Spatially organized multicellular immune hubs in human colorectal cancer.

Immune responses to cancer are highly variable, with mismatch repair-deficient (MMRd) tumors exhibiting more anti-tumor immunity than mismatch repair-proficient (MMRp) tumors. To understand the rules governing these varied responses, we transcriptionally profiled 371,223 cells from colorectal tumors and adjacent normal tissues of 28 MMRp and 34 MMRd individuals. Analysis of 88 cell subsets and their 204 associated gene expression programs revealed extensive transcriptional and spatial remodeling across tumors. To discover hubs of interacting malignant and immune cells, we identified expression programs in different cell types that co-varied across tumors from affected individuals and used spatial profiling to localize coordinated programs. We discovered a myeloid cell-attracting hub at the tumor-luminal interface associated with tissue damage and an MMRd-enriched immune hub within the tumor, with activated T cells together with malignant and myeloid cells expressing T cell-attracting chemokines. By identifying interacting cellular programs, we reveal the logic underlying spatially organized immune-malignant cell networks.

Binimetinib in combination with nivolumab or nivolumab and ipilimumab in patients with previously treated microsatellite-stable metastatic colorectal cancer with RAS mutations in an open-label phase 1b/2 study.

BACKGROUND: In patients with previously treated RAS-mutated microsatellite-stable (MSS) metastatic colorectal cancer (mCRC), a multicenter open-label phase 1b/2 trial was conducted to define the safety and efficacy of the MEK1/MEK2 inhibitor binimetinib in combination with the immune checkpoint inhibitor (ICI) nivolumab (anti-PD-1) or nivolumab and another ICI, ipilimumab (anti-CTLA4). METHODS: In phase 1b, participants were randomly assigned to Arm 1A (binimetinib 45 mg twice daily [BID] plus nivolumab 480 mg once every 4 weeks [Q4W]) or Arm 1B (binimetinib 45 mg BID plus nivolumab 480 mg Q4W and ipilimumab 1 mg/kg once every 8 weeks [Q8W]) to determine the maximum tolerable dose (MTD) and recommended phase 2 dose (RP2D) of binimetinib. The MTD/RP2D was defined as the highest dosage combination that did not cause medically unacceptable dose-limiting toxicities in more than 35% of treated participants in Cycle 1. During phase 2, participants were randomly assigned to Arm 2A (binimetinib MTD/RP2D plus nivolumab) or Arm 2B (binimetinib MTD/RP2D plus nivolumab and ipilimumab) to assess the safety and clinical activity of these combinations. RESULTS: In phase 1b, 21 participants were randomized to Arm 1A or Arm 1B; during phase 2, 54 participants were randomized to Arm 2A or Arm 2B. The binimetinib MTD/RP2D was determined to be 45 mg BID. In phase 2, no participants receiving binimetinib plus nivolumab achieved a response. Of the 27 participants receiving binimetinib, nivolumab, and ipilimumab, the overall response rate was 7.4% (90% CI: 1.3, 21.5). Out of 75 participants overall, 74 (98.7%) reported treatment-related adverse events (AEs), of whom 17 (22.7%) reported treatment-related serious AEs. CONCLUSIONS: The RP2D binimetinib regimen had a safety profile similar to previous binimetinib studies or nivolumab and ipilimumab combination studies. There was a lack of clinical benefit with either drug combination. Therefore, these data do not support further development of binimetinib in combination with nivolumab or nivolumab and ipilimumab in RAS-mutated MSS mCRC. TRIAL REGISTRATION: NCT03271047 (09/01/2017).

Mohs micrographic surgery in the surgical treatment paradigm of melanoma in situ and invasive melanoma: A clinical review of treatment efficacy and ongoing controversies.

