Polyglycerol-Based Biomedical Matrix for Immunomagnetic Circulating Tumor Cell Isolation and Their Expansion into Tumor Spheroids for Drug Screening.

Circulating tumor cells (CTCs) are established as distinct cancer biomarkers for diagnosis, as preclinical models, and therapeutic targets. Their use as preclinical models is limited owing to low purity after isolation and the lack of effective techniques to create 3D cultures that accurately mimic in vivo conditions. Herein, a two-component system for detecting, isolating, and expanding CTCs to generate multicellular tumor spheroids that mimic the physiology and microenvironment of the diseased organ is proposed. First, an antifouling biointerface on magnetic beads is fabricated by adding a bioinert polymer layer and conjugation of biospecific ligands to isolate cancer cells, dramatically enhancing the selectivity and purity of the isolated cancer cells. Next, the isolated cells are encapsulated into self-degradable hydrogels synthesized using a thiol-click approach. The hydrogels are mechanochemically tuned to enable tumor spheroid growth to a size greater than 300 µm and to further release the grown spheroids while retaining their tumor-like characteristics. In addition, drug treatment highlights the need for 3D culture environments rather than conventional 2D culture. The designed biomedical matrix shows potential as a universal method to ensure mimicry of in vivo tumor characteristics in individual patients and to improve the predictability of preclinical screening of personalized therapeutics.

In vivo flow cytometry reveals an anti-metastatic effect of Rujifang in triple-negative breast cancer.

Breast cancer is the most common cancer, and triple-negative breast cancer (TNBC) has the highest metastasis and mortality rate among all breast cancer subtypes. Rujifang is a traditional Chinese medicine formula with many years of clinical application in breast cancer treatment. Here, we aim to investigate the effects of Rujifang on circulating tumor cell (CTC) dynamics and the tumor microenvironment in a ZsGreen/luciferase double-labeled TNBC orthotopic model. We report that the number of CTCs monitored by in vivo flow cytometry (IVFC) strongly correlates with disease progression. Rujifang treatment decreased the number of CTCs and suppressed the distant metastasis of TNBC. Moreover, immunofluorescence analysis revealed that Rujifang treatment could affect the tumor microenvironment by downregulating Kindlin-1, which has been reported to promote metastasis of TNBC. Our study provides evidence of the anti-metastatic effect of Rujifang against TNBC in an animal model using fluorescent cell lines. The results suggest the potential therapeutic value of Rujifang as an anti-metastatic drug, however, further clinical trials are needed to validate these findings in humans.

A new time dimension in the fight against metastasis.

Despite advances in uncovering vulnerabilities, identifying biomarkers, and developing more efficient treatments, cancer remains a threat because of its ability to progress while acquiring resistance to therapy. The circadian rhythm governs most of the cellular functions implicated in cancer progression, and its exploitation therefore opens new promising directions in the fight against metastasis. In this review we summarize the role of the circadian rhythm in tumor development and progression, with emphasis on the circadian rhythm-regulated elements that control the generation of circulating tumor cells (CTCs) and metastasis. We then present data on chronotherapy and discuss how circadian rhythm investigations may open new paths to more effective anticancer treatments.

Specific Aptamer Functionalization of Dense Ceramic by Click Chemistry Towards Circulating Tumor Cells Apheresis.

BACKGROUND/AIM: Circulating tumor cells (CTC) are tumor cells which can be disseminated at distance of the primary tumor and form metastatic niche. Moreover, their quantity is an important parameter which can induce cluster metastasis. A solution, can be the creation of a system that allow the capture and elimination from the blood of patients by using the medical device developed which is an inert bioceramic functionalized by aptamer target to CTC. MATERIALS AND METHODS: Herein we develop chemical reactions to bind a modified MUC1 specific DNA aptamer on an alumina (Al2O3) dense ceramic surface. In fact, MUC1 biomarker is very present on the surface of tumor cells. RESULTS: The specific developed chemical reactions led to the covalent binding of the aptamer while preserving its biological characteristics. CONCLUSION: This functionalization of dense alumina would allow the potential capture of circulating tumor cells.

