insignificant level of β hCG and findings at TVS. She was counselled on the need for prompt localization of any subsequent pregnancy. Conclusion: We presented Mrs AW who was successfully managed for unruptured ectopic pregnancy using the medical/non-surgical approach on out-patient basis. Proper patient selection and follow-up is required for a favourable outcome,

Decoding clinical and molecular pathways of liver dysfunction in Psoriatic Arthritis: Impact of cumulative methotrexate doses

BackgroundThe occurrence of liver abnormalities in Psoriatic Arthritis (PsA) has gained significant recognition. Identifying key factors at the clinical and molecular level can help to detect high-risk patients for non-alcoholic fatty liver disease in PsA. Objectivesto investigate the influence of PsA and cumulative doses of methotrexate on liver function through comprehensive in vivo and in vitro investigations. MethodsA cross-sectional study involving 387 subjects was conducted, 200 patients with PsA, 87 NAFLD-non-PsA patients, and 100 healthy donors (HDs), age and sex-matched. Additionally, a retrospective longitudinal study was carried out, including 83 PsA patients since initiation with methotrexate. Detailed clinical, and laboratory parameters along with liver disease risk were analyzed. In vitro, experiments with hepatocyte cell line (HEPG2) were conducted. ResultsPsA patients present increased liver disease risk associated with the presence of cardiometabolic comorbidities, inflammatory markers, onychopathy, and psoriasis. The treatment with PsA serum on hepatocytes encompassed inflammatory, fibrotic, cell stress, and apoptotic processes. At the molecular level, methotrexate impacts liver biology, although the cumulative doses did not affect those alterations, causing any potential damage to liver function at the clinical level. Finally, anti-PDE-4 or anti-JAK decreased the inflammatory profile induced by PsA serum on hepatocytes. Conclusion1)This study identifies the complex link between liver disease risk, comorbidities, and disease-specific features in PsA patients. 2)Methotrexate dose in PsA patients had no significant effect on liver parameters, confirmed by hepatocyte in vitro studies. 3)Anti-PDE-4 and anti-JAK therapies show promise in reducing PsA serum-induced hepatocyte activation, potentially aiding liver complication management.

Identification of the methotrexate resistance-related diagnostic markers in osteosarcoma via adaptive total variation netNMF and multi-omics datasets.

Osteosarcoma is one of the most common malignant bone tumors with high chemoresistance and poor prognosis, exhibiting abnormal gene regulation and epigenetic events. Methotrexate (MTX) is often used as a primary agent in neoadjuvant chemotherapy for osteosarcoma; However, the high dosage of methotrexate and strong drug resistance limit its therapeutic efficacy and application prospects. Studies have shown that abnormal expression and dysfunction of some coding or non-coding RNAs (e.g., DNA methylation and microRNA) affect key features of osteosarcoma progression, such as proliferation, migration, invasion, and drug resistance. Comprehensive multi-omics analysis is critical to understand its chemoresistant and pathogenic mechanisms. Currently, the network analysis-based non-negative matrix factorization (netNMF) method is widely used for multi-omics data fusion analysis. However, the effects of data noise and inflexible settings of regularization parameters affect its performance, while integrating and processing different types of genetic data is also a challenge. In this study, we introduced a novel adaptive total variation netNMF (ATV-netNMF) method to identify feature modules and characteristic genes by integrating methylation and gene expression data, which can adaptively choose an anisotropic smoothing scheme to denoise or preserve feature details based on the gradient information of the data by introducing an adaptive total variation constraint in netNMF. By comparing with other similar methods, the results showed that the proposed method could extract multi-omics fusion features more effectively. Furthermore, by combining the mRNA and miRNA data of methotrexate (MTX) resistance with the extracted feature genes, four genes, Carboxypeptidase E (CPE), LIM, SH3 protein 1 (LASP1), Pyruvate Dehydrogenase Kinase 1 (PDK1) and Serine beta-lactamase-like protein (LACTB) were finally identified. The results showed that the gene signature could reliably predict the prognostic status and immune status of osteosarcoma patients.

Access to Methotrexate Monitoring in Latin America: A Multicountry Survey of Supportive Care Capacity

High-dose methotrexate (HDMTX) is used to treat a broad spectrum of cancers. Methotrexate (MTX) monitoring and adequate supportive care are critical for safe drug administration; however, MTX level timing is not always possible in low- and middle-income countries. The aim of this study was to evaluate HDMTX supportive care capacity and MTX monitoring practices in Latin America (LATAM) to identify gaps and opportunities for improvement. A multicenter survey was conducted among LATAM pediatric oncologists. Twenty healthcare providers from 20 institutions answered the online questionnaire. HDMTX was used to treat acute lymphoblastic leukemia (ALL; 100%), non-Hodgkin lymphoma (84.2%), diffuse large B-cell lymphoma (47.4%), osteosarcoma (78.9%), and medulloblastoma (31.6%). Delays in starting HDMTX infusion were related to bed shortages (47.4%) and MTX shortages (21.1%). MTX monitoring was performed at an in-hospital laboratory in 52%, at an external/nearby laboratory in 31.6%, and was not available in 10.5%. Median interval between sampling and obtaining MTX levels was ≤ 2 h in 45% and ≥ 6 h in 30%, related to laboratory location. Sites without access to MTX monitoring reduced the MTX dose for patients with high-risk ALL or did not include MTX in the treatment of patients with osteosarcoma. Respondents reported that implementation of point-of-care testing of MTX levels is feasible. In LATAM, highly variable supportive care capacity may affect the safe administration of MTX doses. Improving accessibility of MTX monitoring and the speed of obtaining results should be prioritized to allow delivery of full doses of MTX required by the current protocols.