Mohs Micrographic Surgery (MMS) for treatment of melanoma offers several advantages over wide local excision (WLE), including complete histologic margin evaluation, same-day resection and closure, and sparing of healthy tissue in critical anatomic sites. Recently, a large volume of clinical data demonstrating efficacy in MMS treatment of melanoma was published, leading to emerging patient safety considerations of incurred treatment costs, risk of tumor upstaging, and failure of care coordination for sentinel lymph node biopsy (SLNB). MMS offers a safe, effective, and value-based treatment for both melanoma in situ (MIS) and invasive melanoma (IM), particularly with immunohistochemistry use on frozen sections. Compared to wide local excision, MMS treatment demonstrates similar or improved outcomes for local tumor recurrence, melanoma-specific survival, and overall survival at long-term follow-up. Tumor upstaging risk is low, and if present, alteration to clinical management is minimal. Discussion of SLNB for eligible head and neck IM cases should be done prior to MMS. Though challenging, successful multidisciplinary coordination of SLNB with MMS has been demonstrated. Herein, we provide a detailed clinical review of evidence for MMS treatment of cutaneous melanoma and offer recommendations to address current controversies surrounding the evolving paradigm of surgical management for both MIS and invasive melanoma (IM).

Immunotherapy of microsatellite stable colorectal cancer: resistance mechanisms and treatment strategies.

In recent years, immunotherapy strategies based on immune checkpoint inhibitors have yielded good efficacy in colorectal cancer (CRC)especially in colorectal cancer with microsatellite instability-high. However, microsatellite-stable (MSS) CRCs account for about 85% of CRCs and are resistant to immunotherapy. Previous studies have shown that compared with MSS CRC, high microsatellite instability CRC possesses a higher frequency of mutations and can generate more neoantigens. Therefore, improving the sensitivity of immunotherapy to MSS CRC is a hot topic which is crucial for the treatment of MSS CRC. This review aims to discuss the factors contributing to MSS CRC insensitivity to immunotherapy and explored potential solutions to overcome immunotherapy resistance.

Exploring Interference Issues in the Case of n25 Band Implementation for 5G/LTE Direct-to-Device NTN Services.

This paper delves into an interference analysis, focusing on the forthcoming Starlink Generation 2 satellites, stated to operate within the 1990-1995 MHz frequency band. The aim is to assess the potential interference from this Starlink system to the satellite receivers of mobile satellite systems (MSSs), which are set to function within the 1980-2010 MHz range, and satellite receivers of the NTN systems, which are planned to operate in the n256 bands, defined by the 3GPP specifications. Through simulation-based evaluations, both single-entry and aggregate interference levels from Starlink to MSSs and NTN systems are comprehensively explored. To estimate the interference impact, several protection criteria were used. The study is in line with the Recommendations of International Telecommunication Union (ITU-R) and common approaches that are used when performing compatibility studies between satellite systems. The findings of this study demonstrate the feasibility of utilizing the n25 band for NTN direct-to-device services.

Preliminary clinical study of personalized neoantigen vaccine therapy for microsatellite stability (MSS)-advanced colorectal cancer.

Immunotherapy based on immune checkpoint inhibitors (ICIs) has provided revolutionary results in treating various cancers. However, its efficacy in colorectal cancer (CRC), especially in microsatellite stability-CRC, is limited. This study aimed to observe the efficacy of personalized neoantigen vaccine in treating MSS-CRC patients with recurrence or metastasis after surgery and chemotherapy. Candidate neoantigens were analyzed from whole-exome and RNA sequencing of tumor tissues. The safety and immune response were assessed through adverse events and ELISpot. The clinical response was evaluated by progression-free survival (PFS), imaging examination, clinical tumor marker detection, circulating tumor DNA (ctDNA) sequencing. Changes in health-related quality of life were measured by the FACT-C scale. A total of six MSS-CRC patients with recurrence or metastasis after surgery and chemotherapy were administered with personalized neoantigen vaccines. Neoantigen-specific immune response was observed in 66.67% of the vaccinated patients. Four patients remained progression-free up to the completion of clinical trial. They also had a significantly longer progression-free survival time than the other two patients without neoantigen-specific immune response (19 vs. 11 months). Changes in health-related quality of life improved for almost all patients after the vaccine treatment. Our results shown that personalized neoantigen vaccine therapy is likely to be a safe, feasible and effective strategy for MSS-CRC patients with postoperative recurrence or metastasis.

