C-C motif chemokine ligand 2 regulates prostaglandin synthesis and embryo attachment of the bovine endometrium during implantation.

C-C motif chemokine ligand 2 (CCL2) has been reported to be expressed in the bovine endometrium during pregnancy. However, the details of its functions involved in the implantation mechanism are still not clear. The purpose of this study is to analyze the functional properties of CCL2 in the bovine endometrium and embryos. The expression of CCR2 was not different between the luteal phase and implantation phase of their endometrial tissues, but was significantly high in IFNa treated bovine endometrial stromal (BES) cells in vitro. The expressions of PGES1, PGES2, AKR1C4, and AKR1C4 were high at the implantation stage compared with the luteal stage. On the other hand, PGES2 and AKR1B1 in BEE and PGES3 and AKR1A1 in BES were significantly increased by CCL2 treatment, respectively. The expressions of PCNA and IFNt were found significantly high in the bovine trophoblastic cells (BT) treated with CCL2 compared to the control. CCL2 significantly increased the attachment rate of BT vesicles to BEE in in vitro co-culture system. The expression of OPN and ICAM-1 increased in BEE, and ICAM-1 increased in BT by CCL2 treatment, respectively. The present results indicate that CCL2 has the potential to regulate the synthesis of PGs in the endometrium and the embryo growth. In addition, CCL2 has the possibility to regulate the process of bovine embryo attachment to the endometrium by modulation of binding molecules expression.

Current development of chimeric antigen receptor T-cell therapy for diffuse large B-cell lymphoma and high-grade B-cell lymphoma.

Chimeric antigen receptor (CAR) T-cell therapy has become a commercially available treatment option for relapsed or refractory (r/r) diffuse large B-cell lymphoma (DLBCL) with two or more lines of prior therapies, and recently for high-risk r/r DLBCL with one prior line of therapy. The successful development of CAR T-cell therapy for multiple relapsed DLBCL has led to a boom in subsequent trials that investigated its utility in patients with other r/r B-cell lymphoma subtypes. However, CAR T-cell therapy is a multistep process that includes leukapheresis and manipulation which take several weeks. Therefore, patients with rapidly progressing or bulky disease may not be able to complete the therapeutic regimen involving CAR T-cell products. This raises the question of the generalizability of the results of pivotal studies to the entire population. In this review, we summarize the development of CAR-T cell therapy for B-cell lymphoma and discuss strategies to further improve the clinical outcomes of this treatment.

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.

Feasibility and clinical utility of comprehensive genomic profiling of hematological malignancies.

Identification of genetic alterations through next-generation sequencing (NGS) can guide treatment decision-making by providing information on diagnosis, therapy selection, and prognostic stratification in patients with hematological malignancies. Although the utility of NGS-based genomic profiling assays was investigated in hematological malignancies, no assays sufficiently cover driver mutations, including recently discovered ones, as well as fusions and/or pathogenic germline variants. To address these issues, here we have devised an integrated DNA/RNA profiling assay to detect various types of somatic alterations and germline variants at once. Particularly, our assay can successfully identify copy number alterations and structural variations, including immunoglobulin heavy chain translocations, IKZF1 intragenic deletions, and rare fusions. Using this assay, we conducted a prospective study to investigate the feasibility and clinical usefulness of comprehensive genomic profiling for 452 recurrently altered genes in hematological malignancies. In total, 176 patients (with 188 specimens) were analyzed, in which at least one alteration was detected in 171 (97%) patients, with a median number of total alterations of 7 (0-55). Among them, 145 (82%), 86 (49%), and 102 (58%) patients harbored at least one clinically relevant alteration for diagnosis, treatment, and prognosis, respectively. The proportion of patients with clinically relevant alterations was the highest in acute myeloid leukemia, whereas this assay was less informative in T/natural killer-cell lymphoma. These results suggest the clinical utility of NGS-based genomic profiling, particularly for their diagnosis and prognostic prediction, thereby highlighting the promise of precision medicine in hematological malignancies.

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.

Prophylactic antiviral therapy for hepatitis B virus surface antigen-positive patients with diffuse large B-cell lymphoma treated with rituximab-containing chemotherapy.