Manipulation and elimination of circulating tumor cells using multi-responsive nanosheet for malignant tumor therapy.

Tumor recurrence caused by metastasis is a major cause of death for patients. Thus, a strategy to manipulate the circulating tumor cells (CTCs, initiators of tumor metastasis ) and eliminate them along with the primary tumor has significant clinical significance for malignant tumor therapy. In this study, a magnet-NIR-pH multi-responsive nanosheet (Fe3O4@SiO2-GO-PEG-FA/AMP-DOX, FGPFAD) was fabricated to capture CTCs in circulation, then magnetically transport them to the primary tumor, and finally perform NIR-dependent photothermal therapy as well as acidic-environment-triggered chemotherapy to destroy both the CTCs and the primary tumor. The FGPFAD nanosheet consists of silica-coated ferroferric oxide nanoparticles (Fe3O4@SiO2, magnetic targeting agent), graphene oxide (GO, photothermal therapy agent), polyethylene glycol (PEG, antifouling agent for sustained circulation), folic acid (FA, capturer of CTCs) and antimicrobial-peptide-conjugated doxorubicin (AMP-DOX, agent for chemotherapy), in which the AMP-DOX was bound to the FGPFAD nanosheet via a cleavable Schiff base to achieve acidic-environment-triggered drug release for tumor-specific chemotherapy. Both in vitro and in vivo results indicated that the effective capture and magnetically guided transfer of CTCs to the primary tumor, as well as the multimodal tumor extermination performed by our FGPFAD nanosheet, significantly inhibited the primary tumor and its metastasis.

The Regulators of Peroxisomal Acyl-Carnitine Shuttle CROT and CRAT Promote Metastasis in Melanoma.

Circulating tumor cells are the key link between a primary tumor and distant metastases, but once in the bloodstream, loss of adhesion induces cell death. To identify the mechanisms relevant for melanoma circulating tumor cell survival, we performed RNA sequencing and discovered that detached melanoma cells and isolated melanoma circulating tumor cells rewire lipid metabolism by upregulating fatty acid (FA) transport and FA beta-oxidation‒related genes. In patients with melanoma, high expression of FA transporters and FA beta-oxidation enzymes significantly correlates with reduced progression-free and overall survival. Among the highest expressed regulators in melanoma circulating tumor cells were the carnitine transferases carnitine O-octanoyltransferase and carnitine acetyltransferase, which control the shuttle of peroxisome-derived medium-chain FAs toward mitochondria to fuel mitochondrial FA beta-oxidation. Knockdown of carnitine O-octanoyltransferase or carnitine acetyltransferase and short-term treatment with peroxisomal or mitochondrial FA beta-oxidation inhibitors thioridazine or ranolazine suppressed melanoma metastasis in mice. Carnitine O-octanoyltransferase and carnitine acetyltransferase depletion could be rescued by medium-chain FA supplementation, indicating that the peroxisomal supply of FAs is crucial for the survival of nonadherent melanoma cells. Our study identifies targeting the FA-based cross-talk between peroxisomes and mitochondria as a potential therapeutic opportunity to challenge melanoma progression. Moreover, the discovery of the antimetastatic activity of the Food and Drug Administration‒approved drug ranolazine carries translational potential.

Drug susceptibility testing of circulating lung cancer cells for personalized treatment.

The presence of Circulating tumor cells (CTCs) has been proven to be correlated with disease progression and the patient's response to treatment. However, the culture of CTCs for clinical utility is still a big challenge. We have developed a short-term method that enables CTCs culture and provides an opportunity to monitor drug susceptibility testing in individual patients. In a proof-of-concept study, we established a unique method using Matrigel® coated in 96 well plate to enable cancer cell clusters to attach and proliferate. The culture method using Matrigel® provides in vitro conditions and improves the attachment and differentiation of anchorage-dependent epithelial cells proliferation and mimics the tumor microenvironment. We further treated the cells attached to Matrigel® with the same drug regimen as the patient has undergone. Around 30.7% of the CTCs were viable after the drug treatment. We also correlated the decrease in cell viability after drug treatment with the reduction in the pleural effusion of the patient as seen by the images obtained from CT scans pre-and post-treatment. Moreover, as per the RECIST criterion, the patient had exhibited a positive response to the treatment. The short-term culturing of CTC along with the drug susceptibility testing offers a novel method to predict patient response to the treatment and could be utilized for screening suitable drug combinations for personalized treatment.