Atypical emotion sharing in individuals with mirror sensory synaesthesia.

Being able to empathise with others is a crucial ability in everyday life. However, this does not usually entail feeling the pain of others in our own bodies. For individuals with mirror-sensory synaesthesia (MSS), however, this form of empathic embodiment is a common feature. Our study investigates the empathic ability of adults who experience MSS using a video-based empathy task. We found that MSS participants did not differ from controls on emotion identification and affective empathy; however, they showed higher affect sharing (degree to which their affect matches what they attribute to others) than controls. This finding indicates difficulties with self-other distinction, which our data shows results in fewer signs of prosocial behaviour. Our findings are in line with the self-other control theory of MSS and highlight how the use of appropriate empathy measures can contribute to our understanding of this important socio-affective ability, both in typical and atypical populations.

How to overcome tumor resistance to anti-PD-1/PD-L1 therapy by immunotherapy modifying the tumor microenvironment in MSS CRC.

Immune checkpoint inhibitors (ICIs), including anti-programmed cell death-1/anti-programmed cell death ligand-1 (anti-PD-1/PD-L1) therapy, have elicited impressive clinical outcomes in several malignancies. This is regarded as a pivotal breakthrough in cancer treatment. However, a vast majority of colorectal cancer (CRC) cases are microsatellite stable (MSS) and respond poorly to anti-PD-1/PD-L1 immunotherapies. Since ICIs serve as rescuers for immune cell-mediated cancer cell elimination, the limited efficacy of anti-PD-1/PD-L1 treatments may be attributed to the privileged tumor microenvironment (TME), which is characterized by unavailable immunosurveillance. Thus, it is essential to modify the pre-existing disordered immune system prior to the application of an anti-PD-1/PD-L1 therapy. In this review, to overcome unsatisfactory immunotherapy in CRC patients with MSS, we discussed various combination therapies based on TME reconstruction for improving the susceptibility to anti-PD-1/PD-L1 treatment.

Role of the cGAS-STING pathway in regulating the tumor-immune microenvironment in dMMR/MSI colorectal cancer.

Deficient mismatch repair (dMMR)/microsatellite instability (MSI) colorectal cancer (CRC) has high immunogenicity and better prognosis compared with proficient MMR (pMMR)/microsatellite stable (MSS) CRC. Although the activation of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway has been considered to contribute to the high number of CD8+ TILs, its role in dMMR/MSI CRC is largely unknown. In this study, to examine the role of the cGAS-STING pathway on the recruitment of CD8+ TILs in dMMR/MSI CRC, we used public datasets and clinical tissue samples in our cohorts to evaluate the expression of cGAS, STING, and CD8+ TILs in pMMR/MSS and dMMR/MSI CRCs. According to the analysis of public datasets, the expression of cGAS-STING, CD8 effector gene signature, and CXCL10-CCL5, chemoattractants for CD8+ TILs which regulated by the cGAS-STING pathway, was significantly upregulated in dMMR/MSI CRC, and the expression of cGAS-STING was significantly associated with the expression of CD8 effector gene signature. Immunohistochemistry staining of the clinical tissue samples (n = 283) revealed that cGAS-STING was highly expressed in tumor cells of dMMR CRC, and higher expression of cGAS-STING in tumor cells was significantly associated with the increased number of CD8+ TILs. Moreover, we demonstrated that the downregulation of MMR gene in human CRC cell lines enhanced the activation of the cGAS-STING pathway. Taken together, for the first time, we found that dMMR/MSI CRC has maintained a high level of cGAS-STING expression in tumor cells, which might contribute to abundant CD8+ TILs and immune-active TME.

Mss51 deletion increases endurance and ameliorates histopathology in the mdx mouse model of Duchenne muscular dystrophy.

