Concurrent targeting of GSK3 and MEK as a therapeutic strategy to treat pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies worldwide. However, drug discovery for PDAC treatment has proven complicated, leading to stagnant therapeutic outcomes. Here, we identify Glycogen synthase kinase 3 (GSK3) as a therapeutic target through a whole-body genetic screening utilizing a '4-hit' Drosophila model mimicking the PDAC genotype. Reducing the gene dosage of GSK3 in a whole-body manner or knocking down GSK3 specifically in transformed cells suppressed 4-hit fly lethality, similar to Mitogen-activated protein kinase kinase (MEK), the therapeutic target in PDAC we have recently reported. Consistently, a combination of the GSK3 inhibitor CHIR99021 and the MEK inhibitor trametinib suppressed the phosphorylation of Polo-like kinase 1 (PLK1) as well as the growth of orthotopic human PDAC xenografts in mice. Additionally, reducing PLK1 genetically in 4-hit flies rescued their lethality. Our results reveal a therapeutic vulnerability in PDAC that offers a treatment opportunity for patients by inhibiting multiple targets.

Impact of the quality of resected thyroid cancer tissue sample on next-generation sequencing testing.

Activating rearranged during transfection (RET) proto-oncogene alterations can be identified using next-generation sequencing (NGS) of tumor DNA/RNA. We assessed factors associated with NGS (Oncomine Dx Target Test [ODxTT]) success for resected thyroid cancer (TC) specimens, including sample age, processing conditions, and DNA/RNA quality. TC samples were from three Japanese hospitals, with sample age <1-<10 years, fixative 10%/15% neutralized buffered formalin (NBF), and fixation time ≤48 h/>48 h-≤72 h. NGS success rate was defined as the percentage of samples returning validated NGS results (RET fusion-positive/negative [RNA] or RET mutation-positive/negative [DNA], detected using ODxTT). DNA/RNA quality was assessed with indexes based on electrophoresis (DNA/RNA integrity number, DV200 ) and quantitative polymerase chain reaction (DNA/RNA integrity score [ddCq/ΔCq]). NGS success rate (N = 202) was 90%/93% (DNA/RNA) overall, 98%-100% (DNA and RNA) for samples <3 years old, and 91% (DNA and RNA) for samples ≥3-<5 years old fixed in 10% NBF for ≤48 h. Multivariate logistic regression analysis identified ddCq and ΔCq as significant predictors of DNA and RNA NGS success rates, respectively. Quality assessment of nucleic acid extracted from archival tissue samples is important for achieving high NGS success rates in clinical practice, especially for samples ≥3 years old.

Polymorphisms of the PD-L1 gene 3'-untranslated region are associated with the expression of PD-L1 in non-small cell lung cancer.

Recent results show that polymorphisms of programmed death ligand 1 (PD-L1, also known as CD274 or B7-H1) might be used as a possible marker for effectiveness of chemotherapy and cancer risk. However, the effect of PD-L1 gene variations on PD-L1 expression remain unclear. Given the post-transcriptional machinery in tumor PD-L1 expression, we investigated single nucleotide polymorphisms (SNPs) in the 3'-untranslated region (3'-UTR) of the PD-L1 gene, rs4143815 and rs4742098, using formalin-fixed paraffin-embedded sections of 154 patients with non-small cell lung cancers (NSCLCs). In rs4143815, the GG genotype showed significant association with PD-L1 expression (P = 0.032). In rs4742098, the AA genotype was significantly associated with histology and PD-L1 expression (P = 0.022 and P = 0.008, respectively). In multivariate logistic regression analysis, the AA genotype in rs4742098 was correlated with PD-L1 expression (odds ratio 0.408, P = 0.048). Interestingly, approximately 10% of the NSCLC cases showed somatic mutation when we compared genotypes of these SNPs between NSCLC tissues and non-tumor tissues from the same patients. In addition, cases with somatic mutation showed higher levels of PD-L1 expression than cases with germline mutation in rs4143815 GG. In conclusion, we demonstrated that the rs4143815 and rs4742098 SNPs in the 3'-UTR of PD-L1 were associated with tumor PD-L1 expression in NSCLCs.

