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PurposeAnalysis of circulating tumor DNA (ctDNA) in blood could allow noninvasive genetic analysis of primary tumors. Although there have been unmet needs for noninvasive methods in patients with primary central nervous system lymphoma (PCNSL), it is still not determined whether plasma ctDNA analysis could be useful for patients with PCNSL. Materials and MethodsTargeted deep sequencing of 54 genes was performed in cell-free DNA isolated from plasma samples collected pretreatment, during treatment, and at the end of treatment in 42 consecutively diagnosed PCNSL patients between January 2017 and December 2018.ResultsTargeted sequencing of plasma cell-free DNA detected somatic mutations representing ctDNA in 11 cases (11/41, 27%). The detection of ctDNA was not related to the concentration of cell-free DNA or tumor volume. The mutation profiles of these 11 cases varied between patients. The most frequently mutated gene was PIM1 (4/11, 36.4%), whereas KMT2D, PIK3CA, and MYD88 were each observed in three patients (3/11, 27%). The mutations of 13 genes were concordantly found in primary tumor tissue and plasma ctDNA, giving a detection sensitivity of 45%. During the serial tracking of seven patients with complete response, the disappearance of ctDNA mutations was found in four patients, whereas three patients had detected ctDNA mutation at the end of treatment. ConclusionThe plasma ctDNA mutation analysis still has limited value for surveillance and predicting treatment outcomes of PCNSL because the detection efficiency was lower than other systemic lymphomas. Thus, analytical platforms should be improved to overcome anatomical hurdles associated with PCNSL.

Malignant tumours arising from the sublingual glands are very rare, and the extent and frequency of local invasion or regional spread in malignant sublingual gland tumour (MSLT) has not been fully studied due to the disease rarity. To provide comprehensive features of local and regional spread of MSLT, we reviewed 20 surgical cases for detailed pathological analyses among 26 cases diagnosed as having primary MSLT. Adenoid cystic carcinoma (ACC) was the most common pathological subtype, followed by mucoepidermoid carcinoma. Disease-free and overall survivals at 5 years were 76.1 % and 77.7 %, respectively. High-grade malignant tumours and grade 2-3 ACC accounted for 41.7 % and 85.7 %. Clinical and pathological extraparenchymal extensions were found in 34.6 % and 80.0 %, respectively. Tumour invasion to the lingual nerve and submandibular gland/ductal system were also detected in 40.0 % and 28.6 %. The incidences of lingual nerve invasion in ACC and ACC ≥4 cm were 30.8 % and 42.9 %. Regional nodal involvement occurred in seven of 26 cases, and all metastatic lymph nodes were found in neck levels Ib and IIa. In summary, a significant portion of MSLT cases consisted of high-grade tumours and grade 2-3 ACC; therefore local invasion into adjacent structures should be cautiously evaluated in cases of MSLT.

To investigate whether women with a short cervical length (CL), but delivered at term in the first pregnancy might have increased risks of having short cervix and spontaneous preterm birth (sPTB) in the subsequent pregnancies.This is a retrospective cohort study including singleton gestations who were delivered between Jan 2011 and Dec 2018, who had had one childbirth experience and who had transvaginal sonographic CL assessment performed at mid-trimester (18 ~ 30 weeks) in both pregnancy. The women were divided into four group according to the history of preterm birth and a short cervix ( 25 mm before 30 weeks of gestation): (1) the Preterm-short cervix group, (2) the Preterm-no shortening group, (3) the Term-short cervix group, and (4) the Term-no shortening group. We compared the risk of having short cervix and sPTB during the second pregnancy of women. Secondary outcomes were threatened preterm labor, need for tocolytics, and cerclage placement.A total of 804 women met our inclusion criteria. The rate of having short cervix ( 25 mm before 28 weeks of gestation) during the second pregnancy in women in the Term-short cervix group (43.2%) was significantly higher than those in women in the Term-no shortening group (6.6%), and in the Preterm-no shortening group (8.8%) (all p 0.001 with Bonferroni correction), but not higher than those in women with the Preterm-short cervix group (30.8%, p 0.05 with Bonferroni correction). When compared with women in the Preterm-no shortening group, women in the Term-short cervix group were also at an increased risk of need for need of tocolytics (60.2% vs. 26.5%) and cerclage placement (15.9% vs. 6.1%, all p 0.001). Women in the Term-short cervix group had an increased risk of sPTB ( 37 weeks) during the second pregnancy, as compared to those in the Term-no shortening group (adjusted odds ratio 5.098, 95% CI 2.107-11.874).Women with a history of short cervix in their first pregnancy, but who delivered at term, were at increased risk of having a short cervix and sPTB in their second pregnancy, as compared to women with a history of term delivery without cervical shortening. Thus, short cervix of the previous pregnancy might be a predictive factors for preterm birth in the subsequent pregnancy.

Many surgeons have modified the technique to reduce bleeding from anastomosis line since Bentall and De Bono introduced aortic root replacement using a composite graft. We present a new composite, butterfly wing–shaped, valve graft fixated using hand-made double sewing cuff. The “wings of a butterfly” technique is a method of reinforcement suturing between an added sewing cuff and residual aortic wall at the proximal stump and helps to reduce bleeding after the modified Bentall's procedure.

Owing to rapid advancements in NGS (next generation sequencing), genomic alteration is now considered an essential predictive biomarkers that impact the treatment decision in many cases of cancer. Among the various predictive biomarkers, tumor mutation burden (TMB) was identified by NGS and was considered to be useful in predicting a clinical response in cancer cases treated by immunotherapy. In this study, we directly compared the lab-developed-test (LDT) results by target sequencing panel, K-MASTER panel v3.0 and whole-exome sequencing (WES) to evaluate the concordance of TMB. As an initial step, the reference materials (n = 3) with known TMB status were used as an exploratory test. To validate and evaluate TMB, we used one hundred samples that were acquired from surgically resected tissues of non-small cell lung cancer (NSCLC) patients. The TMB of each sample was tested by using both LDT and WES methods, which extracted the DNA from samples at the same time. In addition, we evaluated the impact of capture region, which might lead to different values of TMB; the evaluation of capture region was based on the size of NGS and target sequencing panels. In this pilot study, TMB was evaluated by LDT and WES by using duplicated reference samples; the results of TMB showed high concordance rate (R2 = 0.887). This was also reflected in clinical samples (n = 100), which showed R2 of 0.71. The difference between the coding sequence ratio (3.49%) and the ratio of mutations (4.8%) indicated that the LDT panel identified a relatively higher number of mutations. It was feasible to calculate TMB with LDT panel, which can be useful in clinical practice. Furthermore, a customized approach must be developed for calculating TMB, which differs according to cancer types and specific clinical settings. [BMB Reports 2021; 54(7): 386-391].

