Germline Variants and Characteristic Features of Hereditary Hematological Malignancy Syndrome.

Due to the proliferation of genetic testing, pathogenic germline variants predisposing to hereditary hematological malignancy syndrome (HHMS) have been identified in an increasing number of genes. Consequently, the field of HHMS is gaining recognition among clinicians and scientists worldwide. Patients with germline genetic abnormalities often have poor outcomes and are candidates for allogeneic hematopoietic stem cell transplantation (HSCT). However, HSCT using blood from a related donor should be carefully considered because of the risk that the patient may inherit a pathogenic variant. At present, we now face the challenge of incorporating these advances into clinical practice for patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) and optimizing the management and surveillance of patients and asymptomatic carriers, with the limitation that evidence-based guidelines are often inadequate. The 2016 revision of the WHO classification added a new section on myeloid malignant neoplasms, including MDS and AML with germline predisposition. The main syndromes can be classified into three groups. Those without pre-existing disease or organ dysfunction; DDX41, TP53, CEBPA, those with pre-existing platelet disorders; ANKRD26, ETV6, RUNX1, and those with other organ dysfunctions; SAMD9/SAMD9L, GATA2, and inherited bone marrow failure syndromes. In this review, we will outline the role of the genes involved in HHMS in order to clarify our understanding of HHMS.

[Expectations and challenges for clinical application of cancer genome panels in acute myeloid leukemia].

The blood cancer field has played a pioneering role in advancing precision medicine, with milestones such as development of ABL1 inhibitors for chronic myeloid leukemia. The significance of gene mutation information in AML treatment has increased, evident in classifications and guidelines from organizations such as WHO and ELN. This article examines the anticipated roles of cancer genome panels (CGPs) in AML treatment from three perspectives: diagnosis, risk stratification, and treatment selection. Use of CGPs enables more accurate diagnosis and risk stratification. In treatment selection, CGPs not only complements but also substitutes existing companion diagnostics, and is expected to be a crucial information source for future drug adoption and investigation of tumor-agnostic therapies. However, various challenges remain to be addressed, including the purpose and timing of CGPs, the time required for the tests, and how to utilize expert panels.

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.

Patients with B-cell malignancies experience reduced antibody responses with class switching defect following BNT162b2 SARS-CoV-2 vaccination.

Vaccines having aided in escaping the majority of the population from immunological naïvety, our strategies are now shifting towards an increased focus on identifying and protecting the extremely vulnerable. We here describe the results of testing 12 patients, those with lymphoid malignancies having been targeted their B-cells for therapy with rituximab-containing regimens or a Bruton tyrosine kinase inhibitor, for anti-SARS-CoV-2 spike antibodies after receiving the BNT162b2 mRNA vaccine doses. The interval from last dosing of B-cell depletion therapy to SARS-CoV-2 vaccination was at median 5.3 (range 3.1-6.6) months. Using the 'seroprotection' threshold of 775 [BAU/mL] for the anti-spike antibody titer, our finding points out the crucial unresponsiveness of the targeted population with 0/12 (0%) achieving 'seroprotection'. Although IgG seroconversion was observed in 4/12 (33%), supporting the overall benefit of vaccination, the figures still point out a potential need for optimization of practice. IgA was further less responsive (unsuccessful 'seroconversion' in 11/12 (92%)), implicating an underlying class switch defect. Those with depletion on B-cells are caught at a dilemma between, being too early and too late on receiving SARS-CoV-2 vaccines. They wish to get over their immunological naïvety at the earliest, while, in order to assure quality immune memory, are also required to hold the patience for their B-cells to repopulate. Although it remains an issue whether intensified vaccine schedules and/or regimens will lead to stronger immunogenicity or more effective boosters for non-responders, we shall take advantage of every increasing evidence in order to optimize current options.

The Immuno-Oncology and Genomic Aspects of DNA-Hypomethylating Therapeutics in Acute Myeloid Leukemia.