Expression Analysis of Early Metastatic Seeding of Colorectal Cancer.

BACKGROUND: Distant metastasis is the leading cause of death in patients with colorectal cancer (CRC). Tumor dissemination for metastasis formation occurs in advanced cancers and also during early stages of tumorigenesis. Here, we investigated the genes involved in early metastatic seeding of CRC using gene expression analysis. PATIENTS AND METHODS: We performed a cDNA microarray using specimens resected from stages I-II CRC with and without metachronous metastatic recurrence. For the candidate genes, we immunohistochemically validated protein expression using a tissue microarray of stages I-III CRC. RESULTS: The expression of TROP2, VWCE, and BMP7 was upregulated in the recurrence group rather than in the non-recurrence group. Protein expression analysis revealed significant association of these genes with distant metastatic recurrence. The specimens with high expression of BMP7 showed worse recurrence-free survival (RFS; p = 0.02). Those with high expression of TROP2 and VWCE showed worse overall survival (OS) and RFS (TROP2: p = 0.01 and p = 0.03; VWCE: p < 0.05 and p < 0.001, respectively). In the multivariate analysis, high expression of VWCE and BMP7 was an independent predictor of recurrence [VWCE: hazard ratio (HR) 3.41, p < 0.001; BMP7: HR 2.93, p = 0.005]. In contrast, TROP2 was an independent prognostic factor for OS (HR 4.58, p = 0.03). CONCLUSIONS: Gene expression analysis revealed that TROP2, VWCE, and BMP7 were involved in early metastatic seeding. The high expression of these genes may warrant careful surveillance or adjuvant therapy, even in stages I-II CRC cases.

Exhaustion, rather than lack of infiltration and persistence, of CAR-T cells hampers the efficacy of CAR-T therapy in an orthotopic PDAC xenograft model.

Chimeric antigen receptor T-cell (CAR-T) therapy has demonstrated impressive success in the treatment of patients with hematologic tumors yet achieved very limited efficacy for solid tumors due to hurdles unique to solid tumors. It is also noted that the tumor microenvironment composition varies between tumor type, which again imposes unique set of hurdles in each solid tumor. Therefore, elucidation of individual hurdles is key to achieving successful CAR-T therapy for solid tumors. In the present study, we employed an orthotopic human PDAC xenograft model, in which quantitative, spatial and functional dynamics of CAR-T cells in tumor tissues were analyzed to obtain insights into ways of overcoming PDAC related hurdles. Contrary to previous studies that demonstrated a limited persistency and infiltration of CAR-T cells in many solid tumors, they persist and accumulated in PDAC tumor tissues. Ex vivo analysis revealed that CAR-T cells that had been recovered at different time points from mice bearing an orthotopic PDAC tumor exhibited a gradual loss of tumor reactivity. This loss of tumor reactivity of CAR-T cells was associated with the increased expression of AMP-activated protein kinase and Mitofusin 1/ Dynamin-related protein 1 ratio.

Targeted next-generation sequencing of Japanese patients with sinonasal mucosal melanomas identifies frequent NRAS and CTNNB1 mutations.