Background Although the World Health Organization and health authorities in most countries recommend that pregnant women receive inactivated influenza virus vaccines, coverage remains low. This study aimed to investigate (1) the proportion of pregnant women who received an influenza vaccination and influencing factors and (2) the proportion of obstetrics and gynecology (OBGYN) doctors who routinely recommend influenza vaccination to pregnant women and influencing factors. Methods Two separate, anonymized questionnaires were developed for physicians and pregnant and postpartum women and were distributed to multicenters and clinics in South Korea. The proportions of women who received influenza vaccination during pregnancy and OBGYN doctors who routinely recommend the influenza vaccine to pregnant women were analyzed. Independent influencing factors for both maternal influenza vaccination and OBGYN doctors’ routine recommendations to pregnant women were analyzed using log-binomial regression analysis. Results The proportion of self-reported influenza vaccination during pregnancy among 522 women was 63.2%. Pregnancy-related independent factors influencing maternal influenza vaccination were “(ever) received information about influenza vaccination during pregnancy” (OR 8.9, 95% CI 4.17–19.01), “received vaccine information about from OBGYN doctors” (OR 11.44, 95% CI 5.46–24.00), “information obtained from other sources” (OR 4.38, 95% CI 2.01–9.55), and “second/third trimester” (OR 2.41, 95% CI 1.21–4.82).. Among 372 OBGYN doctors, 76.9% routinely recommended vaccination for pregnant women. Independent factors effecting routine recommendation were “working at a private clinic or hospital” (OR 5.33, 95% CI 2.44–11.65), “awareness of KCDC guidelines” (OR 3.11, 95% CI 1.11–8.73), and “awareness of the 2019 national free influenza vaccination program for pregnant women” (OR 4.88, 95% CI 2.34–10.17). OBGYN doctors most commonly chose ‘guidelines proposed by the government or public health (108, 46%) and academic committees (59, 25%), as a factor which expect to affect the future recommendation Conclusion This study showed that providing information about maternal influenza vaccination, especially by OBGYN doctors, is crucial for increasing vaccination coverage in pregnant women. Closer cooperation between the government and OBGYN academic societies to educate OBGYN doctors might enhance routine recommendations.

Highlights • H&N radiation treatment plan directives are typically not patient-specific. • Patient-specific directives may facilitate the best-achievable dose distribution. • Use of an AI-guided tool significantly improved achieved dose for nearly all OARs., Background and purpose Volumetric modulated arc therapy (VMAT) planning for head and neck cancer is a complex process. While the lowest achievable dose for each individual organ-at-risk (OAR) is unknown a priori, artificial intelligence (AI) holds promise as a tool to accurately estimate the expected dose distribution for OARs. We prospectively investigated the benefits of incorporating an AI-based decision support tool (DST) into the clinical workflow to improve OAR sparing. Materials and methods The DST dose prediction model was based on 276 institutional VMAT plans. Under an IRB-approved prospective trial, the physician first generated a custom OAR directive for 50 consecutive patients (physician directive, PD). The DST then estimated OAR doses (AI directive, AD). For each OAR, the treating physician used the lower directive to form a hybrid directive (HD). The final plan metrics were compared to each directive. A dose difference of 3 Gray (Gy) was considered clinically significant. Results Compared to the AD and PD, the HD reduced OAR dose objectives by more than 3 Gy in 22% to 75% of cases, depending on OAR. The resulting clinical plan typically met these lower constraints and achieved mean dose reductions between 4.3 and 16 Gy over the PD, and 5.6 to 9.1 Gy over the AD alone. Dose metrics achieved using the HD were significantly better than institutional historical plans for most OARs and NRG constraints for all OARs. Conclusions The DST facilitated a significantly improved treatment directive across all OARs for this generalized H&N patient cohort, with neither the AD nor PD alone sufficient to optimally direct planning.

Previous studies suggested that genetic, environmental factors and their interactions could affect body fat mass (BFM). However, studies describing these effects were performed at a single time point in a population. In this study, we investigated the interaction between genetic and environmental factors in affecting BFM and implicate the healthcare utilization of lifestyle modifications from a personalized and genomic perspective. We examined how nutritional intake or physical activity changes in the individuals affect BFM concerning the genetic composition. We conducted an observational study including 259 adult participants with single nucleotide polymorphism (SNP) genotyping and longitudinal lifestyle monitoring, including food consumption and physical activities, by following lifestyle modification guidance. The participants’ lifelog data on exercise and diet were collected through a wearable device for 3 months. Moreover, we measured anthropometric and serologic markers to monitor their potential changes through lifestyle modification. We examined the influence of genetic composition on body fat reduction induced by lifestyle changes using genetic risk scores (GRSs) of three phenotypes: GRS-carbohydrate (GRS-C), GRS-fat (GRS-F), and GRS-exercise (GRS-E). Our results showed that lifestyle modifications affected BFM more significantly in the high GRS class compared to the low GRS class, indicating the role of genetic factors affecting the efficiency of the lifestyle modification-induced BFM changes. Interestingly, the influence of exercise modification in the low GRS class with active lifestyle change was lower than that in the high GRS class with inactive lifestyle change (P = 0.022), suggesting the implication of genetic factors for efficient body fat control.

Due to the highly immunogenic nature of renal cell carcinoma (RCC), the tumor microenvironment (TME) is enriched with various innate and adaptive immune subsets. In particular, gamma-delta (γδ) T cells can act as potent attractive mediators of adoptive cell transfer immunotherapy because of their unique properties such as non-reliance on major histocompatibility complex expression, their ability to infiltrate human tumors and recognize tumor antigens, relative insensitivity to immune checkpoint molecules, and broad tumor cytotoxicity. Therefore, it is now critical to better characterize human γδ T-cell subsets and their mechanisms in RCCs, especially the stage of differentiation. In this study, we aimed to identify γδ T cells that might have adaptive responses against RCC progression. We characterized γδ T cells in peripheral blood and tumor-infiltrating lymphocytes (TILs) in freshly resected tumor specimens from 20 RCC patients. Furthermore, we performed a gene set enrichment analysis on RNA-sequencing data from The Cancer Genome Atlas (TCGA) derived from normal kidneys and RCC tumors to ascertain the association between γδ T-cell infiltration and anti-cancer immune activity. Notably, RCC-infiltrating CD3low Vγ9Vδ1 T cells with a terminally differentiated effector memory phenotype with up-regulated activation/exhaustion molecules were newly detected as predominant TILs, and the cytotoxic activity of these cells against RCC was confirmed in vitro. In an additional analysis of the TCGA RCC dataset, γδ T-cell enrichment scores correlated strongly with those for CTLs, Th1 cells, “exhausted” T cells, and M1 macrophages, suggesting active involvement of γδ T cells in anti-tumor rather than pro-tumor activity, and Vδ1 cells were more abundant than Vδ2 or Vδ3 cells in RCC tumor samples. Thus, we posit that Vγ9Vδ1 T cells may represent an excellent candidate for adoptive immunotherapy in RCC patients with a high risk of relapse after surgery.

