: Blood services ― which are managed in Japan by the Japanese Red Cross Society ― are a key part of the national medical infrastructure. However, doctorsʼ perception of blood services is often limited to a superficial understanding. Although the role of medical check-up doctors in securing safe and high-quality blood is important, there is a continuing shortage of doctors dedicated to blood centers and medical check-up doctors in terms of actual numbers and knowledge of blood services. The “Nara Regimen” consists of two protocols, namely “blood transfusion education and practical training (14 hours) ” at the Blood Transfusion Division of the Nara Medical University Hospital and “practical training (3 hours), including blood donor experience” at the Nara Red Cross Blood Center, as pre-graduate education for medical students of Nara Medical University; and “On-the-Job Training for blood donation work” for 1 week at the Nara Red Cross Blood Center as post-graduate education for clinical residents. In this study, we examined the results of these programs, focusing on the number of participating medical check-up doctors, and clarified the contribution of doctors who completed the “Nara Regimen” to blood donation medical check-ups. Although training for medical students and clinical residents is a substantial burden on blood centers, the “Nara Regimen” of active and continuous education and training will increase doctorsʼ awareness of blood services, and function effectively and efficiently to train and secure medical check-up doctors who will contribute to blood services.

Inhibitor‐development is a serious complication in patients with haemophilia (PwH). Previous studies reported that therapeutic and genetic factors could be associated with these alloantibodies. Relevant clinical features such as genetic‐background and different treatment regimens in Japan remain unclear, however.

Abstract Some genetic and treatment-related factors are risk factors for inhibitor development in patients with hemophilia A (PwHA). However, the genotype distribution of the factor VIII gene (F8) and genetic impact on inhibitor development in Japanese PwHA remain unknown. In 2007, the Japan Hemophilia Inhibitor Study 2 (J-HIS2) was organized to establish a nationwide registry system for hemophiliacs and to elucidate risk factors for inhibitor development, designed for prospective investigation following a retrospective study (J-HIS1) which had already finished. Patients, newly diagnosed after January 2007, were enrolled in J-HIS2 and followed up for inhibitor development and clinical environments since 2008 onward. In the present study, F8 genotypes of PwHA were investigated in the patients recruited from the J-HIS2 cohort as well as those with inhibitor from the J-HIS1 cohort. F8 variants identified in 59 PwHA with inhibitor in J-HIS1 were: 20 intron-22 inversions, 5 intron-1 inversions, 9 large deletions, 4 nonsense, 8 missense, 11 small in/del, and 2 splice-site variants. F8 variants identified in 267 (67 with inhibitor) PwHA in J-HIS2 were: 76(28) intron–22 inversions, 3(2) intron–1 inversion, 1(0) duplication, 8(5) large deletions, 21(7) nonsense, 109(7) missense, 40(11) small in/del, and 9(7) splice-site variants. Forty variants were novel. The cumulative inhibitor incidence rate in the severe group with null changes was 42.4% (95% confidence interval [CI]: 33.7–50.8), higher than that with nonnull changes (15.6% [95%CI: 6.8–27.8]), in J-HIS2. Relative risk for inhibitor development of null changes was 2.89. The spectrum of F8 genotype and genetic impact on inhibitor development in Japanese PwHA were consistent with the previous reports.

The authors would like to correct the errors in the publication of the original article. The correction details are given below.