OBJECTIVE: Mucosal melanoma is a rare malignancy; however, the reported incidence rate of mucosal melanoma is higher in Asians than in Caucasians. Sinonasal mucosal melanoma (SNMM) is an aggressive malignancy with a poor prognosis due to distant metastasis. Systemic therapy with BRAF inhibitor and MEK inhibitor is one of the standards of care for cutaneous melanoma patients with BRAF V600 mutations. However, no molecular targeted therapy for patients with mucosal melanoma has been established. Relatively few studies have described the genetic mutations associated with mucosal melanoma because of its low frequency. Furthermore, to the best of our knowledge, the genetic mutations among Japanese patients have not been reported. Therefore, in the current study, we evaluated the genetic and clinicopathological characteristics of patients with SNMM. METHODS: A total of 18 tissue samples obtained from patients with SNMM were analyzed for genetic mutations based on targeted next-generation sequencing to investigate the driver of tumorigenesis and/or candidate genes for predicting clinical outcomes in SNMM. We also performed immunohistochemistry for patients identified with CTNNB1 mutations. RESULTS: Eight of the 18 (44 %) patients had genetic mutations. The most frequent mutation was NRAS (6/18, 33 %), followed by CTNNB1 (2/18, 11 %) and BRAF (1/18, 5.6 %). One patient had both NRAS and CTNNB1 mutations. Clinical outcomes did not differ significantly between those with and without genetic mutations. NRAS mutations were associated with relatively higher T classification and worse survival rates, although the differences were not significant. The nuclear translocation of ß-catenin was detected in both tumors with CTNNB1 mutations. The amino acid change in the BRAF mutation was K601R in exon 15. In the current study, no BRAF V600 mutations were detected. CONCLUSION: Genetic mutations were not significantly associated with clinical outcomes. However, NRAS mutations may be a prognostic predictor and CTNNB1 mutation may be a treatment effector for immune check inhibitors. A larger prospective study is required to clarify the clinical importance of genetic mutations in patients with SNMM.

Targeted next-generation sequencing for detection of PIK3CA mutations in archival tissues from patients with Klippel-Trenaunay syndrome in an Asian population : List the full names and institutional addresses for all authors.

BACKGROUND: Klippel-Trenaunay syndrome (KTS) is a rare slow-flow combined vascular malformation with limb hypertrophy. KTS is thought to lie on the PIK3CA-related overgrowth spectrum, but reports are limited. PIK3CA encodes p110α, a catalytic subunit of phosphatidylinositol 3-kinase (PI3K) that plays an essential role in the PI3K/AKT/mammalian target of rapamycin (mTOR) signaling pathway. We aimed to demonstrate the clinical utility of targeted next-generation sequencing (NGS) in identifying PIK3CA mosaicism in archival formalin-fixed paraffin-embedded (FFPE) tissues from patients with KTS. RESULTS: Participants were 9 female and 5 male patients with KTS diagnosed as capillaro-venous malformation (CVM) or capillaro-lymphatico-venous malformation (CLVM). Median age at resection was 14 years (range, 5-57 years). Median archival period before DNA extraction from FFPE tissues was 5.4 years (range, 3-7 years). NGS-based sequencing of PIK3CA achieved an amplicon mean coverage of 119,000x. PIK3CA missense mutations were found in 12 of 14 patients (85.7%; 6/8 CVM and 6/6 CLVM), with 8 patients showing the hotspot variants E542K, E545K, H1047R, and H1047L. The non-hotspot PIK3CA variants C420R, Q546K, and Q546R were identified in 4 patients. Overall, the mean variant allele frequency for identified PIK3CA variants was 6.9% (range, 1.6-17.4%). All patients with geographic capillary malformation, histopathological lymphatic malformation or macrodactyly of the foot had PIK3CA variants. No genotype-phenotype association between hotspot and non-hotspot PIK3CA variants was found. Histologically, the vessels and adipose tissues of the lesions showed phosphorylation of the proteins in the PI3K/AKT/mTOR signaling pathway, including p-AKT, p-mTOR, and p-4EBP1. CONCLUSIONS: The PI3K/AKT/mTOR pathway in mesenchymal tissues was activated in patients with KTS. Amplicon-based targeted NGS could identify low-level mosaicism from low-input DNA extracted from FFPE tissues, potentially providing a diagnostic option for personalized medicine with inhibitors of the PI3K/AKT/mTOR signaling pathway.

Drosophila Screening Identifies Dual Inhibition of MEK and AURKB as an Effective Therapy for Pancreatic Ductal Adenocarcinoma.

