Genetic alterations of Ikaros family zinc finger protein 1 (IKZF1), point mutations in Janus kinase 2 (JAK2), and overexpression of cytokine receptor-like factor 2 (CRLF2) were recently reported to be associated with poor outcomes in pediatric B-cell precursor (BCP)-ALL. Herein, we conducted genetic analyses of IKZF1 deletion, point mutation of JAK2 exon 16, 17, and 21, CRLF2 expression, the presence of P2RY8-CRLF2 fusion and F232C mutation in CRLF2 in 202 pediatric BCP-ALL patients newly diagnosed and registered in Japan Childhood Leukemia Study ALL02 protocol to find out if alterations in these genes are determinants of poor outcome. All patients showed good response to initial prednisolone (PSL) treatment. Ph⁺, infantile, and Down syndrome-associated ALL were excluded. Deletion of IKZF1 occurred in 19/202 patients (9.4%) and CRLF2 overexpression occurred in 16/107 (15.0%), which are similar to previous reports. Patients with IKZF1 deletion had reduced event-free survival (EFS) and overall survival (OS) compared to those in patients without IKZF1 deletion (5-year EFS, 62.7% vs. 88.8%, 5-year OS, 71.8% vs. 90.2%). Our data also showed significantly inferior 5-year EFS (48.6% vs. 84.7%, log rank P = 0.0003) and 5-year OS (62.3% vs. 85.4%, log rank P = 0.009) in NCI-HR patients (n = 97). JAK2 mutations and P2RY8-CRLF2 fusion were rarely detected. IKZF1 deletion was identified as adverse prognostic factor even in pediatric BCP-ALL in NCI-HR showing good response to PSL.

Cysteine can be synthesized by tRNA-dependent mechanism using a two-step indirect pathway, where O-phosphoseryl-tRNA synthetase (SepRS) catalyzes the ligation of a mismatching O-phosphoserine (Sep) to tRNACys followed by the conversion of tRNA-bounded Sep into cysteine by Sep-tRNA:Cys-tRNA synthase (SepCysS). In ancestral methanogens, a third protein SepCysE forms a bridge between the two enzymes to create a ternary complex named the transsulfursome. By combination of X-ray crystallography, SAXS and EM, together with biochemical evidences, here we show that the three domains of SepCysE each bind SepRS, SepCysS, and tRNACys, respectively, which mediates the dynamic architecture of the transsulfursome and thus enables a global long-range channeling of tRNACys between SepRS and SepCysS distant active sites. This channeling mechanism could facilitate the consecutive reactions of the two-step indirect pathway of Cys-tRNACys synthesis (tRNA-dependent cysteine biosynthesis) to prevent challenge of translational fidelity, and may reflect the mechanism that cysteine was originally added into genetic code.

Photosynthetic light-harvesting complexes (LHCs) play a pivotal role in collecting solar energy for photochemical reactions in photosynthesis. One of the major LHCs are fucoxanthin chlorophyll a/c-binding proteins (FCPs) present in diatoms, a group of organisms having important contribution to the global carbon cycle. Here, we report a 2.40-Å resolution structure of the diatom photosystem I (PSI)-FCPI supercomplex by cryo-electron microscopy. The supercomplex is composed of 16 different FCPI subunits surrounding a monomeric PSI core. Each FCPI subunit showed different protein structures with different pigment contents and binding sites, and they form a complicated pigment-protein network together with the PSI core to harvest and transfer the light energy efficiently. In addition, two unique, previously unidentified subunits were found in the PSI core. The structure provides numerous insights into not only the light-harvesting strategy in diatom PSI-FCPI but also evolutionary dynamics of light harvesters among oxyphototrophs.

