Methotrexate Toxicity Research Articles

Effects of SCT genetic polymorphisms on methotrexate concentrations and toxicities in Chinese children with acute lymphoblastic leukemia

Solute carrier (SLC) transporters play a crucial role in facilitating the cellular uptake of various anticancer drugs, such as methotrexate (MTX). This study aimed to analyze the impact of nonsynonymous single nucleotide polymorphisms (SNPs) in SLC19A1, SLCO1B1, and SLCO1B3 on MTX exposure, toxicities, and prognosis in 148 patients with acute lymphoblastic leukemia (ALL). The SLCO1B3 rs7311358 polymorphism was significantly associated with the median dose-normalized MTX concentrations at 24 h (p < .05). There were significant differences in the proportions of patients with serum MTX levels >40 µmol/L at 24 h among SLC19A1 rs1051266 GG, GA, and AA genotype carriers (29.0, 24.7, and 6.2%, respectively, p < .05). The SLC19A1 rs1051266 G > A polymorphism also displayed significant associations with hematological (p < .05) and hepatic toxicities (p < .01). Our findings indicate that the analysis of SNPs in solute carrier transporters (SCTs) could offer valuable insights into the interpatient variability of MTX pharmacokinetics and toxicities in ALL children.

Mucocutaneous Ulcerations and Pancytopenia Secondary to Methotrexate Toxicity in a Patient With Rheumatoid Arthritis: A Case Report.

Toxicity from methotrexate overdosing is life threatening condition that requires prompt recognition and early treatment. A 71-year-old man from rural Bhutan with diabetes mellitus had symmetrical small joint pain for 3 years associated with early morning stiffness. He was evaluated and diagnosed with seropositive rheumatoid arthritis in a hospital in neighbouring India. He was initiated on Methotrexate and Hydroxychloroquine. The patient had inadvertently taken Methotrexate 10 mg OD for 2 weeks. Following this, he developed painful ulcers on the lips preventing him from swallowing and ulcers around the corona on the penis. He also had intermittent fever with chills. On examination, he had fluid responsive hypotension and pancytopenia without bleeding manifestations. He was managed with intravenous folinic acid, subcutaneous granulocyte colony stimulating factor and intravenous cloxacillin. He had an uneventful recovery and methotrexate has been restarted following adequate patient education. Education of patient and family members is key in preventing medication errors. Early identification and timely management of methotrexate toxicity is key in preventing mortality.

A review on toxic epidermal necrolysis-like superficial wound lesions: Issue and challenge on 10 clinical entities.

Toxic epidermal necrolysis (TEN) is a severe skin reaction caused by extensive epidermal and mucosal necrosis. This clinical phenomenon is known as an acute syndrome of apoptotic pan-epidermolysis (ASAP). The ASAP phenomenon is observed in conditions that mimic TEN, highlighting the challenge in distinguishing these conditions. While TEN is a well-recognized entity, distinguishing it from other TEN-like conditions presents significant diagnostic and treatment challenges. These conditions include drug rash with eosinophilia and systemic symptoms (DRESS), generalized bullous fixed drug eruption (GBFDE), acute generalized exanthematous pustulosis (AGEP), TEN-like methotrexate toxicity, mustard gas toxicity, pseudoporphyria, mycoplasma-induced rash and mucositis (MIRM), multisystem inflammatory syndrome in children (MIS-C), graft versus host disease (GVHD), and acute cutaneous lupus erythematosus and subacute cutaneous lupus erythematosus. This review explores these ten separate entities and debates their clinical features, pathophysiology, diagnosis and management.

MIL-101 (Fe)-NH2/multi-walled carbon nanotubes nanocomposite modified glassy carbon electrode for simultaneous voltammetric determination of methotrexate and calcium folinate

