Diagnostic accuracy of liquid biopsy in endometrial cancer

Background: Liquid biopsy is a minimally invasive collection of a patient body fluid sample. In oncology, they offer several advantages compared to traditional tissue biopsies. However, the potential of this method in endometrial cancer (EC) remains poorly explored. We studied the utility of tumor educated platelets (TEPs) and circulating tumor DNA (ctDNA) for preoperative EC diagnosis, including histology determination. Methods: TEPs from 295 subjects (53 EC patients, 38 patients with benign gynecologic conditions, and 204 healthy women) were RNA-sequenced. DNA sequencing data were obtained for 519 primary tumor tissues and 16 plasma samples. Artificial intelligence was applied to sample classification. Results: Platelet-dedicated classifier yielded AUC of 97.5% in the test set when discriminating between healthy subjects and cancer patients. However, the discrimination between endometrial cancer and benign gynecologic conditions was more challenging, with AUC of 84.1%. ctDNA-dedicated classifier discriminated primary tumor tissue samples with AUC of 96% and ctDNA blood samples with AUC of 69.8%. Conclusions: Liquid biopsies show potential in EC diagnosis. Both TEPs and ctDNA profiles coupled with artificial intelligence constitute a source of useful information. Further work involving more cases is warranted.publishedVersio

Detection and localization of early- and late-stage cancers using platelet RNA

Cancer patients benefit from early tumor detection since treatment outcomes are more favorable for less advanced cancers. Platelets are involved in cancer progression and are considered a promising biosource for cancer detection, as they alter their RNA content upon local and systemic cues. We show that tumor-educated platelet (TEP) RNA-based blood tests enable the detection of 18 cancer types. With 99% specificity in asymptomatic controls, thromboSeq correctly detected the presence of cancer in two-thirds of 1,096 blood samples from stage I–IV cancer patients and in half of 352 stage I–III tumors. Symptomatic controls, including inflammatory and cardiovascular diseases, and benign tumors had increased false-positive test results with an average specificity of 78%. Moreover, thromboSeq determined the tumor site of origin in five different tumor types correctly in over 80% of the cancer patients. These results highlight the potential properties of TEP-derived RNA panels to supplement current approaches for blood-based cancer screening

Detection and Characterization of Circulating Tumor Cells Using Imaging Flow Cytometry—A Perspective Study

Tumor dissemination is one of the most-investigated steps of tumor progression, which in recent decades led to the rapid development of liquid biopsy aiming to analyze circulating tumor cells (CTCs), extracellular vesicles (EVs), and circulating nucleic acids in order to precisely diagnose and monitor cancer patients. Flow cytometry was considered as a method to detect CTCs; however, due to the lack of verification of the investigated cells’ identity, this method failed to reach clinical utility. Meanwhile, imaging flow cytometry combining the sensitivity and high throughput of flow cytometry and image-based detailed analysis through a high-resolution microscope might open a new avenue in CTC technologies and provide an open-platform system alternative to CellSearch®, which is still the only gold standard in this field. Hereby, we shortly review the studies on the usage of flow cytometry in CTC identification and present our own representative images of CTCs envisioned by imaging flow cytometry providing rationale that this novel technology might be a good tool for studying tumor dissemination, and, if combined with a high CTC yield enrichment method, could upgrade CTC-based diagnostics

Tumor Heterogeneity at Protein Level as an Independent Prognostic Factor in Endometrial Cancer

