Combination of an aurora kinase inhibitor and the ABL tyrosine kinase inhibitor asciminib against ABL inhibitor-resistant CML cells.

The development of BCR::ABL1-targeting tyrosine kinase inhibitors (TKIs) has improved the prognosis of patients with chronic myeloid leukemia (CML). However, resistance to ABL TKIs can develop in CML patients due to BCR::ABL1 point mutations and CML leukemia stem cell (LSC). Aurora kinases are essential kinases for cell division and regulate mitosis, especially the process of chromosomal segregation. Aurora kinase members also promote cancer cell survival and proliferation. This study analyzed whether aurora kinases were regulated in the progression of CML. It also evaluated the efficacy of the ABL TKI asciminib and the aurora kinase inhibitor LY3295668. The expressions of AURKA and AURKB were higher in the CML cells compared with normal cells using a public database (GSE100026). Asciminib or LY3295668 alone inhibited CML cells after 72 h, and cellular cytotoxicity was increased. The combined use of Asciminib and LY3295668 increased superior efficacy compared with either drug alone. Colony formation was reduced by cotreatment with asciminib and LY3295668. In the cell-cycle analyses, LY3295668 induced G2/M arrest. Cell populations in the sub-G1 phase were observed when cotreating with asciminib and LY3295668. The combination treatment also changed the mitochondrial membrane potential. In addition, AURKA shRNA transfectant cells had increased asciminib sensitivity. Combining asciminib and aurora kinase inhibition enhanced the efficacy and is proposed as a new therapeutic option for patients with CML. These findings have clinical implications for a potential novel therapeutic strategy for CML patients.

Characterization of Asciminib-Resistant Philadelphia Chromosome-Positive Cells.

Background: Asciminib is approved for treating patients with chronic-phase chronic myeloid leukemia who were previously treated with two or more tyrosine kinase inhibitors or those with T315I mutation. However, the mechanisms underlying asciminib resistance remain unclear. Methods: In this study, we established a new asciminib-resistant cell line. We examined BCR::ABL1 gene mutation analysis and the effects of conventional chronic myelogenous leukemia inhibitors. Results: Direct sequencing revealed Y139D and T315I mutations in asciminib-resistant cells. Ponatinib and omacetaxine were effective against asciminib-resistant cells. Conclusions: Y139D and T315I mutations are extremely resistant to asciminib. Ponatinib and omacetaxine show potential for treating asciminib-resistant chronic myeloid leukemia.

[Ovarian hyperstimulation syndrome with controlled ovarian stimulation after induction therapy for Philadelphia chromosome-positive acute lymphoblastic leukemia].

A 27-year-old woman with newly diagnosed Philadelphia chromosome-positive acute lymphoblastic leukemia received induction therapy with dasatinib and prednisolone. From the time of diagnosis, oocyte storage was planned in accordance with the patient's wishes. After progesterone administration for suppression of menstruation, and blood cell recovery, ovarian stimulation was performed and a sufficient number of eggs was collected. The patient was considered at high risk for ovarian stimulation syndrome (OHSS) and received cabergoline and letrozole. However, ovarian enlargement and ascites were observed on ultrasonography 2 days after egg collection, and a diagnosis of moderate OHSS was made. Circulatory management was performed and low-molecular-weight heparin was administered. Dasatinib was discontinued due to the appearance of pleural effusion. Fluid retention improved after menstruation resumed, and the patient was able to continue consolidation with dasatinib and cord blood transplantation. Although tyrosine kinase inhibitors are expected to simplify planning of oocyte storage, the risk of complicating OHSS should be noted.

Robot therapy aids mental health in patients with hematological malignancy during hematopoietic stem cell transplantation in a protective isolation unit.