Circulating tumor cells detected in follow-up predict survival outcomes in tri-modality management of advanced non-metastatic esophageal cancer: a secondary analysis of the QUINTETT randomized trial.

BACKGROUND: Our aim was to establish if presence of circulating tumor cells (CTCs) predicted worse outcome in patients with non-metastatic esophageal cancer undergoing tri-modality therapy. METHODS: We prospectively collected CTC data from patients with operable non-metastatic esophageal cancer from April 2009 to November 2016 enrolled in our QUINTETT esophageal cancer randomized trial (NCT00907543). Patients were randomized to receive either neoadjuvant cisplatin and 5-fluorouracil (5-FU) plus radiotherapy followed by surgical resection (Neoadjuvant) or adjuvant cisplatin, 5-FU, and epirubicin chemotherapy with concurrent extended volume radiotherapy following surgical resection (Adjuvant). CTCs were identified with the CellSearch® system before the initiation of any treatment (surgery or chemoradiotherapy) as well as at 6-, 12-, and 24-months post-treatment. The threshold for CTC positivity was one and the findings were correlated with patient prognosis. RESULTS: CTC data were available for 74 of 96 patients and identified in 27 patients (36.5%) at a median follow-up of 13.1months (interquartile range:6.8-24.1 months). Detection of CTCs at any follow-up visit was significantly predictive of worse disease-free survival (DFS;hazard ratio [HR]: 2.44; 95% confidence interval [CI]: 1.41-4.24; p=0.002), regional control (HR: 6.18; 95% CI: 1.18-32.35; p=0.031), distant control (HR: 2.93; 95% CI: 1.52-5.65;p=0.001) and overall survival (OS;HR: 2.02; 95% CI: 1.16-3.51; p=0.013). After adjusting for receiving neoadjuvant vs. adjuvant chemoradiotherapy, the presence of CTCs at any follow-up visit remained significantly predictive of worse OS ([HR]:2.02;95% [Cl]:1.16-3.51; p=0.013) and DFS (HR: 2.49;95% Cl: 1.43-4.33; p=0.001). Similarly, any observed increase in CTCs was significantly predictive of worse OS (HR: 3.14; 95% CI: 1.56-6.34; p=0.001) and DFS (HR: 3.34; 95% CI: 1.67-6.69; p<0.001). CONCLUSION: The presence of CTCs in patients during follow-up after tri-modality therapy was associated with significantly poorer DFS and OS regardless of timing of chemoradiotherapy.

The role of folate receptor-positive circulating tumor cell analysis in the diagnosis of colorectal cancer: a retrospective cohort study.

OBJECTIVE: To explore the application value of folate receptor-positive circulating tumor cell analysis (FR+-CTC analysis) in the diagnosis of colorectal cancer (CRC). METHODS: Clinical data of CRC patients and healthy subjects admitted to our hospital from January 2019 to October 2019 were retrospectively collected. CTC result and serological and pathological outcomes of the study patients were collected and analyzed. Receiver operating characteristic curve (ROC curve) was drawn. RESULTS: The CTC levels of cancer patients (9.34 ± 3.53 FU/3 ml) were significantly higher than those of healthy subjects (7.00 ± 2.33 FU/3 ml). CTC levels could be related to cancer stage and metastasis in patients. ROC curves were drawn and the area under the ROC curve (AUC) was 0.702. The cutoff value was determined to be 8.87 FU/3 ml. At this cutoff value, the sensitivity and specificity of FR+-CTC analysis in the diagnosis of colorectal cancer were 61.8% and 82.6%, respectively. The diagnostic efficiency of FR+-CTCs in advanced CRC was significantly higher than that in the early stage. And the cutoff value of early and advanced stage CRC was determined to be 9.66 FU/3 ml. CONCLUSION: FR+-CTC analysis has high potential in recurrence diagnosis and decision of adjuvant chemotherapy for CRC.