Hypomethylating agents (HMAs) have been used for decades in the treatment of hematologic neoplasms, and now, have gathered attention again in terms of their combination with potent molecular-targeted agents such as a BCL-6 inhibitor venetoclax and an IDH1 inhibitor ivosidenib, as well as a novel immune-checkpoint inhibitor (anit-CD47 antibody) megrolimab. Several studies have shown that leukemic cells have a distinct immunological microenvironment, which is at least partially due to genetic alterations such as the TP53 mutation and epigenetic dysregulation. HMAs possibly improve intrinsic anti-leukemic immunity and sensitivity to immune therapies such as PD-1/PD-L1 inhibitors and anti-CD47 agents. This review describes the immuno-oncological backgrounds of the leukemic microenvironment and the therapeutic mechanisms of HMAs, as well as current clinical trials of HMAs and/or venetoclax-based combination therapies.

Emerging Targeted Therapy for Specific Genomic Abnormalities in Acute Myeloid Leukemia.

We describe recent updates of existing molecular-targeting agents and emerging novel gene-specific strategies. FLT3 and IDH inhibitors are being tested in combination with conventional chemotherapy for both medically fit patients and patients who are ineligible for intensive therapy. FLT3 inhibitors combined with non-cytotoxic agents, such as BCL-2 inhibitors, have potential therapeutic applicability. The menin-MLL complex pathway is an emerging therapeutic target. The pathway accounts for the leukemogenesis in AML with MLL-rearrangement, NPM1 mutation, and NUP98 fusion genes. Potent menin-MLL inhibitors have demonstrated promising anti-leukemic effects in preclinical studies. The downstream signaling molecule SYK represents an additional target. However, the TP53 mutation continues to remain a challenge. While the p53 stabilizer APR-246 in combination with azacitidine failed to show superiority compared to azacitidine monotherapy in a phase 3 trial, next-generation p53 stabilizers are now under development. Among a number of non-canonical approaches to TP53-mutated AML, the anti-CD47 antibody magrolimab in combination with azacitidine showed promising results in a phase 1b trial. Further, the efficacy was somewhat better in patients with the TP53 mutation. Although clinical evidence has not been accumulated sufficiently, targeting activating KIT mutations and RAS pathway-related molecules can be a future therapeutic strategy.

Molecular Classification and Overcoming Therapy Resistance for Acute Myeloid Leukemia with Adverse Genetic Factors.

The European LeukemiaNet (ELN) criteria define the adverse genetic factors of acute myeloid leukemia (AML). AML with adverse genetic factors uniformly shows resistance to standard chemotherapy and is associated with poor prognosis. Here, we focus on the biological background and real-world etiology of these adverse genetic factors and then describe a strategy to overcome the clinical disadvantages in terms of targeting pivotal molecular mechanisms. Different adverse genetic factors often rely on common pathways. KMT2A rearrangement, DEK-NUP214 fusion, and NPM1 mutation are associated with the upregulation of HOX genes. The dominant tyrosine kinase activity of the mutant FLT3 or BCR-ABL1 fusion proteins is transduced by the AKT-mTOR, MAPK-ERK, and STAT5 pathways. Concurrent mutations of ASXL1 and RUNX1 are associated with activated AKT. Both TP53 mutation and mis-expressed MECOM are related to impaired apoptosis. Clinical data suggest that adverse genetic factors can be found in at least one in eight AML patients and appear to accumulate in relapsed/refractory cases. TP53 mutation is associated with particularly poor prognosis. Molecular-targeted therapies focusing on specific genomic abnormalities, such as FLT3, KMT2A, and TP53, have been developed and have demonstrated promising results.

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.

[Successful treatment with gilteritinib for relapsed acute myeloid leukemia with FLT3-N676K mutation].

The patient was a 68-year-old woman, diagnosed with acute myelomonocytic leukemia with normal karyotype and FLT3-ITD-negative status in May 2019. She had achieved complete remission (CR) after "7+3" intensive induction chemotherapy and maintained CR by consolidation chemotherapy. However, she relapsed with swelling of the lips and gums in January 2020. She did not achieve CR by salvage chemotherapy with cytarabine-aclarubicin-G-CSF regimen. Comprehensive genomic analysis of leukemic cells revealed the presence of FLT3-N676K mutation, which was undetectable by companion diagnostics at the time. Complete remission with incomplete count recovery was obtained on day 28 after initiation of gilteritinib monotherapy, and the lip and gum swelling improved rapidly. However, she relapsed on day 106 after gilteritinib administration, and gilteritinib was discontinued. Genomic analysis at recurrence revealed NRAS mutation for the first time. Finally, the patient died of the uncontrolled primary disease. This is a case in which comprehensive gene mutation analysis was useful in determining a treatment strategy.

