A case report of immune checkpoint inhibitor-related myositis and cholangitis induced by pembrolizumab.

Key Clinical Message: Rare but severe, immune-related adverse events such as myositis and sclerosing cholangitis can occur with immune checkpoint inhibitors in lung cancer treatment. This case report highlights their co-occurrence after pembrolizumab treatment, indicating the need for vigilance and management strategies in immune checkpoint inhibitors therapy. Abstract: Immune checkpoint inhibitors (ICI) are used in advanced treatment of lung cancer but can lead to immune-related adverse events. ICI-related myositis and cholangitis are rare, and their combination has not been previously reported. Here, we report the first case of ICI-related myositis and sclerosing cholangitis. A patient with stage IV lung adenocarcinoma who received one cycle of pembrolizumab with cisplatin and pemetrexed developed myositis. Treatment with prednisolone improved the myositis, but the patient subsequently developed cholangitis. The patient did not respond to a regimen of prednisolone, mycophenolate mofetil, and azathioprine, and eventually died due to worsening lung cancer. An autopsy confirmed the presence of ICI-related myositis and sclerosing cholangitis.

Rapidly Growing Locally Advanced Non-Small Cell Lung Cancer Treated with Definitive Chemoradiotherapy Using Adaptive Volumetric Modulated Arc Therapy Followed by Durvalumab Maintenance: A Case Report.

BACKGROUND It is difficult to reduce lung toxicity in chemoradiotherapy for locally advanced lung cancer. Volume-modulated arc therapy (VMAT) is a useful lung dose-lowering radiation technique, but it is time-consuming because of its complexity. We present a case of a rapidly growing bulky lung cancer treated with VMAT and intensive adaptation to volume change. CASE REPORT A 43-year-old man with chest pain was diagnosed with non-small cell lung cancer, cT4N3M0 stage IIIC (UICC 8th edition). Concurrent chemoradiotherapy with a VMAT of 60 Gy in 30 fractions and carboplatin/paclitaxel was performed. Despite initiating chemoradiation, monitoring with cone-beam computed tomography (CT) revealed tumor progression. The peak tumor volume was 1.5 times larger than that on CT simulation. The VMAT plan was recreated to cover the increased tumor size. After the irradiation field was enlarged, the tumor, on the contrary, shrank rapidly. Therefore, VMAT planning was performed again to further shrink the irradiation field. CT at the end of the treatment showed a good volume reduction response. Durvalumab therapy was continued for 1 year. After that, the patient was alive and showed no sign of progression. Only asymptomatic radiation pneumonitis was observed as a sub-acute adverse event. CONCLUSIONS We present a case in which proper adaptive VMAT and durvalumab for dramatically progressive non-small cell lung cancer were effective, resulting in 1-year progression-free survival. Even when rapid progression of bulky lung cancer is suggested, the combination of VMAT and adaptive radiotherapy with improved target coverage and reduced lung dose can be a treatment option.

Pedunculated pulmonary artery intimal sarcoma with poor uptake in 18F-FDG PET/CT: A case report.

Pulmonary artery intimal sarcoma (PAIS) is a rare tumor with an incidence of 0.001%-0.03% that usually grows along artery walls and absorbs fluorodeoxyglucose. It is difficult to distinguish PAIS from pulmonary thromboembolism due to the similarities of their symptoms. Therefore, contrast-enhanced computed tomography and positron emission tomography-computed tomography (PET-CT) should be used to establish a correct diagnosis. Here we report a case of an extremely rare type of PAIS, pedunculated PAIS, which could not be visualized on PET-CT. Histological features of a tumor with a low accumulation of fluorodeoxyglucose revealed low-cellularity and necrotizing background. Multimodal imaging was useful to diagnose PET-CT negative PAIS accurately. .

Bleeding from pancreatic arteriovenous malformation with duodenal ulcer penetration. Case report and literature review.

Dilated vessels at the ulcer floor of the second part of the duodenum can be signs of pancreatic arteriovenous malformation; contrast-enhanced computed tomography should be performed, and surgical treatment should be considered.

Homeodomain-Interacting Protein Kinase-2: A Critical Regulator of the DNA Damage Response and the Epigenome.

Homeodomain-interacting protein kinase 2 (HIPK2) is a serine/threonine kinase that phosphorylates and activates the apoptotic program through interaction with diverse downstream targets including tumor suppressor p53. HIPK2 is activated by genotoxic stimuli and modulates cell fate following DNA damage. The DNA damage response (DDR) is triggered by DNA lesions or chromatin alterations. The DDR regulates DNA repair, cell cycle checkpoint activation, and apoptosis to restore genome integrity and cellular homeostasis. Maintenance of the DDR is essential to prevent development of diseases caused by genomic instability, including cancer, defects of development, and neurodegenerative disorders. Recent studies reveal a novel HIPK2-mediated pathway for DDR through interaction with chromatin remodeling factor homeodomain protein 1γ. In this review, we will highlight the molecular mechanisms of HIPK2 and show its functions as a crucial DDR regulator.

