DOACs for ProteinS deficiency

Protein S (PS) deficiency is an inherited thrombophilia associated with an increased risk of venous thromboembolism (VTE). In Japan, unfractionated heparin followed by warfarin has been historically applied for the treatment of VTE. Recent evidence showed that direct oral anticoagulants (DOACs) were non-inferior to standard therapy with warfarin, with significantly less bleeding in patients with VTE. However, it is unknown whether DOACs are effective for the treatment of VTE in patients with thrombophilia, including protein S deficiency, due to lack of evidence. Here, we report a case of recurrent venous thrombosis during edoxaban therapy in a patient with protein S deficiency, which was successfully treated using high-dose apixaban therapy

Successful voriconazole treatment of invasive pulmonary aspergillosis in a patient with acute biphenotypic leukemia

A 23-year old woman with acute biphenotypic leukemia (ABL) complained of chest pain with cough, high fever and hemoptysis during induction chemotherapy, although she had been treated with anti-biotics and micafungin. We made a clinical diagnosis of invasive pulmonary aspergillosis (IPA) based on a consolidation in the right upper lung field on a chest radiograph as well as a high level of serum beta-D-glucan (with no evidence of tuberculosis and candidiasis). We changed her treatment from micafungin to voriconazole. Later, we discovered an air-crescent sign by CT scan that supported the diagnosis of IPA. Following voriconazole treatment, clinical symptoms ceased and abnormal chest shadows improved gradually and concurrently with a recovery of neutrophils. IPA must be considered in immunocompromised patients with pulmonary infiltrates who do not respond to broad-spectrum antibiotics. Serological tests and CT findings can aid in early diagnosis of IPA, which, along with treatment for IPA, will improve clinical outcomes.</p

A novel high-throughput (HTP) cloning strategy for site-directed designed chimeragenesis and mutation using the Gateway cloning system

There is an increasing demand for easy, high-throughput (HTP) methods for protein engineering to support advances in the development of structural biology, bioinformatics and drug design. Here, we describe an N- and C-terminal cloning method utilizing Gateway cloning technology that we have adopted for chimeric and mutant genes production as well as domain shuffling. This method involves only three steps: PCR, in vitro recombination and transformation. All three processes consist of simple handling, mixing and incubation steps. We have characterized this novel HTP method on 96 targets with >90% success. Here, we also discuss an N- and C-terminal cloning method for domain shuffling and a combination of mutation and chimeragenesis with two types of plasmid vectors

Polyclonal Immunoglobulin Recovery after Autologous Stem Cell Transplantation Is an Independent Prognostic Factor for Survival Outcome in Patients with Multiple Myeloma

We retrospectively analyzed multiple myeloma (MM) patients who underwent autologous stem cell transplantation (ASCT) without maintenance therapy to assess the impact of recovery of normal immunoglobulin (Ig) on clinical outcomes. The recovery of polyclonal Ig was defined as normalization of all values of serum IgG, IgA, and IgM 1 year after ASCT. Among 50 patients, 26 patients showed polyclonal Ig recovery; 14 patients were in ≥complete response (CR) and 12 remained in non-CR after ASCT. The patients with Ig recovery exhibited a significantly better progression-free survival (PFS, median, 46.8 vs. 26.7 months, p = 0.0071) and overall survival (OS, median, not reached vs. 65.3 months, p < 0.00001) compared with those without Ig recovery. The survival benefits of Ig recovery were similarly observed in ≥CR patients (median OS, not reached vs. 80.5 months, p = 0.061) and non-CR patients (median OS, not reached vs. 53.2 months, p = 0.00016). Multivariate analysis revealed that non-CR and not all Ig recovery were independent prognostic factors for PFS (HR, 4.284, 95%CI (1.868–9.826), p = 0.00059; and HR, 2.804, 95%CI (1.334–5.896), p = 0.0065, respectively) and also for OS (HR, 8.245, 95%CI (1.528–44.47), p = 0.014; and HR, 36.55, 95%CI (3.942–338.8), p = 0.0015, respectively). Therefore, in addition to the depth of response, the recovery of polyclonal Ig after ASCT is a useful indicator especially for long-term outcome and might be considered to prevent overtreatment with maintenance therapy in transplanted patients with MM

Cryptosporidiosis in a transplant recipient with severe intractable diarrhea : Detection of Cryptosporidium oocysts by intestinal biopsies

Disseminated Cryptosporidium infection results in manifestations similar to those of graft‐versus‐host disease (GVHD), which hampers the detection of Cryptosporidium infection after allogeneic hematopoietic stem cell transplantation. Surveillance of oocysts on the surface of intestinal epithelial cells is needed for early and appropriate detection of Cryptosporidium infection in transplant recipients on immunosuppressants with severe intractable diarrhea. We present the first case of Cryptosporidium meleagridis infection in Japan after allogeneic cord blood transplantation

Multiple myeloma with high adenosine deaminase expression

A 50-year-old man with immunoglobulin A type multiple myeloma (MM) was referred to our hospital after bortezomib therapy. He had high alkaline phosphatase and lactate dehydrogenase levels. Computed tomography showed osteolytic and osteoblastic bone lesions. Response to salvage chemotherapy was temporary, and he developed a right pleural effusion with high adenosine deaminase (ADA) levels. He died from bleeding associated with a pelvic bone fracture 9 months later. ADA mRNA expression and ADA secretion of the MM cells from the patient were higher than those from myeloma cell lines tested. Clinical relevance of high ADA expression in MM cells is warranted