Purpose We investigated the feasibility of using an anatomically localized, target-enriched liquid biopsy (TLB) in mouse models of lung cancer.Materials and Methods After irradiating xenograft mouse with human lung cancer cell lines, H1299 (NRAS proto-oncogene, GTPase [NRAS] Q61K) and HCC827 (epidermal growth factor receptor [EGFR] E746-750del), circulating (cell-free) tumor DNA (ctDNA) levels were monitored with quantitative polymerase chain reaction on human long interspersed nuclear element-1 and cell line-specific mutations. We checked dose-dependency at 6, 12, or 18 Gy to each tumor-bearing mouse leg using 6-MV photon beams. We also analyzed ctDNA of lung cancer patients by LiquidSCAN, a targeted deep sequencing to validated the clinical performances of TLB method.Results Irradiation could enhance the detection sensitivity of NRAS Q61K in the plasma sample of H1299-xenograft mouse to 4.5- fold. While cell-free DNA (cfDNA) level was not changed at 6 Gy, ctDNA level was increased upon irradiation. Using double-xenograft mouse with H1299 and HCC827, ctDNA polymerase chain reaction analysis with local irradiation in each region could specify mutation type matched to transplanted cell types, proposing an anatomically localized, TLB. Furthermore, when we performed targeted deep sequencing of cfDNA to monitor ctDNA level in 11 patients with lung cancer who underwent radiotherapy, the average ctDNA level was increased within a week after the start of radiotherapy.Conclusion TLB using irradiation could temporarily amplify ctDNA release in xenograft mouse and lung cancer patients, which enables us to develop theragnostic method for cancer patients with accurate ctDNA detection.

Sequence alterations in microsatellites and an elevated mutational burden are observed in 20% of gastric cancers and associated with clinical response to anti–PD-1 antibodies. However, 50% of microsatellite instability–high (MSI-H) cancers are intrinsically resistant to PD-1 therapies. We conducted a phase II trial of pembrolizumab in patients with advanced MSI-H gastric cancer and included serial and multi-region tissue samples in addition to serial peripheral blood analyses. The number of whole-exome sequencing (WES)–derived nonsynonymous mutations correlated with antitumor activity and prolonged progression-free survival (PFS). Coupling WES to single-cell RNA sequencing, we identified dynamic tumor evolution with greater on-treatment collapse of mutational architecture in responders. Diverse T-cell receptor repertoire was associated with longer PFS to pembrolizumab. In addition, an increase in PD-1+ CD8+ T cells correlated with durable clinical benefit. Our findings highlight the genomic, immunologic, and clinical outcome heterogeneity within MSI-H gastric cancer and may inform development of strategies to enhance responsiveness. Significance: This study highlights response heterogeneity within MSI-H gastric cancer treated with pembrolizumab monotherapy and underscores the potential for extended baseline and early on-treatment biomarker analyses to identify responders. The observed markers of intrinsic resistance have implications for patient stratification to inform novel combinations among patients with intrinsically resistant features. See related commentary by Fontana and Smyth, p. 2126. This article is highlighted in the In This Issue feature, p. 2113

Synthetic small diameter vascular grafts frequently fail owing to intimal hyperplasia, results from mismatched compliance between the vascular graft and native vessels. A vascular graft with a negative Poisson's ratio (NPR, materials expand transversely when pulled axially) was suggested to enhance compliance. We produced a three-layer tubular vascular scaffold with NRP properties. The luminal side consisted of nanosized electrospun fibers for endothelial cell (EC) growth. The middle layer was an NPR structure created using 3D printing, and the outer layer was a microsized electrospun fiber layer for vascular smooth muscle cell (VSMC) growth. The developed multi-layer tubular vascular scaffold contained NPR value. And the NPR vascular scaffold showed 1.7 time higher compliance than the PPR scaffold and 3.8 times higher than that of commercial polytetrafluoroethylene (PTFE) vascular graft. In addition, the ECs and VSMCs were well survived and proliferated on the scaffold during 10 days of culture. From the optimized co-culture condition of the VSMCs and ECs that VSMC phenotype changed was inhibited, we successfully generated a thin luminal layer, which consisted of ECs and the proper thickness of the VSMC layer under the ECs. This scaffold may have a potential to replace conventional artificial vascular graft by providing enhanced compliance and improved cell culture environment.

Objective: to investigate pertussis vaccination rates during pregnancy and the routine recommendation rates by maternity healthcare professionals (HCPs), including influencing factors, in Korea. Materials and methods: Two different questionnaires were developed and conducted anonymously for pregnant or postpartum women and maternity HCPs in 30 multi-centers. Maternal pertussis vaccination rates and maternity HCPs’ recommendation rates were analyzed. Independent influencing factors were analyzed using multivariate logistic regression analysis, respectively. Results: The rate of pertussis vaccination during pregnancy among 466 women was 67%. Among 164 multiparous women, 35.5% received pertussis vaccinations during every pregnancy. However, 27.9% among all pregnant women did not receive information about pertussis and vaccination. The independent influencing factors for maternal pertussis vaccination, given as the tetanus, diphtheria and acellular pertussis (Tdap), were “getting informed” (OR 18.597, 95% CI 11.206–30.861), “informed by OBGYN doctors” (OR 4.426, 95% CI 2.144–9.267), and “metropolitan residence” (OR 3.048, 95% CI 1.419–6.548). Among a total of 373 maternity HCPs, 210 (56.3%) routinely recommended pertussis vaccination, but 21.7% of the total maternity HCP participants did not know the maternal Tdap guideline. The independent factors affecting routine recommendation were the awareness of guideline (OR 9.771, 95% CI 5.227–18.265, p

Background: Young-PEARL study showed median progression-free survival (PFS) of 20·1 months in the palbociclib plus endocrine therapy group versus 14·4 months in the capecitabine group (hazard ratio 0·659 [95% CI 0·437-0·994], log-rank p=0·0235). We conducted exploratory biomarker analysis to predict the efficacy of the trial. Methods: This was a phase II trial that randomized 184 patients with HR+ MBC in premenopausal women to palbociclib plus exemestane with GNRH agonist (Arm A, n=79) versus capecitabine (Arm B, n=62). We performed targeted sequencing (CancerSCANTM) containing 375 cancer-related genes (141 patients) and whole transcriptome sequencing (165 patients) using baseline tumor samples to examine genomic alteration in relation to drug response in terms of PFS. Result: By research-based PAM50 subtyping, 73% of patients classified as Luminal (Luminal A and Luminal B), and showed better prognosis in all patients and Arm A (p Conclusions: The alteration of a few genes including Rb1 loss may be associated with resistance of palbociclib in HR-positive premenopausal population with MBC. Luminal type showed better prognosis, and BRCA2 pathogenic mutation showed worse prognosis regardless luminal/non-luminal type. ESR1 mutation was found in low population frequency because all patients didn’t received AI therapy. Further exploration of molecular variables is warranted to determine and validate biomarkers of efficacy. Clinical trial information: NCT02592746. Citation Format: Kyunghee Park, Gun Min Kim, Kyung Hae Jung, Seok Yun Kang, In Hae Park, Jee Hyun Kim, Hee Kyung Ahn, Woong-Yang Park, Seock-Ah Im, Yeon Hee Park. Exploratory biomarker analysis of Young-PEARL [palbociclib plus exemestane with GnRH agonist versus capecitabine in premenopausal women with HR (hormone receptor)-positive, HER2-negative metastatic breast cancer (MBC)] study [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-19.