Patients with hematological malignancy experience physical and psychological pain, such as a sense of isolation and confinement due to intensive chemotherapy in a protective isolation unit (PIU). We examined whether the intervention of a robotic puppy, aibo (manufactured by Sony), could improve patients' mental health as an alternative therapy for pet therapy, which is not feasible in PIU. This study included 21 patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) (n = 16) or autologous HSCT (n = 5). The patients were randomly divided into the aibo and control groups. Psychological effects were regularly assessed by measuring the levels of salivary stress hormone chromogranin A (CgA), serum oxytocin, and serum cortisol and the quick Inventory of Depressive Symptomatology Self-Report (QIDS-SR) scores. The aibo group demonstrated a significant decrease in CgA level, while the control group showed the opposite trend. In addition, changes in serum oxytocin and cortisol levels indicated that aibo helped reduce stress. There was no significant difference in the QIDS-SR scores between the two groups; however, the psychomotor activity in the aibo group improved significantly. These findings suggest that aibo intervention during a stay in a PIU can improve the mental health of patients with hematological malignancies who have undergone HSCT.

Effect of asciminib and vitamin K2 on Abelson tyrosine-kinase-inhibitor-resistant chronic myelogenous leukemia cells.

BACKGROUND: Abelson (ABL) tyrosine kinase inhibitors (TKIs) are effective against chronic myeloid leukemia (CML); however, many patients develop resistance during ABL TKI therapy. Vitamin K2 (VK2) is a crucial fat-soluble vitamin used to activate hepatic coagulation factors and treat osteoporosis. Although VK2 has demonstrated impressive anticancer activity in various cancer cell lines, it is not known whether VK2 enhances the effects of asciminib, which specifically targets the ABL myristoyl pocket (STAMP) inhibitor. METHOD: In this work, we investigated whether VK2 contributed to the development of CML cell lines. We also investigated the efficacy of asciminib and VK2 by using K562, ponatinib-resistant K562 (K562 PR), Ba/F3 BCR-ABL, and T315I point mutant Ba/F3 (Ba/F3 T315I) cells. RESULTS: Based on data from the Gene Expression Omnibus (GEO) database, gamma-glutamyl carboxylase (GGCX) and vitamin K epoxide reductase complex subunit 1 (VKORC1) were elevated in imatinib-resistant patients (GSE130404). UBIA Prenyltransferase Domain Containing 1 (UBIAD1) was decreased, and K562 PR cells were resistant to ponatinib. In contrast, asciminib inhibited CML cells and ponatinib resistance in a dose-dependent manner. CML cells were suppressed by VK2. Caspase 3/7 activity was also elevated, as was cellular cytotoxicity. Asciminib plus VK2 therapy induced a significantly higher level of cytotoxicity than use of each drug alone. Asciminib and VK2 therapy altered the mitochondrial membrane potential. CONCLUSIONS: Asciminib and VK2 are suggested as a novel treatment for ABL-TKI-resistant cells since they increase treatment efficacy. Additionally, this treatment option has intriguing clinical relevance for patients who are resistant to ABL TKIs.

A cross-sectional survey of symptoms and daily living among patients with polycythemia vera and their treating physicians in Japan.

OBJECTIVES: This Japanese cross-sectional survey evaluated the symptoms, daily living activities, and treatment needs of patients with polycythemia vera (PV), as perceived by patients themselves and their physicians. METHODS: The study was conducted at 112 centers (March to July 2022) and included PV patients aged ≥20 years (n = 265) and their attending physicians (n = 151). The patient and physician questionnaires included 34 and 29 questions, respectively, to assess daily living, PV symptoms, treatment goals, and physician-patient communication. RESULTS: Concerning daily living (primary endpoint), work (13.2%), leisure activities (11.3%), and family life (9.6%) were most affected by PV symptoms. Patients aged <60 years more frequently reported an impact on daily living than patients aged ≥60 years. Some patients (30%) reported anxiety about their future condition. The most common symptoms were pruritus (13.6%) and fatigue (10.9%). Pruritus was ranked as the first treatment need for patients, while physicians ranked it fourth. Concerning treatment goals, physicians prioritized thrombosis/vascular event prevention, while patients prioritized delaying PV progression. Physicians were less satisfied with physician-patient communication than patients. CONCLUSIONS: Patients' daily living was largely affected by PV symptoms. There are differences in physician and patient perceptions of symptoms, daily living, and treatment needs in Japan. TRIAL REGISTRATION: UMIN Japan identifier: UMIN000047047.

