scholarly journals Hsp70 in Liquid Biopsies—A Tumor-Specific Biomarker for Detection and Response Monitoring in Cancer

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3706
Author(s):  
Caroline Werner ◽  
Stefan Stangl ◽  
Lukas Salvermoser ◽  
Melissa Schwab ◽  
Maxim Shevtsov ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Extracellular Vesicles ◽  
Conformational Changes ◽  
Small Cell Lung Carcinoma ◽  
Sandwich Elisa ◽  
Response Monitoring ◽  
Tumor Biomarker ◽  
Liquid Biopsies ◽  
Cell Lung Carcinoma ◽  
Viable Tumor

In contrast to normal cells, tumor cells of multiple entities overexpress the Heat shock protein 70 (Hsp70) not only in the cytosol, but also present it on their plasma membrane in a tumor-specific manner. Furthermore, membrane Hsp70-positive tumor cells actively release Hsp70 in small extracellular vesicles with biophysical characteristics of exosomes. Due to conformational changes of Hsp70 in a lipid environment, most commercially available antibodies fail to detect membrane-bound and vesicular Hsp70. To fill this gap and to assess the role of vesicular Hsp70 in circulation as a potential tumor biomarker, we established the novel complete (comp)Hsp70 sandwich ELISA, using two monoclonal antibodies (mAbs), that is able to recognize both free and lipid-associated Hsp70 on the cell surface of viable tumor cells and on small extracellular vesicles. The epitopes of the mAbs cmHsp70.1 (aa 451–461) and cmHsp70.2 (aa 614–623) that are conserved among different species reside in the substrate-binding domain of Hsp70 with measured affinities of 0.42 nM and 0.44 nM, respectively. Validation of the compHsp70 ELISA revealed a high intra- and inter-assay precision, linearity in a concentration range of 1.56 to 25 ng/mL, high recovery rates of spiked liposomal Hsp70 (>84%), comparable values between human serum and plasma samples and no interference by food intake or age of the donors. Hsp70 concentrations in the circulation of patients with glioblastoma, squamous cell or adeno non-small cell lung carcinoma (NSCLC) at diagnosis were significantly higher than those of healthy donors. Hsp70 concentrations dropped concomitantly with a decrease in viable tumor mass upon irradiation of patients with approximately 20 Gy (range 18–22.5 Gy) and after completion of radiotherapy (60–70 Gy). In summary, the compHsp70 ELISA presented herein provides a sensitive and reliable tool for measuring free and vesicular Hsp70 in liquid biopsies of tumor patients, levels of which can be used as a tumor-specific biomarker, for risk assessment (i.e., differentiation of grade III vs. IV adeno NSCLC) and monitoring of therapeutic outcomes.

scholarly journals Exosomal Hsp70 in Liquid Biopsies - A Biomarker for Prediction and Response Monitoring in Cancer

2021 ◽  
Author(s):  
Caroline Werner ◽  
Stefan Stangl ◽  
Lukas Salvermoser ◽  
Melissa Schwab ◽  
Maxim Shevstov ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Conformational Changes ◽  
Small Cell Lung Carcinoma ◽  
Sandwich Elisa ◽  
Response Monitoring ◽  
Tumor Biomarker ◽  
Liquid Biopsies ◽  
Cell Lung Carcinoma ◽  
Viable Tumor ◽  
Lipid Environment