Background & Aims: Since the first account of the myth of Prometheus, the amazing regenerative capacity of the liver has fascinated researchers because of its enormous medical potential. Liver regeneration is promoted by multiple types of liver cells, including hepatocytes and liver non-parenchymal cells (NPCs), through complex intercellular signaling. However, the mechanism of liver organogenesis, especially the role of adult hepatocytes at ectopic sites, remains unknown. In this study, we demonstrate that hepatocytes alone spurred liver organogenesis to form an organ-sized complex 3D liver that exhibited native liver architecture and functions in the kidneys of mice. Methods: Isolated hepatocytes were transplanted under the kidney capsule of monocrotaline (MCT) and partial hepatectomy (PHx)-treated mice. To determine the origin of NPCs in neo-livers, hepatocytes were transplanted into MCT/PHxtreated green fluorescent protein transgenic mice or wild-type mice transplanted with bone marrow cells isolated from green fluorescent protein-mice. Results: Hepatocytes engrafted at the subrenal space of mice underwent continuous growth in response to a chronic hepatic injury in the native liver. More than 1.5 years later, whole organ-sized liver tissues with greater mass than those of the injured native liver had formed. Most remarkably, we revealed that at least three types of NPCs with similar phenotypic features to the liver NPCs were recruited from the host tissues including bone marrow. The neo-livers in the kidney exhibited liver-specific functions and architectures, including sinusoidal vascular systems, zonal heterogeneity, and emergence of bile duct cells. Furthermore, the neo-livers successfully rescued the mice with lethal liver injury. Conclusion: Our data clearly show that adult hepatocytes play a leading role as organizer cells in liver organogenesis at ectopic sites via NPC recruitment. Lay summary: The role of adult hepatocytes at ectopic locations has not been clarified. In this study, we demonstrated that engrafted hepatocytes in the kidney proliferated, recruited non-parenchymal cells from host tissues including bone marrow, and finally created an organ-sized, complex liver system that exhibited liver-specific architectures and functions. Our results revealed previously undescribed functions of hepatocytes to direct liver organogenesis through non-parenchymal cell recruitment and organize multiple cell types into a complex 3D liver at ectopic sites. Transcript profiling: Microarray data are deposited in GEO (GEO accession: GSE99141). (C) 2017 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Rurioctocog alfa (recombinant factor VIII: Advate®) is available for the control of bleeding among patients with hemophilia A in Japan. To evaluate the perioperative safety and hemostatic efficacy of Advate®, a postmarketing surveillance was conducted in Japanese patients undergoing surgery in a real-world setting. A total of 74 surgical procedures performed in 58 subjects aged 0–75 years, including three females, were studied. A hemostatic efficacy rating of “excellent” or “good” was reported in 73/74 surgical procedures (98.6%). Perioperative bleeding was successfully controlled by Advate® in five subjects with positive FVIII inhibitors (2.4-9.1 BU/mL). Advate® was administered at higher initial bolus doses (114-385 IU/kg) and at higher rates by subsequent initial continuous infusion (8.3-15 IU/kg/hour) in the five subjects with inhibitor than in the subjects without inhibitor (n = 47; mean initial bolus dose: 53.4 IU/kg; subsequent mean initial continuous infusion: 3.8 IU/kg/h). Adverse drug reactions were reported in 7/74 (9.5%) procedures, two of which were the development of de novo FVIII inhibitors. Overall, the perioperative use of Advate® in a real-world setting was found to be safe and effective among Japanese patients with hemophilia A.

The International Immune Tolerance Induction (I-ITI) Study in hemophilia A patients with inhibitors included 16 Japanese patients among a total of 115 test subjects. The results within this group of Japanese patients were 11 cases of I-ITI off-study, three cases of I-ITI on-study, and two cases of tolerance on prophylaxis. There was no significant difference in success rate between the low-dose and high-dose groups (Study I). Successively, independent follow-up survey in Japan was conducted in 14 cases, with consent (Study II). Ten cases were off-study at the end of the I-ITI Study. Of these 10 cases, seven of seven successful cases remained clinical successes at the end of the follow-up study, one partial success became a full success while a second relapsed, and one failure was subsequently evaluated as a partial success. Four cases that were on-study at the end of I-ITI Study were classified as three successes and one failure at the end of the follow-up study. As a result, the status at the end of follow-up study was: 11 ITI successes (78.6 %); one partial success; one failure; and one relapse. Thus, the ITI follow-up study was helpful in providing a long-term prognostic determination of inhibitors.