Methotrexate (MTX), a structural analog of folic acid, known as one of the most potent and widely used medications for the treatment of different types of cancer and autoimmune diseases. However, the long-term treatment with MTX can cause various side effects. Calcium folinate (CF) is usually used with MTX to reduce the toxicity of MTX and enhance the efficacy of treatment. Therefore, the simultaneous determination of MTX and CF is very important. In this study a nanocomposite consisting of Fe-based metal–organic framework (MIL-101 (Fe)-NH2) and multi-walled carbon nanotubes MWCNTs) was synthesized by solvothermal method. The morphology and structure of the MIL-101 (Fe)-NH2/MWCNTs nanocomposite were investigated by various techniques. Then, a glassy carbon electrode modified with synthesized nanocomposite (MIL-101(Fe)-NH2/MWCNTs/GCE) was employed as a working electrode for voltammetric detection of MTX. Compared with un-modified GCE, the MIL-101 (Fe)-NH2/MWCNTs/GCE showed good electro-catalytic performance for oxidation of MTX. Under optimal conditions, the oxidation peak currents of MTX exhibit a linear relationship with its concentration in the range of 0.1–300.0 μM, and the limit of detection (LOD) is 0.04 μM. Also, the MIL-101 (Fe)-NH2/MWCNTs/GCE showed good activity towards MTX determination in the presence of CF. The separation of the oxidation peak potentials (200 mV), indicating its suitability for the simultaneous detection of these two drugs using differential pulse voltammetry (DPV). Finally, applicability of the developed sensor was verified by MTX and CF analysis in the pharmaceutical specimens with excellent results (recovery 96.9–103.4 %, and relative standard deviation (RSD) ≤ 3.6 %).

A case of methotrexate toxicity in induced abortion

Background: Methotrexate is an anticancer drug from the antimetabolite class which is being used in cases of unruptured ectopic pregnancy and as an abortifacient as well. Giving correct dosage according to the body weight and body surface area is of utmost importance to avoid the complications.Case: A 29-year-old, G4P3L3 at 8+3 weeks of gestation with previous three cesarean sections, presented to our hospital with complaints of fever, abdominal pain, multiple episodes of diarrhea and vomiting and oral ulcers for two days. On enquiring, it was found that she received two doses of injection methotrexate (75 and 50 mg) as an abortifacient for an unwanted intrauterine pregnancy from a local doctor nearby. On presentation, her general condition was poor, she was dehydrated, temperature was 101.3°F, pulse rate was 130 beats/minute, blood pressure was 102/64mmHg, respiratory rate was 25/minute and SPO2 was 99% on room air. Her body weight was 55 kgs. Skin pigmentation and multiple oral ulcers were present. Per speculum and per vaginal examination was suggestive of minimal spotting and bulky uterus. Initial blood investigations revealed a hemoglobin level of 8.8 gm/dl, TLC of 1000/mm3, platelet count of 1.45L, INR of 1.37, and normal liver and renal function tests. A provisional diagnosis of methotrexate toxicity was made. Management of the patient was started in consultation with medical oncologist along with supportive treatment. She received IV fluids, antibiotics, and blood products (2-unit PCV and 12-RDP). She received oral leucoverin 15 mg QID and two doses of 300 mg granulocyte stimulating factor each. Gradually, her condition improved and thereafter tablet misoprostol was given for abortion. She was discharged after 13 days of hospital stay with contraceptive advice.Conclusion: The successful management of this case who presented with a constellation of symptoms and deteriorating hematological parameters exhibits the significance of a multidisciplinary approach and vigilant monitoring. Strategies to reduce the side effects and their early recognition include a safety checking procedure like SMART (safety of methotrexate therapy by assessing risks and toxicity) and determining plasma methotrexate concentrations.

Association of SLC19A1 Gene Polymorphisms and Its Regulatory miRNAs with Methotrexate Toxicity in Children with Acute Lymphoblastic Leukemia.

Methotrexate (MTX) is an anti-folate chemotherapeutic agent that is considered to be a gold standard in Acute Lymphoblastic Leukemia (ALL) therapy. Nevertheless, toxicities induced mainly due to high doses of MTX are still a challenge for clinical practice. MTX pharmacogenetics implicate various genes as predictors of MTX toxicity, especially those that participate in MTX intake like solute carrier family 19 member 1 (SLC19A1). The aim of the present study was to evaluate the association between SLC19A1 polymorphisms and its regulatory miRNAs with MTX toxicity in children with ALL. A total of 86 children with ALL were included in this study and were all genotyped for rs2838958, rs1051266 and rs1131596 SLC19A1 polymorphisms as well as the rs56292801 polymorphism of miR-5189. Patients were followed up (48, 72 and 96 h) after treatment with MTX in order to evaluate the presence of MTX-associated adverse events. Our results indicate that there is a statistically significant correlation between the rs1131596 SLC19A1 polymorphism and the development of MTX-induced hepatotoxicity (p = 0.03), but there is no significant association between any of the studied polymorphisms and mucositis or other side effects, such as nausea, emesis, diarrhea, neutropenia, skin rash and infections. In addition, when genotype TT of rs1131596 and genotype AA of rs56292801 are both present in a patient then there is a higher risk of developing severe hepatotoxicity (p = 0.0104).

