Clonal evolution mechanisms in NT5C2 mutant-relapsed acute lymphoblastic leukaemia.

Relapsed acute lymphoblastic leukaemia (ALL) is associated with resistance to chemotherapy and poor prognosis. Gain-of-function mutations in the 5'-nucleotidase, cytosolic II (NT5C2) gene induce resistance to 6-mercaptopurine and are selectively present in relapsed ALL. Yet, the mechanisms involved in NT5C2 mutation-driven clonal evolution during the initiation of leukaemia, disease progression and relapse remain unknown. Here we use a conditional-and-inducible leukaemia model to demonstrate that expression of NT5C2(R367Q), a highly prevalent relapsed-ALL NT5C2 mutation, induces resistance to chemotherapy with 6-mercaptopurine at the cost of impaired leukaemia cell growth and leukaemia-initiating cell activity. The loss-of-fitness phenotype of NT5C2+/R367Q mutant cells is associated with excess export of purines to the extracellular space and depletion of the intracellular purine-nucleotide pool. Consequently, blocking guanosine synthesis by inhibition of inosine-5'-monophosphate dehydrogenase (IMPDH) induced increased cytotoxicity against NT5C2-mutant leukaemia lymphoblasts. These results identify the fitness cost of NT5C2 mutation and resistance to chemotherapy as key evolutionary drivers that shape clonal evolution in relapsed ALL and support a role for IMPDH inhibition in the treatment of ALL.

A case of a pilocytic astrocytoma with histological features of anaplasia and unprecedent genetic alterations.

We report a case of pediatric glioma with uncommon imaging, morphological, and genetic features. A one-year-old boy incidentally presented with a tumor in the fourth ventricle. The tumor was completely resected surgically and investigated pathologically. The mostly circumscribed tumor had piloid features but primitive and anaplastic histology, such as increasing cellularity and mitosis. The Ki-67 staining index was 25% at the hotspot. KIAA1549::BRAF fusion and KIAA1549 partial deletions were detected by direct PCR, supported by Sanger sequencing. To the best of our knowledge, this is the first report of a glioma with both deletion of KIAA1549 p.P1771_P1899 and fusion of KIAA1549::BRAF. The tumor could not be classified using DNA methylome analysis. The present tumor fell into the category of pilocytic astrocytoma with histological features of anaplasia (aPA). Further studies are needed to establish pediatric aPA.

Regular prophylaxis with activated prothrombin complex concentrates in pediatric hemophilia.

BACKGROUND: Regular prophylaxis with activated prothrombin complex concentrates (aPCCs) is effective in adult patients with hemophilia with inhibitors; however, data in children are scarce. METHODS: This was a single-center retrospective study at Saitama Children's Medical Center. Patients with severe and moderate hemophilia with inhibitors aged <15 years at the start of aPCCs prophylaxis were included. Medical records were retrospectively reviewed. RESULTS: We treated nine pediatric patients with hemophilia with inhibitors (median age, 1.9 years; age range, 1.3-12.9 years; inhibitor titers before treatment with aPCCs, 5.9-69 BU/mL) using prophylactic aPCCs (doses, 50-100 U/kg; 2-3 times/week). The median prophylactic period was 13 months (range: 5-31 months). The median annualized bleeding rate (ABR) during prophylactic treatment with aPCCs was 2 (range, 0-17). In four patients, ABR was reduced by 19%-100% with prophylactic aPCCs compared to on-demand aPCCs. An adverse effect of treatment was that a patient with hemophilia B developed nephrotic syndrome 34 months after starting regular prophylaxis with aPCCs. CONCLUSIONS: Regular prophylactic aPCCs reduced the ABR even in younger children with hemophilia A and B. Serious adverse events include nephrotic syndrome, which requires caution.

Alteration of the immune environment in bone marrow from children with recurrent B cell precursor acute lymphoblastic leukemia.