Since the human factor VIII (FVIII) gene was cloned and expressed in mammalian cells by two independent biotechnology groups in 1984, two full-length, recombinant FVIII (rFVIII) preparations have been developed. Subsequently, second-generation preparations have been produced in which the human serum albumin (HAS) in the final therapeutic material is replaced by nonprotein stabilizers. More recent third-generation products have been developed in which no exogenous bovine and/or human protein is added to either the cell cultures or the final material. The production of rFVIII was commercialized more than 20 years ago, and therapeutic products of this nature have now become the major choice for hemostatic treatment in hemophilia. The safety and efficacy of these recombinant products have been confirmed in thorough preclinical studies. Unresolved differences have been reported between recombinant and plasma-derived products related to the incidence of FVIII inhibitors in both previously untreated patients (PUPs) and previously treated patients (PTPs). In addition, the introduction of recombinant products has facilitated regular prophylaxis as the principal type of therapy especially in pediatric and young adult patients. The outcomes of long-term prophylactic use and pharmacokinetic information are also important aspects to be investigated, therefore, as well as inhibitor development. Recently, novel recombinant FVIII and FIX concentrates with a longer half-life have also been developed. The classic concept of hemostatic treatment for patients with hemophilia may not be entirely appropriate for the new products, and major changes in therapeutic protocols seem likely to be required when these longer acting concentrates, especially modified rFIX, are produced commercially on a larger scale. Simple overall protocols may not be practical in view of the wide variation in specific clinical symptoms and individual physical activity. The relationship between inhibitor development and product type in particular remains controversial, and immunogenicity should be carefully and thoroughly investigated in well-organized protocols when the new FVIII or FIX therapeutic materials become more widely available.

BACKGROUND:We previously reported that a humanized anti-factor IXa/X bispecific antibody, hBS23, mimics the function of FVIII even in the presence of FVIII inhibitors, and has preventive hemostatic activity against bleeding in an animal model of acquired hemophilia A. After further molecular engineering of hBS23, we recently identified an improved humanized bispecific antibody, ACE910, for clinical investigation.OBJECTIVES:To elucidate the in vivo hemostatic potency of ACE910 by examining its effect against ongoing bleeds, and to determine its pharmacokinetic parameters for discussion of its potency for prophylactic use.METHODS:A nonhuman primate model of acquired hemophilia A was established by injecting anti-primate FVIII neutralizing antibody. When bleeds emerged following an artificial bleed-inducing procedure, either ACE910 or recombinant porcine FVIII (rpoFVIII) was intravenously administered. rpoFVIII was additionally administered twice daily on the following 2 days. Bleeding symptoms were monitored for 3 days. A pharmacokinetic study and multiple-dosing simulations of ACE910 were also performed.RESULTS:A single bolus of 1 or 3 mg kg⁻¹ ACE910 showed hemostatic activity comparable to that of 10 U kg⁻¹ (twice daily) rpoFVIII against ongoing bleeds. The determined ACE910 pharmacokinetic parameters included a long half-life (3 weeks) and high subcutaneous bioavailability (nearly 100%). The simulation results based on pharmacokinetic parameters indicated that the above hemostatic level could be maintained with once-weekly subcutaneous administration of ACE910, suggesting the possibility of more effective prophylaxis.CONCLUSIONS:ACE910 may offer an alternative on-demand treatment option for patients with hemophilia A, as well as user-friendly and aggressive routine supplementation.