Managing macrophage activation syndrome SLE: Clinical and therapeutic insights

Systemic lupus erythematosus (SLE) is a multisystem autoimmune disease with frequent hematologic complications, including leukopenia, thrombocytopenia, autoimmune hemolytic anemia, and, in rare cases, macrophage activation syndrome (MAS). MAS, a severe hyperinflammatory condition, presents diagnostic challenges due to its overlap with SLE flares, infections, and drug-related adverse effects. This case report presents a 22-year-old female with SLE who experienced clinical deterioration following methotrexate initiation, manifesting as fever, pancytopenia, and transaminitis. Differential diagnoses were considered, including MAS, infection, and methotrexate toxicity. Prompt recognition and management with corticosteroids and immunoglobulins led to clinical improvement, underscoring the complexity of diagnosing and treating MAS in SLE patients.

Microneedles with Implantable Tip-Accumulated Therapeutics for the Long-Term Management of Psoriasis.

Methotrexate is successfully used as the gold standard for managing moderate-to-severe psoriasis. However, the low bioavailability and short half-life of the oral pills and the invasiveness of the parenteral injections make these suboptimal therapeutic options. Microneedles, bridging the advantages of the former forms, are successfully used to deliver methotrexate for different therapeutic purposes. However, the utilized dissolving microneedles demand frequent administration, potentially compromising patients' compliance. Additionally, the high toxicity of methotrexate prompts a quest for safer alternatives. Phloretin, a natural compound with confirmed antipsoriatic potential, emerges as a promising candidate. Herein, microneedle patches with separable, slow-degrading tips are developed for the sustained delivery of methotrexate and phloretin, as a comprehensive solution for long-term psoriasis management. Both compounds are individually loaded at varying doses and display sustained-release profiles. The developed microneedle patches demonstrate high mechanical strength, favorable drug delivery efficiency, and remarkable antipsoriatic potential both in vitro in keratinocytes and in vivo in a psoriasis mouse model. Comparative analysis with two subcutaneous injections reveals a similar antipsoriatic efficacy with a single patch of either compound, with prominent phloretin safety. Therefore, the developed patches present a superior alternative to methotrexate's current marketed forms and provide a viable alternative (phloretin) with comparable antipsoriatic efficacy and higher safety.

Frequency of pharmacogenomic variants affecting safety and efficacy of immunomodulators and biologics in a South Asian population from Sri Lanka

BackgroundImmunomodulators are important for management of autoimmune diseases and hematological malignancies. Significant inter-individual variation in drug response/reactions exists due to genetic polymorphisms. We describe frequency of identified genetic polymorphisms among Sri Lankans.MethodsSri Lankan data were obtained from an anonymized database of 670 participants. Data on variants and global distribution of Minor Allele frequency (MAF) of other populations (South Asian, Ashkenazi-Jewish, East-Asian, European-Finnish, European-non-Finnish, Latino-American, African/African-American) were obtained from pharmGKB online database.ResultsSLC19A1 (rs1051266) variant had a MAF (95% CI) of 63.3% (60.7–65.9). Other common variants included FCGR3A (rs396991), MTHFR (rs1801133), ITPA (rs1127354), CYP2C9*3 (rs1057910) and NUD15*3 (rs116855232), with MAFs of 35.3% (32.7–37.9), 12.2% (10.4–13.9), 10.9% (9.2–12.6), 9.8% (8.2–11.4), 8.3% (6.8–9.8) respectively. Less commonly present variants included CYP2C9*2 (rs1799853) (2.5%[1.7–3.4]), TPMT*3C (rs1142345) (1.9%[1.1–2.6]), TPMT*3B (rs1800460) (0.2%[0–0.5]), CYP3A5*6 (rs10264272) (0.2%[0–0.4]) and CYP3A4*18 (rs28371759) (0.1%[0–0.2]). The SLC19A1 (rs1051266), NUD15*3 (rs116855232), CYP2C9*3 (rs1057910), FCGR3A (rs396991), and ITPA (rs1127354) showed significantly higher frequencies in Sri Lankans compared to many other populations, exceptions include FCGR3A in Ashkenazi-Jewish and ITPA in East-Asians. Conversely, MTHFR (rs1801133), TPMT*3B (rs1800460), and CYP2C9*2 (rs1799853) were significantly less prevalent among Sri Lankans than in many other populations. Sri Lankans exhibited lower prevalence of TPMT*3C (rs1142345) compared to European-non-Finnish, Latino-Americans, and African/African-Americans; CYP3A4*18 (rs28371759) compared to East-Asians; and CYP3A5*6 (rs10264272) compared to African/African-Americans and Latino-Americans.ConclusionSri Lankans exhibit higher frequencies in variants reducing methotrexate efficacy (SLC19A1), increasing azathioprine myelotoxicity (NUDT15), and lower frequencies in variants linked to increased azathioprine toxicity (TPMT*3B, TPMT*3C), reduced tacrolimus efficacy (CYP3A4*18), and methotrexate toxicity risk (MTHFR). Beneficial variants enhancing rituximab efficacy (FCGR3A) are more prevalent, while those reducing tacrolimus dosage (CYP3A5*6) are less common. This highlights need for targeted medication strategies to improve treatment outcomes.