Mitochondrial derangement is an important contributor to the pathophysiology of muscular dystrophies and may be among the earliest cellular deficits. We have previously shown that disruption of Mss51, a mammalian skeletal muscle protein that localizes to the mitochondria, results in enhanced muscle oxygen consumption rate, increased endurance capacity, and improved limb muscle strength in mice with wildtype background. Here, we investigate whether Mss51 deletion in the mdx murine model of Duchenne muscular dystrophy (mdx-Mss51 KO) counteracts the muscle pathology and mitochondrial irregularities observed in mdx mice. We found that mdx-Mss51 KO mice had increased myofiber oxygen consumption rates and an amelioration of muscle histopathology compared to mdx counterparts. This corresponded with greater treadmill endurance and less percent fatigue in muscle physiology, but no improvement in forelimb grip strength or limb muscle force production. These findings suggest that although Mss51 deletion ameliorates the skeletal muscle mitochondrial respiration defects in mdx and improves fatigue resistance in vivo, the lack of improvement in force production suggests that this target alone may be insufficient for a therapeutic effect.

SARS-CoV-2 main protease (3CLpro) interaction with acyclovir antiviral drug/methyl-ß-cyclodextrin complex: Physiochemical characterization and molecular docking.

During the current outbreak of the novel coronavirus disease 2019 (COVID-19), researchers have examined several antiviral drugs with the potential to inhibit the proliferation of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The antiviral drug acyclovir (AVR), which is used to treat COVID-19, in complex with methyl-ß-cyclodextrin (Mß-CD) was examined in the solution and solid phases. UV-visible and fluorescence spectroscopic analyses confirmed that the guest (AVR) was included inside the host (Mß-CD) cavity. A solid inclusion complex of AVR was prepared by co-precipitation, physical mixing, kneading, and bath sonication methods at a 1:1 ratio of Mß-CD:AVR. The prepared Mß-CD:AVR inclusion complex was characterized using Fourier transform infrared spectroscopy (FTIR), X-ray powder diffraction (XRD), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and scanning electron microscopy (SEM) analysis. Phase solubility studies indicated the Mß-CD:AVR inclusion complex exhibited a higher stability constant and linear enhancement in AVR solubility with increasing Mß-CD concentrations. In silico analysis of the Mß-CD/AVR inclusion complex confirmed that AVR drugs show potential as inhibitors of SARS-CoV-2 3C-like protease (3CLpro) receptors. Results obtained using the PatchDock and FireDock servers indicated that the most favorable docking ligand was Mß-CD:AVR, which interacted with SARS-CoV-2 (3CLPro) protease inhibitors with high geometric shape complementarity scores (2522 and 5872) and atomic contact energy (-313.77 and -214.70 kcal mol-1). Our results suggest that the Mß-CD/AVR inclusion complex inhibits the main protease of SARS-CoV-2, although further wet-lab experiments are needed to verify these findings.

A global analysis of IFT-A function reveals specialization for transport of membrane-associated proteins into cilia.

Intraflagellar transport (IFT), which is essential for the formation and function of cilia in most organisms, is the trafficking of IFT trains (i.e. assemblies of IFT particles) that carry cargo within the cilium. Defects in IFT cause several human diseases. IFT trains contain the complexes IFT-A and IFT-B. To dissect the functions of these complexes, we studied a Chlamydomonas mutant that is null for the IFT-A protein IFT140. The mutation had no effect on IFT-B but destabilized IFT-A, preventing flagella assembly. Therefore, IFT-A assembly requires IFT140. Truncated IFT140, which lacks the N-terminal WD repeats of the protein, partially rescued IFT and supported formation of half-length flagella that contained normal levels of IFT-B but greatly reduced amounts of IFT-A. The axonemes of these flagella had normal ultrastructure and, as investigated by SDS-PAGE, normal composition. However, composition of the flagellar 'membrane+matrix' was abnormal. Analysis of the latter fraction by mass spectrometry revealed decreases in small GTPases, lipid-anchored proteins and cell signaling proteins. Thus, IFT-A is specialized for the import of membrane-associated proteins. Abnormal levels of the latter are likely to account for the multiple phenotypes of patients with defects in IFT140.This article has an associated First Person interview with the first author of the paper.