HuR regulates alternative splicing of the TRA2ß gene in human colon cancer cells under oxidative stress.

Hu antigen R (HuR) regulates stress responses through stabilizing and/or facilitating the translation of target mRNAs. The human TRA2ß gene encodes splicing factor transformer 2ß (Tra2ß) and generates 5 mRNA isoforms (TRA2ß1 to -5) through alternative splicing. Exposure of HCT116 colon cancer cells to sodium arsenite stimulated checkpoint kinase 2 (Chk2)- and mitogen-activated protein kinase p38 (p38(MAPK))-mediated phosphorylation of HuR at positions S88 and T118. This induced an association between HuR and the 39-nucleotide (nt) proximal region of TRA2ß exon 2, generating a TRA2ß4 mRNA that includes exon 2, which has multiple premature stop codons. HuR knockdown or Chk2/p38(MAPK) double knockdown inhibited the arsenite-stimulated production of TRA2ß4 and increased Tra2ß protein, facilitating Tra2ß-dependent inclusion of exons in target pre-mRNAs. The effects of HuR knockdown or Chk2/p38(MAPK) double knockdown were also confirmed using a TRA2ß minigene spanning exons 1 to 4, and the effects disappeared when the 39-nt region was deleted from the minigene. In endogenous HuR knockdown cells, the overexpression of a HuR mutant that could not be phosphorylated (with changes of serine to alanine at position 88 [S88A], S100A, and T118A) blocked the associated TRA2ß4 interaction and TRA2ß4 generation, while the overexpression of a phosphomimetic HuR (with mutations S88D, S100D, and T118D) restored the TRA2ß4-related activities. Our findings revealed the potential role of nuclear HuR in the regulation of alternative splicing programs under oxidative stress.

Oxidative stress-inducible truncated serine/arginine-rich splicing factor 3 regulates interleukin-8 production in human colon cancer cells.

Serine/arginine-rich splicing factor 3 (SRSF3) is a member of the SR protein family and plays wide-ranging roles in gene expression. The human SRSF3 gene generates two alternative splice transcripts, a major mRNA isoform (SRSF3-FL) encoding functional full-length protein and a premature termination codon (PTC)-containing isoform (SRSF3-PTC). The latter is degraded through nonsense-mediated mRNA decay (NMD). Treatment of a human colon cancer cell line (HCT116) with 100 µM sodium arsenite increased SRSF3-PTC mRNA levels without changing SRSF3-FL mRNA levels. A chemiluminescence-based NMD reporter assay system demonstrated that arsenite treatment inhibited NMD activity and increased SRSF3-PTC mRNA levels in the cytoplasm, facilitating translation of a truncated SRSF3 protein (SRSF3-TR) from SRSF3-PTC mRNA. SRSF3-TR lacked two-thirds of the Arg/Ser-rich (RS) domain whose phosphorylation state is known to be crucial for subcellular distribution. SRSF3-FL was localized in the nucleus, while overexpressed SRSF3-TR was diffusely distributed in the cytoplasm and the nucleus. A part of SRSF3-TR was also associated with stress granules in the cytoplasm. Interestingly, treatment of HCT116 cells with a small interference RNA specifically targeting SRSF3-PTC mRNA significantly attenuated arsenite-stimulated induction of c-JUN protein, its binding activity to the AP-1 binding site (-126 to 120 bp) in the interleukin (IL)-8 gene promoter, and AP-1 promoter activity, resulting in significant reduction of arsenite-stimulated IL-8 production. Our results suggest that SRSF3-TR may function as a positive regulator of oxidative stress-initiated inflammatory responses in colon cancer cells.

Truncated serine/arginine-rich splicing factor 3 accelerates cell growth through up-regulating c-Jun expression.