Methotrexate induced hepatotoxicity in metabolic dysfunction-associated steatotic liver disease.

Hepatotoxicity is an under-recognized and potentially fatal side effect of high-dose methotrexate (HDMTX) chemotherapy, and this risk is compounded in children with metabolic dysfunction-associated steatotic liver disease and/or metabolic-associated steatohepatitis. We present the case of a 12-year-old obese, Hispanic male with elevated hepatic transaminases of unknown etiology at initiation of high-risk B-cell acute lymphoblastic leukemia chemotherapy. He developed acute kidney injury within 24 hours of receiving intravenous HDMTX which progressed to acute hepatic failure. Liver biopsy confirmed methotrexate toxicity aggravated by undiagnosed metabolic dysfunction-associated steatotic liver disease. Rapid deterioration precluded liver transplantation, and he died 21 days after HDMTX treatment. This case highlights the need for comprehensive hepatic evaluation in patients with known or suspected liver disease when administering HDMTX. Dialysis should be considered if delayed methotrexate clearance occurs due to potential for rapid, irreversible hepatotoxicity.

The Impact of ATIC (rs2372536) Polymorphism on Responsiveness to and Toxicity of Methotrexate (MTX) in Rheumatoid Arthritis

The Impact of ATIC (rs2372536) Polymorphism on Responsiveness to and Toxicity of Methotrexate (MTX) in Rheumatoid Arthritis

Methotrexate toxicity presenting as pancytopenia

Methotrexate toxicity presenting as pancytopenia

Untargeted metabolic profiling of high-dose methotrexate toxicity shows alteration in betaine metabolism

Cardiotoxicity is a well-established adverse effect of several drugs across multiple therapeutic indications. It is particularly prevalent following anticancer therapy. In order to evaluate the changes in cellular metabolism associated with methotrexate cardiotoxicity, we treated Wistar rats with a single high dose of methotrexate (HDMTX), and after five days, the animals were sacrificed. We then analyzed the cardiotoxicity parameters in serum like Cardiac enzymes(CK-MB, Troponin T, ALP), Inflammatory markers (TNF-α and IL-6), oxidative stress markers (NO, NOX-2), histopathology and cardiac tissue with the goal of identifying a metabolic signature of cardiotoxicity using discovery-based metabolomics. The biochemical parameters for cardiac enzymes, oxidative stress and inflammatory markers showed a significant increase in all three categories in rats treated with HDMTX. These findings were mirrored in the histopathological analysis confirming cardiotoxicity due to HDMTX. The results showed a total of 95 metabolites that were found to be significantly (p < 0.05) modulated: either up- or downregulated in the HDMTX-treated group when compared with the control group. Using integrated pathway analysis we found these metabolites were associated with many important cardiac tissue metabolic pathways, such as the malate aspartate shuttle, taurine and hypotaurine metabolism, betaine metabolism, spermidine biosynthesis, and homocysteine degradation. Among them, L-arginine, homocysteine, and betaine were significantly upregulated, suggesting their possible association with cardiac tissue injury. Overall, we provided evidence for using untargeted metabolomics to identify novel metabolites associated with HDMTX cardiac toxicity.