Jinfukang inhibits lung cancer metastasis by upregulating CX3CL1 to recruit NK cells to kill CTCs.

ETHNOPHARMACOLOGICAL RELEVANCE: Circulating tumor cells (CTCs) play an important role in tumor metastasis and may be a target for metastasis prevention. The traditional Chinese medicine Jinfukang functions to improve immunity, prevent metastasis, and prolong lung cancer patient survival periods. Yet, whether Jinfukang prevents metastasis by regulating immune cells to clearance CTCs is still unknown. AIM OF THE STUDY: To explore the anti-metastasis mechanism of Jinfukang from the perspective of regulating NK cells to clear CTCs. MATERIALS AND METHODS: CTC-TJH-01 cell was treated with Jinfukang. Cytokine chip was used to detect cytokines in cell culture supernatant. Lymphocyte recruitment assay was detected by Transwell and flow cytometry. Protein expression was analysis by Western blot. LDH kit was used to detect cytotoxicity. NOD-SCID mice used for tail vein injection to study lung metastasis. RESULTS: Jinfukang could promote the expression and secretion of the chemokine CX3CL1 by CTCs. In addition, Jinfukang could promote the recruitment of natural killer (NK) cells by CTCs and significantly increase the cytotoxic effect of NK cells on CTCs. Moreover, Jinfukang could upregulate the expression of FasL and promote the secretion of TNF-α by NK cells and that NK cells could induce the apoptosis of CTCs through the Fas/FasL signaling pathway. Finally, we confirmed that Jinfukang could promote NK cells to kill CTCs and then inhibit lung cancer metastasis in vivo. The above effects of Jinfukang could be partially reversed by an anti-CX3CL1 mAb. CONCLUSIONS: These results suggest that Jinfukang may prevent lung cancer metastasis by enhancing the clearance of CTCs in the peripheral blood by NK cells, providing evidence for the anti-metastasis effect of Jinfukang.

Multiblock Discriminant Analysis of Integrative 18F-FDG-PET/CT Radiomics for Predicting Circulating Tumor Cells in Early-Stage Non-small Cell Lung Cancer Treated With Stereotactic Body Radiation Therapy.

PURPOSE: The main objective of the present study was to integrate 18F-FDG-PET/CT radiomics with multiblock discriminant analysis for predicting circulating tumor cells (CTCs) in early-stage non-small cell lung cancer (ES-NSCLC) treated with stereotactic body radiation therapy (SBRT). METHODS: Fifty-six patients with stage I NSCLC treated with SBRT underwent 18F-FDG-PET/CT imaging pre-SBRT and post-SBRT (median, 5 months; range, 3-10 months). CTCs were assessed via a telomerase-based assay before and within 3 months after SBRT and dichotomized at 5 and 1.3 CTCs/mL. Pre-SBRT, post-SBRT, and delta PET/CT radiomics features (n = 1548 × 3/1562 × 3) were extracted from gross tumor volume. Seven feature blocks were constructed including clinical parameters (n = 12). Multiblock data integration was performed using block sparse partial least squares-discriminant analysis (sPLS-DA) referred to as Data Integration Analysis for Biomarker Discovery Using Latent Components (DIABLO) for identifying key signatures by maximizing common information between different feature blocks while discriminating CTC levels. Optimal input blocks were identified using a pairwise combination method. DIABLO performance for predicting pre-SBRT and post-SBRT CTCs was evaluated using combined AUC (area under the curve, averaged across different blocks) analysis with 20 × 5-fold cross-validation (CV) and compared with that of concatenation-based sPLS-DA that consisted of combining all features into 1 block. CV prediction scores between 1 class versus the other were compared using the Wilcoxon rank sum test. RESULTS: For predicting pre-SBRT CTCs, DIABLO achieved the best performance with combined pre-SBRT PET radiomics and clinical feature blocks, showing CV AUC of 0.875 (P = .009). For predicting post-SBRT CTCs, DIABLO achieved the best performance with combined post-SBRT CT and delta CT radiomics feature blocks, showing CV AUCs of 0.883 (P = .001). In contrast, all single-block sPLS-DA models could not attain CV AUCs higher than 0.7. CONCLUSIONS: Multiblock integration with discriminant analysis of 18F-FDG-PET/CT radiomics has the potential for predicting pre-SBRT and post-SBRT CTCs. Radiomics and CTC analysis may complement and together help guide the subsequent management of patients with ES-NSCLC.