HaemophiliaVolume 19, Issue 5 p. e301-e304 Letter to the Editor Two haemophilia patients with inhibitors who became ambulatory after physiotherapy under haemostatic cover with bypassing agents M. Kajiwara, Corresponding Author M. Kajiwara Department of Paediatrics, National Hospital Organization Fukui National Hospital, Tsuruga, Japan Correspondence: Masue Kajiwara, Department of Paediatrics, Kano Hospital, 1-2-39 Hananomori, 919-0633 Awara City, Japan. Tel.: +81 0776 73 1001; fax: +81 0776 73 4096; e-mail: [email protected]Search for more papers by this authorM. Shima, M. Shima Department of Paediatrics, Nara Medical University, Kashihara City, JapanSearch for more papers by this authorA. Yoshioka, A. Yoshioka Nara Medical University, Kashihara City, JapanSearch for more papers by this author M. Kajiwara, Corresponding Author M. Kajiwara Department of Paediatrics, National Hospital Organization Fukui National Hospital, Tsuruga, Japan Correspondence: Masue Kajiwara, Department of Paediatrics, Kano Hospital, 1-2-39 Hananomori, 919-0633 Awara City, Japan. Tel.: +81 0776 73 1001; fax: +81 0776 73 4096; e-mail: [email protected]Search for more papers by this authorM. Shima, M. Shima Department of Paediatrics, Nara Medical University, Kashihara City, JapanSearch for more papers by this authorA. Yoshioka, A. Yoshioka Nara Medical University, Kashihara City, JapanSearch for more papers by this author First published: 11 June 2013 https://doi.org/10.1111/hae.12179Citations: 2Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat References 1Blanchette VS, Manco-Johnson MJ. Meeting unmet needs in inhibitor patients. Haemophilia 2010; 16(Suppl. 3): 46–51. 2Hay CR, Brown S, Collins PW, Keeling DM, Liesner R. The diagnosis and management of factor VIII and IX inhibitors: a guideline from the United Kingdom Haemophilia Centre Doctors Organisation. Br J Haematol 2006; 133: 591–605. 3Manco-Johnson MJ, Abshire TC, Shapiro AD et al. Prophylaxis versus episodic treatment to prevent joint disease in boys with severe hemophilia. N Engl J Med 2007; 357: 535–44. 4Caviglia H, Candela M, Galatro G, Neme D, Moretti N, Bianco RP. Elective orthopaedic surgery for haemophilia patients with inhibitors: single centre experience of 40 procedures and review of the literature. Haemophilia 2011; 17: 910–9. 5Young G, Auerswald G, Jimenez-Yuste V et al. When should prophylaxis therapy in inhibitor patients be considered? Haemophilia 2011; 17: e849–57. 6Leissinger C, Gringeri A, Antmen B et al. Anti-inhibitor coagulant complex prophylaxis in hemophilia with inhibitors. N Engl J Med 2011; 365: 1684–92. 7Young G, McDaniel M, Nugent DJ. Prophylactic recombinant factor VIIa in haemophilia patients with inhibitors. Haemophilia 2005; 11: 203–7. 8Konkle BA, Ebbesen LS, Erhardtsen E et al. Randomized, prospective clinical trial of recombinant factor VIIa for secondary prophylaxis in hemophilia patients with inhibitors. J Thromb Haemost 2007; 5: 1904–13. 9Hoots WK, Ebbesen LS, Konkle BA et al. Secondary prophylaxis with recombinant activated factor VII improves health-related quality of life of haemophilia patients with inhibitors. Haemophilia 2008; 14: 466–75. 10Hilgartner MW, Makipernaa A, Dimichele DM. Long-term FEIBA prophylaxis does not prevent progression of existing joint disease. Haemophilia 2003; 9: 261–8. Citing Literature Volume19, Issue5September 2013Pages e301-e304 ReferencesRelatedInformation

インヒビターを保有する先天性血友病患者の出血エピソードにおける遺伝子組換え活性型血液凝固第VII因子製剤（rFVIIa）の10年間の使用成績調査をもとに，その有効性と安全性を解析した．有効性評価が可能な先天性血友病患者142例1,718出血エピソードについて解析した結果，rFVIIa投与から12時間以内に止血効果が認められた割合は65％，最終的に止血効果が認められた割合は88％であった．rFVIIa単独治療群の1回投与量，投与回数，投与日数の中央値はそれぞれ103μg/kg，3.0回，1.0日であった．出血部位は関節内出血が63％と最も多く，次いで筋肉内出血が14％，皮下出血が9％であった．頭蓋内出血，消化管出血，血尿では投与日数が有意に長かった．また安全性については，144例，8,097出血エピソード，全24,778回の投与に対して，4例5件の血栓塞栓性事象が報告された．