The Long Noncoding RNA CCAT2 Induces Chromosomal Instability Through BOP1-AURKB Signaling.

BACKGROUND & AIMS: Chromosomal instability (CIN) is a carcinogenesis event that promotes metastasis and resistance to therapy by unclear mechanisms. Expression of the colon cancer-associated transcript 2 gene (CCAT2), which encodes a long noncoding RNA (lncRNA), associates with CIN, but little is known about how CCAT2 lncRNA regulates this cancer enabling characteristic. METHODS: We performed cytogenetic analysis of colorectal cancer (CRC) cell lines (HCT116, KM12C/SM, and HT29) overexpressing CCAT2 and colon organoids from C57BL/6N mice with the CCAT2 transgene and without (controls). CRC cells were also analyzed by immunofluorescence microscopy, γ-H2AX, and senescence assays. CCAT2 transgene and control mice were given azoxymethane and dextran sulfate sodium to induce colon tumors. We performed gene expression array and mass spectrometry to detect downstream targets of CCAT2 lncRNA. We characterized interactions between CCAT2 with downstream proteins using MS2 pull-down, RNA immunoprecipitation, and selective 2'-hydroxyl acylation analyzed by primer extension analyses. Downstream proteins were overexpressed in CRC cells and analyzed for CIN. Gene expression levels were measured in CRC and non-tumor tissues from 5 cohorts, comprising more than 900 patients. RESULTS: High expression of CCAT2 induced CIN in CRC cell lines and increased resistance to 5-fluorouracil and oxaliplatin. Mice that expressed the CCAT2 transgene developed chromosome abnormalities, and colon organoids derived from crypt cells of these mice had a higher percentage of chromosome abnormalities compared with organoids from control mice. The transgenic mice given azoxymethane and dextran sulfate sodium developed more and larger colon polyps than control mice given these agents. Microarray analysis and mass spectrometry indicated that expression of CCAT2 increased expression of genes involved in ribosome biogenesis and protein synthesis. CCAT2 lncRNA interacted directly with and stabilized BOP1 ribosomal biogenesis factor (BOP1). CCAT2 also increased expression of MYC, which activated expression of BOP1. Overexpression of BOP1 in CRC cell lines resulted in chromosomal missegregation errors, and increased colony formation, and invasiveness, whereas BOP1 knockdown reduced viability. BOP1 promoted CIN by increasing the active form of aurora kinase B, which regulates chromosomal segregation. BOP1 was overexpressed in polyp tissues from CCAT2 transgenic mice compared with healthy tissue. CCAT2 lncRNA and BOP1 mRNA or protein were all increased in microsatellite stable tumors (characterized by CIN), but not in tumors with microsatellite instability compared with nontumor tissues. Increased levels of CCAT2 lncRNA and BOP1 mRNA correlated with each other and with shorter survival times of patients. CONCLUSIONS: We found that overexpression of CCAT2 in colon cells promotes CIN and carcinogenesis by stabilizing and inducing expression of BOP1 an activator of aurora kinase B. Strategies to target this pathway might be developed for treatment of patients with microsatellite stable colorectal tumors.

Clinical, Pathology, Genetic, and Molecular Features of Colorectal Tumors in Adolescents and Adults 25 Years or Younger.