Purpose Intermittent hypoxia (IH) is characterized by hypoxia-reoxygenation, reported to be a critical risk factor for obstructive sleep apnea (OSA). This experiment aimed to evaluate the direct effects of IH on the human nasal mucosa. Methods The direct effects of IH on the human nasal mucosa was evaluated by measuring the ciliary beat frequency (CBF) and expression levels of inflammatory cytokines (granulocyte-macrophage colony-stimulating factor, transforming growth factor-β, interleukin-6, and tumor necrosis factor-α). The normoxia group was exposed to a normoxic condition for 72 h. The IH group was exposed to 288 cycles of IH (1 cycle: hypoxia, 5 min; subsequent normoxia, 10 min) for 72 h. CBF was measured using an automated computer-based video image processing technique. Changes in the expression of cytokines were assessed by real-time reverse transcription-polymerase chain reaction (RT-PCR). Results The normoxia group revealed a persistent CBF pattern and a physiological range of inflammatory cytokines. However, the IH group showed a cyclic decrease in CBF and increased expression of inflammatory cytokines. Cytotoxicity assay indicated no difference in the survival rates between the two groups. Conclusions IH results in increased expression of inflammatory cytokines that adversely affects the mucociliary transport in the upper airway and, consequently, may result in airway inflammation.

Objective We intended to identify the risk factors of intraoperative hemorrhage on occasions of a combined operation of myomectomy with cesarean section for patients with uterine leiomyoma. Materials and methods A retrospective cohort study was done of all patients who underwent cesarean myomectomy for intramural leiomyoma at a single university hospital. Cases identified with subserosal leiomyoma, placental disorder, and comorbid conditions related to coagulopathy were excluded. All the included cases were classified into intraoperative hemorrhage and non-hemorrhage group. Obstetric and demographic factors and parameters of leiomyoma were compared between two groups. Results A total of 302 women underwent cesarean myomectomy during the study period. Among these women, 212 pregnant women met the inclusion criteria. Intraoperative hemorrhage occurred in 43 women (20.3%). There was no significant intergroup difference in the number of removed leiomyomas. Multiple logistic regression analysis demonstrated that lower segmental location (odds ratio [OR], 2.827; 95% confidence interval [CI], 1.033–7.734, P = 0.043) and the diameter (OR, 1.167; 95% CI, 1.044–1.305, P = 0.006) were significant independent risk factors for hemorrhage during cesarean myomectomy. The combination of ≥ 8 cm diameter or lower segmental position of the leiomyoma yielded a specificity of 79.3% for operative hemorrhage during cesarean myomectomy. The negative predictive value of this combination was 88.7% for operative hemorrhage with a prevalence of 20%. Conclusion The large size and lower segmental position of the leiomyoma are significantly risk factors for intraoperative hemorrhage during cesarean myomectomy. If the leiomyoma is located in the uterine fundus or body and its diameter is less than 8 cm, the removal of leiomyoma may be considered at the time of cesarean section.

Background: Macrophages produce many inflammation-associated molecules, released by matrix metalloproteinases, such as adhesion molecules, and cytokines, as well, which play a crucial role in atherosclerosis. In this context, we investigated the relationship between Ninjurin-1 (Ninj1 [nerve injury-induced protein]), a novel matrix metalloproteinase 9 substrate, expression, and atherosclerosis progression. Methods: Ninj1 expression and atherosclerosis progression were assessed in atherosclerotic aortic tissue and serum samples from patients with coronary artery disease and healthy controls, and atheroprone apolipoprotein e–deficient ( Apoe −/− ) and wild-type mice, as well. Apoe −/− mice lacking systemic Ninj1 expression ( Ninj1 −/− Apoe −/− ) were generated to assess the functional effects of Ninj1. Bone marrow transplantation was also used to generate low-density lipoprotein receptor–deficient ( Ldlr −/− ) mice that lack Ninj1 specifically in bone marrow–derived cells. Mice were fed a Western diet for 5 to 23 weeks, and atherosclerotic lesions were investigated. The anti-inflammatory role of Ninj1 was verified by treating macrophages and mice with the peptides Ninj1 1 –56 (ML56) and Ninj1 26 –37 (PN12), which mimic the soluble form of Ninj1 (sNinj1). Results: Our in vivo results conclusively showed a correlation between Ninj1 expression in aortic macrophages and the extent of human and mouse atherosclerotic lesions. Ninj1 -deficient macrophages promoted proinflammatory gene expression by activating mitogen-activated protein kinase and inhibiting the phosphoinositide 3-kinase/Akt signaling pathway. Whole-body and bone marrow–specific Ninj1 deficiencies significantly increased monocyte recruitment and macrophage accumulation in atherosclerotic lesions through elevated macrophage-mediated inflammation. Macrophage Ninj1 was directly cleaved by matrix metalloproteinase 9 to generate a soluble form that exhibited antiatherosclerotic effects, as assessed in vitro and in vivo. Treatment with the sNinj1-mimetic peptides, ML56 and PN12, reduced proinflammatory gene expression in human and mouse classically activated macrophages, thereby attenuating monocyte transendothelial migration. Moreover, continuous administration of mPN12 alleviated atherosclerosis by inhibiting the enhanced monocyte recruitment and inflammation characteristics of this disorder in mice, regardless of the presence of Ninj1. Conclusions: Ninj1 is a novel matrix metalloproteinase 9 substrate in macrophages, and sNinj1 is a secreted atheroprotective protein that regulates macrophage inflammation and monocyte recruitment in atherosclerosis. Moreover, sNinj1-mediated anti-inflammatory effects are conserved in human macrophages and likely contribute to human atherosclerosis.

Background Hereditary cerebellar ataxias exhibit heterogeneous phenotypes and genotypes. To date, advancement of next-generation sequencing technologies have identified many causative genes for ataxia in various population. In this study, whole-exome sequencing (WES) was utilized to explore the genetic cause of ataxia among Korean patients who remained undiagnosed following routine investigation. Methods Patients with ataxia were enrolled in this study. We excluded patients with acquired, degenerative, and trinucleotide repeat ataxias, such as spinocerebellar ataxia 1 (SCA1), SCA2, SCA3, SCA6, SCA7, SCA8, SCA17, Dentatorubral-pallidoluysian atrophy, and Friedreich ataxia. WES was performed. After basic filtering based on population databases, we then performed primary filtering to screen for known ataxia-associated genes, followed by expanded filtering customized for individual patients. Results We enrolled 77 ataxia patients from 68 families. Eighteen families had pathogenic or likely pathogenic variants in 14 different genes, including NEU1, APTX, SPG7, HTRA1, POLG2, SYNE1, CACNA1G, CACNA1A, ITPR1, AHI1, SPG11, ANO10, ATM, and C5orf42, resulting in a diagnostic yield of 26.5%. Hereditary spastic paraplegia was the most common diagnosis. Adult-onset ataxias and those without family history were frequently encountered. Variants of unknown significance were found in 14 (20.6%) families, some of which were highly probable from the clinical perspective. Conclusion Using WES, we explored the molecular etiology of ataxia in patients whom were not diagnosed through routine clinical investigation. This study revealed unexpected rare disorders as well as the known ataxia-associated genes in a Korean population.