Methotrexate toxicity in patients with rheumatic diseases: A 10-year evaluation of manifestations, risk factors, and mortality

Methotrexate (MTX) is a commonly used medication in the treatment of rheumatic diseases. MTX toxicity is a serious and at times fatal complication. In this 10-year cohort, we retrospectively evaluated the cases of MTX toxicity among rheumatic patients to identify demographic, clinical, and laboratory characteristics and risk factors of mortality. Through 10 years, patients older than 17 years with a rheumatic disease and at least one major manifestation of MTX toxicity (mucositis, myelosuppression, pneumonitis) were enrolled in the study. Demographic, clinical, and laboratory data were collected. Comparisons were made between patients who recovered and those who passed away. Of the 66 patients, most had rheumatoid arthritis, followed by systemic lupus erythematous. Most patients were females, and their mean age was 63.3±12.4 years. The most common comorbidity was hypertension followed by diabetes mellitus. Most patients were on polypharmacy and were taking MTX incorrectly. Other potential risk factors of toxicity were hypoalbuminemia, decreased renal function, and infection. The outcome of six patients was death and the predictors of mortality were severe leukopenia at admission, infection, especially sepsis, and severe renal impairment. In this study, we observed some potential risk factors for toxicity, namely incorrect MTX use, polypharmacy, renal impairment, hypoalbuminemia, infection, and comorbid conditions such as diabetes mellitus, hypertension, and ischemic heart disease. We also identified the risk factors of mortality, including infection, sepsis, severe leukopenia, high creatinine levels, and severe renal impairment.

Navigating methotrexate toxicity: Examining the therapeutic roles of folinic acid and glucarpidase.

Methotrexate (MTX) toxicity varies depending on factors such as dosing frequency (acute or repeated), dosage (low or high) and the administration route (oral, parenteral or intrathecal). Renal impairment can trigger or exacerbate MTX toxicity. Acute oral low-dose MTX (LDMTX) overdoses seldom lead to toxicity due to the saturable maximal bioavailable dose, but toxicity risks increase with repeated low doses (>3 days), high-dose MTX (HDMTX) or intrathecal poisoning. Folinic acid shares MTX transporters in the gut and cells and bypasses the MTX-induced dihydrofolate reductase inhibition. The required folinic acid dosage differs for low-dose and high-dose MTX toxicities. Acute LDMTX poisoning rarely requires folinic acid, while chronic LDMTX poisoning needs low-dose folinic acid until cellular function is restored. In HDMTX toxicities, early intravenous folinic acid administration is recommended, with dose and duration being guided by MTX concentrations and clinical improvement. In intrathecal MTX poisoning, folinic acid should be administered intravenously. Glucarpidase, a recombinant bacterial enzyme, has a high affinity for MTX and folate analogues in the intravascular or intrathecal systems. It decreases serum MTX concentrations by 90%-95% within 15 min. Its primary indication is for intrathecal MTX poisoning. It is rarely indicated in HDMTX toxicity unless patients have renal injury. However, there is no literature evidence supporting its use in HDMTX poisoning. Its use is limited by its significant cost and lack of availability. Haemodialysis can be potentially useful for MTX removal in cases where glucarpidase is not available. Additionally, fluid hydration, renal support and urine alkalinization are important adjunctive therapies for managing MTX toxicities.

Hyaluronic acid-conjugated methotrexate and 5-fluorouracil for targeted drug delivery

The delivery of chemotherapeutical drugs via nanomaterials has become a focus of pharmaceutical research over several decades due to improved drug delivery to cancer cells, decreased side effects on normal tissues, and increased therapeutic efficacy. Herein, a novel hyaluronic acid-conjugated methotrexate and 5-fluorouracil nanodrug system has been developed to address the critical limitations associated with the high toxicity and side effects of methotrexate and 5-fluorouracil. Furthermore, this nanodrug system enhances the targeting capacity of drug molecules and facilitates the potential integration of multimodal drug therapies. Concomitantly, the synergistic effects of MTX with 5-fluorouracil have been shown to improve the therapeutic index of MTX while attenuating the associated toxicities of MTX. The structure and micromorphology of the novel nanodrug can be confirmed by 1HNMR, FT-IR, UV–Vis, DLS, TEM, and AFM. Due to the ability of HA to bind to CD44 receptors activated on the surface of cancer cells and its enhanced permeability and retention (EPR) effect, the novel nanodrug we designed and synthesized can effectively target cancer cells. Cell counting Kit-8 (CCK8), flow cytometry, and live-dead staining assays in vitro showed that this nanodrug system had high targeting and antitumor activity against CD44 receptors. By using drugs to act on patient-derived colorectal, liver, and breast cancer organoids, the anticancer effect of the nanodrug was identified and verified. These results showed that the nanodrug system developed in this study may have great potential as a targeted therapy for cancer.

