Targeting a mitochondrial E3 ubiquitin ligase complex to overcome AML cell-intrinsic Venetoclax resistance.

To identify molecules/pathways governing Venetoclax (VEN) sensitivity, we performed genome-wide CRISPR/Cas9 screens using a mouse AML line insensitive to VEN-induced mitochondrial apoptosis. Levels of sgRNAs targeting March5, Ube2j2 or Ube2k significantly decreased upon VEN treatment, suggesting synthetic lethal interaction. Depletion of either Ube2j2 or Ube2k sensitized AML cells to VEN only in the presence of March5, suggesting coordinate function of the E2s Ube2j2 and Ube2k with the E3 ligase March5. We next performed CRISPR screens using March5 knockout cells and identified Noxa as a key March5 substrate. Mechanistically, Bax released from Bcl2 upon VEN treatment was entrapped by Mcl1 and Bcl-XL and failed to induce apoptosis in March5 intact AML cells. By contrast, in March5 knockout cells, liberated Bax did not bind to Mcl1, as Noxa likely occupied Mcl1 BH3-binding grooves and efficiently induced mitochondrial apoptosis. We reveal molecular mechanisms underlying AML cell-intrinsic VEN resistance and suggest a novel means to sensitize AML cells to VEN.

Genome-wide CRISPR-Cas9 screen identifies rationally designed combination therapies for CRLF2-rearranged Ph-like ALL.

Acute lymphoblastic leukemia (ALL) harboring the IgH-CRLF2 rearrangement (IgH-CRLF2-r) exhibits poor clinical outcomes and is the most common subtype of Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL). While multiple chemotherapeutic regimens, including ruxolitinib monotherapy and/or its combination with chemotherapy, are being tested, their efficacy is reportedly limited. To identify molecules/pathways relevant for IgH-CRLF2-r ALL pathogenesis, we performed genome-wide CRISPR-Cas9 dropout screens in the presence or absence of ruxolitinib using 2 IgH-CRLF2-r ALL lines that differ in RAS mutational status. To do so, we employed a baboon envelope pseudotyped lentiviral vector system, which enabled, for the first time, highly efficient transduction of human B cells. While single-guide RNAs (sgRNAs) targeting CRLF2, IL7RA, or JAK1/2 significantly affected cell fitness in both lines, those targeting STAT5A, STAT5B, or STAT3 did not, suggesting that STAT signaling is largely dispensable for IgH-CRLF2-r ALL cell survival. We show that regulators of RAS signaling are critical for cell fitness and ruxolitinib sensitivity and that CRKL depletion enhances ruxolitinib sensitivity in RAS wild-type (WT) cells. Gilteritinib, a pan-tyrosine kinase inhibitor that blocks CRKL phosphorylation, effectively killed RAS WT IgH-CRLF2-r ALL cells in vitro and in vivo, either alone or combined with ruxolitinib. We further show that combining gilteritinib with trametinib, a MEK1/2 inhibitor, is an effective means to target IgH-CRLF2-r ALL cells regardless of RAS mutational status. Our study delineates molecules/pathways relevant for CRLF2-r ALL pathogenesis and could suggest rationally designed combination therapies appropriate for disease subtypes.

Listeria monocytogenes Bacteremia During Isatuximab Therapy in a Patient with Multiple Myeloma.

An elderly patient with multiple myeloma (MM) was being treated with several regimens and developed a severe drug eruption, necessitating the use of atovaquone instead of trimethoprim-sulfamethoxazole for pneumocystis pneumonia (PCP) prophylaxis. For progressive MM, treatment with isatuximab, an anti-CD38 monoclonal antibody, was started. During the treatment, he developed Listeria monocytogenes bacteremia and recovered quickly with ampicillin administration. CD38 is closely related to the innate immune response against L. monocytogenes, and isatuximab may increase the risk of infection. Therefore, trimethoprim-sulfamethoxazole may be useful in the prevention of not only PCP but also L. monocytogenes infection.

Successful allogeneic stem cell transplantation in a case with acute myeloid leukemia and invasive Schizophyllum commune rhinosinusitis.

Schizophyllum commune, a basidiomycete fungus, is a quite rare cause of invasive sinusitis for which no standard treatment has yet been established. We report herein a 59-year-old woman who developed S. commune rhinosinusitis after remission induction chemotherapy for her acute myeloid leukemia. No causative microorganisms were identified in the sinus lavage fluid culture, whereas nucleotide sequencing of the internal transcribed spacer region using endoscopic sinus biopsy specimen could confirm the pathogen as S. commune. Liposomal amphotericin B and voriconazole (VRCZ) treatment ameliorated both her clinical symptoms and laboratory findings. The patient was successfully treated with allogeneic stem cell transplantation, under continuous VRCZ administration, without aggravation of S. commune sinusitis. Molecular diagnosis and prompt intervention with suitable antifungal drugs may be crucial to manage this rare infectious complication.