Significant progress has been made in understanding the pathogenesis of pancreatic ductal adenocarcinoma (PDAC) by generating and using murine models. To accelerate drug discovery by identifying novel therapeutic targets on a systemic level, here we generated a Drosophila model mimicking the genetic signature in PDAC (KRAS, TP53, CDKN2A, and SMAD4 alterations), which is associated with the worst prognosis in patients. The '4-hit' flies displayed epithelial transformation and decreased survival. Comprehensive genetic screening of their entire kinome revealed kinases including MEK and AURKB as therapeutic targets. Consistently, a combination of the MEK inhibitor trametinib and the AURKB inhibitor BI-831266 suppressed the growth of human PDAC xenografts in mice. In patients with PDAC, the activity of AURKB was associated with poor prognosis. This fly-based platform provides an efficient whole-body approach that complements current methods for identifying therapeutic targets in PDAC. SIGNIFICANCE: Development of a Drosophila model mimicking genetic alterations in human pancreatic ductal adenocarcinoma provides a tool for genetic screening that identifies MEK and AURKB inhibition as a potential treatment strategy.

Deep fibrous histiocytoma of the index finger: a case report.

Our patient presented with an elastic soft mass of his left index finger. Hematoxylin and eosin staining showed a high cellular density with spindle-shaped cells in a storiform pattern. Immunohistochemical staining was positive for CD68, factor XIIIa and α-smooth muscle actin, and negative for CD34, STAT6, S100 protein, and desmin.

Post-transplant donor-specific anti-HLA antibodies with a higher mean fluorescence intensity are associated with graft fibrosis in pediatric living donor liver transplantation.

The roles of post-transplant anti-HLA donor specific antibody (DSA) in pediatric liver transplantation (LT), including therapeutic strategies, remain controversial. This study aimed to identify the risks of post-transplant DSA for graft fibrosis progression in pediatric living donor LT (LDLT). We retrospectively evaluated 88 LDLT pediatric cases between December 1995 and November 2019. DSAs were assessed with single antigen bead test. Graft fibrosis was histopathologically scored with METAVIR and the centrilobular sinusoidal fibrosis system. Post-transplant DSAs were detected in 37 (52.9%) cases at 10.8 (1.3-26.9) years post-LDLT. The histopathological examination of 32 pediatric cases with post-transplant DSA revealed that 7 (21.9%) with a high DSA-MFI (≥9,378) showed graft fibrosis progression (≥F2). No graft fibrosis was observed in the subjects with a low DSA-MFI. The risk factors for developing graft fibrosis in pediatric cases with post-transplant DSA were an older graft age (>46.5 years old), lower platelet count (<10.7 × 104/ml) and higher Fib4 index (>0.7807, recipient age; >1.8952, donor age). Limited efficacy of additional immunosuppressants was observed in DSA positive pediatric cases. In conclusion, pediatric cases with a high DSA-MFI and risk factors should undergo a histological examination. The appropriate treatment for post-transplant DSA in pediatric LT needs to be determined.

Assessment for the timing of comprehensive genomic profiling tests in patients with advanced solid cancers.

Comprehensive genomic profiling (CGP) tests have been covered by public insurance in Japan for patients with advanced solid tumors who have completed or are completing standard treatments or do not have them. Therefore, genotype-matched drug candidates are often unapproved or off-label, and improving clinical trial access is critical, involving the appropriate timing of CGP tests. To address this issue, we analyzed the previous treatment data for 441 patients from an observational study on CGP tests discussed by the expert panel at Hokkaido University Hospital between August 2019 and May 2021. The median number of previous treatment lines was two; three or more lines accounted for 49%. Information on genotype-matched therapies was provided to 277 (63%). Genotype-matched clinical trials were ineligible because of an excess number of previous treatment lines or use of specific agents were found in 66 (15%) patients, with the highest proportion in breast and prostate cancers. Many patients met the exclusion criteria of one to two or more treatment lines across cancer types. In addition, previous use of specific agents was a frequent exclusion criterion for breast, prostate, colorectal, and ovarian cancers. The patients with tumor types with a low median number (two or fewer) of previous treatment lines, including most rare cancers, primary unknown cancers, and pancreatic cancers, had significantly fewer ineligible clinical trials. The earlier timing of CGP tests may improve access to genotype-matched clinical trials, with their proportion varying by cancer type. Each relevant society needs to advocate the desirable timing of CGP testing nationwide.

Programmed death ligand 1-positive immune cells in primary tumor or metastatic axillary lymph nodes can predict prognosis of triple-negative breast cancer even when present at < 1% in the tumor region.