BACKGROUND & AIMS: Colorectal cancers (CRCs) are rare in adolescents and adults ages 25 years or younger. We analyzed clinical, pathology, and molecular features of colorectal tumors from adolescents and young adults in an effort to improve genetic counseling, surveillance, and, ultimately, treatment and outcomes. METHODS: We analyzed clinical data and molecular and genetic features of colorectal tumor tissues from 139 adolescents or young adults (age, ≤25 y; median age, 23 y; 58% male), collected from 2000 through 2017; tumor tissues and clinical data were obtained from the nationwide network and registry of histopathology and cytopathology and The Netherlands Cancer Registry, respectively. DNA samples from tumors were analyzed for microsatellite instability, mutations in 56 genes, and genome-wide somatic copy number aberrations. RESULTS: Mucinous and/or signet ring cell components were observed in 33% of tumor samples. A genetic tumor risk syndrome was confirmed for 39% of cases. Factors associated with shorter survival time included younger age at diagnosis, signet ring cell carcinoma, the absence of a genetic tumor risk syndrome, and diagnosis at an advanced stage of disease. Compared with colorectal tumors from patients ages 60 years or older in the Cancer Genome Atlas, higher proportions of tumors from adolescents or young adults were microsatellite stable with nearly diploid genomes, or contained somatic mutations in TP53 and POLE, whereas lower proportions contained mutations in APC. CONCLUSIONS: We found clinical, molecular, and genetic features of CRCs in adolescents or young adults to differ from those of patients older than age 60 years. In 39% of patients a genetic tumor risk syndrome was identified. These findings provide insight into the pathogenesis of CRC in young patients and suggest new strategies for clinical management. Performing genetic and molecular analyses for every individual diagnosed with CRC at age 25 years or younger would aid in this optimization.

T-cell immunoglobulin and ITIM domain, as a potential immune checkpoint target for immunotherapy of colorectal cancer.

The importance of the tumor microenvironment in cancer progression has been well studied for many years. Immune checkpoint inhibitors (ICIs) are regarded as potential strategies in enhancing the immune responses in patients with cancer, particularly colorectal cancer (CRC). Notably, CRCs are extraordinarily heterogeneous and mostly are microsatellite-stable (MSS) or cold tumors, which means that the immune response is not usually as strong as that of foreign cells. T-cell immunoglobulin and ITIM domain (TIGIT) is a new immune checkpoint receptor overexpressed inside the CRC tumor-immune microenvironments. Moreover, several studies have shown that TIGIT in combination with other ICIs and/or conventional treatments, can lead to a robust anti-tumor response in CRC. This review looks deep inside TIGIT expression patterns, their various functions, and possible immunotherapy strategies to increase survival rates and decrease immune-related adverse events.

Microsatellite Instability and Effectiveness of Adjuvant Treatment in pT1N1 Gastric Cancer: A Multicohort Study.

BACKGROUND: Microsatellite status is a prognostic biomarker in advanced gastric cancer. This retrospective study aimed to investigate the usefulness of microsatellite status in predicting prognosis and response to adjuvant treatment in pT1N1 gastric cancer. PATIENTS AND METHODS: Among 875 patients who underwent radical gastrectomy for pT1N1 gastric cancer at two tertiary hospitals, 838 with available microsatellite instability (MSI) data were included and classified into two groups according to microsatellite status: microsatellite stable (MSS) and MSI-high (MSI-H). Recurrence-free survival rate and risk factors for tumor recurrence were analyzed. RESULTS: Of 838 gastric cancer patients, 100 (11.9%) were MSI-H and 307 (36.6%) received adjuvant treatment. During median follow-up of 70 months, 42 (5.0%) patients experienced gastric cancer recurrence; hematogenous recurrences were the most common (45.2%). Recurrence-free survival was similar in the MSS and MSI-H groups (p = 0.27), and adjuvant treatment did not show an oncological benefit over surgery alone for pT1N1 gastric cancer (p = 0.53). On univariate analysis, age, operation period, and Lauren classification were significantly associated with tumor recurrence, while adjuvant treatment and MSI status were not associated with tumor recurrence. On multivariate analysis, MSI status was not associated with tumor recurrence, and adjuvant treatment worsened the tumor recurrence risk [hazard ratio (HR) 2.373, 95% confidence interval (CI) 1.125-5.006, p = 0.023). CONCLUSION: MSI status may not be a prognostic factor for tumor recurrence or a predictor of response to adjuvant treatment in pT1N1 gastric cancer patients. Considering that the effect of adjuvant treatment to decrease the risk of tumor recurrence is not clear, it may not be indicated in pT1N1 patients.