Gilteritinib Affects the Selection of Dominant Clones in Clonal Hematopoiesis: Sequential Genetic Analysis of an FLT3-ITD Positive AML Patient with Long-Term Gilteritinib Therapy.

We performed sequential molecular analyses of a 75-year-old woman with de novo FLT3-ITD positive acute myeloid leukemia (AML) who had received gilteritinib therapy for 43 months. At the time of diagnosis, her karyotype was normal; however, FLT3-ITD, NPM1, DNMT3A, and IDH2 mutations were detected. She received induction therapy with daunorubicin and cytarabine and achieved hematological complete remission (HCR). After attaining HCR, she underwent consolidation therapy with azacytidine or cytarabine, aclarubicin, and granulocyte-colony stimulating factor. However, AML relapsed eight months after the first HCR. FLT3-ITD and NPM1 mutations were persistently positive, and the patient received gilteritinib therapy. Although the FLT3-ITD clone was not detected during gilteritinib treatment, a clone harboring monosomy 7 and CBL mutations emerged. Bone marrow examinations at 15, 24, and 32 months after gilteritinib treatment revealed multi-lineage blood cell dysplasia without an increase in myeloblasts. After 33 months of treatment, gilteritinib was discontinued for two months because to ileus development, and the FLT3-ITD clone was detected again. Gilteritinib treatment was restarted, and FLT3-ITD became negative. Our analysis demonstrated that: (1) hematopoiesis derived from gilteritinib-resistant clones was generated by long-term gilteritinib treatment, and (2) FLT3-ITD clones regained clonal dominance in the absence of FLT3 inhibition. These findings suggest that gilteritinib affects the selection of dominant clones during clonal hematopoiesis.

Interim analysis of post-marketing surveillance of ravulizumab for paroxysmal nocturnal hemoglobinuria in Japan.

Ravulizumab is a long-acting C5 inhibitor available for treating paroxysmal nocturnal hemoglobinuria (PNH). Post-marketing surveillance (PMS) was implemented following its approval in September 2019 in Japan. We report safety data obtained through to December 2021 for 218 patients and effectiveness data for 194 patients (182 switched from eculizumab and 12 complement inhibitor-naïve). Over a median follow-up of 74.4 weeks, 193 adverse events (AEs) were reported in 66/218 patients (30.3%; incidence 72.73/100 patient-years). The two most frequent AEs were anemia and pyrexia (each 3.01/100 patient-years). The incidence of serious AEs was 36.93/100 patient-years. In patients who switched from eculizumab, lactate dehydrogenase (LDH) and hemoglobin (Hb) levels were maintained over 26 weeks of ravulizumab treatment. In complement inhibitor-naïve patients, LDH decreased significantly and Hb increased significantly over 26 weeks of ravulizumab treatment. These data for Japanese patients with PNH who were naïve to complement inhibitors and patients who switched from eculizumab show that the safety and effectiveness of ravulizumab are consistent with the published clinical trial data. However, transfusion independence was less likely in patients with bone marrow failure. Further follow-up data from this PMS will help to elucidate the long-term clinical safety and effectiveness of ravulizumab for treating PNH.

WEE1 and PARP-1 play critical roles in myelodysplastic syndrome and acute myeloid leukemia treatment.