Serine/arginine-rich splicing factor 3 (SRSF3), a member of the SRSF family, plays a wide-ranging role in gene expression. The human SRSF3 gene generates a major mRNA isoform encoding a functional, full-length protein and a PTC-containing isoform (SRSF3-PTC). The latter is expected to be degraded through the nonsense-mediated mRNA decay system. However, it was reported that SRSF3-PTC mRNA was produced under stressful conditions and translated into a truncated SRSF3 protein (SRSF3-TR). To disclose unknown functions of SRSF3-TR, we established Flp-In-293 cells stably expressing SRSF3-TR. The SRSF3-TR-expressing cells increased mRNA and protein levels of positive regulators for G1 to S phase transition (cyclin D1, cyclin D3, CDC25A, and E2F1) and accelerated their growth. c-Jun is required for progression through the G1 phase, the mechanism by which involves transcriptional control of the cyclin D1 gene. We also found that the JUN promoter activity was significantly increased in the Flp-In-293 cells stably expressing SRSF3-TR, compared with mock-transfected control cells. The SRSF3-TR-expressing cells increased c-Jun and Sp-1 levels, which are important for the positive autoregulation and basal transcription of JUN, respectively. Our results suggest that stress-inducible SRSF3-TR may participate in the acceleration of cell growth through facilitating c-Jun-mediated G1 progression under stressful conditions.

Chronic academic stress increases a group of microRNAs in peripheral blood.

MicroRNAs (miRNAs) play key roles in regulation of cellular processes in response to changes in environment. In this study, we examined alterations in miRNA profiles in peripheral blood from 25 male medical students two months and two days before the National Examination for Medical Practitioners. Blood obtained one month after the examination were used as baseline controls. Levels of seven miRNAs (miR-16, -20b, -26b, -29a, -126, -144 and -144*) were significantly elevated during the pre-examination period in association with significant down-regulation of their target mRNAs (WNT4, CCM2, MAK, and FGFR1 mRNAs) two days before the examination. State anxiety assessed two months before the examination was positively and negatively correlated with miR-16 and its target WNT4 mRNA levels, respectively. Fold changes in miR-16 levels from two days before to one month after the examination were inversely correlated with those in WNT4 mRNA levels over the same time points. We also confirmed the interaction between miR-16 and WNT4 3'UTR in HEK293T cells overexpressing FLAG-tagged WNT4 3'UTR and miR-16. Thus, a distinct group of miRNAs in periheral blood may participate in the integrated response to chronic academic stress in healthy young men.

Ets1 and heat shock factor 1 regulate transcription of the Transformer 2ß gene in human colon cancer cells.

BACKGROUND: Transformer (Tra) 2ß is a member of the serine/arginine-rich (SR)-like protein family that regulates alternative splicing of numerous genes in a concentration-dependent manner. Several types of cancer cells up-regulate Tra2ß expression, while the regulatory mechanism of Tra2ß expression remains to be elucidated. In this study, we examined the transcriptional regulation and possible functions of Tra2ß in human colon cancer cells. METHODS: We cloned 959 bp-upstream of the human TRA2ß 5'-flank into luciferase constructs. Chromatin immunoprecipitation (ChIP) was employed to identify crucial cis element(s) and trans activator(s) of the TRA2ß promoter. Tra2ß expression in the human colon and colon cancer tissues was examined by immunohistochemistry. RESULTS: In response to sodium arsenite, colon cancer cells (HCT116) increased levels of TRA2ß1 mRNA encoding a functional, full-length Tra2ß with a peak around 6 h without changing its mRNA stability. Transient expression assays using a reporter gene driven by serially truncated TRA2ß promoters and Chip assay demonstrated that an Ets1-binding site present at -64 to -55 bp was crucial for basal transcription, while three heat shock elements (HSEs) located at -145 to -99 bp mediated the oxidant-induced transactivation of TRA2ß. Tra2ß knockdown caused apoptosis of HCT116 cells. Tra2ß were preferentially expressed in proliferative compartment of normal human colonic glands and adenocarcinomas, where Ets1 and heat shock factor 1 were also highly expressed. CONCLUSIONS: Our results suggest that oxidative stress-responsive Tra2ß may play an important role in colon cancer growth.

MicroRNAs miR-144/144* and miR-16 in peripheral blood are potential biomarkers for naturalistic stress in healthy Japanese medical students.

Non-coding microRNAs (miRNAs) are suggested to serve fundamental roles in cellular stress responses and in coping with sudden environmental changes in experimental animals. We examined whether naturalistic stressor-responsive miRNAs were detectable in whole blood. Blood and saliva were collected between 16:00 and 17:00 from 10 healthy medical students (5 males and 5 females; aged 22.4±0.8 years, mean±SD) 7 weeks before, one day before, immediately after, and one week after a nationally administered examination for academic promotion. Samples obtained one week after the examination were used as baseline controls. State anxiety and salivary cortisol levels reached maximum levels the day before the examination. Eleven candidate miRNAs (miR-144, -144*, -16, -15a, -19a, -19b, -26b, -30b, -106b, -126, and -142-3p) were extracted using a human miRNA microarray, and quantitative real-time reverse transcription PCR confirmed significant elevation of miR-144/144* and miR-16 levels immediately after finishing the examination. miR-16 levels in individual students were positively correlated with those of serum tumor necrosis factor (TNF)-α measured immediately after the examination. Percentage changes in miR-144* and miR-16 levels from immediately after to one week after the examination were significantly correlated with percentage changes in circulating interferon-γ and/or TNF-α levels over the same time points. Our results suggest that miR-144/144* and miR-16 may constitute a part of an integrated response to naturalistic stressors in healthy young adults.