Intratumor heterogeneity implies heterogeneous protein function, facilitating tumor adaptation which results in therapeutic failure. We hypothesized that tumor heterogeneity at protein level may influence the course of the disease. As a single biopsy might not represent the full biologic complexity of the tumor, we have analyzed immunohistochemically four different cores obtained from each primary tumor within the cohort of 364 patients with endometrial cancer (EC). The following proteins were examined: estrogen receptor 1 (ESR1), progesterone receptor, epidermal growth factor receptor, v-erb-b2 erythroblastic leukemia viral oncogene homolog 2, receptor tyrosine-protein kinase erbB-3, v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4, phosphatidylinositol-4,5-bisphosphate 3-kinase, phosphorylated v-akt murine thymoma viral oncogene homolog 1, v-myc avian myelocytomatosis viral oncogene homolog, DNA topoisomerase II alpha 170 kDa (TOP2A), cyclin-dependent kinase inhibitor 2A (CDKN2A), tumor protein p53, RAD21 homolog, S. pombe, and runt-related transcription factor 1. Particularly strong correlation was found between TOP2A and CDKN2A heterogeneity and higher stage of the disease (P = .0002 and P = .0003, respectively). Most correlations with clinicopathologic data were observed for ESR1 heterogeneity that correlated with non-endometrioid carcinomas (P=.02), higher stage (P=.005), grade (P=.01), and the presence of metastases (P = .01). Thirty-nine (11.0%) patients were classified as “globally heterogeneous”. Cumulative tumor heterogeneity strongly correlated with the presence of metastases, higher stage, and higher grade of the disease (all P b .05). It also carried negative prognostic value (P=.0008). We show that the degree of heterogeneity in EC might serve as a clinically valid molecular marker

Expression of epithelial to mesenchymal transition-related markers in lymph node metastases as a surrogate for primary tumor metastatic potential in breast cancer

Abstract Background Breast cancers are phenotypically and genotypically heterogeneous tumors containing multiple cancer cell populations with various metastatic potential. Aggressive tumor cell subpopulations might more easily be captured in lymph nodes metastases (LNM) than in primary tumors (PT). We evaluated mRNA and protein levels of master EMT regulators: TWIST1, SNAIL and SLUG, protein levels of EMT-related markers: E-cadherin, vimentin, and expression of classical breast cancer receptors: HER2, ER and PgR in PT and corresponding LNM. The results were correlated with clinicopathological data and patients outcomes. Methods Formalin-fixed paraffin-embedded samples from PT and matched LNM from 42 stage II-III breast cancer patients were examined. Expression of TWIST1, SNAIL and SLUG was measured by reverse-transcription quantitative PCR. Protein expression was examined by immunohistochemistry on tissue microarrays. Kaplan-Meier curves for disease-free survival (DFS) and overall survival (OS) were compared using F-Cox test. Hazard ratios (HRs) with 95% confidence intervals (95% CI) were computed using Cox regression analysis. Results On average, mRNA expression of TWIST1, SNAIL and SLUG was significantly higher in LNM compared to PT (P TWIST1 and SNAIL in LNM was associated with shorter OS (P = 0.04 and P = 0.02, respectively) and DFS (P = 0.02 and P = 0.01, respectively), whereas their expression in PT had no prognostic impact. Negative-to-positive switch of SNAIL protein correlated with decreased OS and DFS (HR = 4.6; 1.1-18.7; P = 0.03 and HR = 3.8; 1.0-48.7; P = 0.05, respectively). Conclusions LNM are enriched in cells with more aggressive phenotype, marked by elevated levels of EMT regulators. High expression of TWIST1 and SNAIL in LNM, as well as negative-to-positive conversion of SNAIL confer worse prognosis, confirming the correlation of EMT with aggressive disease behavior. Thus, molecular profiling of LNM may be used as surrogate marker for aggressiveness and metastatic potential of PT.</p

Prognostic Significance of <i>ESR1</i> Amplification and <i>ESR1</i> PvuII, <i>CYP2C19*2</i>, <i>UGT2B15*2</i> Polymorphisms in Breast Cancer Patients