Screening for potential interaction partners with surface plasmon resonance imaging coupled to MALDI mass spectrometry.

We coupled SPR imaging (SPRi) with matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) to identify new potential RNA binders. Here, we improve this powerful method, especially by optimizing the proteolytic digestion (type of reducing agent, its concentration, and incubation time), to work with complex mixtures, specifically a lysate of the rough mitochondrial fraction from yeast. The advantages of this hyphenated method compared to column-based or separate analyses are (i) rapid and direct visual readout from the SPRi array, (ii) possibility of high-throughput analysis of different interactions in parallel, (iii) high sensitivity, and (iv) no sample loss or contamination due to elution or micro-recovery procedures. The model system used is a catalytically active RNA (group IIB intron from Saccharomyces cerevisiae, Sc.ai5γ) and its cofactor Mss116. The protein supports the RNA folding process and thereby the subsequent excision of the intronic RNA from the coding part. Using the novel approach of coupling SPR with MALDI MS, we report the identification of potential RNA-binding proteins from a crude yeast mitochondrial lysate in a non-targeted approach. Our results show that proteins other than the well-known cofactor Mss116 interact with Sc.ai5γ (Dbp8, Prp8, Mrp13, and Cullin-3), suggesting that the intron folding and splicing are regulated by more than one cofactor in vivo.

Tumor mutation burden is correlated with response and prognosis in microsatellite-stable (MSS) gastric cancer patients undergoing neoadjuvant chemotherapy.

BACKGROUND: Neoadjuvant chemotherapy (NACT) before radical gastrectomy is preferred for locally advanced gastric cancer (GC). However, clinical practices demonstrate that a considerable proportion of GC patients do not benefit from NACT, largely due to the lack of biomarkers for patient selection and prognosis prediction. A recent study revealed that patients with microsatellite instability-high (MSI-H) may be resistant to NACT, however, most tumors in Chinese GC patients (~ 95%) are characterized by microsatellite stability (MSS). Here, we aimed to discover new molecular biomarkers for this larger population. METHODS: We performed whole-exome sequencing on 46 clinical samples (pre- and post-treatment) from 30 stage II/III MSS GC patients whose response to NACT was rigorously defined. Serum tumor markers (TMs), including AFP, CEA, CA199, CA724 and CA242 were measured during the course. RESULTS: High tumor mutation burden (TMB-H) and 19q12 amplification (19q12 +) were positively associated with the NACT response. When TMB and 19q12 amplification were jointly analyzed, those with TMB-H or 19q12 + showed favorable response to NACT (p = 0.035). Further, TMB-H was negatively correlated with ypN stage, lymph node metastasis, and macrophage infiltration. Patients with TMB-H showed better disease-free survival (DFS) than those with TMB-L (P = 0.025, HR = 0.1331), and this was further validated using two larger GC datasets: TCGA-STAD (p = 0.004) and ICGC-CN (p = 0.045). CONCLUSION: The combination of TMB-H and 19q12 + can serve as an early indicator of response to NACT. Superior to traditional clinical indicators, TMB-H is a robust and easily accessible candidate biomarker associated with better DFS, and can be evaluated at the time of diagnosis.

Quantitative evaluation of range and metabolic activity of hepatic alveolar echinococcosis lesion microenvironment using PET/CT and multi-site sampling method.