This study was a second multicenter trial on childhood ALL by the Japan Childhood Leukemia Study Group (JACLS) to improve outcomes in non-T ALL. Between April 2002 and March 2008, 1138 children with non-T ALL were enrolled in the JACLS ALL-02 trial. Patients were stratified into three groups using age, white blood cell count, unfavorable genetic abnormalities, and treatment response: standard risk (SR), high risk (HR), and extremely high risk (ER). Prophylactic cranial radiation therapy (PCRT) was abolished except for CNS leukemia. Four-year event-free survival (4yr-EFS) and 4-year overall survival (4yr-OS) rates for all patients were 85.4% ± 1.1% and 91.2% ± 0.9%, respectively. Risk-adjusted therapy resulted in 4yr-EFS rates of 90.4% ± 1.4% for SR, 84.9% ± 1.6% for HR, and 66.5% ± 4.0% for ER. Based on NCI risk classification, 4yr-EFS rates were 88.2% in NCI-SR and 76.4% in NCI-HR patients, respectively. Compared to previous trial ALL-97, 4yr-EFS of NCI-SR patients was significantly improved (88.2% vs 81.2%, log rank p = 0.0004). The 4-year cumulative incidence of isolated (0.9%) and total (1.5%) CNS relapse were significantly lower than those reported previously. In conclusion, improved EFS in NCI-SR patients and abolish of PCRT was achieved in ALL-02.

This multicenter, open-label, phase I study assessed the safety and antitumor activity of acalabrutinib in Japanese patients with relapsed/refractory (r/r) B-cell malignancies. Parts 1 (dose confirmation) and 2 (dose expansion) of this three-part study are reported. Treatment was a single dose of 100 mg acalabrutinib (day 1), followed by a washout period and then twice daily 100 mg acalabrutinib in part 1, or twice daily 100 mg acalabrutinib in part 2. Patients from parts 1 and 2 with r/r chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL), and r/r mantle cell lymphoma (MCL) were assessed as r/r CLL/SLL and r/r MCL cohorts, respectively. Twenty-five patients received treatment (part 1, n = 6). Median age was 71.0 years. Nine (one patient from part 1) and 13 (two patients from part 1) patients were included in the r/r CLL/SLL and r/r MCL cohorts, respectively. Treatment-related adverse events (AEs) occurred in 88% of patients (grade ≥3, 36%); the most common were headache (28%) and purpura (24%), both grade 1/2. No AEs resulted in treatment discontinuation or death. Median duration of treatment was 31, 20, and 7 months for part 1, r/r CLL/SLL cohort, and r/r MCL cohort, respectively. Overall response rate (ORR) was 89% and 62% for the r/r CLL/SLL and r/r MCL cohorts, respectively. The median progression-free survival (PFS) was not reached for the r/r CLL/SLL cohort and was 7 months for the r/r MCL cohort. Acalabrutinib (100 mg twice daily) was generally safe and well-tolerated in adult Japanese patients with B-cell malignancies.

To assess the efficacy and safety of upadacitinib (UPA), an oral Janus kinase inhibitor, as monotherapy or in combination with non-biologic DMARDs (nbDMARDs) in patients with PsA.

We identified neutralizing monoclonal antibodies against severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) variants (including Omicron variants BA.5 and BA.2.75) from individuals who received two doses of mRNA vaccination after they had been infected with the D614G virus. We named them MO1, MO2, and MO3. Among them, MO1 showed particularly high neutralizing activity against authentic variants: D614G, Delta, BA.1, BA.1.1, BA.2, BA.2.75, and BA.5. Furthermore, MO1 suppressed BA.5 infection in hamsters. A structural analysis revealed that MO1 binds to the conserved epitope of seven variants, including Omicron variants BA.5 and BA.2.75, in the receptor-binding domain of the spike protein. MO1 targets an epitope conserved among Omicron variants BA.1, BA.2, and BA.5 in a unique binding mode. Our findings confirm that D614G-derived vaccination can induce neutralizing antibodies that recognize the epitopes conserved among the SARS-CoV-2 variants. IMPORTANCE Omicron variants of SARS-CoV-2 acquired escape ability from host immunity and authorized antibody therapeutics and thereby have been spreading worldwide. We reported that patients infected with an early SARS-CoV-2 variant, D614G, and who received subsequent two-dose mRNA vaccination have high neutralizing antibody titer against Omicron lineages. It was speculated that the patients have neutralizing antibodies broadly effective against SARS-CoV-2 variants by targeting common epitopes. Here, we explored human monoclonal antibodies from B cells of the patients. One of the monoclonal antibodies, named MO1, showed high potency against broad SARS-CoV-2 variants including BA.2.75 and BA.5 variants. The results prove that monoclonal antibodies that have common neutralizing epitopes among several Omicrons were produced in patients infected with D614G and who received mRNA vaccination.

