Construction and drug screening of Co-culture system using extrahepatic cholangiocarcinoma organoids and tumor-associated macrophages.

Patient-derived organoids (PDOs) have been proposed as a novel in vitro tumor model that can be applied to tumor research and drug screening. However, current tumor organoid models lack components of the tumor microenvironment, particularly tumor-associated macrophages(TAMs).We collected peripheral blood and tumor samples from 6 patients with extrahepatic cholangiocarcinoma(eCCA). Monocytes were induced into TAMs by cytokine and conditioned medium, and then co-cultured with tumor organoids. Our comprehensive analysis and comparison of histopathology and genomics results confirmed that this co-culture model can better capture intra- and inter-tumor heterogeneity retain the specific mutations of the original tumor. Drug sensitivity data in vitro revealed that gemcitabine and cisplatin are effective drugs for cholangiocarcinoma, but TAMs in the tumor microenvironment promote organoids growth and chemotherapy resistance. In conclusion, our organoid model of cholangiocarcinoma co-cultured with TAMs can not only shorten the model construction cycle, but also preserve the heterogeneity of original tumors to improve the accuracy of drug screening, and can also be applied to the researches of TAMs and tumors.

Biological impact and therapeutic potential of a novel camptothecin derivative (FLQY2) in pancreatic cancer through inactivation of the PDK1/AKT/mTOR pathway.

BACKGROUND: Camptothecin (CPT), a pentacyclic alkaloid with antitumor properties, is derived from the Camptotheca acuminata. Topotecan and irinotecan (CPT derivatives) were first approved by the Food and Drug Administration for cancer treatment over 25 years ago and remain key anticancer drugs today. However, their use is often limited by clinical toxicity. Despite extensive development efforts, many of these derivatives have not succeeded clinically, particularly in their effectiveness against pancreatic cancer which remains modest. AIM OF THE STUDY: This study aimed to evaluate the therapeutic activity of FLQY2, a CPT derivative synthesized in our laboratory, against pancreatic cancer, comparing its efficacy and mechanism of action with those of established clinical drugs. METHODS: The cytotoxic effects of FLQY2 on cancer cells were assessed using an MTT assay. Patient-derived organoid (PDO) models were employed to compare the sensitivity of FLQY2 to existing clinical drugs across various cancers. The impact of FLQY2 on apoptosis and cell cycle arrest in Mia Paca-2 pancreatic cancer cells was examined through flow cytometry. Transcriptomic and proteomic analyses were conducted to explore the underlying mechanisms of FLQY2's antitumor activity. Western blotting was used to determine the levels of proteins regulated by FLQY2. Additionally, the antitumor efficacy of FLQY2 in vivo was evaluated in a pancreatic cancer xenograft model. RESULTS: FLQY2 demonstrated (1) potent cytotoxicity; (2) superior tumor-suppressive activity in PDO models compared to current clinical drugs such as gemcitabine, 5-fluorouracil, cisplatin, paclitaxel, ivosidenib, infinitinib, and lenvatinib; (3) significantly greater tumor inhibition than paclitaxel liposomes in a pancreatic cancer xenograft model; (4) robust antitumor effects, closely associated with the inhibition of the TOP I and PDK1/AKT/mTOR signaling pathways. In vitro studies revealed that FLQY2 inhibited cell proliferation, colony formation, induced apoptosis, and caused cell cycle arrest at nanomolar concentrations. Furthermore, the combination of FLQY2 and gemcitabine exhibited significant inhibitory and synergistic effects. CONCLUSION: The study confirmed the involvement of topoisomerase I and the PDK1/AKT/mTOR pathways in mediating the antitumor activity of FLQY2 in treating Mia Paca-2 pancreatic cancer. Therefore, FLQY2 has potential as a novel therapeutic option for patients with pancreatic cancer.

Analysis of a new therapeutic target and construction of a prognostic model for breast cancer based on ferroptosis genes.