Emerging Immunotherapy for Acute Myeloid Leukemia.

Several immune checkpoint molecules and immune targets in leukemic cells have been investigated. Recent studies have suggested the potential clinical benefits of immuno-oncology (IO) therapy against acute myeloid leukemia (AML), especially targeting CD33, CD123, and CLL-1, as well as immune checkpoint inhibitors (e.g., anti-PD (programmed cell death)-1 and anti-CTLA4 (cytotoxic T-lymphocyte-associated protein 4) antibodies) with or without conventional chemotherapy. Early-phase clinical trials of chimeric antigen receptor (CAR)-T or natural killer (NK) cells for relapsed/refractory AML showed complete remission (CR) or marked reduction of marrow blasts in a few enrolled patients. Bi-/tri-specific antibodies (e.g., bispecific T-cell engager (BiTE) and dual-affinity retargeting (DART)) exhibited 11-67% CR rates with 13-78% risk of cytokine-releasing syndrome (CRS). Conventional chemotherapy in combination with anti-PD-1/anti-CTLA4 antibody for relapsed/refractory AML showed 10-36% CR rates with 7-24 month-long median survival. The current advantages of IO therapy in the field of AML are summarized herein. However, although cancer vaccination should be included in the concept of IO therapy, it is not mentioned in this review because of the paucity of relevant evidence.

Immune-Checkpoint Blockade Therapy in Lymphoma.

Tumor cells use immune-checkpoint pathways to evade the host immune system and suppress immune cell function. These cells express programmed cell-death protein 1 ligand 1 (PD-L1)/PD-L2, which bind to the programmed cell-death protein 1 (PD-1) present on cytotoxic T cells, trigger inhibitory signaling, and reduce cytotoxicity and T-cell exhaustion. Immune-checkpoint blockade can inhibit this signal and may serve as an effective therapeutic strategy in patients with solid tumors. Several trials have been conducted on immune-checkpoint inhibitor therapy in patients with malignant lymphoma and their efficacy has been reported. For example, in Hodgkin lymphoma, immune-checkpoint blockade has resulted in response rates of 65% to 75%. However, in non-Hodgkin lymphoma, the response rate to immune-checkpoint blockade was lower. In this review, we evaluate the biology of immune-checkpoint inhibition and the current data on its efficacy in malignant lymphoma, and identify the cases in which the treatment was more effective.

Epstein-Barr Virus-Positive Pyothorax-Associated Lymphoma Arising from a Posttraumatic Empyema.

Pyothorax-associated lymphoma (PAL) develops from a pyothorax caused by an artificial pneumothorax created during the treatment of pulmonary tuberculosis or tuberculous pleuritis. We report the first case of Epstein-Barr virus (EBV)-positive PAL arising from a posttraumatic empyema. A 75-year-old woman with chronic posttraumatic empyema presented with a tumor, which was connected to the wall of a pyothorax in the right thoracic cavity. She had a history of trauma to the right chest, which had occurred at the age of 45 years and had caused the chronic posttraumatic empyema. Pathological features of the resected tumor were conclusive for a diagnosis of EBV-positive PAL. Although neither postoperative chemotherapy nor radiotherapy was performed, remission was maintained for 3 years until recurrence in the liver. Combination chemotherapy led to complete remission, and 9 years after the initial diagnosis of PAL, the patient is still alive. An intriguing finding is the phenotypic alteration during the disease course. Although the primary tumor was negative for CD20 and CD3, the recurrent tumor expressed both of these molecules. We discuss this case of PAL, which was not a complication of lung tuberculosis, and the aberrant chronological phenotypic change observed in the lymphoma cells, and compare it with a usual case of PAL.