<div><p>Introduction</p><p>Amplification of the <i>ESR1</i> gene, coding for estrogen receptor alpha, was shown to predict responsiveness to tamoxifen, however its prognostic impact in breast cancer patients has not been thoroughly investigated. Other factors that could contribute to responsiveness to tamoxifen treatment are polymorphisms in <i>ESR1</i> gene and genes involved in tamoxifen metabolism.</p> <p>The aim of this study was to assess the prognostic role of <i>ESR1</i> gene dosage in a consecutive group of breast cancer patients and to correlate this feature with clinico-pathological factors. Additionally, <i>ESR1</i> PvuII, <i>CYP2C19*2</i> and <i>UGT2B15*2</i> polymorphisms were analyzed in the tamoxifen-treated subgroup of patients.</p> <p>Materials and Methods</p><p>Primary tumor samples from 281 stage I-III consecutive breast cancer patients were analyzed for <i>ESR1</i> gene dosage using real-time PCR with locked nucleic acids hydrolysis probes. In the tamoxifen-treated subgroup of patients, <i>ESR1</i> PvuII, <i>CYP2C19*2</i> and <i>UGT2B15*2</i> polymorphism in leukocytes genomic DNA were analyzed. Results were correlated with clinico-pathological factors and with disease-free survival (DFS) and overall survival (OS).</p> <p>Results</p><p><i>ESR1</i> amplification (with a cut-off level of 2.0) was found in 12% of the entire group of breast cancer patients, and in 18% of the ER-negative subgroup. This feature was associated with decreased DFS both in the entire group (<i>P</i>=0.007) and in the ER-negative subgroup (<i>P</i>=0.03), but not in the tamoxifen-treated patients.</p> <p>Patients with <i>ESR1</i> PvuII wt/wt genotype and at least one <i>UGT2B15</i> wt allele had a worse DFS (<i>P</i>=0.03) and showed a trend towards decreased Os (<i>P</i>=0.08) in comparison to patients with <i>ESR1</i> PvuII wt/vt or vt/vt genotype and <i>UGT2B15</i> *2/*2 genotype.</p> <p>Conclusions</p><p><i>ESR1</i> amplification can occur in ER-negative tumors and may carry poor prognosis. In the tamoxifen-treated subgroup, poor prognosis was related to the combined presence of <i>ESR1</i> PvuII wt/wt and <i>UGT2B15</i>wt/wt or wt/*2 genotype.</p> </div

Kaplan-Meier survival curves according to <i>ESR1</i> gene amplification status.

<p>Probability of disease-free survival (A, C) and overall survival (B, D) in all patients (A, B) and in ER-negative patients (C, D).</p

Patients survival in relation to <i>ESR1</i> PvuII and <i>UGT2B15*2</i> polymorphisms in tamoxifen-treated patients.

<p>Disease-free survival (A) and overall survival (B) are shown in respective panels.</p

Schematic representation of blood samples analysis process.

<p>Collected blood sample was layered on density gradient. After centrifugation fraction containing tumor cells (marked by green box) was collected and subjected to negative selection. After CD45-depletetion, CTC-enriched blood sample was subjected to RNA isolation, reverse transcription and gene expression analysis with qPCR.</p

Diagnostic accuracy of liquid biopsy in endometrial cancer

Background: Liquid biopsy is a minimally invasive collection of a patient body fluid sample. In oncology, they offer several advantages compared to traditional tissue biopsies. However, the potential of this method in endometrial cancer (EC) remains poorly explored. We studied the utility of tumor educated platelets (TEPs) and circulating tumor DNA (ctDNA) for preoperative EC diagnosis, including histology determination. Methods: TEPs from 295 subjects (53 EC patients, 38 patients with benign gynecologic conditions, and 204 healthy women) were RNA-sequenced. DNA sequencing data were obtained for 519 primary tumor tissues and 16 plasma samples. Artificial intelligence was applied to sample classification. Results: Platelet-dedicated classifier yielded AUC of 97.5% in the test set when discriminating between healthy subjects and cancer patients. However, the discrimination between endometrial cancer and benign gynecologic conditions was more challenging, with AUC of 84.1%. ctDNA-dedicated classifier discriminated primary tumor tissue samples with AUC of 96% and ctDNA blood samples with AUC of 69.8%. Conclusions: Liquid biopsies show potential in EC diagnosis. Both TEPs and ctDNA profiles coupled with artificial intelligence constitute a source of useful information. Further work involving more cases is warranted