Combining serum inflammation indexes at baseline and post treatment could predict pathological efficacy to anti­PD­1 combined with neoadjuvant chemotherapy in esophageal squamous cell carcinoma.

BACKGROUND: The neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), platelet-to-lymphocyte ratio (PLR), and systemic immune-inflammation index (SII) have been used to predict therapeutic response in different tumors. However, no assessments of their usefulness have been performed in esophageal squamous cell carcinoma (ESCC) patients receiving anti­PD­1 combined with neoadjuvant chemotherapy. METHODS: The respective data of 64 ESCC patients receiving anti­PD­1 combined with neoadjuvant chemotherapy were analyzed. Whether NLR, LMR, PLR, and SII at baseline and post-treatment might predict pathological response to anti­PD­1 plus neoadjuvant chemotherapy, and cutoff values of these parameters were all determined by ROC curve analysis. RESULTS: NLR (cutoff = 3.173, AUC = 0.644, 95% CI 0.500-0.788, P = 0.124, sensitivity = 1.000, specificity = 0.373), LMR (cutoff = 1.622, AUC = 0.631, 95% CI 0.477-0.784, P = 0.161, sensitivity = 0.917, specificity = 0.137), PLR (cutoff = 71.108, AUC = 0.712, 95% CI 0.575-0.849, P = 0.023, sensitivity = 1.000, specificity = 0.059), and SII at baseline (cutoff = 559.266, AUC = 0.681, 95% CI 0.533-0.830, P = 0.052, sensitivity = 0.373, specificity = 1.000) seemed to be a useful predictor for distinguishing responders from non-responders. Combining NLR with SII at baseline (AUC = 0.729, 95% CI 0.600-0.858, P = 0.014, sensitivity = 0.917, specificity = 0.510), LMR and SII at baseline (AUC = 0.735, 95% CI 0.609-0.861, P = 0.012, sensitivity = 1.000 specificity = 0.471), PLR and SII at baseline (AUC = 0.716, 95% CI 0.584-0.847, P = 0.021, sensitivity = 1.000 specificity = 0.431), and LMR and PLR at post-treatment in the third period (AUC = 0.761, 95% CI 0.605-0.917, P = 0.010, sensitivity = 0.800, specificity = 0.696) might slightly increase the prediction ability to determine patients who have response or no response. Finally, combining LMR at baseline, SII at post-treatment in the second period with PLR at post-treatment in the third period could be considered a better predictor for discriminating responders and non-responders than single or dual biomarkers (AUC = 0.879, 95% CI 0.788-0.969, P = 0.0001, sensitivity = 0.909, specificity = 0.800). CONCLUSIONS: The models we constructed allowed for the accurate and efficient stratification of ESCC patients receiving anti-PD-1 plus chemotherapy and are easily applicable for clinical practice at no additional cost.

G-quadruplex-enhanced circular single-stranded DNA (G4-CSSD) adsorption of miRNA to inhibit colon cancer progression.

BACKGROUND: Chromosomal heterogeneity leads to the abnormal expression and mutation of tumor-specific genes. Drugs targeting oncogenes have been extensively developed. However, given the random mutation of tumor suppressor genes, the development of its targeted drugs is difficult. METHODS: Our early research revealed that artificial circular single-stranded DNA (CSSD) can restore multiple tumor suppressor genes to inhibit tumor malignant progression by adsorbing miRNA. Here, we improved CSSD to a fully closed single-stranded DNA with G quadruplex DNA secondary structure (G4-CSSD), which made G4-CSSD with higher acquisition rate and decreased degradation. The Cancer Genome Atlas (TCGA) and Human Protein Atlas database were used to predict tumour suppressor genes in colon cancer. Cellular and animal experiments were performed to validate the role of G4-CSSD in cancer cell progression. RESULTS: In colon cancer, we observed the simultaneous low expressions of chloride channel accessory 1 (CLCA1), UDP-GlcNAc:betaGal beta-1,3-N-acetylglucosaminyltransferase 6 (B3GNT6) and UDP glucuronosyltransferase family 2 member A3 (UGT2A3), which indicated an favourable prognosis. After repressing miR-590-3p with G4-CSSD590, the upregulation of CLCA1, B3GNT6 and UGT2A3 inhibited the proliferation and metastasis of colon cancer cells. CONCLUSIONS: This study may provide basis for new treatment methods for colon cancer by restoration of tumor suppressor genes.