Exosome-based liquid biopsies in cancer: opportunities and challenges.

Liquid biopsy in cancer has gained momentum in clinical research and is experiencing a boom for a variety of applications. There are significant efforts to utilize liquid biopsies in cancer for early detection and treatment stratification, as well as residual disease and recurrence monitoring. Although most efforts have used circulating tumor cells and circulating tumor DNA for this purpose, exosomes and other extracellular vesicles have emerged as a platform with potentially broader and complementary applications. Exosomes/extracellular vesicles are small vesicles released by cells, including cancer cells, into the surrounding biofluids. These exosomes contain tumor-derived materials such as DNA, RNA, protein, lipid, sugar structures, and metabolites. In addition, exosomes carry molecules on their surface that provides clues regarding their origin, making it possible to sort vesicle types and enrich signatures from tissue-specific origins. Exosomes are part of the intercellular communication system and cancer cells frequently use them as biological messengers to benefit their growth. Since exosomes are part of the disease process, they have become of tremendous interest in biomarker research. Exosomes are remarkably stable in biofluids, such as plasma and urine, and can be isolated for clinical evaluation even in the early stages of the disease. Exosome-based biomarkers have quickly become adopted in the clinical arena and the first exosome RNA-based prostate cancer test has already helped >50 000 patients in their decision process and is now included in the National Comprehensive Cancer Network guidelines for early prostate cancer detection. This review will discuss the advantages and challenges of exosome-based liquid biopsies for tumor biomarkers and clinical implementation in the context of circulating tumor DNA and circulating tumor cells.

Jinfukang regulates integrin/Src pathway and anoikis mediating circulating lung cancer cells migration.

ETHNOPHARMACOLOGICAL RELEVANCE: Metastasis is the main cause of death in lung cancer patients. Circulating tumor cells (CTCs) may be an important target of metastasis intervention. Previous studies have shown that Jinfukang could prevent the recurrence and metastasis of lung cancer, and we have established a circulating lung tumor cell line CTC-TJH-01. However, whether Jinfukang inhibition of lung cancer metastasis is related to CTCs is still unknown. AIM OF THE STUDY: To further explore the mechanism of Jinfukang in anti-metastasis of lung cancer from the perspective of intervention of CTCs. MATERIALS AND METHODS: CTC-TJH-01 and H1975 cells were treated with Jinfukang. Cell viability was detected by CCK8, and the cell apoptosis was detected by flow cytometry. Transwell was used to detected cell migration and invasion. Cell anoikis was detected by anoikis detection kit. Protein expression was analysis by Western blot. RESULTS: Jinfukang could inhibit the proliferation, migration and invasion of CTC-TJH-01 and H1975 cells. Besides, Jinfukang could also induce anoikis in CTC-TJH-01 and H1975 cells. Analysis of the mRNA expression profile showed ECM-receptor interaction and focal adhesion were regulated by Jinfukang. Moreover, it was also find that Jinfukang significantly inhibited integrin/Src pathway in CTC-TJH-01 and H1975 cells. When suppress the expression of integrin with ATN-161, it could promote Jinfukang to inhibit migration and induce anoikis in CTC-TJH-01 and H1975 cells. CONCLUSIONS: Our results indicate that the migration and invasion of CTCs are inhibited by Jinfukang, and the mechanism may involve the suppression of integrin/Src axis to induce anoikis. These data suggest that Jinfukang exerts anti-metastatic effects in lung cancer may through anoikis.

Effects of dietary patterns combined with dietary phytochemicals on breast cancer metastasis.