Breast cancer, which is the most common malignant tumor among women worldwide and an important cause of death in women. The existing prognostic model for patients with breast cancer is not accurate as breast cancer is resistant to commonly used antitumor drugs. Ferroptosis is a novel mechanism of programmed cell death that depends on iron accumulation and lipid peroxidation. Various studies have confirmed the role of ferroptosis in tumor regulation and ferroptosis is now considered to play an important role in breast cancer development. At present, the association between breast cancer prognosis and ferroptosis-related gene expression remains unclear. Further exploration of this research area may optimize the evaluation and prediction of prognosis of patients with breast cancer and finding of new therapeutic targets. In this study, clinical factors and the expression of multiple genes were evaluated in breast cancer samples from the Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database database. Eleven prognostication-related genes (TP63, IFNG, MT3, ANO6, FLT3, PTGS2, SLC1A4, JUN, SLC7A5, CHAC1, and TF) were identified from differentially expressed genes to construct a survival prediction model, which showed a good prediction ability. KEGG pathway analysis revealed that immune-related pathways were the primary pathways. ssGSEA analysis showed significant differences in the distribution of certain immune-related cell subsets, such as CD8+T cells and B cells, and in the expression of multiple immune genes, including type II IFN response and APC coinhibition. In addition, 10 immune targets related to ferroptosis in breast cancer were found: CD276, CD80, HHLA2, LILRA2, NCR3LG1, NECTIN3, PVR, SLAMF9,TNFSF4, and BTN1A1. Using TCGA, new ferroptosis genes related to breast cancer prognosis were identified, a new reliable and accurate prognosis model was developed, and 10 new potential therapeutic targets different from the traditional targeted drugs were identified to provide a reference for improving the poor prognosis of patients with breast cancer.

Effectiveness of no drainage after elective day-case laparoscopic cholecystectomy, even with intraoperative gallbladder perforation: a randomized controlled trial.

BACKGROUND: Laparoscopic cholecystectomy (LC) has been carried out as day-case surgery. Current guidelines do not mention the role of drainage after LC. In particular, data stay blank with no prospective study on drainage management when gallbladder perforation (GP) accidentally occurs intraoperatively. METHODS: A randomized controlled trial was conducted to compare clinical outcomes of drainage and no drainage after elective day-case LC. Intraoperative GP was recorded. The primary and secondary outcomes were major and minor complications, respectively. RESULTS: Two hundred patients were randomized. No major complications occurred in either group. In secondary outcomes, nausea/vomiting, pain, hospital stay, and cost were similar in the drainage group and no drainage group; postoperative fever, WBC, and CRP levels were significantly lower in the no drainage group. GP occurred in 32 patients. Male patients with higher BMI and CRP and abdominal pain within 1 month were more likely to occur GP. Subgroup analysis of GP, primary outcomes, and most secondary outcomes had no difference. Postoperative WBC and CRP were higher in the drainage group. Postoperative fever occurred in 63 patients. Univariate analysis of fever showed that blood loss, drainage, postoperative WBC, CRP, and hospital stay were significant. Multivariable logistic regression analysis demonstrated that drainage was an independent risk factor for fever after LC (OR 3.418, 95% CI 1.392-8.390; p = 0.007). CONCLUSIONS: No drainage after elective day-case LC is safe and associated with fewer complications, even in intraoperative GP. The trial proves that drainage is an independent risk factor for postoperative fever. The use of a drain after LC may lead to an unsuccessful day-case procedure by causing fever, elevated CRP, and extended hospital stay (NCT03909360).

Is it justified to assess the resectability of pancreatic cancer combined with biological and conditional factors?

Background: This study aimed to investigate the biological and conditional resectability criteria for pancreatic ductal adenocarcinoma (PDAC), as proposed by the International Association of Pancreatology (IAP), as well as to identify the role of biological and conditional factors in assessing the resectability of PDAC. Methods: The clinical data of PDAC patients who underwent upfront open/laparoscopic pancreaticoduodenectomy (PD/LPD) or distal pancreatectomy (DP/LDP) at our hospital between January 2013 to June 2019 were retrospectively analyzed. Patients who were diagnosed with anatomically resectable PDAC, as defined by National Comprehensive Cancer Network (NCCN) guideline of PDAC guideline Version 1.2020, were enrolled. Based on IAP-criteria, these patients were divided into two groups, including IAP-resectable (IAP-R) and IAP Borderline Resectable (IAP-BR). Clinical characteristics and outcomes were compared between the two groups. In order to identify independent biological and conditional predictors of recurrence-free survival (RFS) and overall survival (OS) of enrolled patients, an analysis was performed through the use of a Cox proportional-hazard model. Results: Overall, 97 patients were included in this study. Among them, 38 patients were IAP-R and 59 patients were IAP-BR. Compared to the IAP-R group, the IAP-BR group had a higher early recurrence rate (62.7% vs. 42.1%; P=0.047), and the median RFS (9.2 vs. 18.3 months, P<0.01) and OS (19.1 vs. 30.6 months, P<0.05) were also significantly worse. Preoperative CA19-9 serum levels that exceeded 500 U/mL and/or an imaging diagnosis of regional lymph nodes metastasis were independently associated with OS and RFS of anatomically resectable PDAC. Conclusions: The prognosis of patients with PDAC that undergo resection can be predicted more accurately by assessing the resectability of pancreatic cancer combined with anatomical and biological factors according to IAP criteria. Whether conditional factors should be included in the resectability criteria needs to be validated by prospective and large cohorts.