BACKGROUND: The efficacy of pre-operative systemic treatment (PST) combined with immune checkpoint inhibition (ICI) for triple-negative breast cancer (TNBC) has been recognized recently as being independent of the degree of programmed death ligand-1 (PD-L1) positivity of infiltrating immune cells, especially for patients with axillary lymph node metastasis (ALNM). METHODS: TNBC patients with ALNM were treated surgically between 2002 and 2016 in our facility (n = 109), of whom 38 received PST before resection. The presence of tumor-infiltrating lymphocytes (TILs) expressing CD3, CD8, CD68, PD-L1 (detected by antibody SP142) and FOXP3 at primary and metastatic LN sites was quantified. RESULTS: The size of invasive tumor and the number of metastatic axillary LN were confirmed as prognostic markers. The numbers of both CD8+ and FOXP3+ TILs at primary sites were also recognized as prognostic markers, especially for overall survival (OS) (CD8, p = 0.026; FOXP3, p < 0.001). The presence of CD8+, FOXP3+ and PD-L1+ cells was better maintained in LN after PST and may contribute to improved antitumor immunity. Provided they were present as clusters of ≥ 70 positive cells, even < 1% of immune cells expressing PD-L1 at primary sites predicted a more favorable prognosis for both disease-free survival (DFS) (p = 0.004) and OS (p = 0.020). This was the case not only for 30 matched surgical patients, but also in all 71 surgical only patients (DFS: p < 0.001 and OS: p = 0.002). CONCLUSIONS: PD-L1+ , CD8+ or FOXP3+ immune cells in the tumor microenvironment (TME) at both primary and metastatic sites are significant on prognosis, which could be a clue to expect the potential for better responses to the combination of chemotherapy and ICI, especially for patients with ALNM.

CCR7 Mediates Cell Invasion and Migration in Extrahepatic Cholangiocarcinoma by Inducing Epithelial-Mesenchymal Transition.

The epithelial-mesenchymal transition (EMT) contributes to the metastatic cascade in various tumors. C-C chemokine receptor 7 (CCR7) interacts with its ligand, chemokine (C-C motif) ligand 19 (CCL19), to promote EMT. However, the association between EMT and CCR7 in extrahepatic cholangiocarcinoma (EHCC) remains unknown. This study aimed to elucidate the prognostic impact of CCR7 expression and its association with clinicopathological features and EMT in EHCC. The association between CCR7 expression and clinicopathological features and EMT status was examined via the immunohistochemical staining of tumor sections from 181 patients with perihilar cholangiocarcinoma. This association was then investigated in TFK-1 and EGI-1 EHCC cell lines. High-grade CCR7 expression was significantly associated with a large number of tumor buds, low E-cadherin expression, and poor overall survival. TFK-1 showed CCR7 expression, and Western blotting revealed E-cadherin downregulation and vimentin upregulation in response to CCL19 treatment. The wound healing and Transwell invasion assays revealed that the activation of CCR7 by CCL19 enhanced the migration and invasion of TFK-1 cells, which were abrogated by a CCR7 antagonist. These results suggest that a high CCR7 expression is associated with an adverse postoperative prognosis via EMT induction and that CCR7 may be a potential target for adjuvant therapy in EHCC.

Guidelines for Handling of Cytological Specimens in Cancer Genomic Medicine.

Rapid advances are being made in cancer drug therapy. Since molecularly targeted therapy has been introduced, personalized medicine is being practiced, pathological tissue from malignant tumors obtained during routine practice is frequently used for genomic testing. Whereas cytological specimens fixed mainly in alcohol are considered to be more advantageous in terms of preservation of the nucleic acid quality and quantity. This article is aimed to share the information for the proper handling of cytological specimens in practice for genomic medicine based on the findings established in "Guidelines for Handling of Cytological Specimens in Cancer Genomic Medicine (in Japanese)" published by the Japanese Society of Clinical Cytology in 2021. The three-part practical guidelines are based on empirical data analyses; Part 1 describes general remarks on the use of cytological specimens in cancer genomic medicine, then Part 2 describes proper handling of cytological specimens, and Part 3 describes the empirical data related to handling of cytological specimens. The guidelines indicated proper handling of specimens in each fixation, preparation, and evaluation.