Th1-involved immune infiltrates improve neoadjuvant chemoradiotherapy response of esophageal squamous cell carcinoma.

Neoadjuvant chemoradiotherapy (NCRT) followed by surgery is recommended for locally advanced esophageal squamous cell carcinoma (ESCC) treatment. Patients who achieve a pathological complete response (pCR) have better survival. Our study aimed to discover immune-associated predictors of pCR in ESCC. Herein, we found that Th1-cell infiltration inferred from RNA sequencing was higher in the pCR group than in the non-pCR group. Multiplexed immunohistochemistry (mIHC) confirmed that Th1-, CD8+ T-, NK-, NKT-, and dendritic-cell infiltration was positively associated with pCR. The spatial relationships between Th1 cells and CD8+ T, NK, NKT, dendritic, or ESCC cells were significant pCR predictors. The active and desert subtypes were identified based on immune cell infiltration, and showed different pCR rates. In vitro experiments confirmed that Th1 cells inhibited the proliferation and improved the chemosensitivity and radiosensitivity of ESCC cells. Th1 cells upregulated interferon-gamma response signaling and antigen presentation pathways and downregulated lipid metabolism and MAPK pathways of ESCC cells. These findings highlight the important role of Th1 cells as the predictor of pCR and the regulator of chemosensitivity and radiosensitivity of ESCC, and suggest elevating Th1-infiltration as a strategy to improve NCRT response.

Immune Microenvironment Characteristics of Urachal Carcinoma and Its Implications for Prognosis and Immunotherapy.

Urachal carcinoma (UrC) is an exceedingly rare tumor and lacks effective treatment. Herein, we characterized an immune microenvironment characteristic of UrC in detail and identified its implications for prognosis and immunotherapy. In total, 37 resections of UrC were stained for CD20, CD3, CD4, CD8, FOXP3, CD68, HLA-DR, CD163, PD1, and PD-L1, as well as mismatch repair protein including MSH2, MSH6, MLH1, and PMS2 by immunohistochemistry. Intratumoral and peritumoral immune cell densities or the proportion of PD1 and PD-L1 expression alongside MSH2, MSH6, MLH1, and PMS2 status were manually evaluated using the whole slide. UrC patients with the number of tertiary lymphoid structures (TLS) per slide tended to be higher in tumors with dMMR (p = 0.1919), and tumors with higher number of TLS tended to have longer OS (p = 0.0940) and DFS (p = 0.0700). High densities of CD3+ T, CD8+ T, and CD68+ cells were significantly associated with worse OS and DFS (both p<0.05). Increased intratumoral (p = 0.0111) and peritumoral (p = 0.0485) CD8+ T cell densities were significantly associated with PD-L1 expression or increasing proportion of PD-L1 expression on immune cells. Similarly, increased intratumoral (p = 0.0008) and peritumoral (p = 0.063) CD8+ T cell densities were significantly associated with increasing proportion of PD1 expression on immune cells. Tumors with PD-L1 positive expression on immune cells had a significantly increased proportion of PD1 expression (p = 0.0121). High peritumoral CD8+ T cell density (>73.7/mm2) was significantly associated with worse OS (p = 0.0120) and DFS (p = 0.00095). The number of TLS seems to be considered not only as histopathological characteristics in predicting MMR status of UrC, but also as a prognostic or therapeutic biomarker, and we also provide some important suggestions for targeting PD-1/PD-L1 checkpoint in UrC.

[Determination of polychlorinated naphthalenes in soil using accelerated solvent extraction-molecular sieves solid-phase extraction coupled with gas chromatography-tandem mass spectrometry].