Role of miR-19b and its target mRNAs in 5-fluorouracil resistance in colon cancer cells.

BACKGROUND: Drug resistance in colorectal cancers is assumed to be mediated by changes in the expression of microRNAs, but the specific identities and roles of microRNAs are largely unclear. We examined the effect of 5-fluorouracil (5-FU) resistance on microRNA expression. METHODS: Two types of 5-FU-resistant colon cancer cells were derived from the DLD-1 and KM12C cell lines. The expressions of microRNAs were profiled with a microarray containing 723 microRNAs and validated by quantitative real-time polymerase chain reaction (qRT-PCR). To survey the downstream mediators of microRNA, we used a microRNA:mRNA immunoprecipitation (RIP)-Chip and pathway analysis tool to identify potential direct targets of microRNA. RESULTS: In response to 5-FU, miR-19b and miR-21 were over-expressed in 5-FU-resistant cells. Of note, miR-19b was up-regulated 3.47-fold in the DLD-1 resistant cells, which exhibited no alteration in cell cycle profiles despite exposure to 5-FU. After transfection of miR-19b, specific mRNAs were recruited to microRNA:mRNA complexes isolated with Ago2 antibody and subjected to whole-genome transcriptional analysis. In this analysis, 66 target mRNAs were enriched by at least 5.0-fold in the microRNA:mRNA complexes from DLD-1 resistant cells. Ingenuity pathway analysis of mRNA targets significantly (P < 0.05) indicated the category "Cell Cycle" as a probable area of the molecular and cellular function related with 5-FU resistance. Among candidate mRNA targets, SFPQ and MYBL2 have been linked to cell cycle functions. CONCLUSIONS: We revealed up-regulation of miR-19b in response to 5-FU and potential targets of miR-19b mediating the cell cycle under treatment with 5-FU. Our study provides an important insight into the mechanism of 5-FU resistance in colorectal cancers.

Skipping of an alternative intron in the srsf1 3' untranslated region increases transcript stability.

The srsf1 gene encodes serine/arginine-rich splicing factor 1 (SRSF1) that participates in both constitutive and alternative splicing reactions. This gene possesses two ultraconserved elements in the 3' untranslated region (UTR). Skipping of an alternative intron between the two elements has no effect on the protein-coding sequence, but it generates a premature stop codon (PTC)-containing mRNA isoform, whose degradation is considered to depend on nonsense-mediated mRNA decay (NMD). However, several cell lines (HCT116, RKO, HeLa, and WI38 cells) constitutively expressed significant amounts of the srsf1 PTC variant. HCT116 cells expressed the PTC variant nearly equivalent to the major isoform that includes the alternative intron in the 3' UTR. Inhibition of NMD by silencing a key effecter UPF1 or by treatment with cycloheximide failed to increase amounts of the PTC variant in HCT116 cells, and the PTC variant was rather more stable than the major isoform in the presence of actinomycin D. Our results suggest that the original stop codon may escape from the NMD surveillance even in skipping of the alternative intron. The srsf1 gene may produce an alternative splice variant having truncated 3' UTR to relief the microRNA- and/or RNA-binding protein-mediated control of translation or degradation.

Effects of chronic academic stress on mental state and expression of glucocorticoid receptor α and ß isoforms in healthy Japanese medical students.

Chronic academic stress responses were assessed by measuring mental state, salivary cortisol levels, and the glucocorticoid receptor (GR) gene expression in healthy Japanese medical students challenging the national medical license examination. Mental states of 17 male and 9 female medical undergraduates, aged 25.0 ± 1.2 years (mean ± SD), were assessed by the State and Trait Anxiety Inventory (STAI) and the Self-Rating Depression Scale (SDS) 2 months before, 2 days before, and 1 month after the examination. At the same time points, saliva and blood were collected. STAI-state scores peaked 2 days before the examination. Scores on STAI-trait and SDS, and salivary cortisol levels were consistently higher during the pre-examination period. One month after the examination, all these measures had significantly decreased to baseline levels. Real-time reverse transcription PCR showed that this chronic anxious state did not change the expression of the functional GRα mRNA isoform in peripheral leukocytes, while it resulted in reduced expression of the GRß isoform 2 days before the examination. Our results replicate and extend a significant impact of chronic academic stressors on the mental state of healthy Japanese medical students and suggest a possible association of GRß gene in response to psychological stress.