A unique expression pattern of LAG3 distinct from that of other immune checkpoints in diffuse large B-cell lymphoma.

BACKGROUND: Although some patients with diffuse large B-cell lymphoma (DLBCL) show a response to immunotherapy, there are still many who do not respond. This suggests that various immune checkpoints are complicatedly intertwined in the composition of the tumor microenvironment of DLBCL. PATIENTS AND METHODS: To comprehensively understand the expression of various immune checkpoint genes in DLBCL, we performed NanoString assay in 98 patients to investigate 579 genes. In addition, we performed immunohistochemistry for LAG-3 and PD-L1 to compare the results with expression in NanoString assay. RESULTS: As a result of hierarchical clustering of NanoString assay, 98 DLBCLs were classified into three tumor immune microenvironment clusters. Most immune checkpoint genes showed the highest expression in cluster A and the lowest in cluster C. However, the expression of LAG3 was the highest in cluster C and the lowest in cluster A, showing an expression pattern opposite to that of other immune checkpoint genes. In Cluster A, the expression of genes related to T-cell activity such as CD8A and GZMB was increased. In Cluster C, the expression of genes related to major histocompatibility complex molecules was the highest. Immunohistochemical stains showed modest agreement with the NanoString results but did not help clustering. CONCLUSION: Our results show that the unique expression pattern of LAG3 in DLBCL contrasts with that of other immune checkpoints. We suggest that the combination of anti-PD-1/PD-L1 and anti-LAG-3 blockades in the immunotherapy of DLBCL patients can have a synergistic effect, improving the immunotherapy efficacy and outcome in DLBCL patients.

Mesonephric Adenocarcinoma of the Vagina Harboring TP53 Mutation.

Mesonephric adenocarcinoma (MA) of the female genital tract is a rare but distinct entity, exhibiting unique morphological, immunophenotypical, and molecular characteristics. Vaginal MA is hypothesized to arise from the mesonephric remnants located in the lateral vaginal wall. A 52-year-old woman presented with vaginal bleeding. Physical examination revealed a protruding mass in the left vaginal wall. Pelvic magnetic resonance imaging revealed a 2.5-cm mass arising from the left upper vagina and extending posterolaterally to the extravaginal tissue. The punch biopsy was diagnosed as poorly differentiated adenocarcinoma. She received radical surgical resection. Histologically, the tumor displayed various architectural patterns, including compactly aggregated small tubules, solid cellular sheets, endometrioid-like glands and ducts, intraluminal micropapillae, cribriform structure, and small angulated glands accompanied by prominent desmoplastic stroma. The tubules and ducts possessed hyaline-like, densely eosinophilic intraluminal secretions. The tumor extended to the subvaginal soft tissue and had substantial perineural invasion. Immunostaining revealed positivity for the mesonephric markers, including GATA3, TTF1, and PAX2, while showing very focal and weak positivity for estrogen receptor and negativity for progesterone receptor. Additionally, we observed a complete absence of p53 immunoreactivity. Targeted sequencing analysis revealed that the tumor harbored both activating KRAS p.G12D mutation and truncating TP53 p.E286* mutation. A thorough review of the previous literature revealed that 4.5% (3/67) of vaginal/cervical MAs and 0.9% (1/112) of uterine/ovarian mesonephric-like adenocarcinomas harbor TP53 mutations, indicating that this is very uncommon in malignant mesonephric lesions. In summary, we presented a rare case of vaginal MA uniquely harboring pathogenic TP53 mutation, resulting in p53 aberration.

RNase H is an exo- and endoribonuclease with asymmetric directionality, depending on the binding mode to the structural variants of RNA:DNA hybrids.

RNase H is involved in fundamental cellular processes and is responsible for removing the short stretch of RNA from Okazaki fragments and the long stretch of RNA from R-loops. Defects in RNase H lead to embryo lethality in mice and Aicardi-Goutieres syndrome in humans, suggesting the importance of RNase H. To date, RNase H is known to be a non-sequence-specific endonuclease, but it is not known whether it performs other functions on the structural variants of RNA:DNA hybrids. Here, we used Escherichia coli RNase H as a model, and examined its catalytic mechanism and its substrate recognition modes, using single-molecule FRET. We discovered that RNase H acts as a processive exoribonuclease on the 3' DNA overhang side but as a distributive non-sequence-specific endonuclease on the 5' DNA overhang side of RNA:DNA hybrids or on blunt-ended hybrids. The high affinity of previously unidentified double-stranded (ds) and single-stranded (ss) DNA junctions flanking RNA:DNA hybrids may help RNase H find the hybrid substrates in long genomic DNA. Our study provides new insights into the multifunctionality of RNase H, elucidating unprecedented roles of junctions and ssDNA overhang on RNA:DNA hybrids.

Implantable Photothermal Agents based on Gold Nanorods-Encapsulated Microcube.