Emerging pollutants (EPs) are chemical substances that are commonly not regulated and can be detected at low or very low concentrations. However, EPs have triggered special concern because their long-term adverse effects on the environment and human health remain unknown. Most EPs show biological toxicity, environmental persistence, and bioaccumulation. Even at low concentrations in the environment, EPs may pose significant environmental and health risks. Therefore, their treatment has been explicitly included in the 14th Five Year Plan for National Economic and Social Development of the People's Republic of China and the Outline of the Long-term Goals for 2035. Soil is a source of pollutants, and its quality is directly related to economic development, ecological security, and people's livelihood. At present, China's soil environmental monitoring system is not perfect, and the ability to monitor these new organic pollutants is lagging. Therefore, to strengthen the supervision of construction and agricultural land soil environments, it is essential to strengthen the soil environment monitoring ability for these EPs and establish a reliable, steady, and economic analysis method, including their separation and analysis methods in soil. Polychlorinated naphthalenes (PCNs) have received considerable attention as emerging halogenated compounds. They were listed in Annexes A and C of the Stockholm Convention on persistent organic pollutants (POPs) in 2015 because of their persistence, multimedia fate, and toxicity. PCNs have now been detected in the surrounding soils. Owing to their trace levels in complex soil, high requirements have been put forward for the pretreatment and instrument analysis of PCNs. This study aims to develop a new method for the selective purification of PCNs in soil, which can not only effectively remove lipids and other interferences in soil but also effectively reduce time, labor, and material costs in the pre-treatment process. Based on the physicochemical properties of the 13X molecular sieve, it was explored to purify soil-extracts as solid-phase extraction (SPE) sorbents. With n-hexane as the loading and rinsing solvent, 10 mL of a dichloromethane/n-hexane mixture (2∶15, v/v) was used to elute the PCNs. Moreover, selective separation of target substances from lipid macromolecules and other interferences could be achieved simultaneously. For the selective separation of PCNs, the average recovery of the internal standard could reach 56.1% to 88.0%. 13X molecular sieves are superior to gel permeation chromatography (GPC) and Florisil SPE, and they exhibit good cleanup efficiency similar to a multilayer silica gel/alumina column (53.0%-117.0%). Although the obtained recoveries are not as high as those obtained with a multilayer silica gel/alumina column, 13X molecular sieves have advantages in terms of simple operation, environmental friendliness, and low cost. Based on these fundamental experiments, accelerated solvent extraction was used to extract targets in soil, molecular sieves were used as SPE sorbents for purification, and GC-MS/MS was employed for PCN analysis. This method was developed as a systematic analytical method for PCNs determination. The method detection limits (MDLs) for PCN homologs were in the range of 0.009-0.6 ng/g. The precision and accuracy of the method were evaluated using spiked matrices. At three spiked levels (4, 10, and 18 ng), the recoveries of PCNs (CN-3, 13, 42, 46, 52, 53, 73, and 75) were 70%-128%, 71%-115%, and 61%-114%, respectively, and the corresponding relative standard derivations were 4.2%-23%, 6.5%-31% and 4.7%-22%. Thus, this method meets the requirements of trace analysis and shows acceptable parallelism, sensitivity, accuracy, and precision, thus being feasible for the analysis of emerging pollutant. The method is expected to play an important role in sample pretreatment in the future, especially for the nationwide investigation of soil pollution.

Body mass index, as a novel predictor of hepatocellular carcinoma patients treated with Anti-PD-1 immunotherapy.

Immunocheckpoint inhibitors have shown significant efficacy in the treatment of hepatocellular carcinoma (HCC), but there are individual differences. The aim of this study was to explore body mass index (BMI) as a predictor of anti-PD-1 efficacy in patients with HCC. We retrospectively analyzed 101 HCC patients who treated with anti-PD-1 at Sun Yat-sen University Cancer Center from July 2018 to November 2019 and divided them into overweight (BMI > 24.9) and non-overweight (BMI ≤ 24.9) groups based on baseline BMI levels. BMI > 24.9 accounted for 22 cases (21.8%) and BMI ≤ 24.9 accounted for 79 cases (78.2%) in the study cohort. Overweight patients had higher disease control rates than non-overweight patients (P = 0.019, respectively). The mean progression-free survival (PFS) in overweight patients (10.23 months) was significantly longer than that of non-overweight patients (6.85 months; P = 0.027). Among patients with immune-related adverse events (irAEs), the mean PFS was also significantly longer in overweight patients (7.72 months) than in non-overweight patients (5.31 months, P = 0.034). Multivariate analysis showed that BMI was an independent prognostic factor for PFS in HCC patients treated with anti-PD-1 (hazard ratio: 0.47, P = 0.044). Thus, higher BMI predicts a better prognosis among HCC patients treated with anti-PD-1. In clinical practice, patients' BMI can provide a useful tool for predicting the efficacy of anti-PD-1 therapy.

Impact of WTAP in small HCC and paired adjacent non-neoplastic liver tissue on recurrence: A cohort study with external extension analysis.