Japan College ofRheumatology guidance forthe use ofmethotrexate inpatients with rheumatoid arthritis: Secondary publication.

Methotrexate (MTX), the anchor drug in the current treatment strategy for rheumatoid arthritis (RA), was first approved for the treatment of RA in Japan in 1999 at a recommended dose of 6-8 mg/week. The approved maximum dose of MTX has been 16 mg/week since February 2011 when MTX was approved as a first-line drug in the treatment of RA. Recent evidence of MTX-polyglutamate concentration in the red blood cells of Japanese patients with RA justifies the current daily use of MTX in Japan. Additionally, after a nationwide clinical trial, a subcutaneous MTX injection formula (7.5-15 mg/week) was approved for RA treatment in September 2022. Therefore, in March 2023, a subcommittee of the Japan College of Rheumatology updated the guidance (formerly 'guidelines') for the use of MTX in Japanese patients with RA. This article, an abridged English translation summarizing the 2023 update of the Japan College of Rheumatology guidance for the use of MTX and management of patients with RA, will be helpful to both Japanese and global rheumatology communities.

Prediction of Tubal Rupture in Ectopic Pregnancy Using Methotrexate Treatment Protocols and Hematological Markers.

Ectopic pregnancy is a pregnancy complication in which the embryo implants outside the uterine cavity. Although medical treatment is chosen first, sometimes a rupture may occur, and surgical treatment may be required. The parameters to predict rupture have been the subject of many studies. This study aimed to compare the efficacy of different methotrexate protocols in the treatment of ectopic pregnancy and determine the parameters and methotrexate treatment protocols that can predict the risk of rupture. A total of 128 patients diagnosed with ectopic pregnancy were included in this study. Patients were separated into three categories based on their treatment protocols. Regarding the occurrence of rupture, all three groups were compared. The hematological parameters and methotrexate treatment protocols were analyzed and compared between groups. The mean age was 31.9 years. Parity was significantly higher in patients who received multiple doses of methotrexate compared to the other groups. There were significant variations observed among the groups regarding parity, initial β-hCG values, hematocrit (HTC), and mean corpuscular volume (MCV) (p = 0.048, p < 0.001, p = 0.019, and p = 0.047, respectively). According to receiver operating characteristic analysis, neutrophil-to-lymphocyte ratio (NLR) levels were significantly associated with histopathologically confirmed tubal rupture (p < 0.05). NLR levels should be examined in ectopic pregnancy, and the possibility of rupture should be considered in cases with high NLR levels. The potential of NLR to predict ectopic pregnancy rupture should be explored in multicenter prospective studies.

Vitamin D: An effective therapy against methotrexateinduced cardiotoxicity

Purpose: To determine the potential cardioprotective effect of vitamin D (VD) against methotrexate (MTX) induced cardiotoxicity.Methods: A total of thirty-five (35) rats were randomly assigned to five equal groups. Control groupreceived no treatment; MTX group received intraperitoneal (IP) injection of MTX in a single dose of 20 mg/kg on day 8; VD group received 200 IU/kg VD daily dissolved in sunflower oil orally; sunflower oil (SO) group received 1 mL/kg/day SO orally; MTX + VD group received a single dose of MTX (20 mg/kg, IP) on day 8 and VD (200 IU/kg, orally) for 21 days. Myocardial tissue samples were harvested and used for clinical chemistry, histopathological, and ultrastructural evaluation.Results: Histopathological damage in MTX group was more severe than in control group under both light and electron microscopy. Expression of transient receptor potential melastatin 2 (TRPM2) and caspase-3 markers was significantly higher in MTX group (p &lt; 0.05). Glutathione peroxidase (GSH-Px) enzyme activity in cardiac tissue was lower in MTX group, whereas malondialdehyde (MDA) levels increased significantly (p &lt; 0.05). In MTX + VD group, VD treatment alleviated histopathological damage and significantly lowered TRPM2 and caspase-3 expressions (p &lt; 0.05). Vitamin D also reduced tissue MDA levels, and increased GSH-Px activity albeit non-significantly (p &gt; 0.05),Conclusion: These findings suggest that VD exerts an ameliorative effect on MTX-induced cardiotoxicity in rats. Therefore, TRPM2 channel affords a novel therapeutic approach for treatment of diseases related to chemotherapy-induced oxidative stress.