Solitary fibrous tumor of the liver: A case report and review of the literature.

BACKGROUND: Hepatic solitary fibrous tumor (SFT) is a rare neoplasm. Up to now, only 90 cases have been reported in the English language literature. This report describes a case of SFT of the liver misdiagnosed as hepatocellular carcinoma. CASE SUMMARY: A 42-year-old male had a two-year history of a gradually enlarging intrahepatic nodule. The preoperative imaging revealed a mass with a size of 2.7 cm × 2.3 cm located in the segment IV of the liver. The patient was subjected to the resection of the segment IV, such as the medial segment of the left lobe of the liver. The histological examination of the mass showed various spindled cells irregularly arranged in the stroma. The immunohistochemistry of this mass revealed a positive staining for CD34 and STAT6. The history of intracranial tumor and postoperative pathological results led to the diagnosis of SFT of the liver (SFTL) due to a metastasis from the brain. CONCLUSION: SFTL is an uncommon mesenchymal neoplasm that can be easily overlooked or misdiagnosed. The best treatment choice is the complete surgical resection of the mass. A regular follow-up after the surgery should be performed due to the poor prognosis of metastatic or recurrent SFT.

Lactate promotes the growth of patient-derived organoids from hepatopancreatobiliary cancers via ENO1/HIF1α pathway and does not affect their drug sensitivities.

The long culture duration of patient-derived organoids (PDOs) have severely limited their clinical applications. The aim of this study was to determine the effect of lactate supplementation on the growth, genetic profiles and drug sensitivities of PDOs from hepatopancreatobiliary tumors. LM3, Huh7, Panc02, and RBE cell lines were cultured as organoids in the presence or absence of lactate, and total protein was extracted to measure the expression of α-enolase (ENO1), hypoxia-inducible factor-1α (HIF1α), AKT, and PI3 kinase (PI3K). Thirteen hepatopancreatobiliary tumor specimens were collected during surgical resection and cultured as PDOs with or without L-lactate. Hematoxylin and eosin (H&E) staining and immunohistochemical staining were performed on the original tissues and PDOs to compare their pathological structures, and their genetic profiles were analyzed by whole-exome sequencing (WES). The sensitivity of the PDOs to gemcitabine, 5-fluorouracil, cisplatin, paclitaxel, ivosidenib, infigratinib, and lenvatinib were evaluated in terms of cell viability. Peripheral blood mononuclear cells (PBMCs) were isolated and co-cultured with PDOs to test the sensitivity of PDOs to tislelizumab. The addition of 20 mM lactate significantly promoted the growth of LM3 and Huh 7 organoids by 217% and 36%, respectively, compared to the control group, and the inhibition of lactate transporter decreased their growth. The HIF1α/ENO1/AKT/PI3K pathway was also activated by lactate. The inhibition of enolase also partly decreased the growth of organoids treated with lactate. Furthermore, 20 mM lactate increased the viability of 9 PDOs from 135% to 317% without affecting their pathological features. The genetic similarity, in terms of single nucleotide variations, insertions, and deletions, between original tissues and lactate-treated PDOs ranged from 83.2% to 94.1%, and that between the untreated and lactate-treated PDOs was at least 93.2%. Furthermore, the addition of lactate did not significantly change the dose-response curves of the PDOs to chemotherapeutic drugs, targeted drugs, and immune checkpoint inhibitor, especially for the drugs to which the cells were sensitive. Thus, lactate can be added to the culture medium of PDOs to promote their growth without altering their genetic profiles and drug sensitivities.

Genomic and clinical features of endoplasmic reticulum stress factor in digestive system pan-cancer studies.