Metabolic alterations, especially in the mitochondria, play important roles in several kinds of cancers, including acute myeloid leukemia (AML). However, AML-specific molecular mechanisms that regulate mitochondrial dynamics remain elusive. Through the metabolite screening comparing CD34+ AML cells and healthy hematopoietic stem/progenitor cells, we identified enhanced lysophosphatidic acid (LPA) synthesis activity in AML. LPA is synthesized from glycerol-3-phosphate by glycerol-3-phosphate acyltransferases (GPATs), rate-limiting enzymes of the LPA synthesis pathway. Among the four isozymes of GPATs, glycerol-3-phosphate acyltransferases, mitochondrial (GPAM) was highly expressed in AML cells, and the inhibition of LPA synthesis by silencing GPAM or FSG67 (a GPAM-inhibitor) significantly impaired AML propagation through the induction of mitochondrial fission, resulting in the suppression of oxidative phosphorylation and the elevation of reactive oxygen species. Notably, inhibition of this metabolic synthesis pathway by FSG67 administration did not affect normal human hematopoiesis in vivo. Therefore, the GPAM-mediated LPA synthesis pathway from G3P represents a critical metabolic mechanism that specifically regulates mitochondrial dynamics in human AML, and GPAM is a promising potential therapeutic target.

Thg1-like protein (TLP) catalyzes the addition of a nucleotide to the 5'-end of truncated transfer RNA (tRNA) species in a Watson-Crick template-dependent manner. The reaction proceeds in two steps: the activation of the 5'-end by adenosine 5'-triphosphate (ATP)/guanosine 5'-triphosphate (GTP), followed by nucleotide addition. Structural analyses of the TLP and its reaction intermediates have revealed the atomic detail of the template-dependent elongation reaction in the 3'-5' direction. The enzyme creates two substrate binding sites for the first- and second-step reactions in the vicinity of one reaction center consisting of two Mg(2+) ions, and the two reactions are executed at the same reaction center in a stepwise fashion. When the incoming nucleotide is bound to the second binding site with Watson-Crick hydrogen bonds, the 3'-OH of the incoming nucleotide and the 5'-triphosphate of the tRNA are moved to the reaction center where the first reaction has occurred. That the 3'-5' elongation enzyme performs this elaborate two-step reaction in one catalytic center suggests that these two reactions have been inseparable throughout the process of protein evolution. Although TLP and Thg1 have similar tetrameric organization, the tRNA binding mode of TLP is different from that of Thg1, a tRNA(His)-specific G-1 addition enzyme. Each tRNA(His) binds to three of the four Thg1 tetramer subunits, whereas in TLP, tRNA only binds to a dimer interface and the elongation reaction is terminated by measuring the accepter stem length through the flexible β-hairpin. Furthermore, mutational analyses show that tRNA(His) is bound to TLP in a similar manner as Thg1, thus indicating that TLP has a dual binding mode.

Approximately 1 percent of healthy individuals carry human herpesvirus-6 within a host chromosome. This is referred to as chromosomally integrated herpesvirus-6 (CIHHV-6). In this study, we investigated the chromosomal integration site in six individuals harboring CIHHV-6B. Using FISH, we found that HHV-6B signals are consistently located at the telomeric region. The proximal endpoints of the integrated virus were mapped at one of two telomere-repeat-like sequences (TRSs) within the DR-R in all cases. In two cases, we isolated junction fragments between the viral TRS and human telomere repeats. The distal endpoints were mapped at the distal TRS in all cases. The size of the distal TRS was found to be ~5 kb which is sufficient to fulfill cellular telomeric functions. We conclude that the viral TRS in the DR regions fulfill dual functions for CIHHV-6: homology-mediated integration into the telomeric region of the chromosome and neo-telomere formation that is then stably transmitted.

The transport of pyruvate into mitochondria requires a specific carrier, the mitochondrial pyruvate carrier (MPC). The MPC represents a central node of carbon metabolism, and its activity is likely to play a key role in bioenergetics. Until now, investigation of the MPC activity has been limited. However, the recent molecular identification of the components of the carrier has allowed us to engineer a genetically encoded biosensor and to monitor the activity of the MPC in real time in a cell population or in a single cell. We report that the MPC activity is low in cancer cells, which mainly rely on glycolysis to generate ATP, a characteristic known as the Warburg effect. We show that this low activity can be reversed by increasing the concentration of cytosolic pyruvate, thus increasing oxidative phosphorylation. This biosensor represents a unique tool to investigate carbon metabolism and bioenergetics in various cell types.