BACKGROUND: Myelodysplastic syndrome (MDS) is a clonal bone marrow disorder defined by cytopenia and is associated with an increased risk of transformation to acute myeloid leukemia (AML). The outcome of MDS is poor, so alternative therapeutic approaches are needed to improve survival. The inhibition of the DNA damage response pathway, including poly (ADP-ribose) polymerase-1 (PARP-1), has been approved to treat several cancers. In addition, WEE1, a nuclear kinase, is overexpressed in many cancers. Therefore, a WEE1 inhibitor combined with a PARP-1 inhibitor could inhibit the proliferation of MDS and AML. METHODS: We analyzed whether WEE1 was regulated in the progression of MDS and AML. We also evaluated the efficacy of MK-1775 (WEE1 inhibitor) and talazoparib (PARP-1 inhibitor). RESULTS: PARP-1 expression was higher in the AML cells than in the MDS cells. However, WEE1 expression remained unchanged. MK-1775 or talazoparib alone inhibited MDS and AML cells after 72 h, and cellular cytotoxicity and caspase 3/7 activity were increased. The combined use of MK-1775 and talazoparib produced superior efficacy than either drug alone and SKM-1 colony formation was reduced. Significant cell populations in the sub-G1 phase were found in the cell-cycle analyses. Additionally, γ-H2AX expression and caspase 3 activity were increased. The combined treatment also changed the mitochondrial membrane potential. CONCLUSIONS: The combination of a WEE1 inhibitor and PARP-1 inhibitor had enhanced efficacy and is proposed as a new therapeutic option for patients with MDS or AML. Our findings have clinical implications for a potential novel therapeutic strategy for MDS and AML patients.

[RS3PE syndrome with angioimmunoblastic T-cell lymphoma early after the start of immunosuppressive therapy].

A 75-year-old man visited our Collagen Disease Department because of a fever, edema in the lower legs, and arthralgia. He presented with peripheral arthritis of the extremities and was negative for rheumatoid factor, leading to a diagnosis of RS3PE syndrome. A search for malignancy was performed, but no obvious malignant findings were found. After starting treatment with steroid, methotrexate, and tacrolimus, the patient's joint symptoms improved, but after five months, enlarged lymph nodes throughout the body were observed. A lymph node biopsy revealed a diagnosis of other iatrogenic immunodeficiency-associated lymphoproliferative disorders/angioimmunoblastic T-cell lymphoma (OI-LPD/AITL). After discontinuation of methotrexate and follow-up, no lymph node shrinkage was observed, and the patient had strong general malaise, so chemotherapy was started for AITL. After the start of chemotherapy, the patient's general symptoms improved quickly. RS3PE syndrome is a polyarticular, rheumatoid factor-negative, polyarticular synovitis with symmetric dorsolateral hand-palmar symmetric indentation edema that occurs mainly in elderly patients. It is also noted as a paraneoplastic syndrome, with 10%-40% of patients having malignant tumors. When our patient was diagnosed with RS3PE syndrome, a search for malignancy was performed, but there were no findings suggestive of malignant disease. However, after methotrexate and tacrolimus administration was started, the patient developed rapid lymph node enlargement, and the pathology showed AITL. The possibility of AITL as an underlying disease and RS3PE syndrome as a paraneoplastic syndrome, or conversely, OI-LPD/AITL associated with immunosuppressive therapy for RS3PE syndrome is considered. We herein report this case, as sufficient recognition is required for a proper diagnosis to be made and treatment of RS3PE syndrome to be performed.

[Japanese patient preferences between ravulizumab and eculizumab for the treatment of paroxysmal nocturnal hemoglobinuria].

Ravulizumab is the first long-acting complement inhibitor approved for paroxysmal nocturnal hemoglobinuria (PNH) treatment. We evaluated patient preference for ravulizumab or eculizumab among Japanese adults with PNH. The ALXN1210-PNH-301 (NCT02946463) and ALXN1210-PNH-302 (NCT03056040) studies included 23 Japanese adults who are enrolled in complement inhibitor treatment-naive and eculizumab (≥6 months) treatment. Patient preference was assessed using the PNH-specific patient preference questionnaire (PNH-PPQ©). Most patients preferred ravulizumab (19/23, 82.6%), none preferred eculizumab, and four (17.4%) reported no preference (χ2 test, p<0.005). The preference for ravulizumab was driven by its lower infusion frequency (every 8 weeks) compared with eculizumab (every 2 weeks). The included Japanese patients with PNH preferred ravulizumab because of its reduced infusion frequency, which increases activity planning ability, treatment convenience, and overall quality of life, as compared with eculizumab. These data provide useful insight into patient perspectives and may aid decision-making for PNH treatment.