Abstract Abstract 42 Background: Hemophilia A is treated by intravenous replacement therapy with factor VIII (FVIII), either on demand to resolve bleeding or as a prophylactic to prevent bleeding. Recently, routine prophylactic treatment is recommended to effectively prevent bleeding and to reduce bleeding-related chronic joint damage. However, the need for frequent intravenous injections of FVIII negatively affects patients' quality of life and their adherence to the routine prophylactic regimen. More importantly, approximately 30% of severe hemophilia A patients develop inhibitory antibodies toward the injected FVIII, rendering the replacement therapy ineffective. To overcome these drawbacks, we generated a bispecific antibody (termed ACE910) against activated factor IX (FIXa) and factor X (FX), which mimics the cofactor function of FVIII. Objectives: The aims of the present study were to examine the FVIII-mimetic cofactor activity of ACE910 in vitro and its hemostatic activity in vivo. Methods: The FVIII-mimetic cofactor activity of ACE910 was evaluated by a thrombin generation assay in human FVIII-deficient plasma as well as by an enzymatic assay using purified coagulation factors. For in vivo studies, an acquired hemophilia A model was established in cynomolgus monkeys by a single intravenous injection of mouse monoclonal anti-FVIII neutralizing antibody, which was cross-reactive to cynomolgus monkey FVIII but not to porcine FVIII. After artificial bleeding had been induced, ACE910 or porcine FVIII was intravenously administered in a single dose or in twice-daily repeated doses, respectively. Bleeding symptoms, including anemia and skin bruising, were monitored for three days. A pharmacokinetic study of ACE910 was also performed with a single intravenous or subcutaneous administration to cynomolgus monkeys. Results: ACE910 concentration-dependently showed FVIII-mimetic cofactor activity in the enzymatic assay and improved thrombin generation parameters in human FVIII-deficient plasma. Intravenous administration of ACE910 (a single dose of 3 mg/kg) significantly reduced the bleeding symptoms in the acquired hemophilia A model of a non-human primate. This hemostatic effect was comparable to twice-daily intravenous administration of porcine FVIII (repeated doses of 10 U/kg). The half-life of ACE910 was approximately three weeks for both single intravenous and subcutaneous administrations. The subcutaneous bioavailability of ACE910 was nearly 100%. Conclusion: The bispecific antibody against FIXa and FX, ACE910, exerted FVIII-mimetic cofactor activity in vitro. Furthermore, a single dose of ACE910 demonstrated hemostatic activity comparable to twice-daily repeated doses of 10 U/kg porcine FVIII in vivo. Moreover, ACE910 exhibited high subcutaneous bioavailability and approximately three-week half-life in a non-human primate. Our bispecific antibody against FIXa and FX is a subcutaneously injectable, long-acting agent that removes the need to consider the induction or presence of FVIII inhibitors and may establish a novel principle for the prophylactic treatment of hemophilia A patients. Disclosures: Muto: Chugai Pharmaceutical Co., Ltd.: Employment. Kitazawa:Chugai Pharmaceutical Co., Ltd.: Employment. Yoshihasi:Chugai Pharmaceutical Co., Ltd.: Employment. Takeda:Chugai Pharmaceutical Co., Ltd.: Employment. Soeda:Chugai Pharmaceutical Co., Ltd.: Employment. Igawa:Chugai Pharmaceutical Co., Ltd.: Employment. Sampei:Chugai Pharmaceutical Co., Ltd.: Employment. Sakamoto:Chugai Pharmaceutical Co., Ltd.: Employment. Okuyama-Nishida:Chugai Pharmaceutical Co., Ltd.: Employment. Saito:Chugai Pharmaceutical Co., Ltd.: Employment. Kawabe:Chugai Pharmaceutical Co., Ltd.: Employment. Shima:Chugai Pharmaceutical Co., Ltd.: Consultancy, Honoraria, Research Funding. Hattori:Chugai Pharmaceutical Co., Ltd.: Employment.