We conducted a nationwide retrospective analysis of 116 hepatitis B virus (HBV) surface antigen (HBsAg)-positive patients with diffuse large B-cell lymphoma (DLBCL) and 278 HBsAg-negative patients with DLBCL, as a control cohort, who received rituximab-containing regimens as an induction chemotherapy at 30 Japanese medical centers between January 2004 and December 2014. Hepatitis was defined as an absolute serum alanine aminotransferase (ALT) level of ≥100 U/L. HBV reactivation-related hepatitis was defined as hepatitis with an absolute serum HBV DNA level of ≥3.3 log IU/mL or an absolute increase of ≥2 log compared with the baseline value. HBsAg-positive patients were divided into three groups based on anti-HBV prophylactic therapy: no nucleos(t)ide analogue (non-NA, n = 9), lamivudine (LAM, n = 20), and entecavir (ETV, n = 87). The 4-year cumulative incidence (CI) of hepatitis in HBsAg-positive and HBsAg-negative patients was 21.1% and 14.6% (P = .081), respectively. The 4-year CI of HBV reactivation-related hepatitis was higher in HBsAg-positive patients than in HBsAg-negative patients (8.0% vs 0.4%; P < .001). Among HBsAg-positive patients, the 4-year CI of HBV reactivation-related hepatitis was the highest in the non-NA group (33.3%), followed by the LAM (15.0%) and ETV (3.8%) groups (P < .001). Of note, 3 non-NA patients (33%) and 1 LAM patient (5%) (but no ETV patients) died due to HBV hepatitis. Based on Cox multivariate analysis, HBsAg positivity was not associated with poor overall survival. Prophylactic use of ETV would reduce the occurrence of HBV reactivation-related hepatitis and mortality in HBsAg-positive DLBCL patients receiving rituximab-containing chemotherapy.

Gene expression of bovine endometrial epithelial cells cultured in matrigel.

Glandular epithelial cells (GE) in the endometrium are thought to support the elongation and survival of ruminant embryos by secreting histotrophs. In the present study, the gene expression of bovine endometrial epithelial cells cultured in matrigel was analyzed and examined whether it could be an in vitro model of GE. Bovine endometrial epithelial cells (BEE) and stromal cells (BES) were isolated from the slaughterhouse uteri and cultured in DMEM/F12 + 10% FBS. BEE showed the gland-like structure morphological changes when cultured in 15% matrigel but could not be identified in higher concentrations of the matrigel (30% or 60%). The expression of typical genes expressed in GE, SERPINA14 and GRP, was substantially high in matrigel-cultured BEE than in monolayer (P  <  0.05). P4 and INFα have no significant effect on the SERPINA14 expression of BEE cultured in matrigel without co-culture with BES. On the other hand, when BEE were co-cultured with BES in matrigel culture, the expression of FGF13 was increased by the P4 treatment (P  <  0.05). Furthermore, SERPINA14 and TXN expressions were increased by P4 + IFNα treatment (P  <  0.05). These results demonstrate the appropriate conditions for BEE to form glandular structures in matrigel and the effect of co-culture with BES. The present study highlighted the possible use of matrigel for the culture of BEE to investigate the expression of cell-specific glandular epithelial genes as well as P4 and type-I IFN as factors controlling endometrial function during the implantation period.

The role of local radiotherapy following rituximab-containing chemotherapy in patients with transformed indolent B-cell lymphoma.

OBJECTIVES: This study aimed to evaluate the outcomes of local radiotherapy (LRT) in patients with histologic transformation (HT) following rituximab-containing chemotherapy. METHODS: We retrospectively analysed 92 patients with biopsy-confirmed HT undergoing rituximab-containing chemotherapy at our institution between 2003 and 2015. RESULTS: Of the 36 patients with limited-stage disease at diagnosis of HT, 29 (78%) received LRT. The estimated 5-year progression-free survival (PFS) rate was significantly better in patients who underwent LRT than in those who did not (93% and 42%, respectively; P < 0.05). Multivariate analyses employing age, sex, performance status, LRT and treatment response demonstrated that LRT was an independent prognostic factor for PFS (hazard ratio [HR]: 11.8; 95% confidence interval [CI]: 1.28-108.1; P < 0.05). Of the 32 patients who underwent LRT for HT lesion treatment, 31 (97%) did not show disease progression within radiation fields; among them, 27 patients (84%) survived without disease progression during the follow-up period. One patient developed hypothyroidism due to LRT; the others had no acute or late-onset complications of LRT. CONCLUSIONS: Our data support the recommendation of LRT for HT lesion treatment following rituximab-containing chemotherapy in select patients with localised HT, as a rational treatment approach with potentially limited toxicity.

[Successful treatment with silver nitrate chemical cauterization for paronychia and granulation in a patient with chronic lymphocytic leukemia undergoing ibrutinib therapy].