BASIGIN/CD147/EMMPRIN is a multifunctional transmembrane glycoprotein strongly expressed in tumours. BASIGIN controls tumour metabolism, particularly glycolysis by facilitating lactic acid export through the two monocarboxylate transporters MCT1 and hypoxia-inducible MCT4. However, before being recognized as a co-carrier of MCTs, BASIGIN was described as an inducer of extracellular matrix metalloproteases (MMPs). Early on, a model emerged in which, tumour cells use the extracellular domain of BASIGIN to recognize and stimulate neighbouring fibroblasts to produce MMPs. However, this model has remained hypothetical since a direct link between BASIGIN and MMPs production has not yet been clearly established. To validate the BASIGIN/MMP hypothesis, we developed BASIGIN knockouts in three human tumour cell lines derived from glioma, colon, and lung adenocarcinoma. By using co-culture experiments of either human or mouse fibroblasts and tumour cell lines we showed, contrary to what has been abundantly published, that the disruption of BASIGIN in tumour cells and in MEFs has no action on the production of MMPs. Our findings do not support the notion that the pro-tumoural action of BASIGIN is mediated via induction of MMPs. Therefore, we propose that to date, the strongest pro-tumoural action of BASIGIN is mediated through the control of fermentative glycolysis.

To elucidate dynamic changes in native BCR-ABL and alternatively spliced tyrosine kinase inhibitor (TKI)-resistant but function-dead BCR-ABLIns35bp variant, following commencement or discontinuation of TKI therapy, each transcript was serially quantified in patients with chronic myeloid leukemia (CML) by deep sequencing. Because both transcripts were amplified together using conventional PCR system for measuring International Scale (IS), deep sequencing method was used for quantifying such BCR-ABL variants. At the initial diagnosis, 7 of 9 patients presented a small fraction of cells possessing BCR-ABLIns35bp , accounting for 0.8% of the total IS BCR-ABL, corresponding to actual BCR-ABLIns35bp value of 1.1539% IS. TKI rapidly decreased native BCR-ABL but not BCR-ABLIns35bp , leading to the initial increase in the proportion of BCR-ABLIns35bp . Thereafter, both native BCR-ABL and BCR-ABLIns35bp gradually decreased in the course of TKI treatment, whereas small populations positive for TKI-resistant BCR-ABLIns35bp continued fluctuating at low levels, possibly underestimating the molecular response (MR). Following TKI discontinuation, sequencing analysis of 54 patients revealed a rapid relapse, apparently derived from native BCR-ABL+ clones. However, IS fluctuating at low levels around MR4.0 marked a predominant persistence of cells expressing function-dead BCR-ABLIns35bp , suggesting that TKI resumption was unnecessary. We clarified the possible mechanism underlying mis-splicing BCR-ABLIns35bp , occurring at the particular pseudo-splice site within intron8, which can be augmented by TKI treatment through inhibition of RNA polymerase II phosphorylation. No mutations were found in spliceosomal genes. Therefore, monitoring IS functional BCR-ABL extracting BCR-ABLIns35bp would lead us to a correct evaluation of MR status, thus determining the adequate therapeutic intervention.