Due to the considerable success of cancer immunotherapy for leukemia, the tumor immune environment has become a focus of intense research; however, there are few reports on the dynamics of the tumor immune environment in leukemia. Here, we analyzed the tumor immune environment in pediatric B cell precursor acute lymphoblastic leukemia by analyzing serial bone marrow samples from nine patients with primary and recurrent disease by mass cytometry using 39 immunophenotype markers, and transcriptome analysis. High-dimensional single-cell mass cytometry analysis elucidated a dynamic shift of T cells from naïve to effector subsets, and clarified that, during relapse, the tumor immune environment comprised a T helper 1-polarized immune profile, together with an increased number of effector regulatory T cells. These results were confirmed in a validation cohort using conventional flow cytometry. Furthermore, RNA transcriptome analysis identified the upregulation of immune-related pathways in B cell precursor acute lymphoblastic leukemia cells during relapse, suggesting interaction with the surrounding environment. In conclusion, a tumor immune environment characterized by a T helper 1-polarized immune profile, with an increased number of effector regulatory T cells, could contribute to the pathophysiology of recurrent B cell precursor acute lymphoblastic leukemia. This information could contribute to the development of effective immunotherapeutic approaches against B cell precursor acute lymphoblastic leukemia relapse.

Successful perioperative management using prothrombin complex concentrates in patients with severe congenital protein C deficiency.

Perioperative management of severe congenital protein C deficiency remains unestablished. This deficiency is often treated with anticoagulants, such as warfarin. Although anticoagulants need to be perioperatively discontinued, there are few methods for the management of such patients. We adopted a method for administering prothrombin complex concentrates (PCC), which includes intermittent administration of inactive protein C (PPSB-HT), and examined its outcome as a perioperative management approach for severe congenital protein C deficiency. Three patients underwent our perioperative management six times. We monitored activity levels of protein C, factor IX, and so forth. These patients could be perioperatively managed with PCC treatment.

[The role of CRISPR screening in leukemia research].

Recent advances in comprehensive genome analysis have contributed to the discovery of several leukemia-related genes; however, the role of mutated genes in the pathogenesis of leukemia remains unknown. Clustered regularly interspaced short palindromic repeats-associated nuclease 9 (CRISPR-Cas9) technology is a powerful tool for genome-wide editing in both coding and noncoding regions. CRISPR screening has enabled high-throughput validation of gene function in diverse tumor processes, including tumor growth and survival, synthetic lethal interactions, therapeutic resistance, and response to immunotherapy, and is actively used in leukemia research. Herein, we discuss recent advances in CRISPR screening in cancer research, focusing on leukemia, and outline application strategies and prospects for CRISPR screening.

[Angioimmunoblastic T-cell lymphoma after immune checkpoint inhibitor-combined chemotherapy for lung cancer].

A 68-year-old male patient with lung adenocarcinoma, who was treated with chemotherapy and immune checkpoint inhibitors (ICIs), developed lymphadenopathy during treatment. His para-aortic lymph nodes increased to 2.0 cm in diameter. Both inguinal lymph nodes were 1.5 cm in diameter, and multiple hepatic masses appeared. After the ICI readministration, both inguinal lymph nodes increased to 2.0 cm in diameter, but the para-aortic lymph nodes and hepatic masses remained. Angioimmunoblastic T-cell lymphoma (AITL) diagnosis was established after the right inguinal lymph node biopsy, which was accompanied by an infiltration of Epstein-Barr virus (EBV)-encoded small ribonucleic acid-positive B-cells. After the ICI discontinuation, the inguinal lymph nodes decreased to 1.5 cm in diameter, but the para-aortic lymph nodes remained, and hepatic masses increased. Hepatic lesions were possibly lung cancer metastasis. The ICI administration and EBV reactivation were potentially associated with AITL development in the present case. The natural shrinkage of lymphoma after the ICI cessation implied the immunological mechanism like that of the methotrexate-related lymphoproliferative disease.

[Thrombopoietin receptor agonists for pediatric refractory chronic immune thrombocytopenia].