In contrast to normal cells, tumor cells of multiple entities overexpress the Heat Shock Protein 70 (Hsp70) not only in the cytosol, but also present it on their plasma membrane in a tumor-specific manner. Furthermore, membrane-Hsp70 positive tumor cells actively release Hsp70 into lipid microvesicles termed exosomes into the blood. Due to conformational changes of Hsp70 in the lipid environment, most commercially available antibodies fail to detect membrane-bound and exosomal Hsp70. To fill this gap and to assess the role of exosomal Hsp70 in the circulation as a potential tumor biomarker, we established the novel complete Hsp70 (compHsp70) sandwich ELISA using two monoclonal antibodies (mAbs) that are able to recognize both, free and lipid-associated Hsp70 on the cell surface of viable tumor cells and exosomes. The epitopes of the mAbs cmHsp70.1 (aa 451-461) and cmHsp70.2 (aa 614-623) that are conserved among different species reside in the substrate-binding domain of Hsp70, with measured affinities of 0.42 nM and 0.44 nM, respectively. Validation of the compHsp70 ELISA revealed a high intra- and inter-assay precision, linearity in a concentration range of 1.56 to 25 ng/ml, high recovery rates of ‘spiked’ liposomal Hsp70 (>84%), comparable values between human serum and plasma samples, and no interference by food intake or age of the donors. Hsp70 concentrations in the circulation of patients with glioblastoma, squamous cell or adeno non-small cell lung carcinoma (NSCLC) at diagnosis were significantly higher than those of healthy volunteers. Hsp70 concentrations dropped concomitantly with the decrease in viable tumor mass on irradiation of patients with approximately 20 Gy (range 18 – 22.5 Gy) or after completion of radiotherapy (60 - 70 Gy). In summary, the compHsp70 ELISA presented herein provides a highly sensitive and reliable tool for measuring free and exosomal Hsp70 in liquid biopsies of tumor patients, levels of which can be used as a predictive tumor-specific biomarker, risk assessment and for monitoring therapeutic outcome.

scholarly journals Molecular and Circulating Biomarkers of Brain Tumors

2021 ◽  
Vol 22 (13) ◽  
pp. 7039
Author(s):  
Wojciech Jelski ◽  
Barbara Mroczko
Keyword(s):  
Brain Tumors ◽  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
Extracellular Vesicles ◽  
Dna Methyltransferase ◽  
Growth Factor Receptor ◽  
Response To Treatment ◽  
Liquid Biopsies ◽  
Methylguanine Dna Methyltransferase ◽  
The Central Nervous System

Brain tumors are the most common malignant primary intracranial tumors of the central nervous system. They are often recognized too late for successful therapy. Minimally invasive methods are needed to establish a diagnosis or monitor the response to treatment of CNS tumors. Brain tumors release molecular information into the circulation. Liquid biopsies collect and analyze tumor components in body fluids, and there is an increasing interest in the investigation of liquid biopsies as a substitute for tumor tissue. Tumor-derived biomarkers include nucleic acids, proteins, and tumor-derived extracellular vesicles that accumulate in blood or cerebrospinal fluid. In recent years, circulating tumor cells have also been identified in the blood of glioblastoma patients. In this review of the literature, the authors highlight the significance, regulation, and prevalence of molecular biomarkers such as O6-methylguanine-DNA methyltransferase, epidermal growth factor receptor, and isocitrate dehydrogenase. Herein, we critically review the available literature on plasma circulating tumor cells (CTCs), cell-free tumors (ctDNAs), circulating cell-free microRNAs (cfmiRNAs), and circulating extracellular vesicles (EVs) for the diagnosis and monitoring of brain tumor. Currently available markers have significant limitations.While much research has been conductedon these markers, there is still a significant amount that we do not yet understand, which may account for some conflicting reports in the literature.

scholarly journals Cytokeratin-positive interstitial reticulum cell (CIRC) tumor in the lymph node: a case report of the transformation from the epithelioid cell type to the spindle cell type

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Sachiko Kaji ◽  
Nobuyuki Hiruta ◽  
Daisuke Sasai ◽  
Makoto Nagashima ◽  
Rintaro Ohe ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Lung Carcinoma ◽  
Spindle Cell ◽  
Small Cell Lung Carcinoma ◽  
Epithelioid Cell ◽  
Small Cell ◽  
Cell Type ◽  
Small Cell Lung ◽  
Cell Lung Carcinoma ◽  
Spindle Cell Type