Methotrexate Induced Oral Mucositis and Bone Marrow Suppression in Psoriatic Patient

Methotrexate is an antimetabolite which binds to the dihydrofolate reductase enzyme and used in the treatment of malignant disorders and autoimmune diseases. Myelosuppression is a serious complication of methotrexate toxicity. The following case describes a case of 67 year old male patient diagnosed with psoriasis and wrongly taken methotrexate 7.5 mg daily for 3 days and twice daily for 4 days. He presented with oral mucositis and his lab reports showed myelosuppression. Our case emphasizes the significance of effective patient counselling to ensure the understanding of prescription and to prevent drug toxicity. The key learning points of our case report include the importance of adhering to prescribed medication regimens, awareness of MTX toxicity, early initiation of folinic acid as an antidote, vigilant monitoring of toxicity, and effective patient education on proper medication use. Adherence to established MTX therapy guidelines in dermatological conditions is crucial to minimize the risk of patient.

Polymorphisms for the Clinical Efficacy and Toxicity of Methotrexate in Patients with Rheumatoid Arthritis: Systemic Review

Polymorphisms for the Clinical Efficacy and Toxicity of Methotrexate in Patients with Rheumatoid Arthritis: Systemic Review

Mucocutaneous Ulcers Unmasking Severe Systemic Methotrexate Toxicity – A Case-Series and Review of Literature

Abstract Background: Methotrexate is a widely used immunosuppressant with good efficacy and cost-effectiveness. However, one of the drawbacks of methotrexate has been toxicity due to accidental overdose. During the COVID pandemic, there was an alarming increase in the number of patients with methotrexate toxicity which prompted us to do this study. Objective: To evaluate the clinical features and contributing factors in patients presenting with methotrexate toxicity. Materials and Methods: A detailed evaluation of the clinical features, laboratory indices, contributing factors, and outcomes of the patients presenting with methotrexate toxicity was analyzed. Results: A total of 19 cases were seen during the study period. All of the patients had oral mucositis and several developed cutaneous ulcerations. Laboratory abnormalities included cytopenia, transaminitis, and renal impairment. While sixteen patients recovered successfully, three people died as a result of delays in medical assistance. In addition to comorbidities, pandemic-induced restrictions played a major role in patients accidentally overdosing with methotrexate. Conclusion: This study highlights the fact that even low-dose methotrexate taken incorrectly can result in a lethal outcome, which is preventable.

Clinical outcomes of patients receiving three versus four doses of methotrexate with concomitant antithymocyte globulin in match unrelated donor allogeneic stem cell transplant: A single-center experience.

Methotrexate (MTX) doses on days +1, +3, +6, and +11 after match unrelated donor allogeneic stem cell transplant (MUD HSCT) is a common graft-versus-host disease (GVHD) prophylaxis regimen. However, the overlapping toxicity of MTX with conditioning chemotherapy sometimes warrants the omission of the fourth dose of MTX. Prior single-institution studies showed conflicting results comparing the outcomes of patients who received three versus four doses of MTX, but to our knowledge, the effect of concomitant antithymocyte globulin (ATG) has not been reported. Charts of patients who underwent MUD HSCT between 2009 and 2023 were reviewed. Patients received rabbit ATG (Thymoglobulin), given at 0.5mg/kg on day -3, 2mg/kg on day -2, and 2.5mg/kg on day -1. MTX is given at 15mg/m2 on day +1 and 10mg/m2 on days +3, +6, and +11. Severe mucositis was the most common indication for day +11 MTX omission (82%). We identified 292 patients (116 in 3 dose cohort and 176 in 4 dose cohort). Median follow-up was 23 months (range 1-151). Patients in the 4 doses cohort were more frequently male (68% vs. 50%, p<0.01), received a reduced intensity conditioning regimen (38.0% vs. 22%, p<0.01), were older (median 58 vs. 54 years, p=0.02), and received a transplant in the earlier era (median HSCT year 2014 vs. 2018, p<0.01). A statistically significant difference was not evidenced between the cohorts for the following outcomes: acute GVHD (aGVHD) (HR 1.1, 95% CI 0.9-1.5), chronic GVHD (cGVHD) (HR 1.3, 95% CI 0.8-1.6), relapse-free survival (RFS) (HR 1.0, 95% CI 0.6-1.5), non-relapse mortality (NRM) (HR 1.4, 95% CI 0.9-2.2), and overall survival (OS) (HR 1.2, 95% CI 0.9-1.7). Both cohorts had similar median time to neutrophil engraftment at 14 days. When ATG is incorporated, omission of day +11 MTX does not significantly impact the rate of engraftment or cumulative incidence of aGVHD, cGVHD, RFS, NRM, and OS.