BACKGROUND: Alveolar echinococcosis (AE) lesion microenvironment (LME) is crucial site where parasite-host interactions happen and of great significance during surgery and obtaining liver samples for basic research. However, little is known about quantification of LME range and its' metabolic activity regarding different lesion characteristics. METHODS: A prospective and retrospective analysis of LME from surgical AE patients was performed. Patients (n = 75) received abdominal computed tomography (CT) and position emission tomography/computed tomography using 18F-fluodeoxyglucose (18F-FDG-PET/CT) within 1 week prior to surgery. Semiquantitatively, calcification was clustered with 0%, < 50% and ≥ 50% degrees at lesion periphery; liquefaction was clustered with 0%, < 50%, 50 ~ 75%, ≥75% degrees at lesion center using volumetric ratio. Tumor to background ratio (TBR) of 18F-FDG standard uptake value (SUV, n = 75) was calculated, and range of 18F-FDG uptake area was measured; Multi-site sampling method (MSS, n = 35) was introduced to obtain histological slides to evaluate immune cell infiltrative ranges. RESULTS: Altogether six major lesion groups have been identified (A: 0% calcified, 0% liquefied; B: ≥50% calcified, 0% liquefied; C: < 50% calcified, < 50% liquefied; D: ≥50% calcified, < 50% liquefied; E: < 50% calcified, 50 ~ 75% liquefied; F: ≥50% calcified, ≥75% liquefied). Statistically, TBR values respectively were 5.1 ± 1.9, 2.7 ± 1.2, 4.2 ± 1.2, 2.7 ± 0.7, 4.6 ± 1.2, 2.9 ± 1.1 in groups A ~ F, and comparisons showed A > B, A > D, A > F, E > B, E > D, E > F, C > B, C > D, C > F (P < 0.05); LME ranges indicated by PET/CT respectively were 14.9 ± 3.9, 10.6 ± 1.5, 12.3 ± 1.1, 7.8 ± 1.6, 11.1 ± 2.3, 7.0 ± 0.4 mm in groups A ~ F, and comparisons showed A > B, A > D, A > F, A > E, C > B, C > D, C > F, E > D, E > F, B > D, B > F (P < 0.05); LME ranges indicated by MSS respectively were 17.9 ± 4.9, 13.0 ± 2.7, 11.9 ± 2.6, 6.0 ± 2.2, 11.0 ± 4.1, 6.0 ± 2.2 mm in groups A ~ F, and comparisons showed A > C, A > D, A > F, B > D, B > F, C > D, C > F (P < 0.05). Generally, less calcifications indicated higher TBR values and wider LME ranges; and, severer liquefactions indicated smaller LME ranges. Additionally, patients with previous medication history had lower TBR values. CONCLUSIONS: PET/CT and MSS method showed distinct TBRs and LME ranges for different calcifications and liquefactions. This study would be able to provide references for both surgical resections of lesions and more accurate sample acquisitions for basic research targeted to immunology.

Statistical Evaluation of Total Expiratory Breath Samples Collected throughout a Year: Reproducibility and Applicability toward Olfactory Sensor-Based Breath Diagnostics.

The endogenous volatile organic compounds (VOCs) in exhaled breath can be promising biomarkers for various diseases including cancers. An olfactory sensor has a possibility for extracting a specific feature from collective variations of the related VOCs with a certain health condition. For this approach, it is important to establish a feasible protocol for sampling exhaled breath in practical conditions to provide reproducible signal features. Here we report a robust protocol for the breath analysis, focusing on total expiratory breath measured by a Membrane-type Surface stress Sensor (MSS), which possesses practical characteristics for artificial olfactory systems. To assess its reproducibility, 83 exhaled breath samples were collected from one subject throughout more than a year. It has been confirmed that the reduction of humidity effects on the sensing signals either by controlling the humidity of purging room air or by normalizing the signal intensities leads to reasonable reproducibility verified by statistical analyses. We have also demonstrated the applicability of the protocol for detecting a target material by discriminating exhaled breaths collected from different subjects with pre- and post-alcohol ingestion on different occasions. This simple yet reproducible protocol based on the total expiratory breath measured by the MSS olfactory sensors will contribute to exploring the possibilities of clinical applications of breath diagnostics.