We previously reported that cell-based therapies using isolated hepatocytes including hepatocyte transplantation and liver tissue engineering approaches provide therapeutic benefits to hemophilia. For clinical application of these approaches, it is important to establish an active hepatocyte proliferation system that enables providing a sufficient number of hepatocytes. We also reported that human hepatocytes, which were transplanted into the liver of urokinase-type plasminogen activator transgenic severe combined immunodeficiency (uPA/SCID) mice, were able to proliferate while retaining their ability to produce coagulation factor IX. The objective of this study was to explore the functionalities of other coagulation and anticoagulation factors of the propagated human hepatocytes in uPA/SCID mice. Human hepatocytes were transplanted into the liver of uPA/SCID mice, and the propagation status of human hepatocytes in the mice was monitored by the increase in serum human albumin levels and immunohistochemical evaluation on the liver sections. Using uPA/SCID livers with various stages of human hepatocyte propagation, we analyzed the gene expression levels of coagulation factors (prothrombin, factor VII, factor X, and factor VIII) and anticoagulation factors (protein C and protein S) by real-time polymerase chain reaction (PCR) using human-specific primers. As a result, the total amount of raw messenger RNA expression levels increased in all genes analyzed according to the progress of hepatocyte propagation and proliferation. Except for factor VIII, the gene expression levels of the highly repopulated uPA/SCID mouse livers with human hepatocyte showed higher levels than those of normal human livers, indicating that propagated human hepatocytes in the uPA/SCID system possess full functions to produce most of the coagulation-related factors. The current work demonstrated that human hepatocytes can be propagated in experimental animals while maintaining normal gene expression levels of coagulation-related factors. It could be speculated that the propagated cells serve as a cell source for the treatment of various types of coagulation factor deficiencies.

Individuals with hemophilia A lack the coagulation factor FVIII and are treated with frequent intravenous injections of FVIII agents. However, many individuals develop antibodies to FVIII and can no longer be treated by FVIII injection. Takehisa Kitazawa and his colleagues report the development of a bispecific antibody to FIXa and FX that mimics the function of FVIII. This antibody reduces bleeding in a nonhuman primate model of hemophilia A, is resistant to the inhibitory effects of FVIII-specific antibodies and has a long half-life after subcutaneous injection.

BACKGROUND/AIM:Hemophilia A patients with inhibitors are treated effectively with immune tolerance induction (ITI) therapy. Although anti-idiotypic antibodies may play a certain role in the underlying mechanism, the detailed mechanism by which ITI produces a curative effect remains unknown. The aim of this study was to clarify the immunological aspect of ITI.METHODS:Longitudinal T-cell receptor (TCR) analysis was performed during ITI. TCR variable region alpha-chain and beta-chain repertoires were serially analyzed for peripheral blood mononuclear cells (PBMCs), CD4 T cells, and CD8 T cells from 2 hemophilia inhibitor patients treated with ITI (Patients 1 and 2). Furthermore, to see whether skewing observed in TCR analysis resulted from clonality alterations, T cell clonality was investigated using complementarity-determining region 3 (CDR3) size spectratyping.RESULTS:In the patient 1, inhibitor titer remained to be 19.6 BU/mL for 596 days after ITT commencement, and ITI was unsuccessful. In the patient 2, inhibitor titer disappeared 434 days after ITI commencement, and ITI was successful. In both cases, skewing of TCR variable region alpha/beta-chain repertoires was observed in CD8 T cell subset, whereas not in CD4 T cell subset.CONCLUSION:Alteration of TCR repertoires, especially TCR variable region beta-chain repertoire of CD8 T cells, was distinct between successful and unsuccessful cases, suggesting that immunological response in the early phase affected the ITI outcomes.