Background: To evaluate prognostic value of WTAP levels in tumor and paired adjacent non-neoplastic liver tissues (PANLT) for cases of hepatitis B virus (HBV)-positive Asian small hepatocellular carcinoma (sHCC) patients who received curative partial hepatectomy. Method: The investigation with two external cohorts were included. Associations between hazard risk of recurrence and continuous WTAP levels were investigated with restricted cubic spline models. Cox and inverse probability weighting models were established for survival analysis. Based on interaction effects, further stratification analysis was performed. Landmark analysis was employed to analyze cases of late recurrence. Finally, sensitivity analysis was performed to assess unmeasured confounders. Findings: In an investigation cohort of 307 patients, restricted cubic spline models indicated that hazard risk of recurrence increases with elevated WTAP levels for sHCC and PANLT. However, using Cox and inverse probability weighting models, no significant differences were observed in recurrence-free survival (RFS) between groups with different WTAP levels in sHCC. Multivariate analysis showed that patients with high PANLT WTAP levels had significantly worse RFS (HR 1.567, 95% CI 1.065-2.307; p = 0.023). Based on the significant interaction effect between WTAP levels in sHCC and PANLT, stratification analysis revealed that recurrence risk is more pronounced in patients with high WTAP levels in both PANLT and sHCC. Landmark analysis showed that late recurrence was more likely to occur in patients with high PANLT WTAP levels (HR 2.058, 95% CI 1.113-3.805; p = 0.021). Moreover, the detrimental effects of elevated PANLT WTAP levels on RFS were validated with two external cohorts. Sensitivity analysis confirmed the robustness of results. Conclusion: Increased PANLT WTAP expression levels independently predict high recurrence risk in HBV-positive Asian sHCC patients. Both tumor tissues and PANLT need to be considered together in future clinical practice to obtain a more comprehensive and accurate evaluation for recurrence risk.

Pan-Cancer Analysis of PARP1 Alterations as Biomarkers in the Prediction of Immunotherapeutic Effects and the Association of Its Expression Levels and Immunotherapy Signatures.

Background: Poly (ADP-ribose) polymerases-1 (PARP1) alterations are associated with PARP1 inhibitor resistance, regulating the function of Treg cells and PDL1 expression in tumor cells, and high PARP1 expression is significantly associated with aggressive behavior and chemotherapeutic resistance in several tumors. However, a comprehensive analysis of the predictive values of PARP1 alteration for immune checkpoint inhibitor (ICI) effectiveness in tumors remains unclear, and the associations between its expression and immunotherapy signatures also needs to be explored further. Methods: We performed some analyses with the cBioPortal online database (https://www.cbioportal.org), TIMER2.0 (Tumor Immune Estimation Resource 2.0, http://timer.comp-genomics.org/) and TCGA database (https://xenabrowser.net or https://portal.gdc.cancer.gov/). Survival analysis was conducted using Kaplan-Meier method, and the associations between PARP1 transcription levels and immune checkpoint gene expression, the number of neoantigens, tumor mutation burden (TMB) levels, and microsatellite instability (MSI) event are analyzed by spearman correlation analysis and visualization of those mentioned above is performed using R, version 3.6.3 (http://www.r-project.org/). Results: We found that PARP1 was altered in 1338 (2.9%) out of 45604 patients with diverse tumors, which was associated with markedly higher TMB levels in a variety of tumors (P < 0.01). Impressively, patients with PARP1 alterations in advanced tumors showed better overall survival (OS) in the ICI-treated cohort (P = 0.016). PARP1 altered group was substantially correlated with higher immune infiltrates across most tumors, including CD8+ T cells in colorectal adenocarcinoma (P = 0.0061), endometrial carcinoma (P = 0.0033), stomach cancer (P = 0.033), and cervical cancer (P = 0.026), respectively. The PARP1 altered group showed high expression in transcription (P < 0.001), and higher expression of LAG3, PDCD1, CTLA-4, and TIGIT (P < 0.05). Higher PARP1 expression was present in 27 tumor compared the corresponding normal tissues using the GTEx and TCGA databases and it had a worse OS in several tumors (P < 0.05). Further, high PARP1 expression was significantly associated with six immune cells (B cells, CD4+ T cells, CD8+ T cells, macrophages, neutrophils, and dendritic cells) in most tumors, including colon adenocarcinoma (COAD), head and neck squamous cell carcinoma (HNSC), kidney renal clear cell carcinoma (KIRC), and liver hepatocellular carcinoma (LIHC) (P < 0.05). In particular, CD8+T cell infiltration, was also positively correlated with high PARP1 expression in bladder urothelial carcinoma (BLCA), breast invasive carcinoma (BRCA), kidney renal papillary cell carcinoma (KIRP), brain lower grade glioma (LGG), LIHC, pancreatic adenocarcinoma (PAAD), pheochromocytoma and paraganglioma (PCPG), prostate adenocarcinoma (PRAD), rectum adenocarcinoma (READ), testicular germ cell tumors (TGCT), thymoma (THYM), uterine corpus endometrial carcinoma (UCEC), uveal melanoma (UVM) (P < 0.05, no data shown), and PARP1 expression was significantly positively correlated with the transcription levels of some of the 47 immune checkpoint genes, such as CD274, CTLA4, and PDCD1 in several tumors, including PAAD, LIHC, KIRC, HNSC, and BLCA (P < 0.05). A significant positive association between PARP1 expression and the number of immune neoantigen was found within COAD, KIRC, lung adenocarcinoma (LUAD), PAAD and THYM (P < 0.05), and there were also significantly positive correlations between PARP1 expression and TMB in many tumors like adrenocortical carcinoma (ACC), COAD, kidney chromophobe (KICH), LGG, LUAD, READ, skin cutaneous melanoma (SKCM) and stomach adenocarcinoma (STAD) (P < 0.05). In addition, high PARP1 expression was positively associated with microsatellite instability event in COAD, KIRP, BRCA, glioblastoma multiforme (GBM), lung squamous cell carcinoma (LUSC), LGG, READ, UCEC, SKCM and LUAD (P < 0.05). Conclusions: Our results highlight the significance of PARP1 alterations as pan-cancer predictive biomarkers for ICI treatment, and its expression levels seem to be correlated with the status of immunotherapy-associated signatures, thus may be a promising biomarker for predicting ICI response in several tumors.