Tumor Growth Suppression of Pancreatic Cancer Orthotopic Xenograft Model by CEA-Targeting CAR-T Cells.

Chimeric antigen receptor engineered T cell (CAR-T) therapy has high therapeutic efficacy against blood cancers, but it has not shown satisfactory results in solid tumors. Therefore, we examined the therapeutic effect of CAR-T therapy targeting carcinoembryonic antigen (CEA) in pancreatic adenocarcinoma (PDAC). CEA expression levels on the cell membranes of various PDAC cell lines were evaluated using flow cytometry and the cells were divided into high, medium, and low expression groups. The relationship between CEA expression level and the antitumor effect of anti-CEA-CAR-T was evaluated using a functional assay for various PDAC cell lines; a significant correlation was observed between CEA expression level and the antitumor effect. We created orthotopic PDAC xenograft mouse models and injected with anti-CEA-CAR-T; only the cell line with high CEA expression exhibited a significant therapeutic effect. Thus, the therapeutic effect of CAR-T therapy was related to the target antigen expression level, and the further retrospective analysis of pathological findings from PDAC patients showed a correlation between the intensity of CEA immunostaining and tumor heterogeneity. Therefore, CEA expression levels in biopsies or surgical specimens can be clinically used as biomarkers to select PDAC patients for anti-CAR-T therapy.

Evaluation of 95-Gene Classifier of Formalin-fixed Paraffin-embedded Tissues in ER-positive, HER2-negative, and Node-negative Breast Cancer.

BACKGROUND/AIM: A subset of patients with estrogen receptor (ER)-positive, HER2-negative, and node-negative breast cancer experience recurrences. Predicting patients who will have recurrences within 5 years of surgery is essential so that patients can be selected to receive adjuvant chemotherapy. The 95-gene classifier (95-GC) has been validated as a method to differentiate patients into high and low-risk groups for early recurrence. PATIENTS AND METHODS: In this study, we performed 95-GC analysis on 56 formalin-fixed paraffin-embedded (FFPE) tissue samples from patients who underwent surgery for ER-positive, HER2-negative, and node-negative breast cancer and did not receive adjuvant chemotherapy. We associated the obtained high- and low-risk groups with clinicopathological characteristics and recurrence-free survival (RFS). RESULTS: We classified 12 out of 56 patients into the high-risk recurrence group. We found significantly higher KI67 scores in patients in the high-risk group. Other clinicopathological characteristics were not associated with the 95-GC risk groups. Patients in the 95-GC low-risk group had a significantly better prognosis than those in the high-risk group (p=0.0387). The 5-year RFS rate was 97.6% in the low-risk group and 74.1% in the high-risk group, while the 10-year RFS rates were 90.1% and 74.1%, respectively. CONCLUSION: The 95-GC score can accurately predict RFS within 5 years of surgery for ER-positive, HER2-negative, and node-negative breast cancer using FFPE tissue samples. These prediction models could help assign patients to the most effective treatment regimen.

Notch pathway regulates osimertinib drug-tolerant persistence in EGFR-mutated non-small-cell lung cancer.

Osimertinib is a third-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI) that has shown marked antitumor activity in patients with EGFR-mutated non-small-cell lung cancer (NSCLC). However, these effects are transient and most patients develop resistance. Reversible drug-tolerant persister (DTP) cells are defined as a small subpopulation of cells with markedly reduced sensitivity and non-genetic acquired resistance to EGFR-TKIs. Notch is a transmembrane receptor that plays an important role in tumorigenesis. We previously reported that there is significant crosstalk between the Notch and EGFR pathways in NSCLC. Moreover, the Notch pathway is associated with resistance to previous-generation EGFR-TKIs. However, the role of Notch in osimertinib resistance is not fully understood. In this study, we evaluated whether Notch is involved in osimertinib resistance. We show that NOTCH1 and Notch target genes are upregulated in osimertinib DTP cells, and that the addition of a γ-secretase inhibitor (GSI), a Notch inhibitor, impairs drug-tolerant persistence in vitro and in vivo. Compared with osimertinib, combined GSI and osimertinib suppress phospho-ERK partly by enhancing DUSP1 expression. Furthermore, Notch1 and HES1 were upregulated after EGFR-TKI treatment in half of human EGFR-mutated NSCLC tumor tissues. These results suggest that the combination of GSI and osimertinib may be a potential therapy for EGFR-mutated NSCLC.