Haploidentical hematopoietic stem cell transplantation with one-day posttransplant cyclophosphamide and anti-thymocyte globulin for graft failure.

Patients with myelofibrosis usually have poor prognosis. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains the only curative therapy, which has graft failure as a life-threatening complication. However, no consensus is available with regard to therapeutic options for patients with graft failure. Here we report a patient with myelofibrosis who underwent successful salvage haploidentical hematopoietic stem cell transplantation (haplo-HSCT) with one-day posttransplant cyclophosphamide (PTCy) and low-dose anti-thymocyte globulin (ATG) for graft failure. A 39-year-old Japanese male patient with rapidly progressing primary myelofibrosis underwent cord blood transplantation (CBT). Unfortunately, both the first and second CBT resulted in primary graft failures. Therefore, emergent haplo-HSCT from a sibling donor was performed with one-day PTCy (50 mg/kg on day ＋3) after conditioning with etoposide (60 mg/m2 on days -3 and -2) and rabbit anti-human thymocyte globulin (1 mg/kg on days -2 and -1). Neutrophil engraftment was achieved on day ＋13 after haplo-HSCT, and no severe infection or regimen-related toxicity was observed. Skin stage 3, gut stage 1 total grade II acute graft-versus-host disease developed. His posttransplant course had been uneventful with cutaneous chronic graft-versus-host disease (NIH score 2) and suppressed relapse. We believe that haplo-HSCT with one-day PTCy and low-dose ATG is one of the successful therapeutic options for graft failure.

Conditioning regimen with a 75% dose of standard busulfan/cyclophosphamide plus fludarabine before cord blood transplantation in older patients with AML and MDS.

In this retrospective study, we aimed to establish a conditioning regimen for older patients receiving cord blood transplantation (CBT). This study included 21 older patients [median age 65 (58-73) years] with acute myeloid leukemia and myelodysplastic syndrome who underwent single CBT following a conditioning regimen comprising fludarabine (FLU) 125-175 mg/m2, busulfan (BU) 9.6 mg/kg, and cyclophosphamide (CY) 90 mg/kg. Twelve patients (57.1%) were considered high or very high risk according to the disease risk index. Nineteen achieved neutrophil engraftment at a median of 19 days (range 14-29 days) after CBT (cumulative incidence 90.5%). During a median observation period of 24.3 months, the overall survival (OS) rates at 100 days and 2 years were 76.2% and 47.6%, respectively, with cumulative 2-year relapse and non-relapse mortality (NRM) rates of 19.0% and 38.1%, respectively. Infectious disease was the leading cause of NRM (n = 5) and occurred within 100 day post-transplantation in two patients. This suggested that the administration of a reduced BU/CY plus FLU regimen to older patients receiving CBT enables an early recovery with high neutrophil engraftment, relapse suppression, and acceptable NRM rates.

[Toxoplasmosis-associated central nervous system vasculitis accompanied by multiple cerebral hemorrhages developing subsequent to cord blood transplantation].

A 57-year-old man with high-risk myelodysplastic syndrome underwent umbilical cord blood transplantation. He began receiving steroids on day 14 for acute graft-versus-host disease, and experienced dizziness on day 75 during gradual dose reduction. Multiple hemorrhages were observed in the cerebrum, cerebellum, and brainstem. His bleeding increased, and he underwent a brain biopsy on day 91. Subsequently, he was diagnosed with central nervous system vasculitis (CNSV) on the basis of the observed aggregation of mature CD3+ lymphocytes around small vessels and vascular wall invasion by lymphocytes and macrophages. After receiving high-dose steroid therapy, cerebral hemorrhage stopped; however, dysphasia occurred on day 113 and the patient died of cerebral edema on day 128. Toxoplasma DNA and tachyzoites were detected in the brain biopsy specimen during additional examinations; therefore, we suspected that the toxoplasmosis was related to the onset of CNSV. CNSV is a rare, rapidly progressing disease that may present as a fatal post-transplantation central nervous system complication. Investigating the causes of CNSV, including CNSV associated with toxoplasmosis, is critically important for improving the prognosis of patients with CNSV.