Busulfan for lymphoma with CNS involvement

The prognosis of relapsed or refractory lymphoma with central nervous system (CNS) involvement remains poor because of the lack of anticancer drugs with sufficient CNS penetration. [Case 1] A 65-year-old man was diagnosed with Stage IV mantle cell lymphoma. After two courses of chemotherapy and autologous hematopoietic stem cell (HSC) collection, urinary retention with fever developed. Cerebrospinal fluid analysis revealed leptomeningeal involvement, which was refractory to high-dose methotrexate therapy. Autologous peripheral blood stem cell transplantation (ASCT) was performed, followed by intravenous busulfan (ivBU), cyclophosphamide, and etoposide ; thereafter, no relapse has been detected for over six years. [Case 2] A 40-year-old woman with right lower hemiplegia was diagnosed with primary CNS lymphoma. Although four courses of high-dose methotrexate therapy were administered, the cerebral tumor increased in size. HSCs were collected after methotrexate therapy, and ASCT was performed in addition to conditioning using ivBU, cyclophosphamide, and etoposide, followed by whole-brain and local boost irradiation. She achieved complete remission, but relapsed two years after ASCT. High-dose ivBU-containing conditioning regimens with ASCT may be useful for refractory B-cell lymphoma with CNS involvement

ドウケイ コツズイ イショク ガ チョコウ シタ サイジュウショウ サイセイ フリョウセイ ヒンケツ ノ 1レイ

Aplastic anemia is a bone marrow failure caused by severely curtailed hematopoietic stem cells （HSCs） and dysregulation of ambient immune cells. Immumo-suppressive conditioning followed by allogeneic HSC transplantation is currently a mainstay in treatment for patients at a younger age or those refractory to conventional immunosuppressive remedies. Syngeneic HSC transplantation appears to be promising, but has been very rarely performed ; therefore, its impact on a long-term outcome as well as the best preparative measures for HSC engraftment and immune amelioration are still largely unknown. Here, we reported a successful and beneficial syngeneic HSC transplantation for a refractory case with very severe aplastic anemia. A 30-year-old female presented high fever after tooth extraction, and was diagnosed with very severe aplastic anemia. Cyclosporine and anti-thymocyte globulin were initiated, but showed no hematological effects. After obtaining an informed consent, she underwent bone marrow transplantation from a genetically identical twin following an non-myeloablative conditioning regimen consisted of cyclophosphamide （750mg/m２, 4 days）, fludarabine phosphate （25mg/m２, 4 days）. Cyclosporine was given for acute GvHD prophylaxis. Her neutrophils recovered over 500/μl on 12 days after the transplantation, and her blood counts have been maintained in a normal range over 7 years thereafter. Although a fate of HSCs from a genetically identical twin and an immune response of ambient cells in the bone marrow in recipients remains largely unknown, from the present case and previously reported cases, we dare to recommend immunoablative conditioning and acute GvHD prophylaxis in syngeneic HSC transplantation for a refractory case with aplastic anemia for better engraftment and sustained onward recovery of hematopoiesis

臍帯血移植が奏効した肝脾型T細胞リンパ腫

Cord blood for transplant is collected from the umbilical cord and donated cord blood is tested, frozen and stored for future use. Cord blood stem cell transplantation（CBT）do not have to be as closely matched as bone marrow or peripheral blood stem cell transplantations, therefore we are able to perform CBT for refractory patients at optimal timing. Here, we report a ３３ years old woman with refractory hepatosplenic T-cell lymphoma（HSTL）who achieved complete response ; CR after unrelated CBT. She complained of fever and abdominal pain, she was diagnosed with HSTL. She was refractory to several chemotherapy regimens, we planned to allogenic transplantation for her. However, she had no HLA-matched sibling donors and we were not able to find favorable unrelated donors from Japan Marrow Donor Program. Therefore, we decided to perform CBT for her. We used fludarabine（１８０mg/m2）, busulfan（１２．８mg/kg）and melphalan（８０mg/m2） as conditioning regimens and we chose tacrolimus and MMF for graft versus host disease（GVHD） prophylaxis. On day ２２ after transplantation, her neutrophil count engrafted, she suffered from acute GVHD（skin, gastrointestinal tract, grade２）, she was improved by medical treatment. She achieved CR her disease status maintained over two years after CBT. HSTL is often refractory to chemotherapy, the clinical efficacy of hematopoietic stem cell transplantation may be expected. Our case suggests that CBT may be effective and feasible option for refractory HSTL who has no favorable HLA-matched donors

Novel antimyeloma therapeutic option with inhibition of the HDAC1-IRF4 axis and PIM kinase

Multiple myeloma (MM) preferentially expands and acquires drug resistance in the bone marrow (BM). We herein examined the role of histone deacetylase 1 (HDAC1) in the constitutive activation of the master transcription factor IRF4 and the prosurvival mediator PIM2 kinase in MM cells. The knockdown or inhibition of HDAC1 by the class I HDAC inhibitor MS-275 reduced the basal expression of IRF4 and PIM2 in MM cells. Mechanistically, the inhibition of HDAC1 decreased IRF4 transcription through histone hyperacetylation and inhibiting the recruitment of RNA polymerase II at the IRF4 locus, thereby reducing IRF4-targeting genes, including PIM2. In addition to the transcriptional regulation of PIM2 by the HDAC1-IRF4 axis, PIM2 was markedly upregulated by external stimuli from BM stromal cells and interleukin-6 (IL-6). Upregulated PIM2 contributed to the attenuation of the cytotoxic effects of MS-275. Class I HDAC and PIM kinase inhibitors cooperatively suppressed MM cell growth in the presence of IL-6 and in vivo. Therefore, the present results demonstrate the potential of the simultaneous targeting of the intrinsic HDAC1-IRF4 axis plus externally activated PIM2 as an efficient therapeutic option for MM fostered in the BM