PURPOSE ERBB2 amplification (human epidermal growth factor receptor 2 positivity [HER2+]) in metastatic colorectal cancer (mCRC) has important therapeutic implications that necessitate the need for accurate diagnostics. The study purpose was to establish and validate harmonized diagnostic criteria for HER2+ mCRC among 3 groups (GI-SCREEN-Japan, NCTN-SWOG-USA, and Korea). PATIENTS AND METHODS We assessed HER2 status by immunohistochemistry (IHC), ERBB2/CEP17 ratio, gene copy number (GCN) by fluorescence in situ hybridization (FISH), and copy number variation (CNV) using two targeted next-generation sequencing (NGS) panels. Tumor samples from 475 and 16 patients with mCRC in exploratory and validation cohorts, respectively, were used for cross-validation of the NGS panels. RESULTS Consensus diagnostic criteria among the 3 groups for HER2+ mCRC were established as follows: IHC 3+ or IHC 2+ and ERBB2/CEP17 ratio by FISH ≥ 2.0, tumor content > 10% for surgically resected specimens, and necessary tumor content not defined for biopsy specimens. The median GCN and CNV for HER2+ patients were 10.9 and 27.7 compared with 2.5 ( P < .0001) and 3.5 ( P < .0001), respectively, in HER2-negative patients. These findings were validated in a validation cohort (GCN, 16.2 v 2.4 [ P = .0002]; CNV, 42.5 v 2.0 [ P = .0003]). GCN correlated with CNV in both cohorts (exploratory, r = 0.90; validation, r = 0.97; P < .0001). CNV in cross-validation of the 2 NGS panels also showed a strong correlation ( r = 0.98; P < .0001). CNV in patients who fulfilled the consensus criteria was > 4.0 in all, which demonstrates the accuracy of the IHC/FISH criteria and cross-validation of NGS panels. CONCLUSION We have established and verified harmonized diagnostic criteria for HER2+ and demonstrated consistency between IHC/FISH and CNV determined by NGS in mCRC.

Salivary duct carcinoma (SDC) is one of the most aggressive subtypes of salivary gland cancers. Conventional chemotherapy and/or radiation have shown only limited clinical efficacy in the treatment of recurrent or metastatic SDC. Currently, clinically approved targeted-therapeutics are not generally applicable except in very limited cases, and there exists a strong need for the development of treatment against this unique tumor type. To further interrogate genomic features of SDC, we have conducted multi-omic profiling of the SDC to describe the genomic alterations prevalent in this disease. Whole-genome sequencing, whole exome-sequencing and transcriptome sequencing were performed on a discovery cohort of 10 SDC samples. Targeted genomic profiling was performed in additional 32 SDC samples to support the findings obtained from the original discovery cohort. The cancer cohort was characterized by an average mutation burden of 85 somatic exonic mutations per tumor sample. The cohort harbored a mutational signature of BRCA and APOBEC/AID. Several genes, including TP53, RB1, SMAD4, HRAS, APC, PIK3CA and GNAQ were recurrently somatically altered in SDC. A novel fusion gene, generated by genomic rearrangement, MYB-NHSL1, was also noted. Our findings represent a significant layer in the systematic understanding of potentially clinically useful genomic and molecular targets for a subset of recurrent/metastatic SDC.

Background Glioblastoma (GBM) is a complex disease with extensive molecular and transcriptional heterogeneity. GBM can be subcategorized into four distinct subtypes; tumors that shift towards the mesenchymal phenotype upon recurrence are generally associated with treatment resistance, unfavorable prognosis, and the infiltration of pro-tumorigenic macrophages. Results We explore the transcriptional regulatory networks of mesenchymal-associated tumor-associated macrophages (MA-TAMs), which drive the malignant phenotypic state of GBM, and identify macrophage receptor with collagenous structure (MARCO) as the most highly differentially expressed gene. MARCOhigh TAMs induce a phenotypic shift towards mesenchymal cellular state of glioma stem cells, promoting both invasive and proliferative activities, as well as therapeutic resistance to irradiation. MARCOhigh TAMs also significantly accelerate tumor engraftment and growth in vivo. Moreover, both MA-TAM master regulators and their target genes are significantly correlated with poor clinical outcomes and are often associated with genomic aberrations in neurofibromin 1 (NF1) and phosphoinositide 3-kinases/mammalian target of rapamycin/Akt pathway (PI3K-mTOR-AKT)-related genes. We further demonstrate the origination of MA-TAMs from peripheral blood, as well as their potential association with tumor-induced polarization states and immunosuppressive environments. Conclusions Collectively, our study characterizes the global transcriptional profile of TAMs driving mesenchymal GBM pathogenesis, providing potential therapeutic targets for improving the effectiveness of GBM immunotherapy.

Background A single institution retrospective analysis of 124 non‐small cell lung carcinoma (NSCLC) patients was performed to identify whether disease‐free survival (DFS) achieves incremental values when radiomic and genomic data are combined with clinical information. Methods Using the least absolute shrinkage and selection operator (LASSO) Cox regression method, radiomic and genetic features were reduced in number for selection of the most useful prognostic feature. We created four models using only baseline clinical data, clinical data with selected genetic features, clinical data with selected radiomic features, and clinical data with selected genetic and radiomic features together. Multivariate Cox proportional hazards analysis was performed to determine predictors of DFS. Receiver operating characteristic (ROC) calculation was made to compare the discriminative performance for DFS prediction by four constructed models at the five‐year time point. Results On precontrast scan, improved discrimination performance was obtained in a merging of selected radiomics and genetics (AUC = 0.8638), compared with clinical data only (AUC = 0.7990), selected genetic features (AUC = 0.8497), and selected radiomic features (AUC = 0.8355). On post‐contrast scan, discrimination performance was improved (AUC = 0.8672) compared with the clinical variables (AUC = 0.7913), and selected genetic features (AUC = 0.8376) and selected radiomic features (AUC = 0.8399) were considered. Conclusions The combination of selected radiomic and genomic features improved stratification of NSCLC patients upon survival. Thus, integrating clinicopathologic model with radiomic and genomic features may lead to improved prognostic accuracy compared to conventional clinicopathological data alone. Key points Significant findings of the study Receiver operating characteristic (ROC) calculation was made to compare the discriminative performance for disease‐free survival (DFS). The discriminative performance for DFS was better when combining radiomic and genetic features compared to clinical data only, selected genetic features, and selected radiomic features. What this study adds The combination of selected radiomic and genomic features improved stratification of NSCLC patients upon survival. Thus, integrating a clinicopathological model with radiomic and genomic features may lead to improved prognostic accuracy compared to conventional clinicopathological data alone., We tried to compare the results of disease‐free survival (DFS) in patients with non‐small‐cell lung carcinoma (NSCLC) through radiomics with those of traditional staging systems or genetic analysis to determine if incremental values can be obtained when combining them. The addition of selected radiomics and genomic features using the LASSO method improved the stratification of lung cancer patients upon survival. Our results show that integration of radiomics and genomic features with the current clinicopathologic model may lead to improved prognostic accuracy compared to conventional clinicopathological data alone.