Impact of RNF213 c.14576G>A Variant on the Development of Direct and Indirect Revascularization in Pediatric Moyamoya Disease.

BACKGROUND: Superficial temporal artery (direct) and encephalomyosynangiosis (indirect) revascularization may develop after combined bypass surgery in pediatric patients with moyamoya disease (MMD). However, arterial development varies widely among patients, and the underlying mechanisms remain unknown. OBJECTIVES: We evaluated the relationship between the development of donor arteries after bypass surgery in pediatric patients with MMD and the MMD-susceptibility gene variant c.14576G>A of ring finger protein (RNF) 213. METHODS: The data of pediatric patients with MMD (age <16 years at the time of surgery) treated with combined bypass surgery between September 2013 and April 2019 were consecutively analyzed. Quantitative measurements of the superficial temporal artery (STA), deep temporal artery (DTA), and middle meningeal artery (MMA) diameters with magnetic resonance angiography (MRA) source imaging were performed preoperatively and at 6-12 months postoperatively. The postoperative caliber change ratios (CCRs) were calculated. The relationship between CCRs and RNF213 c.14576G>A status was examined. RESULTS: Forty-eight hemispheres from 28 pediatric patients with MMD were examined. Three hemispheres belonged to patients with the AA genotype; 33 to patients with the AG genotype (AA/AG group); and 12 to patients with the GG genotype (GG group; wild type). The CCRs for the DTA were significantly higher in patients with RNF213 variant (AA/AG group; 2.5 ± 0.1) than in the GG group (2.0 ± 0.2) (p = 0.03), whereas the CCRs for the STA were significantly higher in the GG (1.6 ± 0.1) than in the AA/AG group (1.3 ± 0.6) (p = 0.02). There was no significant difference in the CCRs for the MMA and basilar artery between the groups. Other factors, including sex, age, and MRA grading, were not associated with the development of specific bypass development. CONCLUSIONS: The extent of collateral development associated with direct or indirect bypass was found to differ between the genotypes of the RNF213 c.14576G>A associated with pediatric MMD. This genetic variant correlates with the development of the disease and affects revascularization after bypass surgery in pediatric patients with MMD.

Germline PRDM1 Variant rs2185379 in Long-Term Recurrence-Free Survivors of Advanced Ovarian Cancer.

Purpose: To identify the germline genetic characteristics of long-term recurrence-free survivors that can be applied to establishing a new strategy for curing advanced cancer, we investigated the whole-genome single nucleotide variants of ovarian cancer patients. Patients and Methods: DNA specimens were obtained from rare long-term recurrence-free survivors with FIGO stage III-IV ovarian cancer with no recurrence for 8-23 years after primary treatments for a whole-genome analysis of approximately 660,000 single nucleotide variants. We then established a mouse model with a notable gene alteration by CRISPR/Cas9 to confirm the biological role. Results: The long-term recurrence-free survivors more frequently had germline heterozygous variant rs2185379 of the PRDM1 gene exon than patients with early recurrence (6.8-fold, P=0.013) and the general population. In the mouse model, primary intraperitoneal disseminated tumors of allograft ID8 were significantly smaller in the germline heterozygous rs2185379 group than in the wild-type group (57.4% decrease, P=0.008). Immunohistochemistry showed that the area of distribution of infiltrating T lymphocytes with positive CD8 staining was significantly increased in the germline heterozygous rs2185379 group in comparison to the wild-type group. Conclusion: Germline heterozygous rs2185379 in PRDM1 is correlated with an excellent prognosis and can be used to establish a new strategy for treating advanced ovarian cancer.