Abstract Background Cytokeratin-positive interstitial reticulum cells (CIRCs), which are a subgroup of fibroblastic reticular cells (FRCs), are known to be present in the lymph nodes. There have been only a few cases of tumors derived from CIRCs. Case presentation We have reported a new case involving a CIRC tumor in a 75-year-old man and reviewed the literature. The resected mediastinal lymph nodes showed epithelial-like proliferation of large atypical round and polygonal epithelioid cells. The tumor cells expressed CK8, CK18, CAM5.2, AE1/AE3, epithelial membrane antigen, vimentin, fascin, and some FRC markers, which is consistent with the diagnosis of a CIRC tumor. Following chemotherapy, the CIRC tumor was observed to have responded very well and became difficult to confirm on imaging, but a small cell lung carcinoma developed 12 months later. Chemoradiotherapy was performed, but the patient passed away 29 months after the initial diagnosis. The autopsy revealed the recurrence of the CIRC tumor, residual small cell lung carcinoma, and a very small latent carcinoma of the prostate. The relapsed CIRC tumor cells had a spindle shape; they were highly pleomorphic and had invaded the superior vena cava. Conclusion We first reported autopsy findings of CIRC tumors and demonstrated the transformation of the tumor from the epithelioid cell type to the spindle cell type.

scholarly journals Detection of EGFR Mutations Using Bronchial Washing-Derived Extracellular Vesicles in Patients with Non-Small-Cell Lung Carcinoma

Cancers ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 2822
Author(s):  
Juhee Park ◽  
Chaeeun Lee ◽  
Jung Seop Eom ◽  
Mi-Hyun Kim ◽  
Yoon-Kyoung Cho
Keyword(s):  
Lung Cancer ◽  
Extracellular Vesicles ◽  
Egfr Mutation ◽  
Small Cell Lung Carcinoma ◽  
Resistance Mutation ◽  
Detection Sensitivity ◽  
Egfr Mutations ◽  
Tissue Samples ◽  
Cell Lung Carcinoma ◽  
Bronchial Washing

The detection of epidermal growth factor receptor (EGFR) mutation, based on tissue biopsy samples, provides a valuable guideline for the prognosis and precision medicine in patients with lung cancer. In this study, we aimed to examine minimally invasive bronchial washing (BW)-derived extracellular vesicles (EVs) for EGFR mutation analysis in patients with lung cancer. A lab-on-a-disc equipped with a filter with 20-nm pore diameter, Exo-Disc, was used to enrich EVs in BW samples. The overall detection sensitivity of EGFR mutations in 55 BW-derived samples was 89.7% and 31.0% for EV-derived DNA (EV-DNA) and EV-excluded cell free-DNA (EV-X-cfDNA), respectively, with 100% specificity. The detection rate of T790M in 13 matched samples was 61.5%, 10.0%, and 30.8% from BW-derived EV-DNA, plasma-derived cfDNA, and tissue samples, respectively. The acquisition of T790M resistance mutation was detected earlier in BW-derived EVs than plasma or tissue samples. The longitudinal analysis of BW-derived EVs showed excellent correlation with the disease progression measured by CT images. The EGFR mutations can be readily detected in BW-derived EVs, which demonstrates their clinical potential as a liquid-biopsy sample that may aid precise management, including assessment of the treatment response and drug resistance in patients with lung cancer.

scholarly journals Chemotherapeutic drugs stimulate the release and recycling of extracellular vesicles to assist cancer cells in developing an urgent chemoresistance

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
Xiaokun Wang ◽  
Dongjuan Qiao ◽  
Likun Chen ◽  
Meng Xu ◽  
Shupeng Chen ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Small Cell Lung Carcinoma ◽  
Underlying Mechanism ◽  
Chemotherapeutic Agents ◽  
Chemotherapeutic Drugs ◽  
Xenograft Tumor ◽  
Tumor Models ◽  
Cell Lung Carcinoma