Clinical utility of genomic profiling of AML using paraffin-embedded bone marrow clots: HM-SCREEN-Japan 01.

Next-generation sequencing of AML has identified specific genetic mutations in AML patients. Hematologic Malignancies (HM)-SCREEN-Japan 01 is a multicenter study to detect actionable mutations using paraffin-embedded bone marrow (BM) clot specimens rather than BM fluid in AML patients for whom standard treatment has not been established. The purpose of this study is to evaluate the presence of potentially therapeutic target gene mutations in patients with newly diagnosed unfit AML and relapsed/refractory AML (R/R-AML) using BM clot specimens. In this study, 188 patients were enrolled and targeted sequencing was undertaken on DNA from 437 genes and RNA from 265 genes. High-quality DNA and RNA were obtained using BM clot specimens, with genetic alterations successfully detected in 177 patients (97.3%), and fusion transcripts in 41 patients (23.2%). The median turnaround time was 13 days. In the detection of fusion genes, not only common fusion products such as RUNX1-RUX1T1 and KMT2A rearrangements, but also NUP98 rearrangements and rare fusion genes were observed. Among 177 patients (72 with unfit AML, 105 with R/R-AML), mutations in KIT and WT1 were independent factors for overall survival (hazard ratio = 12.6 and 8.88, respectively), and patients with high variant allele frequency (≥40%) of TP53 mutations had a poor prognosis. As for the detection of actionable mutations, 38% (n = 69) of patients had useful genetic mutation (FLT3-ITD/TKD, IDH1/2, and DNMT3AR822 ) for treatment selection. Comprehensive genomic profiling using paraffin-embedded BM clot specimens successfully identified leukemic-associated genes that can be used as therapeutic targets.

von Willebrand factor activity levels are influenced by driver mutation status in polycythemia vera and essential thrombocythemia patients with well-controlled platelet counts.

von Willebrand factor ristocetin cofactor (vWF activity) and platelet count (PLT) are negatively correlated in patients with polycythemia vera (PV) and essential thrombocythemia (ET). However, vWF activity does not always normalize upon controlling PLT in those patients. To address this issue, we investigated the correlation between vWF activity and PLT in PV and ET patients. The negative correlation between vWF activity and PLT was stronger in calreticulin mutation-positive (CALR+) ET than in Janus kinase 2 mutation-positive (JAK2+) PV or ET groups. When PLT were maintained at a certain level (<600 × 109 /L), low vWF activity (<50%) was more frequently observed in JAK2+ PV patients than in JAK2+ ET (p = .013) or CALR+ ET (p = .013) groups, and in PV and ET patients with ≥50% JAK2+ allele burden than in those with allele burden <50% (p = .015). High vWF activity (>150%) was more frequent in the JAK2+ ET group than in the CALR+ ET group (p = .005), and often associated with vasomotor symptoms (p = .002). This study suggests that some patients with JAK2+ PV or ET have vWF activity outside the standard range even with well-controlled PLT, and that the measurement of vWF activity is useful for assessing the risk of thrombosis and hemorrhage.

Tyrosine Kinase Inhibitors Do Not Promote a Decrease in SARS-CoV-2 Anti-Spike IgG after BNT162b2 Vaccination in Chronic Myeloid Leukemia: A Prospective Observational Study.