Gold nanorods (GNRs) are of great interest in cancer therapy given their ability to ablate tumor cells using deep tissue-penetrating near-infrared light. GNRs coated with tumor-specific moieties have the potential to target tumor tissue to minimize damage to normal tissue. However, perfect targeting is difficult to achieve given that nanoparticles could be broadly dispersed inside the body. Moreover, interaction between targeting groups and biological molecules could lower targeting abilities, resulting in off-target accumulation which might produce nanotoxicity. Here we introduce GNR-encapsulated microcubes (GNR@MCs) that can be utilized as implantable photothermal agents. GNR@MCs are created by encapsulating GNRs in polymeric networks via stop flow lithography (SFL), a one-phase synthesis technique which allows for creation of surfactant-free, uniform particles, and injection of GNR@MCs into the body after a simple rinse step. GNRs are highly packed and firmly encapsulated inside MCs, and entrapped GNRs exhibit optical properties comparable to that of unbound GNRs and photothermal efficiency (58%) in line with that of nano-sized agents (51-95%). Photothermal ablation in murine models is achieved using GNR@MCs stably implanted into the tumor tissue, which suggests that GNR@MCs can be a safe and effective platform for cancer therapy.

Fabrication of dual stimuli-responsive multicompartmental drug carriers for tumor-selective drug release.

There has been increasing attention to the development of multi-stimuli-responsive drug carriers for precisely controlled drug release at target disease areas. In this study, pH- and redox-responsive hybrid drug carriers were fabricated by using both ketal-based acid-cleavable precursors and disulfide-based reducible precursors via stop-flow lithography. pH- and redox-sensitive drug release of the dual stimuli-responsive hybrid particles was confirmed, demonstrating their feasibility for selective and efficient drug release into tumor tissues in acidic and highly reductive environments. It was also found that the drug release rate of the particles was fine-tuned by modulating monomer compositions in the precursor. Importantly, the dual stimuli-responsive hybrid particles exhibited synergistic, controlled drug release in complex stimuli (both pH and redox stimuli) environments. To achieve tumor-selective combination chemotherapy, multicompartmental drug carriers consist of an acid-degradable compartment and a reducible compartment, which can separately encapsulate individual model drugs in each of the compartments. The multicompartmental particles exhibited independent drug release upon exposure to the corresponding stimulus. The dual stimuli-responsive, multicompartmental particles are effective drug carriers for tumor-selective release of a drug cocktail, leading to synergistic combination chemotherapy.

Effects of CYP2C9 genetic polymorphisms on the pharmacokinetics of zafirlukast.

Zafirlukast, a cysteinyl leukotriene receptor antagonist, is indicated for the treatment of patients with mild to moderate asthma. Zafirlukast is metabolized mainly by CYP3A4 and CYP2C9. We investigated the effects of the major CYP2C9 variant alleles in Asian populations, CYP2C9*3 and CYP2C9*13, on the pharmacokinetics of zafirlukast in healthy Korean subjects. A single 20-mg oral dose of zafirlukast was given to 23 Korean male subjects divided into two genotype groups according to CYP2C9 genotypes, CYP2C9EM (n = 11; CYP2C9*1/*1) and CYP2C9IM (n = 12; 9 and 3 carriers of CYP2C9*1/*3 and *1/*13, respectively). Zafirlukast concentrations were determined using a validated HPLC-MS/MS analytical method in plasma samples collected after the drug intake. Compared with the CYP2C9EM group, the Cmax and AUCinf of zafirlukast in the CYP2C9IM group were 1.44- and 1.70-fold higher, respectively (p < 0.01 and p < 0.0001). The CL/F of zafirlukast was 42.8 % lower in the CYP2C9IM group compared with the CYP2C9EM group (p < 0.001). Slightly higher Cmax and AUC, and lower CL/F of zafirlukast were observed in subjects with the CYP2C9*1/*13 genotype compared with the CYP2C9*1/*3 genotype subjects. CYP2C9*3 and CYP2C9*13 alleles significantly affected the plasma concentrations of zafirlukast.

Genome-wide identification of haploinsufficiency in fission yeast.

Abnormal phenotypes resulting from haploinsufficiency (HI) are due to the loss of one allele. Recent studies in budding yeast have shown that HI originates from insufficient protein levels or from a stoichiometric imbalance between subunits of protein complexes. In humans, however, HI often involves transcription factors. Therefore, the species differences in HI and the molecular mechanisms of species-specific HI remain under investigation. In this study, HI in fission yeast was systematically surveyed. HI in fission yeast affected genes related to signaling and to basic cellular processes, as observed in budding yeast. These results suggest that there are species differences in HI and that the HI that occurs in fission yeast is intermediate to and HI in budding yeast and humans.

Cerulenin-mediated apoptosis is involved in adenine metabolic pathway.

Cerulenin, a fatty acid synthase (FAS) inhibitor, induces apoptosis of variety of tumor cells. To elucidate mode of action by cerulenin, we employed the proteomics approach using Schizosaccharomyces pombe. The differential protein expression profile of S. pombe revealed that cerulenin modulated the expressions of proteins involved in stresses and metabolism, including both ade10 and adk1 proteins. The nutrient supplementation assay demonstrated that cerulenin affected enzymatic steps transferring a phosphoribosyl group. This result suggests that cerulenin accumulates AMP and p-ribosyl-s-amino-imidazole carboxamide (AICAR) and reduces other necessary nucleotides, which induces feedback inhibition of enzymes and the transcriptional regulation of related genes in de novo and salvage adenine metabolic pathway. Furthermore, the deregulation of adenine nucleotide synthesis may interfere ribonucleotide reductase and cause defects in cell cycle progression and chromosome segregation. In conclusion, cerulenin induces apoptosis through deregulation of adenine nucleotide biosynthesis resulting in nuclear division defects in S. pombe.