A 72-year-old man with leukocytosis, anemia, and lymphadenopathy was diagnosed with chronic lymphocytic leukemia (CLL) in August 2017 and was carefully monitored in a "watch-and-wait" manner until it became an "active disease." Ibrutinib (IBR) was initiated orally in July 2018 at a dose of 420 mg/day after disease progression due to chromosome 17p deletion (del 17p). The patient showed partial response after transient lymphocytosis while on IBR treatment. IBR induces paronychia and skin disorder due to the disruption of disulfide bonds between cysteine and inhibition of epidermal growth factor receptor due to the off-target effect. This results in reduced quality of life. In February 2019, paronychia (grade 1) developed in the patient's right foot's first toe; hence, topical gentamicin and taping therapy were performed. However, the symptoms persisted without any improvements. In July 2019, paronychia/granulation (grade 2) was aggravated and successfully treated with silver nitrate chemical cauterization and taping therapy. The patient was continuously treated with 420 mg/day IBR without dose reduction or discontinuation, resulting in successful disease control of CLL with del 17p.

[Disseminated aspergillosis due to Aspergillus udagawae during immunosuppressive treatment for myelodysplastic syndrome].

An 80 year old male who had received immunosuppressive therapy for myelodysplastic syndrome presented with fever, fatigue, and elevated serum Aspergillus antigen. Computed tomography revealed infiltrative shadows in the left lower lung and subcutaneous nodules. A polymerase chain reaction assay from lung and subcutaneous nodule samples identified the presence Aspergillus udagawae. A. udagawae is a cryptic species that shares similar morphological characteristics with A. fumigatus but genetically differs from the latter in its susceptibility to antifungal drugs. When immunosuppressed patients with hematological malignancies develop disseminated aspergillosis, biopsy and fungal tests are crucial to identify the causative fungus, including cryptic species, for deciding the appropriate therapeutic intervention.

R-CHOP-14 versus R-CHOP-14/CHASER for upfront autologous transplantation in diffuse large B-cell lymphoma: JCOG0908 study.

The efficiency of upfront consolidation with high-dose chemotherapy/autologous stem-cell transplantation (HDCT/ASCT) for newly diagnosed high-risk diffuse large B-cell lymphoma (DLBCL) may be influenced by induction chemotherapy. To select better induction chemotherapy regimens for HDCT/ASCT, a randomized phase II study was conducted in high-risk DLBCL patients having an age-adjusted International Prognostic Index (aaIPI) score of 2 or 3. As induction chemotherapy, 6 cycles of R-CHOP-14 (arm A) or 3 cycles of R-CHOP-14 followed by 3 cycles of CHASER (arm B) were planned, and patients who responded proceeded to HDCT with LEED and ASCT. The primary endpoint was 2-y progression-free survival (PFS), and the main secondary endpoints included overall survival, overall response rate, and adverse events (AEs). In total, 71 patients were enrolled. With a median follow-up of 40.3 mo, 2-y PFS in arms A and B were 68.6% (95% confidence interval [CI], 50.5%-81.2%) and 66.7% (95% CI: 48.8%-79.5%), respectively. Overall survival at 2 y in arms A and B was 74.3% (95% CI: 56.4%-85.7%) and 83.3% (95% CI: 66.6%-92.1%). Overall response rates were 82.9% in arm A and 69.4% in arm B. During induction chemotherapy, 45.7% and 75.0% of patients in arms A and B, respectively, had grade ≥ 3 non-hematologic toxicities. One patient in arm A and 6 in arm B discontinued induction chemotherapy due to AEs. In conclusion, R-CHOP-14 showed higher 2-y PFS and less toxicity compared with R-CHOP-14/CHASER in patients with high-risk DLBCL, suggesting the former to be a more promising induction regimen for further investigations (UMIN-CTR, UMIN000003823).

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.

Mucosa-associated lymphoid tissue lymphoma with t(11;18)(q21;q21) translocation: long-term follow-up results.