We performed a prospective observational study of chronic myeloid leukemia (CML) patients after anti-SARS-CoV-2 BNT162b2 vaccination (VC). In total, 32 CML patients with tyrosine kinase inhibitor (TKI) therapy, 10 CML patients with treatment-free remission, and 16 healthy subjects participated in the study. From April 2021 to September 2021, all cases (median age = 58 years) were vaccinated twice. Immunoglobulin G for SARS-CoV-2 spike protein (S-IgG) was measured at three timepoints (before the first VC, 1−5 weeks after the second VC (T1), and approximately 6 months after the second VC (T2)). S-IgG was not observed before the first VC in any participant. At T1, all cases had acquired S-IgG. There were no significant differences in S-IgG levels among groups. A paired sample comparison of median S-IgG titers between T1 and T2 in all groups showed a significant reduction in T2 S-IgG titers. There were no significant differences in S-IgG levels among groups. When all patients were analyzed, those aged ≥58 years had significantly lower S-IgG levels than those aged <58 years at T1. The BNT162b2 vaccine was highly effective in CML patients with or without TKIs, and S-IgG levels were as persistent as those in healthy individuals.

Clinical characteristics of Japanese patients with polycythemia vera: results of the JSH-MPN-R18 study.

The presence of a JAK2 V617F or JAK2 exon 12 mutation is one of the three major criteria listed for the diagnosis of polycythemia vera (PV) in the 2017 World Health Organization Classification. However, a nationwide study has not yet been conducted in Japan since the discovery of JAK2 mutations. Therefore, the Japanese Society of Hematology (JSH) retrospectively analyzed the clinical characteristics of 596 Japanese patients with PV diagnosed between April 2005 and March 2018. Among the 473 patients with complete data on JAK2 mutations available, 446 (94.3%) and 10 (2.1%) were positive for the JAK2 V617F and JAK2 exon 12 mutations, respectively. During a median follow-up of 46 months (range: 0-179 months), 47 (7.9%) deaths occurred. The major causes of death were secondary malignancies (23.4%), acute leukemia (12.8%), non-leukemic progressive disease (10.6%) and thrombotic (6.4%) and hemorrhagic complications (6.4%). Thrombotic and hemorrhagic events occurred during the clinical course in 4.0% (n = 24) and 3.5% (n = 21) of patients, respectively. These results show that the international PV prognostic score (age, venous thrombosis and leukocytosis) is applicable to Japanese patients with PV.

Cholecystectomy in a patient with paroxysmal nocturnal haemoglobinuria undergoing ravulizumab maintenance treatment.

A 47-year-old male with paroxysmal nocturnal haemoglobinuria (PNH) controlled with routine ravulizumab administration suffered a massive haemolytic crisis due to choledocholithiasis. Laparoscopic cholecystectomy was performed 6 weeks after a regular ravulizumab infusion. After surgery, the patient presented with anaemia without marked elevation in lactate dehydrogenase and required two blood transfusions. Tumour necrosis factor-α increased more than twofold with reticulocyte suppression after surgery, suggesting the involvement of myelosuppressive cytokines. This case suggests that laparoscopic surgery may be safely performed in patients with PNH receiving ravulizumab maintenance treatment. However, attention should be paid to postoperative anaemia, regardless of breakthrough haemolysis.

Mutated KIT Tyrosine Kinase as a Novel Molecular Target in Acute Myeloid Leukemia.

KIT is a type-III receptor tyrosine kinase that contributes to cell signaling in various cells. Since KIT is activated by overexpression or mutation and plays an important role in the development of some cancers, such as gastrointestinal stromal tumors and mast cell disease, molecular therapies targeting KIT mutations are being developed. In acute myeloid leukemia (AML), genome profiling via next-generation sequencing has shown that several genes that are mutated in patients with AML impact patients' prognosis. Moreover, it was suggested that precision-medicine-based treatment using genomic data will improve treatment outcomes for AML patients. This paper presents (1) previous studies regarding the role of KIT mutations in AML, (2) the data in AML with KIT mutations from the HM-SCREEN-Japan-01 study, a genome profiling study for patients newly diagnosed with AML who are unsuitable for the standard first-line treatment (unfit) or have relapsed/refractory AML, and (3) new therapies targeting KIT mutations, such as tyrosine kinase inhibitors and heat shock protein 90 inhibitors. In this era when genome profiling via next-generation sequencing is becoming more common, KIT mutations are attractive novel molecular targets in AML.