Abstract Background Chemotherapy is a widely used treatment for cancer. However, the development of acquired multidrug resistance (MDR) is a serious issue. Emerging evidence has shown that the extracellular vesicles (EVs) mediate MDR, but the underlying mechanism remains unclear, especially the effects of chemotherapeutic agents on this process. Methods Extracellular vesicles isolation was performed by differential centrifugation. The recipient cells that acquired ATP-binding cassette sub-family B member 1 (ABCB1) proteins were sorted out from co-cultures according to a stringent multi-parameter gating strategy by fluorescence-activated cell sorting (FACS). The transfer rate of ABCB1 was measured by flow cytometry. The xenograft tumor models in mice were established to evaluate the transfer of ABCB1 in vivo. Gene expression was detected by real-time PCR and Western blotting. Results Herein, we show that a transient exposure to chemotherapeutic agents can strikingly increase Rab8B-mediated release of extracellular vesicles (EVs) containing ABCB1 from drug-resistant cells, and accelerate these EVs to circulate back onto plasma membrane of sensitive tumor cells via the down-regulation of Rab5. Therefore, intercellular ABCB1 transfer is significantly enhanced; sensitive recipient cells acquire a rapid but unsustainable resistance to evade the cytotoxicity of chemotherapeutic agents. More fascinatingly, in the xenograft tumor models, chemotherapeutical drugs also locally or distantly increase the transfer of ABCB1 molecules. Furthermore, some Non-small-cell lung carcinoma (NSCLC) patients who are undergoing primary chemotherapy have a rapid increase of ABCB1 protein in their monocytes, and this is obviously associated with poor chemotherapeutic efficacy. Conclusions Chemotherapeutic agents stimulate the secretion and recycling of ABCB1-enriched EVs through the dysregulation of Rab8B and Rab5, leading to a significant increase of ABCB1 intercellular transfer, thus assisting sensitive cancer cells to develop an urgent resistant phenotype. Our findings provide a new molecular mechanism of how chemotherapeutic drugs assist sensitive cancer cells in acquiring an urgent resistance.

scholarly journals Induction of CD8 T-cell-Ifn-γ response and positive clinical outcome after immunization with gene-modified allogeneic tumor cells in advanced non-small-cell lung carcinoma

2003 ◽  
Vol 10 (11) ◽  
pp. 850-858 ◽  
Author(s):  
Luis E Raez ◽  
Peter A Cassileth ◽  
James J Schlesselman ◽  
Swaminathan Padmanabhan ◽  
Eva Z Fisher ◽  
...  
Keyword(s):  
T Cell ◽  
Clinical Outcome ◽  
Tumor Cells ◽  
Lung Carcinoma ◽  
Small Cell Lung Carcinoma ◽  
Cd8 T Cell ◽  
Small Cell ◽  
Small Cell Lung ◽  
Cell Lung Carcinoma ◽  
Ifn Γ

scholarly journals Antigen-dependent CD28 Signaling Selectively Enhances Survival and Proliferation in Genetically Modified Activated Human Primary T Lymphocytes

1998 ◽  
Vol 188 (4) ◽  
pp. 619-626 ◽  
Author(s):  
Anja Krause ◽  
Hong-Fen Guo ◽  
Jean-Baptiste Latouche ◽  
Cuiwen Tan ◽  
Nai-Kong  V. Cheung ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Tumor Cells ◽  
Small Cell Lung Carcinoma ◽  
Interleukin 2 ◽  
Culture Conditions ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Cell Lung Carcinoma ◽  
Antiidiotypic Antibody ◽  
Antigen Presenting

Most tumor cells function poorly as antigen-presenting cells in part because they do not express costimulatory molecules. To provide costimulation to T lymphocytes that recognize tumor cells, we constructed a CD28-like receptor specific for GD2, a ganglioside overexpressed on the surface of neuroblastoma, small-cell lung carcinoma, melanoma, and other human tumors. Recognition of GD2 was provided by a single-chain antibody derived from the GD2-specific monoclonal antibody 3G6. We demonstrate that the chimeric receptor 3G6-CD28 provides CD28 signaling upon specific recognition of the GD2 antigen on tumor cells. Human primary T lymphocytes retrovirally transduced with 3G6-CD28 secrete interleukin 2, survive proapoptotic culture conditions, and selectively undergo clonal expansion in the presence of an antiidiotypic antibody specific for 3G6-CD28. Polyclonal CD8+ lymphocytes expressing 3G6-CD28 are selectively expanded when cultured with cells expressing allogeneic major histocompatibility complex class I together with GD2. Primary T cells given such an antigen-dependent survival advantage should be very useful to augment immune responses against tumor cells.