AIMS: Dietary phytochemicals and diet types (e.g., the Mediterranean diet) have been shown to have anti-cancer properties. However, the effects of combined treatment with dietary phytochemicals and different diet types on primary and metastatic tumor growth have yet to be investigated. The purpose of this study is to investigate the effects of phytochemicals combined with diet types on breast cancer metastasis. MAIN METHODS: The inhibitory effects on breast cancer metastasis of three phytochemicals (allicin, hesperidin, astragalus polysaccharides) and two diet types (Mediterranean diet, restricted diet), separately or in combination, were evaluated based on: (i) detection of circulating tumor cells (CTCs) using an in vivo capture method; and (ii) primary tumor growth. KEY FINDINGS: All dietary factors significantly inhibited the growth of primary tumors and metastases, with combinations showing enhancing the effects. SIGNIFICANCE: Dietary phytochemicals and diet types should be further evaluated as adjunct therapies and lifestyle modifications in cancer patients. Furthermore, the in vivo CTC capture method allows dynamic monitoring of cancer metastasis over time, providing a useful approach to evaluating treatment effects in real-time.

Pharmacological ascorbate inhibits pancreatic cancer metastases via a peroxide-mediated mechanism.

Pharmacological ascorbate (P-AscH-, high-dose, intravenous vitamin C) is cytotoxic to tumor cells in doses achievable in humans. Phase I studies in pancreatic cancer (PDAC) utilizing P-AscH- have demonstrated increases in progression free survival, suggesting a reduction in metastatic disease burden. The purpose of this study was to determine the effects of P-AscH- on metastatic PDAC. Several in vitro and in vivo mechanisms involved in PDAC metastases were investigated following treatment with P-AscH-. Serum from PDAC patients in clinical trials with P-AscH- were tested for the presence and quantity of circulating tumor cell-derived nucleases. P-AscH- inhibited invasion, basement membrane degradation, decreased matrix metalloproteinase expression, as well as clonogenic survival and viability during exposure to fluid shear stress. In vivo, P-AscH- significantly decreased formation of ascites, tumor burden over time, circulating tumor cells, and hepatic metastases. Both in vitro and in vivo findings were reversed with the addition of catalase suggesting that the effect of P-AscH- on metastatic disease is mediated by hydrogen peroxide. Finally, P-AscH- decreased CTC-derived nucleases in subjects with stage IV PDAC in a phase I clinical trial. We conclude that P-AscH- attenuates the metastatic potential of PDAC and may prove to be effective for treating advanced disease.

Gold nanoparticle-modified black phosphorus nanosheets with improved stability for detection of circulating tumor cells.

Gold nanoparticle (AuNP)-anchored BP nanosheets were synthesized through in situ growth of AuNPs onto BP. Due to the strong chelating ability of P or phosphorus oxides with AuNPs, the stability of BP is improved. As proof-of-concept demonstration of the functionalized BP, electrochemical detection of circulating tumor cells (CTCs) based on BP@AuNPs@aptamer as a probe combined with immunomagnetic separation is reported. The aptamer can specifically bind with CTCs, while the phosphorus oxides including phosphite ion and phosphate ion (PxOy species) on BP and aptamer can react with molybdate to generate an electrochemical current, leading to dual signal amplification. The biosensor is applied to MCF-7 cell detection and displays good analytical performance with a detection limit of 2 cell mL-1. Furthermore, the practicality of this biosensor was validated through sensitive determination of MCF-7 cells in human blood. Therefore, the reported biosensor could be applied to detect other biomarkers, offering an ultrasensitive strategy for clinical diagnostics. Graphical abstract Electrochemical detection of circulating tumor cells based on gold nanoparticle-modified black phosphorus nanosheets is reported.

Circulating tumor-cell-targeting Au-nanocage-mediated bimodal phototherapeutic properties enriched by magnetic nanocores.