To elucidate the epigenetic mechanisms of drug resistance, epigenetically reprogrammed H460 cancer cells (R-H460) were established by the transient introduction of reprogramming factors. Then, the R-H460 cells were induced to differentiate by the withdrawal of stem cell media for various durations, which resulted in differentiated R-H460 cells (dR-H460). Notably, dR-H460 cells differentiated for 13 days (13dR-H460 cells) formed a significantly greater number of colonies showing drug resistance to both cisplatin and paclitaxel, whereas the dR-H460 cells differentiated for 40 days (40dR-H460 cells) lost drug resistance; this suggests that 13dR-cancer cells present short-term resistance (less than a month). Similarly, increased drug resistance to both cisplatin and paclitaxel was observed in another R-cancer cell model prepared from N87 cells. The resistant phenotype of the cisplatin-resistant (CR) colonies obtained through cisplatin treatment was maintained for 2–3 months after drug treatment, suggesting that drug treatment transforms cells with short-term resistance into cells with medium-term resistance. In single-cell analyses, heterogeneity was not found to increase in 13dR-H460 cells, suggesting that cancer cells with short-term resistance, rather than heterogeneous cells, may confer epigenetically driven drug resistance in our reprogrammed cancer model. The epigenetically driven short-term and medium-term drug resistance mechanisms could provide new cancer-fighting strategies involving the control of cancer cells during epigenetic transition., Cancer chemotherapy: treatment prolongs resistance caused by DNA modification Cancer cells that are transiently resistant to drug therapies owing to changes in their gene expression patterns can become resistant for longer durations if exposed to the drug treatments. A team led by Kyeong-Man Hong and Hyonchol Jang from the National Cancer Center in Goyang, South Korea, used cellular reprogramming technologies to induce changes in the DNA markers that regulate gene expression. Working with lung and gastric cancer cell lines, the researchers found that such epigenetic alterations caused many cells to become resistant to the chemotherapy drugs cisplatin and paclitaxel. In the absence of treatment, the cells soon lost their drug resistance. In the presence of the chemotherapeutics, however, the resistance trait lasted longer, a finding that could inform best practice for how to administer cancer-fighting agents in the face of epigenetic-driven drug resistance.

Background Immune checkpoint inhibitors (ICIs) have been shown to be beneficial for some patients with advanced non-small-cell lung cancer (NSCLC). However, the underlying mechanisms mediating the limited response to ICIs remain unclear. Patients and methods We carried out whole-exome sequencing on 198 advanced NSCLC tumors that had been sampled before anti-programmed cell death 1 (anti-PD-1)/programmed death-ligand 1 (PD-L1) therapy. Detailed clinical characteristics were collected on these patients. We designed a new method to estimate human leukocyte antigen (HLA)-corrected tumor mutation burden (TMB), a modification which considers the loss of heterozygosity of HLA from conventional TMB. We carried out external validation of our findings utilizing 89 NSCLC samples and 110 melanoma samples from two independent cohorts of immunotherapy-treated patients. Results Homology-dependent recombination deficiency was identified in 37 patients (18.7%) and was associated with longer progression-free survival (PFS; P = 0.049). Using the HLA-corrected TMB, non-responders to ICIs were identified, despite having a high TMB (top 25%). Ten patients (21.3% of the high TMB group) were reclassified from the high TMB group into the low TMB group. The objective response rate (ORR), PFS, and overall survival (OS) were all lower in these patients compared with those of the high TMB group (ORR: 20% versus 59%, P = 0.0363; PFS: hazard ratio = 2.91, P = 0.007; OS: hazard ratio = 3.43, P = 0.004). Multivariate analyses showed that high HLA-corrected TMB was associated with a significant survival advantage (hazard ratio = 0.44, P = 0.015), whereas high conventional TMB was not associated with a survival advantage (hazard ratio = 0.63, P = 0.118). Applying this approach to the independent cohorts of 89 NSCLC patients and 110 melanoma patients, TMB-based survival prediction was significantly improved. Conclusion HLA-corrected TMB can reconcile the observed disparity in relationships between TMB and ICI responses, and is of predictive and prognostic value for ICI therapies.

Background A high percentage of small cell lung cancer (SCLC) cases harbor cell cycle-related gene mutations and RICTOR amplification. Based on underlying somatic mutations, the authors have conducted a phase 2 biomarker-driven, multiarm umbrella study. Methods The SCLC Umbrella Korea StudiES (SUKSES) is an adaptive platform trial that undergoes continual modification according to the observed outcomes. This study included 286 patients with SCLC who failed platinum therapy and who had known genomic profiles based on a predesigned screening trial. Patients with MYC amplification or CDKN2A and TP53 co-alterations were allocated to adavosertib (SUKSES protocol C [SUKSES-C]; 7 patients) and those with RICTOR amplification were allocated to vistusertib (SUKSES-D; 4 patients). Alternatively, patients who were without any predefined biomarkers were assigned to a non-biomarker-selected arm: adavosertib (SUKSES-N1; 21 patients) or AZD2811NP (SUKSES-N3; 15 patients). Results Patients in the SUKSES-C and SUKSES-N1 arms demonstrated no objective response. Three patients presented with stable disease (SD) in SUKSES-C and 6 patients in SUKSES-N1. The median progression-free survival (PFS) was 1.3 months (95% confidence interval, 0.9 months to not available) for SUKSES-C and 1.2 months (95% CI, 1.1-1.4 months) for SUKSES-N1. Patients in the SUKSES-D arm demonstrated no objective response and no SD, with a PFS of 1.2 months (95% CI, 1.0 months to not available). The SUKSES-N3 arm had 5 patients with SD and a PFS of 1.6 months (95% CI, 0.9-1.7 months), without an objective response. Grade≥3 adverse events (graded according to National Cancer Institute Common Terminology Criteria for Adverse Events [version 4.03]) were observed as follows: 3.2% in the SUKSES-C and SUKSES-N1 arms and 50.0% in the SUKSES-D arm. Target-related neutropenia (grade≥3) was observed in approximately 60.0% of patients in the AZD2811NP arm using the current dosing schedule. Conclusions To the best of the authors' knowledge, the current study is the first biomarker-driven umbrella study conducted in patients with recurrent SCLC. Although the current study demonstrated the limited clinical efficacy of monotherapy, novel biomarker approaches using other cell cycle inhibitor(s) or combinations warrant further investigation.

Immunotherapy for metastatic colorectal cancer is effective only for mismatch repair-deficient tumors with high microsatellite instability that demonstrate immune infiltration, suggesting that tumor cells can determine their immune microenvironment. To understand this cross-talk, we analyzed the transcriptome of 91,103 unsorted single cells from 23 Korean and 6 Belgian patients. Cancer cells displayed transcriptional features reminiscent of normal differentiation programs, and genetic alterations that apparently fostered immunosuppressive microenvironments directed by regulatory T cells, myofibroblasts and myeloid cells. Intercellular network reconstruction supported the association between cancer cell signatures and specific stromal or immune cell populations. Our collective view of the cellular landscape and intercellular interactions in colorectal cancer provide mechanistic information for the design of efficient immuno-oncology treatment strategies.