Tazemetostat is a selective, reversible, small-molecule inhibitor of the histone methyltransferase enzyme, enhancer of zest homolog 2 (EZH2). In this multicenter, open-label, phase II study, we assessed the efficacy and safety of tazemetostat in Japanese patients with relapsed or refractory (R/R) B-cell non-Hodgkin lymphoma harboring the EZH2 mutation. Tazemetostat (800 mg twice daily) was given orally (28-day cycle) until disease progression or unacceptable toxicity. Among the 20 eligible patients, 17 were enrolled in cohort 1 (follicular lymphoma [FL]), and three were enrolled in cohort 2 (diffuse large B-cell lymphoma). At data cut-off, the objective response rate in cohort 1 was 76.5%, including six patients (35.3%) with complete response and seven patients (41.2%) with partial response (PR). All three patients in cohort 2 achieved PR. In cohort 1, median progression-free survival (PFS) was not reached at the median follow-up of 12.9 months. The estimated PFS rate at 12 and 15 months was 94.1% and 73.2%, respectively. The most common grade 3 treatment-emergent adverse event (TEAE) was lymphopenia (n = 2). Grade 4 TEAEs included hypertriglyceridemia and pneumonia aspiration (n = 1 each), which were not related to tazemetostat. Treatment-emergent adverse events leading to study drug discontinuation were reported in four of the 20 patients, indicating that the safety profile of tazemetostat was acceptable and manageable. Tazemetostat 800 mg twice daily showed encouraging efficacy in patients with R/R EZH2 mutation-positive FL with a manageable safety profile in the overall population. Thus, tazemetostat could be a potential treatment for R/R EZH2 mutation-positive FL.

Bacterial cellulose (BC) is synthesized and exported through the cell membrane via a large protein complex (terminal complex) that consists of three or four subunits. BcsC is a little-studied subunit considered to export BC to the extracellular matrix. It is predicted to have two domains: a tetratrico peptide repeat (TPR) domain and a β-barrelled outer membrane domain. Here we report the crystal structure of the N-terminal part of BcsC-TPR domain (Asp24-Arg272) derived from Enterobacter CJF-002. Unlike most TPR-containing proteins which have continuous TPR motifs, this structure has an extra α-helix between two clusters of TPR motifs. Five independent molecules in the crystal had three different conformations that varied at the hinge of the inserted α-helix. Such structural feature indicates that the inserted α-helix confers flexibility to the chain and changes the direction of the TPR super-helix, which was also suggested by structural analysis of BcsC-TPR (Asp24-Leu664) in solution by size exclusion chromatography-small-angle X-ray scattering. The flexibility at the α-helical hinge may play important role for exporting glucan chains.

Fucoxanthin chlorophyll (Chl) a/c-binding proteins (FCPs) function as light harvesters in diatoms. The structure of a diatom photosystem II-FCPII (PSII-FCPII) supercomplex have been solved by cryo-electron microscopy (cryo-EM) previously; however, the FCPII subunits that constitute the FCPII tetramers and monomers are not identified individually due to their low resolutions. Here, we report a 2.5 Å resolution structure of the PSII-FCPII supercomplex using cryo-EM. Two types of tetrameric FCPs, S-tetramer, and M-tetramer, are identified as different types of hetero-tetrameric complexes. In addition, three FCP monomers, m1, m2, and m3, are assigned to different gene products of FCP. The present structure also identifies the positions of most Chls c and diadinoxanthins, which form a complicated pigment network. Excitation-energy transfer from FCPII to PSII is revealed by time-resolved fluorescence spectroscopy. These structural and spectroscopic findings provide insights into an assembly model of FCPII and its excitation-energy transfer and quenching processes.

Melphalan is widely used for hematopoietic stem cell transplantation (HSCT) conditioning. However, the relationship between its pharmacokinetic (PK) and transplantation outcomes in children has not been thoroughly investigated. We prospectively analyzed the relationship between melphalan area under the curve (AUC) and transplantation outcome and examined the development of a predictive model for melphalan clearance in children. This study included 43 children aged 0 to 19 years who underwent HSCT following a melphalan-based conditioning regimen from 2017 to 2021. In univariable analysis, high-melphalan AUC resulted in a significantly lower cumulative incidence of acute graft-versus-host disease and a higher cumulative incidence of thrombotic microangiopathy, although no significant difference was observed in survival. Regression analysis of a randomly selected derivation cohort (n = 21) revealed the following covariate PK model: predicted melphalan clearance (mL/min) = 6.47 × 24-h urinary creatinine excretion rate (CER, g/day) × 24-h creatinine clearance rate (CCR, mL/min) + 92.8. In the validation cohort (n = 22), the measured melphalan clearance values were significantly correlated with those calculated based on the prediction equation (R2 = 0.663). These results indicate that melphalan exposure may be optimized by adjusting the melphalan dose according to CER and CCR.