Various treatments have been used to treat chronic immune thrombocytopenic purpura in children; however, none of it has been established as the standard of care. The administration of thrombopoietin receptor agonists (TPO-RAs) has been approved as a new treatment option in Japan. In this case series, TPO-RAs were administered to 16 patients (eltrombopag, n=9; romiplostim, n=7). Excluding the data of two patients who underwent splenectomy immediately after starting treatment with these medicines, platelet counts increased to ≥50,000/µl in seven patients. The adverse events recorded were grade 2 liver dysfunction (n=1), according to the common terminology criteria for adverse events version 4, and myelofibrosis (classified as MF1 or mild reticulin fibrosis), as observed on bone marrow biopsy (n=2). We continued the administration of TPO-RAs at the same dose in these patients because the complications they experienced were mild. The risk of adverse events associated with long-term use of TPO-RAs in this pediatric population remains unclear, and a prospective evaluation is needed.

[A case of Endocrine Active Retroperitoneal Ganglioneuroma, Difficult to Differentiate From Adrenal Pheochromocytoma].

A 66-year-old woman who had been receiving medication for hypertension and hyperlipidemia was referred to our hospital for evaluation of a left adrenal tumor (12×8 mm) that was incidentally detected on computed tomography. Her 24-hour urinary catecholamine level was elevated, and metaiodobenzylguanidine (MIBG) scintigraphy revealed increased uptake in the area around the left adrenal gland, necessitating laparoscopic adrenalectomy for preoperative diagnosis of left adrenal pheochromocytoma. Intraoperatively, we detected a para-aortic tumor behind the adrenal gland, and this lesion was excised together with the adrenal gland. However, manipulation of the para-aortic tumor led to elevation in the blood pressure to 170 mmHg. Histopathological examination of the resected specimens revealed an adrenocortical adenoma and a para-aortic ganglioneuroma, consisting of ganglion cells, nerve fibers, and Schwann cells. The patient's blood pressure normalized immediately postoperatively, and MIBG scintigraphy revealed a negative result. Endocrine active ganglioneuromas are rare, and to our knowledge, currently only 8 cases (including ours) have been reported in the Japanese and English literature.

[Interleukin-6-producing paraganglioma mimicking multicentric Castleman disease].

Multicentric Castleman disease (MCD) comprises a heterogeneous group of lymphoproliferative disorders. Interleukin 6 (IL-6) plays an important role in the MCD pathophysiology. Here, we report the case of a 17-year-old Japanese man who presented with fever, headache, fatigue, and weight loss, with normal blood pressure. A movable mass was palpated in his lower abdomen. Laboratory tests revealed microcytic anemia and hypoalbuminemia, with elevated IL-6, sIL-2R, and vascular endothelial growth factor. Computed tomography of the abdomen demonstrated a 55-mm-diameter pelvic tumor and enlarged mesenteric lymph nodes. MCD was suspected, and the pelvic tumor resected. After the operation, his blood pressure rose slowly, and resulted to seizures of posterior reversible encephalopathy syndrome. Evaluation of hypertension revealed that plasma norepinephrine and normetanephrine concentrations were elevated, and pathological examinations showed that the resected tumor was positive for IL-6 and chromogranin-A. Therefore, we diagnosed the patient with IL-6-producing paraganglioma with MCD-mimicking symptoms. Moreover, IL-6-producing pheochromocytoma and paraganglioma should be included in differential diagnoses of MCD, even in normotensive patients.

[Gilteritinib for pediatric FLT3 internal tandem duplication-positive recurrent acute myeloid leukemia].