Colorectal cancer liver metastases (CRCLM) that present with a replacement histopathological growth pattern (HGP) are resistant to neoadjuvant anti-angiogenic therapy. Surrogate biomarkers are not available to preoperatively identify patients with these tumors. Here we identify differentially expressed genes between CRCLM with a replacement HGP and those with a desmoplastic HGP using RNA sequencing. We demonstrate that LOXL4 is transcriptionally upregulated in replacement HGP CRCLM compared with desmoplastic HGP CRCLM and the adjacent normal liver. Interestingly, lysyl oxidase-like 4 (LOXL4) protein was expressed by neutrophils present in the tumor microenvironment in replacement HGP CRCLM. We further demonstrate that LOXL4 expression is higher in circulating neutrophils of cancer patients compared with healthy control patients and its expression can be induced by stimulation with lipopolysaccharide and TNF-α. Our study is the first to show the expression of LOXL4 in neutrophils and reveals the potential for LOXL4-expressing neutrophils to support the replacement HGP phenotype and to serve as a surrogate biomarker for this subtype of CRCLM. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Purpose of review The gut microbiota has been speculated to underpin metabolic changes associated with chronic antipsychotic use. The change in the gut microbiota can also cause abnormal absorbtion from the gut into the blood stream (leaky gut syndrome) that can lead to inflammatory reactions, and thus, secondary damage to the brain and central nervous system. Our article aims to highlight relevant research on antipsychotic's effect on the microbiota and to point out future directions. Recent findings Antipsychtoic use can result in specific microbiota changes, and it is important to differentiate this from the innate microbiota of the patient. It is important to treat these microbiota changes, as they are correlated with obesity, which is a negative contributor to the cardiovascular health of those suffering with schizophrenia. Ways to prevent antipsychotic-induced side-effects include antibiotic treatment, histamine 3 receptor blockade and metformin use. Summary Given the dearth of current literature, more research is needed, however, to determine, which comes first in people with schizophrenia--an abnormal gut microbiota that elevates one's risk for schizophrenia or psychopharmacologic treatment of schizophrenia leading to secondary microbiota abnormalities or the negative symptoms of schizophrenia leading to obesity and its associated microbiota changes.

Advanced metastatic cancer poses utmost clinical challenges and may present molecular and cellular features distinct from an early-stage cancer. Herein, we present single-cell transcriptome profiling of metastatic lung adenocarcinoma, the most prevalent histological lung cancer type diagnosed at stage IV in over 40% of all cases. From 208,506 cells populating the normal tissues or early to metastatic stage cancer in 44 patients, we identify a cancer cell subtype deviating from the normal differentiation trajectory and dominating the metastatic stage. In all stages, the stromal and immune cell dynamics reveal ontological and functional changes that create a pro-tumoral and immunosuppressive microenvironment. Normal resident myeloid cell populations are gradually replaced with monocyte-derived macrophages and dendritic cells, along with T-cell exhaustion. This extensive single-cell analysis enhances our understanding of molecular and cellular dynamics in metastatic lung cancer and reveals potential diagnostic and therapeutic targets in cancer-microenvironment interactions., Understanding the mechanisms that lead to lung adenocarcinoma metastasis is important for identifying new therapeutics. Here, the authors document the changes in the transcriptome of human lung adenocarcinoma using single-cell sequencing and link cancer cell signatures to immune cell dynamics.

Cell-free DNA (cfDNA) can be released from tumor cells during proliferation and apoptosis; thus, a fraction of the cfDNA in patients with cancer is tumor-derived. However, the prognostic value of cfDNA in aggressive non-Hodgkin lymphoma (NHL) has not been determined. Between March 2017 and April 2019, plasma cfDNA was obtained from 158 patients with aggressive NHL who were registered in a prospective Samsung Medical Center lymphoma cohort (diffuse large B cell lymphoma (DLBCL), n = 51; T cell lymphoma (TCL), n = 51; NK/T cell lymphoma (NKTCL), n = 56). The concentration of cfDNA was estimated in longitudinal samples collected from patients with NHL before and during various chemotherapy regimens. In pretreatment samples, the median cfDNA concentration of all patients with aggressive lymphoma was 13.7 ng/dl (range 1.7–1792), which was significantly higher than that of healthy volunteers (median 7.4 ng, range 3.7–14.4, p

Extranodal NK/T-cell lymphoma is an aggressive lymphoma that is strongly associated with Epstein–Barr virus infection. Although some extranodal NK/T-cell lymphoma patients have shown responses to immune checkpoint blockade, biomarkers for predicting extranodal NK/T-cell lymphoma patient response to immunotherapy have not yet been defined. To understand the tumor immune microenvironment, we analyzed the expression of 579 immune-related genes and characterized the immune cells using immunohistochemistries and in situ hybridization for EBER. Based on comprehensive analyses, we developed an immune subtyping model that classifies extranodal NK/T-cell lymphoma patients into four tumor immune microenvironment subgroups using three immunohistochemical markers (FoxP3, PD-L1, and CD68). The four tumor immune microenvironment subgroups were named immune tolerance, immune evasion-A, immune evasion-B, and immune silenced. The immune tolerance group was characterized by high-Treg counts and was frequently observed in early stage, and nasal extranodal NK/T-cell lymphoma. The immune evasion group showed high cytotoxic T-cell counts and high PD-L1 expression but low Treg counts. In the immune-silenced group, almost all immune responses were exhausted, most patients were at an advanced stage, and had the poorest disease prognosis among the tumor immune microenvironment subgroups. In some patients (n = 3), a shift in the tumor immune microenvironment subgroup classification was observed in sequential biopsies. The response rate to pembrolizumab, an anti-PD-1 antibody, was 100% (1/1) in the immune tolerance group, 60% (3/5) in the immune evasion group, and 0% (0/5) in the immune-silenced group. We classified extranodal NK/T-cell lymphoma into four tumor immune microenvironment subgroups using a new classification system. In conclusion, we propose that the tumor immune microenvironment of extranodal NK/T-cell lymphoma may change during disease progression and may serve as a useful biomarker for immunotherapy.

Normal-range sleep duration is an important factor for general health and metabolism, and insufficient or excessive sleep is associated with chronic metabolic disease. Among the many factors that affect sleep duration, sun exposure plays an important role in maintaining regular circadian rhythm and is also involved in the production and activation of 25-hydroxyvitamin D [25(OH)D], which regulates various functions in the body. However, 25(OH)D is available through food and various nutritional supplements without sun exposure, so it is important to find out the complex relationship among sun exposure, vitamin D status, and sleep duration. The relationship between sun exposure, vitamin D status, and sleep duration was analyzed in the nationwide survey and examination of 25,534 study populations, after adjusting for demographic characteristics, physical characteristics, lifestyle status, and socio-demographic variables. Vitamin D status alone did not show the relationship with sleep duration, although there were statistical relationships in the various factors including sun exposure with sleep duration. There was a statistical difference in 25(OH)D according to sleep duration, only in low sun exposure group. Subjects with low sun exposure and excessive sleep duration comparatively lower 25(OH)D than those with normal-range sleep, even after adjustment for potentially confounding factors. Individuals with limited exposure to sunlight should maintain adequate vitamin D status to have an appropriate sleep duration for health.