scholarly journals Early detection of the major male cancer types in blood-based liquid biopsies using a DNA methylation panel

2019 ◽  
Vol 11 (1) ◽  
Author(s):  
Vera Constâncio ◽  
Sandra P. Nunes ◽  
Catarina Moreira-Barbosa ◽  
Rui Freitas ◽  
Jorge Oliveira ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Early Detection ◽  
Minimally Invasive ◽  
Lung Carcinoma ◽  
Small Cell Lung Carcinoma ◽  
Small Cell ◽  
Small Cell Lung ◽  
Liquid Biopsies ◽  
Cell Lung Carcinoma ◽  
Cancer Types

Abstract Background Lung (LC), prostate (PCa) and colorectal (CRC) cancers are the most incident in males worldwide. Despite recent advances, optimal population-based cancer screening methods remain an unmet need. Due to its early onset, cancer specificity and accessibility in body fluids, aberrant DNA promoter methylation might be a valuable minimally invasive tool for early cancer detection. Herein, we aimed to develop a minimally invasive methylation-based test for simultaneous early detection of LC, PCa and CRC in males, using liquid biopsies. Results Circulating cell-free DNA was extracted from 102 LC, 121 PCa and 100 CRC patients and 136 asymptomatic donors’ plasma samples. Sodium-bisulfite modification and whole-genome amplification was performed. Promoter methylation levels of APCme, FOXA1me, GSTP1me, HOXD3me, RARβ2me, RASSF1Ame, SEPT9me and SOX17me were assessed by multiplex quantitative methylation-specific PCR. SEPT9me and SOX17me were the only biomarkers shared by all three cancer types, although they detected CRC with limited sensitivity. A “PanCancer” panel (FOXA1me, RARβ2me and RASSF1Ame) detected LC and PCa with 64% sensitivity and 70% specificity, complemented with “CancerType” panel (GSTP1me and SOX17me) which discriminated between LC and PCa with 93% specificity, but with modest sensitivity. Moreover, a HOXD3me and RASSF1Ame panel discriminated small cell lung carcinoma from non-small cell lung carcinoma with 75% sensitivity, 88% specificity, 6.5 LR+ and 0.28 LR–. An APCme and RASSF1Ame panel independently predicted disease-specific mortality in LC patients. Conclusions We concluded that a DNA methylation-based test in liquid biopsies might enable minimally invasive screening of LC and PCa, improving patient compliance and reducing healthcare costs. Moreover, it might assist in LC subtyping and prognostication.

Imaging of the P-glycoprotein in normal and multidrug-resistant small-cell lung carcinoma cells with the Atomic-Force Microscope(AFM)

1994 ◽  
Vol 52 ◽  
pp. 1074-1075
Author(s):  
J. M. Tsai ◽  
D. Sadava ◽  
S. A. C. Gould
Keyword(s):  
Tumor Cells ◽  
Small Cell Lung Carcinoma ◽  
Protein A ◽  
Multidrug Resistant ◽  
Extracellular Medium ◽  
Contact Mode ◽  
Cell Lung Carcinoma ◽  
Lung Carcinoma Cells ◽  
Atomic Force ◽  
P Glycoprotein

Cancer is characterized by the often rapid and uncontrolled rate of cell growth. This alteration in growth pattern causes normal cells to become tumor cells. After undergoing a period of chemotherapy, some tumor cells become resistant to a variety of drugs, a phenomenon known as the multidrug resistance (MDR). One explanation for this change is the overexpression of P-glycoprotein in the drug-resistant cells. This membrane protein is capable of pumping drugs into the extracellular medium. Since the drugs do not accumulate, the tumor cells are not killed. In order to examine this protein, a contact mode AFM was used to image the cell membranes of both the normal and the MDR tumor cells. The four figures provided show the topographical information of the membranes and suggest that there are differences between the cell lines.

Sign in / Sign up

Export Citation Format

Share Document