Gilteritinib is an FMS-like tyrosine kinase 3 (FLT3) inhibitor that has shown efficacy in patients with refractory or recurrent adult acute myeloid leukemia (AML) with FLT3 mutations. However, there are limited data for pediatric patients treated with this drug. Herein, we report the clinical courses of two children with FLT3-mutated recurrent AML who received gilteritinib. Case 1: An 11-year-old boy with secondary relapsed AML presented with an FLT3 internal tandem duplication (ITD) since the first recurrence. One week after gilteritinib initiation, blasts, which had comprised 90% of the white blood cells before treatment, almost disappeared from the peripheral blood without tumor lysis syndrome. The patient developed multiple adverse effects and died from the disease 2.5 months after gilteritinib initiation. Case 2: A 12-year-old girl diagnosed with AML was positive for FLT3 ITD. She received gilteritinib during her first relapse post-stem cell transplantation. After the drug was administered, the recipient cell counts increased, as determined by molecular tests (i.e., FISH), whereas microscopically, there was a complete response for 5 months with good performance status. Gilteritinib treatment in children with FLT3-mutated recurrent AML is feasible and effective. As a patient experienced several adverse effects with gilteritinib treatment, clinical trials are required to determine the appropriate pediatric dose of this medication.

Reticular dysgenesis: international survey on clinical presentation, transplantation, and outcome.

Reticular dysgenesis (RD) is a rare congenital disorder defined clinically by the combination of severe combined immunodeficiency (SCID), agranulocytosis, and sensorineural deafness. Mutations in the gene encoding adenylate kinase 2 were identified to cause the disorder. Hematopoietic stem cell transplantation (HSCT) is the only option to cure this otherwise fatal disease. Retrospective data on clinical presentation, genetics, and outcome of HSCT were collected from centers in Europe, Asia, and North America for a total of 32 patients born between 1982 and 2011. Age at presentation was <4 weeks in 30 of 32 patients (94%). Grafts originated from mismatched family donors in 17 patients (55%), from matched family donors in 6 patients (19%), and from unrelated marrow or umbilical cord blood donors in 8 patients (26%). Thirteen patients received secondary or tertiary transplants. After transplantation, 21 of 31 patients were reported alive at a mean follow-up of 7.9 years (range: 0.6-23.6 years). All patients who died beyond 6 months after HSCT had persistent or recurrent agranulocytosis due to failure of donor myeloid engraftment. In the absence of conditioning, HSCT was ineffective to overcome agranulocytosis, and inclusion of myeloablative components in the conditioning regimens was required to achieve stable lymphomyeloid engraftment. In comparison with other SCID entities, considerable differences were noted regarding age at presentation, onset, and type of infectious complications, as well as the requirement of conditioning prior to HSCT. Although long-term survival is possible in the presence of mixed chimerism, high-level donor myeloid engraftment should be targeted to avoid posttransplant neutropenia.

Mutational landscape, clonal evolution patterns, and role of RAS mutations in relapsed acute lymphoblastic leukemia.

Although multiagent combination chemotherapy is curative in a significant fraction of childhood acute lymphoblastic leukemia (ALL) patients, 20% of cases relapse and most die because of chemorefractory disease. Here we used whole-exome and whole-genome sequencing to analyze the mutational landscape at relapse in pediatric ALL cases. These analyses identified numerous relapse-associated mutated genes intertwined in chemotherapy resistance-related protein complexes. In this context, RAS-MAPK pathway-activating mutations in the neuroblastoma RAS viral oncogene homolog (NRAS), kirsten rat sarcoma viral oncogene homolog (KRAS), and protein tyrosine phosphatase, nonreceptor type 11 (PTPN11) genes were present in 24 of 55 (44%) cases in our series. Interestingly, some leukemias showed retention or emergence of RAS mutant clones at relapse, whereas in others RAS mutant clones present at diagnosis were replaced by RAS wild-type populations, supporting a role for both positive and negative selection evolutionary pressures in clonal evolution of RAS-mutant leukemia. Consistently, functional dissection of mouse and human wild-type and mutant RAS isogenic leukemia cells demonstrated induction of methotrexate resistance but also improved the response to vincristine in mutant RAS-expressing lymphoblasts. These results highlight the central role of chemotherapy-driven selection as a central mechanism of leukemia clonal evolution in relapsed ALL, and demonstrate a previously unrecognized dual role of RAS mutations as drivers of both sensitivity and resistance to chemotherapy.

Human AK2 links intracellular bioenergetic redistribution to the fate of hematopoietic progenitors.