Advancements in minimal residual disease detection: a practical approach using single-cell droplet PCR for comprehensive monitoring in hematological malignancy.

The identification of chromosomal abnormalities accompanied by copy number alterations is important for understanding tumor characteristics. Testing methodologies for copy number abnormality have limited sensitivity, resulting in their use only for the sample provided at the time of diagnosis or recurrence of malignancy, but not for the monitoring of minimal residual disease (MRD) during and after therapy. We developped the "DimShift" technology which enable to measure the copy number of target gene/chromosome in each cell, which is given by the single cell droplet PCR. Qualitative result of DimShift given by peripheral blood was perfectly concordant with that of bone marrow. These findings and performances are promising to be the new methodology for MRD detection in malignant diseases utilizing bone marrow as well as peripheral blood.

Effect of recombinant human soluble thrombomodulin on clinical outcomes of patients with coagulopathy after hematopoietic stem cell transplantation.

From 2001 to 2012, 71 individuals with hematological diseases received HSCT in our institution. Of these, 41 developed disseminated intravascular coagulation (DIC) in association with various underlying conditions. The patients who developed DIC after 2008 (n = 23) were treated by recombinant human soluble thrombomodulin (rTM), and the others (n = 11) were treated by either heparin and/or antithrombin III concentrate. Seven patients did not receive any anticoagulant therapy. Of note, treatment for coagulopathy by rTM significantly improved clinical outcomes of patients at day 100 and dramatically prolonged their overall survival (P = 0.044). Taken together, rTM is useful to improve clinical outcomes of transplant recipients with coagulopathy.

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.

FLT3-targeted treatment for acute myeloid leukemia.

Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene are detected in approximately 30% of acute myeloid leukemia (AML). The high frequency of FLT3 mutations, along with their adverse effect on prognosis, makes FLT3 a promising therapeutic target, and has spurred development of FLT3 inhibitors. First-generation inhibitors, including midostaurin and sorafenib, lack specificity for FLT3 and act on multiple kinases, whereas second-generation inhibitors, including gilteritinib, and quizartinib, are highly specific to FLT3 and are more potent than first-generation inhibitors. Several FLT3 inhibitors have recently gained regulatory approval worldwide, and several others are under development. The advent of FLT3 inhibitors has changed the standard treatment for FLT3-mutated AML in the frontline and relapsed/refractory settings and contributed to improved outcomes for this formidable AML subtype. However, numerous unresolved issues remain owing to rapid changes in practice. These include identification of optimum FLT3 inhibitors and combination therapies, the role of maintenance therapy, and the indication for allogeneic hematopoietic cell transplantation. Furthermore, strategies to overcome resistance to FLT3 inhibitors must be pursued. Results of ongoing and future studies will improve our ability to use FLT3 inhibitors more effectively, which should provide significant benefits to a wider range of patients.

Molecular-Targeted Therapy for Tumor-Agnostic Mutations in Acute Myeloid Leukemia.

Comprehensive genomic profiling examinations (CGPs) have recently been developed, and a variety of tumor-agnostic mutations have been detected, leading to the development of new molecular-targetable therapies across solid tumors. In addition, the elucidation of hereditary tumors, such as breast and ovarian cancer, has pioneered a new age marked by the development of new treatments and lifetime management strategies required for patients with potential or presented hereditary cancers. In acute myeloid leukemia (AML), however, few tumor-agnostic or hereditary mutations have been the focus of investigation, with associated molecular-targeted therapies remaining poorly developed. We focused on representative tumor-agnostic mutations such as the TP53, KIT, KRAS, BRCA1, ATM, JAK2, NTRK3, FGFR3 and EGFR genes, referring to a CGP study conducted in Japan, and we considered the possibility of developing molecular-targeted therapies for AML with tumor-agnostic mutations. We summarized the frequency, the prognosis, the structure and the function of these mutations as well as the current treatment strategies in solid tumors, revealed the genetical relationships between solid tumors and AML and developed tumor-agnostic molecular-targeted therapies and lifetime management strategies in AML.