Metastasis resulting from circulating tumor cells (CTCs) is associated with 90% of all cancer mortality. To disrupt cancer dissemination, therapeutic targeting of CTCs by extracorporeal photodynamic therapy (PDT) has emerged; however, it still remains impractical due to its limited therapeutic window. Herein, we developed a photosensitive and magnetic targeted core-satellite nanomedicine (TCSN) to augment the light-induced damage to the targeted cells. The magnetic nanocore (MNC) with multiple iron oxide nanoparticles stabilized using thiolated polyvinyl alcohol can magnetize the CTCs to achieve magnetic enrichment under a magnetic field. Multiple gold nanocage (AuNC) satellites were conjugated on the MNC to facilitate bimodal photothermal therapy and PDT. Adjusting the thiol content in the MNC allows manipulating the AuNC density on TCSNs, which has been found to demonstrate a density-dependent bimodal phototherapeutic effect under laser irradiation at 808 and 940 nm. Moreover, with the immobilization of anti-epithelial cell adhesion molecule (anti-EpCAM), TCSN exhibited an enhanced affinity toward EpCAM-expressing 4T1 cells. We demonstrate that TCSN-labeled 4T1 cells can be isolated and photo-eradicated in a microfluidic channel with a dynamic flow. Our studies showed that TCSN with the complementary properties of MNC and AuNCs can largely augment the therapeutic window by magnetic enrichment and bimodal phototherapy, serving as an advanced extracorporeal strategy to remove CTCs.

Advances in the Characterization of Circulating Tumor Cells in Metastatic Breast Cancer: Single Cell Analyses and Interactions, and Patient-Derived Models for Drug Testing.

Metastasis is the major cause of breast cancer death worldwide. In metastatic breast cancer, circulating tumor cells (CTCs) can be captured from patient blood samples sequentially over time and thereby serve as surrogates to assess the biology of surviving cancer cells that may still persist in solitary or multiple metastatic sites following treatment. CTCs may thus function as potential real-time decision-making guides for selecting appropriate therapies during the course of disease or for the development and testing of new treatments. The heterogeneous nature of CTCs warrants the use of single cell platforms to better inform our understanding of these cancer cells. Current techniques for single cell analyses and techniques for investigating interactions between cancer and immune cells are discussed. In addition, methodologies for growing patient-derived CTCs in vitro or propagating them in vivo to facilitate CTC drug testing are reviewed. We advocate the use of CTCs in appropriate microenvironments to appraise the effectiveness of cancer chemotherapies, immunotherapies, and for the development of new cancer treatments, fundamental to personalizing and improving the clinical management of metastatic breast cancer.

Proteomics analysis of tumor exosomes reveals vital pathways of Jinfukang inhibiting circulating tumor cells metastasis in lung cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Jinfukang has long been used for the clinical treatment of lung cancer. Previous studies have shown that Jinfukang can induce the apoptosis of circulating tumor cells by intervening ROS-mediated DNA damage pathway. However, whether Jinfukang can inhibit the metastasis of circulating tumor cells and its mechanism are still unclear. AIM OF THE STUDY: To further investigate the mechanism of Jinfukang in anti-metastasis of lung cancer from the perspective of intervention of tumor exosomes. MATERIALS AND METHODS: The invadopodia formation was determined with immunofluorescence. Invasion and migration were detected using the Transwell assay. Ultracentrifugation was used to isolate exosomes. Exosomes were characterized by electron microscopy, nanoparticle tracking analysis and immunoblotting, and the protein profile was evaluated by proteomic analysis. The molecular functions, biological processes and signaling pathway enrichment analysis were performed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Key differentially expressed proteins were verified by Western blot. RESULTS: Jinfukang can inhibit the expression of MMP14, cortactin, Tks5 and the formation of invadopodia of CTC-TJH-01 cells. Furthermore, Jinfukang can significantly inhibit the invasion and migration of CTC-TJH-01 cells. The diameter of exosomes extracted from the CTC-TJH-01 cells treated by Jinfukang was 30-100 nm, and the exosomal markers CD63, CD81 and TSG101 were expressed. We identified 680 deferentially expressed proteins. Gene oncology analysis indicated that exosomes were mostly derived from plasma membrane and mainly involved in protein localization and intracellular signaling. The ingenuity pathway analysis showed that the EGF pathway was significantly inhibited, whereas the GP6 signaling pathway was significantly activated. We also confirmed that Jinfukang inhibited the expression of EGF pathway-related proteins in CTC-TJH-01 cells. Besides, when EGF was used to activate EGF signaling pathway, the inhibition of Jinfukang on CTC cell metastasis was reversed. CONCLUSION: Jinfukang inhibits the metastasis of CTC-TJH-01 cells through the EGF pathway.