AK2 is an adenylate phosphotransferase that localizes at the intermembrane spaces of the mitochondria, and its mutations cause a severe combined immunodeficiency with neutrophil maturation arrest named reticular dysgenesis (RD). Although the dysfunction of hematopoietic stem cells (HSCs) has been implicated, earlier developmental events that affect the fate of HSCs and/or hematopoietic progenitors have not been reported. Here, we used RD-patient-derived induced pluripotent stem cells (iPSCs) as a model of AK2-deficient human cells. Hematopoietic differentiation from RD-iPSCs was profoundly impaired. RD-iPSC-derived hemoangiogenic progenitor cells (HAPCs) showed decreased ATP distribution in the nucleus and altered global transcriptional profiles. Thus, AK2 has a stage-specific role in maintaining the ATP supply to the nucleus during hematopoietic differentiation, which affects the transcriptional profiles necessary for controlling the fate of multipotential HAPCs. Our data suggest that maintaining the appropriate energy level of each organelle by the intracellular redistribution of ATP is important for controlling the fate of progenitor cells.

[The Usefulness of 11C-Choline PET/CT for Detecting Metastasis and Treatment Monitoring of Renal Cell Carcinoma : Report of Two Cases].

Although fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) has been widely used as a powerful tool in clinical oncologic imaging, its application in urological malignancies is limited. Recently, choline PET/CT has been successfully used for prostate cancer restaging. Here, we report the utility of choline PET/CT for the detection and monitoring of metastatic disease in two cases of renal cell carcinoma (RCC). A 53-year-old woman and a 45-year-old woman underwent FDG and choline PET/CT for evaluation of metastatic lesions in lymph nodes and bone following left and right RCC, respectively, and choline PET/CT demonstrated significantly higher uptake when compared with FDGPET/CT in both cases. Choline PET/CT accurately reflected the remission and progression of diseases in their clinical course, indicating that choline PET/CT could be a useful imaging modality in metastatic RCC.

Peripheral T-cell lymphoma, not otherwise specified: a retrospective single-center analysis.

We retrospectively analyzed clinical and pathological features, treatments, and prognoses in 28 patients with newly diagnosed peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS) in Niigata University Medical and Dental Hospital. Of them, 16 were males and 12 were females, and their median age was 62.5 (range, 26-88) years. The International Prognostic Index was high-intermediate/high in 68% of patients. Twelve patients were treated with CHOP/THP-COP and nine with third-generation chemotherapy regimens. At a median follow-up period of 30 (range: 1-164) months, the 2-year overall survival and progression-free survival rates were 61% and 44%, respectively. Further investigation of novel agents for treating PTCL-NOS is warranted.

X-linked agammaglobulinemia: Twenty years of single-center experience from North West India.

BACKGROUND: X-linked agammaglobulinemia (XLA) is an X-linked genetic defect in maturation of B lymphocytes that results in the absence of B lymphocytes in the peripheral blood and profound hypogammaglobulinemia. It is caused by a mutation in the BTK gene located on the X chromosome. There are no large series describing XLA from the developing world. OBJECTIVE: To analyze the clinical features, immunologic and genetic characteristics, and outcomes of 36 patients with XLA diagnosed and managed for a period of 2 decades. METHODS: Diagnosis of XLA was made on the basis of presence of BTK gene mutation or marked reduction of B lymphocytes in peripheral blood with a family history of an affected male relative. The diagnosis was confirmed by genetic mutation studies in 28 patients with 25 unique mutations in the BTK gene. RESULTS: There was a significant delay in diagnosis in most of the patients. The mean (SD) delay in the diagnosis was 4.2 (3.5) years. Point mutations were the most common mutations detected, accounting for 68% of all mutations. Deletions and insertions were also seen in a few cases. Four of the mutations are novel mutations that have not been previously reported. Seven of the 36 patients (19%) were dead at the time of analysis in the present cohort. The mean survival was 137 months (95% confidence interval, 13-163 months). CONCLUSION: The present study is perhaps the largest series of patients with XLA from any developing country so far.