Introduction: Digestive system pan-cancer is one of the lethal malignant tumors, which have the propensity for poor prognosis and difficult treatment. Endoplasmic reticulum (ER) stress has served as a pivotal role in the progression of the tumor, while the implication of ER stress on digestive system pan-cancers still needs elucidation, especially from the perspective of clinical outcome and that of genomic features. Methods: First, Among the ER STRESS factors from the REACTOME_UNFOLDED_PROTEIN_RESPONSE_UPR (113 genes) and HALLMARK_UNFOLDED_PROTEIN_RESPONSE (92 genes) terms, 153 ER STRESS regulators were identified after removing replicates. The somatic mutation data and copy number variation data of gastrointestinal pan-cancer were downloaded from The Cancer Genome Atlas (TCGA) database. Then, we explored the clinical outcome and genetic mutation of ER stress-related differentially expressed genes (DEGs) by multiple bioinformatics analysis. Subsequently, we analyzed the Spearman correlation between the drug sensitivity of 179 gastrointestinal anticancer drugs and the transcriptional expression of 153 ER stress factors in 769 cancer cell lines of the GDSC2 cohort. Next, ssGSEA method was used to quantify the immune cell infiltration scores in the tumor microenvironment, and Spearman correlation was used to calculate the correlation between ER stress scores and immune cell infiltration. Finally, we analyzed the cellular origin of ER stress factor dysregulation. Results: We analyzed the genomic changes and clinical outcomes of ER stress factors in different tumors of gastrointestinal pan-cancer. Endoplasmic reticulum stress factor (ER) in digestive tract tumors showed high SNV mutation frequency, less methylation dysregulation and was associated with multiple oncogenic pathways. Endoplasmic reticulum stress factor (ER) is a risk factor for many cancers, but the effect on overall survival in rectal adenocarcinoma is opposite to that in other gastrointestinal tumors. And ER stress factors are highly correlated with drugs that target important pathways. Discussion: Based on the clinical prognosis and genomic analysis of ER stress-related factors in patients with gastrointestinal pan-cancer, this study provides a new direction for further research on gastrointestinal pan-cancer.

Treatment of microvascular invasion in hepatocellular carcinoma with drug-loaded nanocomposite platform under synergistic effect of magnetic field/near-infrared light.

Despite progress in clinical treatment, microvascular invasion (MVI) remains a major factor for frequent recurrence and metastasis of hepatocellular carcinoma (HCC) after liver resection and surgery. Thus, this study constructed a target nanoplatform (αCD97-USPIO-Au-DDP) with magnetic field/near-infrared (NIR) light response using ultrasmall superparamagnetic iron oxide-gold nanoporous spheres (USPIO-Au) as multifunctional nanocarrier. Anticancer drug cisplatin (DDP) was loaded, and specifically expressed CD97 protein in MVI was taken as the therapeutic target. The αCD97-USPIO-Au-DDP showed favorable photothermal and stable properties under the NIR light at 808 nm wavelength. As suggested by in vitro and in vivo research, this composite nanopreparation could effectively reduce damage to normal organs and showed good biocompatibility. Excellent magnetic targeting function of nanocarrier and modification of αCD97 strengthened accumulation of composite nanodrug in tumor to inhibit tumor growth. This system may have important ramifications for treatment of MVI in HCC.

Establishment and drug screening of patient-derived extrahepatic biliary tract carcinoma organoids.

BACKGROUND: Patient-derived organoids (PDO) have been proposed as a novel in vitro method of drug screening for different types of cancer. However, to date, extrahepatic biliary tract carcinoma (eBTC) PDOs have not yet been fully established. METHODS: We collected six samples of gallbladder carcinoma (GBC) and one sample of extrahepatic cholangiocarcinoma (eCCA) from seven patients to attempt to establish eBTC PDOs for drug screening. We successfully established five GBC and one eCCA PDOs. Histological staining was used to compare structural features between the original tissues and cancer PDOs. Whole exome sequencing (WES) was performed to analyze the genetic profiles of original tissues and cancer PDOs. Drug screening, including gemcitabine, 5-fluorouracil, cisplatin, paclitaxel, infigratinib, and ivosidenib, was measured and verified by clinical effects in certain cases. RESULTS: Different PDOs exhibited diverse growth rates during in vitro culture. Hematoxylin and eosin staining demonstrated that the structures of most cancer PDOs retained the original structures of adenocarcinoma. Immunohistological and periodic acid-schiff staining revealed that marker expression in cancer PDOs was similar to that of the original specimens. Genetic profiles from the four original specimens, as well as paired cancer PDOs, were analyzed using whole exome sequencing. Three of the four PDOs exhibited a high degree of similarity when compared to the original specimens, except for GBC2 PDO, which only had a concordance of 74% in the proportion of single nucleotide polymorphisms in the coding sequence. In general, gemcitabine was found to be the most efficient drug for eBTC treatment, as it showed moderate or significant inhibitory impact on cancer growth. Results from drug screening were confirmed to a certain extent by three clinical cases. CONCLUSIONS: Our study successfully established a series of eBTC PDOs, which contributed to the field of eBTC PDOs. Additional enhancements should be explored to improve the growth rate of PDOs and to preserve their immune microenvironment.