The degree of microsatellite instability predicts response to PD-1 blockade immunotherapy in mismatch repair-deficient/microsatellite instability-high colorectal cancers.

The development of programmed cell death-1 inhibitor (PD-1) has shed light on the treatment of tumors with deficiencies in DNA mismatch repair system or microsatellite instability (dMMR/MSI). However, predicting the subset in this group that will benefit from PD-1 blockade remains a challenge. In this study, we aimed to investigate the relationship between the degree of microsatellite instability and the responses to anti-PD-1 immunotherapy. 33 patients with colorectal adenocarcinoma who had a known MSI status and received anti-PD-1 immunotherapy were included. PCR results for MSI of the whole cohort were collected and treatment response was evaluated. Our data indicated that objective response rate (ORR) in instability-high group (instability loci ≥ 3) was significantly higher than ORR in instability-intermediate group (13/16 versus 6/17, P = 0.008). Besides, patients in instability-high group had significant longer progression-free survival (log-rank test, P = 0.004), and a significant increase in T lymphocyte infiltration and cytolytic activity in tumors. Future study might implement the intensity of microsatellite instability for more delicate selection for anti-PD-1 therapy in patient with dMMR/MSI-H tumors.

Neoadjuvant programmed death-1 blockade plus chemotherapy in locally advanced esophageal squamous cell carcinoma.

BACKGROUND: Immunotherapy is effective in treating unresectable esophageal squamous cell carcinoma (ESCC), but little is known about its role in the preoperative setting. The aim of this study was to evaluate the safety, feasibility and efficacy of neoadjuvant treatment with camrelizumab plus chemotherapy in locally advanced ESCC. METHODS: Patients diagnosed with locally advanced ESCC were retrospectively included if they had received neoadjuvant camrelizumab plus nab-paclitaxel and S1 capsule followed by radical esophagectomy between November, 2019 and June, 2020 at Sun Yat-sen University Cancer Center. Primary endpoints were safety and feasibility. In addition, pathological response and the relationship between tumor immune microenvironment (TIME)/tumor mutational burden (TMB) and treatment response were also investigated. RESULTS: Twelve patients were included and they all received three courses of preoperative treatment with camrelizumab plus nab-paclitaxel/S1. No grade 3 or higher toxicities occurred. No surgical delay or perioperative death was reported. Nine patients (75%) responded to the treatment, four with a complete pathological response (pCR) and five with a major pathological response (MPR). Neither programmed death-ligand 1 (PD-L1) expression nor TMB was correlated with treatment response. TIME analysis revealed that a higher abundance of CD56dim natural killer cells was associated with better pathological response in the primary tumor, while lower density of M2-tumor-associated macrophages was associated with better pathological response in the lymph nodes (LNs). CONCLUSIONS: Neoadjuvant camrelizumab plus nab-paclitaxel and S1 is safe, feasible and effective in locally advanced ESCC and is worth further investigation.