Mechanisms Underlying Resistance to FLT3 Inhibitors in Acute Myeloid Leukemia.

FLT3-ITD and FLT3-TKD mutations were observed in approximately 20 and 10% of acute myeloid leukemia (AML) cases, respectively. FLT3 inhibitors such as midostaurin, gilteritinib and quizartinib show excellent response rates in patients with FLT3-mutated AML, but its duration of response may not be sufficient yet. The majority of cases gain secondary resistance either by on-target and off-target abnormalities. On-target mutations (i.e., FLT3-TKD) such as D835Y keep the TK domain in its active form, abrogating pharmacodynamics of type II FLT3 inhibitors (e.g., midostaurin and quizartinib). Second generation type I inhibitors such as gilteritinib are consistently active against FLT3-TKD as well as FLT3-ITD. However, a "gatekeeper" mutation F691L shows universal resistance to all currently available FLT3 inhibitors. Off-target abnormalities are consisted with a variety of somatic mutations such as NRAS, AXL and PIM1 that bypass or reinforce FLT3 signaling. Off-target mutations can occur just in the primary FLT3-mutated clone or be gained by the evolution of other clones. A small number of cases show primary resistance by an FL-dependent, FGF2-dependent, and stromal CYP3A4-mediated manner. To overcome these mechanisms, the development of novel agents such as covalently-coupling FLT3 inhibitor FF-10101 and the investigation of combination therapy with different class agents are now ongoing. Along with novel agents, gene sequencing may improve clinical approaches by detecting additional targetable mutations and determining individual patterns of clonal evolution.

[Geriatric syndrome: slightly reduced visual and hearing impairments reduce activities daily living (ADL) and quality of life (QOL) in the community-dwelling elderly].

Although it is well-known that moderate and severe visual and/or hearing impairments in elderly persons reduce their activities of daily living (ADL) and their quality of life (QOL), most elderly people, their caregivers and even nurses/doctors do not care about those disturbances considering them as normal aging. We studied 1,874 community-dwelling elderly (813 men, 1,061 women, mean age; 76+/-9 yrs.) and demonstrated that apparently healthy older persons with slightly reduced function clarified by self-reported questionnaires do not only have lower scores of ADL and VAS (QOL), but also higher score of GDS 15, that is, they have a more depressive state, compared to those without visual and/or hearing impairments. Therefore, because visual and/or hearing functional impairments in the elderly, even if slight, affect their ADL impairments, QOL and mood, we should assess whether the older persons have visual/hearing disturbance(s) in functional screening and should give them some kinds of assistance to succeed in achieving on enjoyable elderly life.

In vitro studies on the role of recombinant human soluble thrombomodulin in the context of retinoic acid mediated APL differentiation syndrome.

Recombinant human soluble thrombomodulin (rTM) is a newly developed anti-coagulant approved for treatment of disseminated intravascular coagulation (DIC) in Japan. rTM exerts anti-inflammatory and cytoprotective functions via its lectin-like and epidermal growth factor-like domains, respectively. In this study, we retrospectively reviewed the treatment of 21 consecutive patients with coagulopathy, complicated by acute promyelocytic leukemia (APL), with all-trans retinoic acid (ATRA) with or without combination with rTM. Surprisingly, none of the 14 rTM-treated patients developed retinoic acid (RA)-related differentiation syndrome (DS). The co-culture of vascular endothelial cell-derived EA.hy926 and APL-derived NB4 cells in the presence of RA increased production of tumor necrosis factor alpha (TNF-α) in culture media, in parallel with activation of p38 mitogen-activated protein kinase and increased levels of intracellular adhesion molecule 1 (ICAM1) in EA.hy926 cells. This was also associated with increased levels of the phosphorylated forms of VE-cadherin and enhanced vascular permeability of EA.hy926 monolayers. Importantly, addition of rTM to this co-culture system inhibited the RA-induced phosphorylation of p38 and VE-cadherin and decreased ICAM1 and vascular permeability in EA.hy926 cells, without a decrease inthe levels of TNF-α. Taken together, use of rTM may be a promising treatment strategy to prevent DS in APL patients who receive ATRA.