Translocation (11;18)(q21;q21) is found in mucosa-associated lymphoid tissue (MALT) lymphoma, resulting in API2/MALT1 gene fusion. It is known that t(11;18)-positive MALT lymphoma shows a tendency to disseminate and be resistant to Helicobacter pylori eradication by antibiotics. However, the prognostic features including recurrence and histological transformation (HT) remain unknown. We conducted a single-institute retrospective analysis of 464 patients with newly diagnosed MALT lymphoma, evaluating the impact of t(11;18) on clinical outcomes. One hundred and six patients were screened for the translocation by fluorescence in situ hybridization and/or reverse transcriptase-polymerase chain reaction. Of these patients, 26 patients (25%) were diagnosed as MALT lymphoma with t(11;18). The patients had a significantly shortened progression-free survival (PFS at 10 years; 26% v 57%; P = 0.004) compared to those without t(11;18). However, this did not translate into overall survival or incidence of HT. We confirmed previous reports stating that t(11;18)-positive MALT lymphoma showed disseminated disease and refractoriness to H. pylori eradication therapy. Patients with t(11;18) had more frequent monoclonal gammopathy, especially of IgM subtype (31% v 8%; P = 0.008), some of which developed class switch. These findings characterize the features of t(11;18)-positive MALT lymphoma, suggesting that it comprises a distinct clinical entity of MALT lymphoma.

[Vascular adverse events of ponatinib during treatment of Philadelphia chromosome-positive leukemia: a retrospective single-institution analysis].

Ponatinib (PON) is a key drug for patients with second tyrosine kinase inhibitor (TKI)-resistant/intolerant Philadelphia chromosome-positive leukemia (Ph+ leukemia); however, the occurrence of vascular adverse events (VAEs) in patients treated with PON should be carefully monitored. A retrospective analysis involving seven patients treated with PON was conducted to elucidate the incidence rate and risk factor for the development of VAEs. In the present study, risk assessment and monitoring of VAEs were performed using SCORE Risk Chart and Suita Score (10-year risk for fatal cardiovascular event), respectively. Despite the prophylactic use of aspirin, cerebral infarction and unstable angina occurred in two patients. By contrast, deep vein thrombosis did not improve in a patient treated with edoxaban. Our data suggest that patients with Ph+ leukemia possessing risk factors, medical history of lifestyle diseases, and administration of long-term second TKI treatment require careful monitoring of VAEs and therapeutic intervention to lifestyle diseases.

[Hairy cell leukemia complicated by bone marrow necrosis following cladribine administration].

Classic hairy cell leukemia (classic HCL) is a rare disease associated with indolent mature B-cell lymphoma. A 50-year-old man presented with pancytopenia for 3 years and was diagnosed with classic HCL because his lymphoid cells showed a hairy morphology with oval nuclei and indistinct nucleoli both in the peripheral blood and bone marrow (BM) smears. Flow cytometric analysis revealed that these cells expressed CD11c, CD25, and CD103, and the Sanger sequence method detected BRAF V600E mutation. Cladribine (0.09 mg/kg/day) was initiated for 7 days via continuous intravenous injection. On day 13, the patient died from bloodstream infection caused by methicillin-resistant Staphylococcus epidermidis. Autopsy findings revealed BM necrosis without residual leukemia cells caused by classic HCL, severe infection, and agents, such as cladribine and granulocyte-colony stimulating factor; however, its cause remained undetermined. Both early diagnosis and immediate clinical intervention are required to improve the clinical outcomes in classic HCL. The cause of hematopoiesis disturbance should also be identified using BM biopsy or magnetic resonance imaging before initiating treatment in classic HCL with severe pancytopenia.

Development of an in vitro model to study uterine functions and early implantation using rat uterine explants.

This study was conducted to develop an in vitro model using rat uterine explants to explore complex uterine functions. Rat uterine explants (1-2 mm) were isolated, cultured and further characterized. Steroid hormone treatment of cultured explants showed that both Muc1 and Pr were significantly up-regulated (P < 0.05) by E2. Areg was significantly up-regulated (P < 0.05) by P4 and Igfbp1 was significantly up-regulated (P < 0.05) by the combination of E2 and P4, although, in rat, Igfbp1 is E2-dependent. In vitro decidualization of cultured explants was induced and two potential markers of decidualization, Prl8a2 and Bmp2, were examined. Real-time quantitative PCR data revealed that both Prl8a2 and Bmp2 were significantly up-regulated (P < 0.05) in MPA- and db-cAMP-treated explants compared to the control group of explants. Then, an individual hatched blastocyst and cultured explant was placed in a 96-well (round-bottom U-shaped) plate. Co-culture results showed that stable attachments were observed after 48 h, where embryos were stably attached to the explants and could not be dislodged after mild shaking and/or pipetting. The rates of attachment of embryos to the explants were increased significantly in the P4-treated group (63.6%) compared to the control group (35.5%), after steroid hormone treatment. The rates of attachment were reduced significantly in the E2-treated group (0.0%), where no stable attachments were observed. Despite the necessity of comprehensive investigation, our results suggest that the cultured rat uterine explants can be a useful in vitro model to study uterine functions and early implantation.