Polatuzumab vedotin (pola) is a CD79b-targeted antibody-drug conjugate delivering a potent antimitotic agent (monomethyl auristatin E) to B cells. This was an open-label, single-arm study of pola 1.8 mg/kg, bendamustine 90 mg/m2 , rituximab 375 mg/m2 (pola + BR) Q3W for up to six cycles in patients with relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL) who received ≥1 prior line of therapy and were ineligible for autologous stem cell transplantation (ASCT) or experienced treatment failure with prior ASCT. Primary endpoint was complete response rate (CRR) at the end of the treatment (EOT) by positron emission tomography-computed tomography (PET-CT) using modified Lugano Response Criteria. Secondary endpoints included efficacy, safety, and pharmacokinetics. Thirty-five patients (median age 71 [range 46-86] years) were enrolled. Twenty-three (66%) patients had refractory disease, and 23 (66%) had ≥2 prior lines of therapy. At a median follow-up of 5.4 (0.7-11.9) months, patients received a median of five treatment cycles. CRR was 34.3% (95% confidence interval [CI] 19.1-52.2) at EOT. Overall response rate was 42.9% at EOT, and median progression-free survival was 5.2 months (95% CI 3.6-not evaluable). Median overall survival was not reached. No fatal adverse events (AEs) were observed. Grade 3-4 AEs were mainly hematological: anemia (37%), neutropenia (31%), white blood cell count decreased (23%), thrombocytopenia/platelet count decreased/neutrophil count decreased (20% each), and febrile neutropenia (11%). Grade 1-2 peripheral neuropathy (PN; sensory and/or motor) was reported in 14% of patients; there were no ≥grade 3 PN events. This study (JapicCTI-184048) demonstrated the efficacy and safety of pola + BR in Japanese patients with R/R DLBCL who were ineligible for ASCT.

Hepatitis C virus (HCV) is a single-stranded RNA virus that not only affects hepatocytes, by B cells as well. It is thought that HCV is involved in the onset of B-cell lymphoma. The clinicopathological characteristics of HCV-positive diffuse large B-cell lymphoma (DLBCL) and HCV-positive splenic marginal zone lymphoma (SMZL) are known, but there has been no report on HCV-positive follicular lymphoma (FL). In this study, the clinicopathological characteristics of HCV-positive FL were examined in 263 patients with FL who were classified into a HCV-positive group with HCV antibody and negative groups without one. The number of patients with HCV-positive FL and HCV-negative FL was 10 (3.8%) and 253 (96.2%), respectively. The patients with HCV-positive FL commonly had more than one region of lymphadenopathy, Ann Arbor stage III/IV, hemoglobin <120 g/l, elevated lactate dehydrogenase level, and high-risk categorization of Follicular Lymphoma International Prognostic Index (FLIPI) than in patients with HCV-negative FL. Overall survival and progression-free survival were poorer in patients with HCV-positive FL than in those with HCV-negative FL (p < 0.0001 and 0.006, respectively). Also, multivariate analysis revealed that positive HCV antibody was a poor prognostic factor of OS. In conclusion, HCV-positive FL has unique clinical features and may have a great impact on the overall survival of affected patients.

The defensive-offensive associations between algae and herbivores determine marine ecology. Brown algae utilize phlorotannin as their chemical defense against the predator Aplysia kurodai, which uses β-glucosidase (akuBGL) to digest the laminarin in algae into glucose. Moreover, A. kurodai employs Eisenia hydrolysis-enhancing protein (EHEP) as an offense to protect akuBGL activity from phlorotannin inhibition by precipitating phlorotannin. To underpin the molecular mechanism of this digestive-defensive-offensive system, we determined the structures of the apo and tannic acid (TNA, a phlorotannin analog) bound forms of EHEP, as well as the apo akuBGL. EHEP consisted of three peritrophin-A domains arranged in a triangular shape and bound TNA in the center without significant conformational changes. Structural comparison between EHEP and EHEP-TNA led us to find that EHEP can be resolubilized from phlorotannin precipitation at an alkaline pH, which reflects a requirement in the digestive tract. akuBGL contained two GH1 domains, only one of which conserved the active site. Combining docking analysis, we propose the mechanisms by which phlorotannin inhibits akuBGL by occupying the substrate-binding pocket, and EHEP protects akuBGL against this inhibition by binding with phlorotannin to free the akuBGL pocket.