BackgroundYoung patients with colorectal cancer (CRC) exhibit poor prognoses compared to older patients due to the difficulty in early diagnosis and treatment. However, the underlying molecular characteristics are still unclear.MethodsWe conducted a comprehensive analysis of 49 CRC patients without hereditary CRC using the whole-exome and RNA sequencing with tumor and matched normal samples. A total of 594 TCGA samples and 4 patient-derived cells were utilized for validation.ResultsConsensus molecular subtype 4 (CMS4) (53.85%) and CMS2 (38.46%) were enriched in the young (≤ 40 years) and old (> 60 years) age groups, respectively. A CMS4-associated gene, platelet-derived growth factor receptor α (PDGFRA), was significantly upregulated in young patients with CRC (FC = 3.21, p = 0.0001) and was negatively correlated with age (p = 0.0001, R = − 0.526). Moreover,PDGFRAshowed a positive co-expression with metastasis-related genes in young CRC patients. In vitro validation confirmed that young patient-derived cells (PDCs) showed an enriched expression ofPDGFRAcompared to old PDCs and a reduced proliferation rate by knockdown ofPDGFRA. Furthermore, young CRC patients were more sensitive to regorafenib, a PDGFRA-targeting drug, than old CRC patients.ConclusionsOur study suggests that CRC in young patients is associated with CMS4 and PDGFRA. In addition, PDGFRA may serve potential of novel therapeutic strategies and represent a predictive biomarker of response to regorafenib for young CRC patients.

Brain organoid research is advancing, but generation of organoids with proper axis formation, which could lead to spatially ordered structures for complex brain structure and function, still remains a challenge. Axis formation and related spatial cell organization in the CNS are initiated by the symmetry breaking during the early embryo development. It has been demonstrated that the geometrically confined culture of human pluripotent stem cells (hPSCs) can be used to induce symmetry breaking and regionalized cell differentiation. In this study, we generated a polarized spinal cord organoid with a self-organized dorsoventral (DV) organization, using 2D cell patterning by geometric confinement. Initially, the application of caudalization signals to hPSCs promoted the regionalized cell differentiation along the radial axis and sprouting-like protrusion morphogenesis in cell colonies confined to ECM protein micropatterns. Detachment of colonies turned them into extended spinal cord-like organoids which maintained center- and edge-derived two poles. Further analyses including single cell RNA sequencing and spatial transcriptome analysis unveiled that these organoids contained rich repertoire of developing spinal cord cells and exhibited the spatially ordered DV domain formation along the long axis without external organizing signals. Modulation of BMP and Shh signaling can control the extent of DV coverage in organoids following the principles of embryo patterning. Our study provides a simple, and precisely controllable method to generate spatially-ordered organoids for understanding of biological principles of cell patterning and axis formation during neural development.

CHARGE syndrome has a clinically broad spectrum of phenotypes, including partial or atypical type. CHD7 mutation is related to CHARGE syndrome that shows various phenotypes according to the CHD7 variant. Developments in genetic analysis techniques, such as next-generation sequencing (NGS), are helping both diagnosis and treatment of diseases. We report the case of a preterm infant diagnosed with atypical CHARGE who has a novel and de novo CHD7 variant that was identified using whole-genome sequencing (WGS). Neonatologists tend to be reluctant to diagnose infants with multiple malformations because they have to focus on treating life-threatening complications; however, NGS is considered helpful for the early diagnosis of broad-spectrum anomalies during the neonatal period.

BackgroundDiaphanous-related formin 1 (DIA1), which assembles the unbranched actin microfilament and microtubule cytoskeleton, is encoded byDIAPH1. Constitutive activation by the disruption of autoinhibitory interactions between the N-terminal diaphanous inhibitory domain (DID) and C-terminal diaphanous autoregulatory domain (DAD) dysregulates DIA1, resulting in both hearing loss and blood cell abnormalities.Methods and resultsHere, we report the first constitutively active mutant in the DID (p.A265S) of humans with only hearing loss and not blood cell abnormality through whole exome sequencing. The previously reported DAD mutants and our DID mutant (p.A265S) shared the finding of diminished autoinhibitory interaction, abnormally upregulated actin polymerisation activity and increased localisations at the plasma membrane. However, the obvious defect in the DIA1-driven assembly of cytoskeleton ‘during cell division’ was only from the DAD mutants, not from p.A265S, which did not show any blood cell abnormality. We also evaluated the five DID mutants in the hydrophobic pocket since four of these five additional mutants were predicted to critically disrupt interaction between the DID and DAD. These additional pathogenic DID mutants revealed varying degrees of defect in the DIA1-driven cytoskeleton assembly, including nearly normal phenotype during cell division as well as obvious impaired autoinhibition, again coinciding with our key observation in DIA1 mutant (p.A265S) in the DID.ConclusionHere, we report the first mutant in the DID of humans with only hearing loss. The differential cell biological phenotypes of DIA1 during cell division appear to be potential determinants of the clinical severity ofDIAPH1-related cytoskeletopathy in humans.

Autosomal dominant polycystic kidney disease (ADPKD) is one of the main causes of end-stage renal disease (ESRD). Genetic information is of the utmost importance in understanding pathogenesis of ADPKD. Therefore, this study aimed to demonstrate the genetic characteristics of ADPKD and their effects on renal function in 749 Korean ADPKD subjects from 524 unrelated families. Genetic studies of PKD1/2 were performed using targeted exome sequencing combined with Sanger sequencing in exon 1 of the PKD1 gene and a multiple ligation probe assay. The mutation detection rate was 80.7% (423/524 families, 331 mutations) and 70.7% was novel. PKD1 protein-truncating (PKD1-PT) genotype was associated with younger age at diagnosis, larger kidney volume, lower renal function compared to PKD1 non-truncating and PKD2 genotypes. The PKD1 genotype showed earlier onset of ESRD compared to PKD2 genotype (64.9 vs. 72.9 years old, P PKD2 genotype had 0.2 times lower risk for reaching ESRD than PKD1-PT genotype (p = 0.037). In conclusion, our results suggest that genotyping can contribute to selecting rapid progressors for new emerging therapeutic interventions among Koreans.

Aims Immune checkpoint inhibitors (ICIs) induce durable responses, but their clinical benefits apply to only a subset of patients. Therefore, precisely predicting a patient's response before ICI treatment is crucial. Methods A total of 248 patients with anti-Programmed cell death protein 1/Programmed death-ligand 1 (PD1/PD-L1)–treated advanced non–small cell lung cancer were enrolled, and clinical outcomes were collected with a minimum 6-month follow-up period. Tumour tissues were used for PD-L1 staining, targeted sequencing of 380 cancer-related genes and whole-exome sequencing (WES). Results The tumour mutation burden (TMB) obtained from targeted sequencing was higher among patients with a partial response (PR) than those with progressive disease (PD)/stable disease (SD) (P = 0.01) and in those with durable clinical benefit (DCB) than nondurable benefit (NDB) (P = 0.05). The somatic copy number alteration (SCNA) was lower in patients with a PR than those with PD/SD (P = 0.02) and in those with DCB than NDB (P = 0.02). The accuracy of the TMB and SCNA results from the targeted sequencing was confirmed by testing the correlation of the TMB and SCNA results from the targeted sequencing against those results from WES (r = 0.87, r = 0.62, respectively). To improve prediction score, TMB, SCNA and PD-L1 were integrated. New prediction scores reached Area under the ROC Curve (AUC) = 0.71 from TMB (AUC = 0.63), SCNA (AUC = 0.52) or PD-L1 (AUC = 0.57) with our cohort, and validation set from other cohorts also showed improved prediction scores with our new model. Conclusion We report TMB, SCNA and PD-L1 as ICI biomarkers. Combining all these factors improved the prediction accuracy of ICI response compared with using individual factors. Tumour molecular features, TMB and SCNA, were efficiently obtained by targeted sequencing.