Ligand-targeted Polymerase Chain Reaction Research Articles

The role of folate receptor-positive circulating tumor cells in the prognosis analysis of esophageal squamous cell carcinomas.

e15026 Background: Patients with ESCC always be discovered at an advanced stage and have a poor prognosis. The Circulating Tumor Cells (CTCs) were investigated for use as a biomarker for diagnosis and assessing the effectiveness of treatment. This study aims to investigate the predictive value of the baseline level of folate receptor-positive circulating tumor cells (FR+CTCs) for disease-free survival (DFS) and overall survival (OS) in patients with esophageal squamous cell carcinomas (ESCC). Methods: A retrospective observational study was conducted from January 2018 to February 2021 and followed up to June 2022 with ESCC patients in Sichuan Cancer Hospital & Institute Esophageal Cancer Case Management Database (SCCH-ECCM Database). Three ml of peripheral blood were obtained from each patient, and FR+CTCs were quantified using ligand-targeted polymerase chain reaction (LT-PCR). Traditional techniques were employed to measure other serum markers. Clinicopathological features were collected from the hospital medical record system, and survival data (DFS and OS) were obtained by outpatient follow-up or phone calls. The correlation between clinicopathological characteristics, survival data, and FR+CTCs was analyzed. Cox regression analysis was used to assess risk factors that might have an impact on DFS and OS. Results: Analysis was conducted on 123 ESCC patients. Of them, 86% were men, and their average age was 62.0. FR+CTCs did not significantly correlate with patient age, sex, albumin, pre-albumin, C-reactive protein (CRP), ferritin, tumor size, grade of differentiation, metastasis of lymph nodes, perineural invasion/vessel invasion, or tumor size (all P>0.05). But for DFS (HR 3.29; 95% CI 1.52-7.15, P=0.003) and OS (HR 3.85; 95% CI 1.53-9.72, P=0.004), preoperative FR+CTCs were an independent prognostic factor. Patients with post-operative FR+CTCs ≥17.42 FU/3ml had a substantially shorter DFS than patients with FR+CTCs < 17.42 FU/3ml (P=0.0012). Patients with FR+CTCs ≥17.42 FU/3ml had a shorter OS than patients with FR+CTCs <17.42 FU/3ml, although the difference was not statistically significant (P=0.51). Conclusions: Patients with ESCC who have elevated FR+CTCs typically have a poorer prognosis. We anticipate that FR+CTCs will be able to guide clinical treatment, assess patient prognosis, and monitor cancer recurrence and metastasis in real-time.

Clinical value of folate receptor-positive circulating tumor cells in patients with esophageal squamous cell carcinomas: a retrospective study

BackgroundThe aim of the study is to explore the role of preoperative folate receptor-positive circulating tumor cell (FR+CTC) levels in predicting disease-free survival (DFS) and overall survival (OS) in patients with esophageal squamous cell carcinomas (ESCC).MethodsThree ml blood samples were prospectively drawn from ESCC patients, and ligand-targeted polymerase chain reaction (LT-PCR) was used for the quantification of FR+CTCs. Other serum indicators were measured by traditional methods. Clinicopathological characteristics were obtained from the hospital medical record system, DFS and OS data were obtained by follow-up. The correlation between clinico-pathological characteristics, DFS, and OS and FR+CTCs were analyzed, respectively. Risk factors potentially affecting DFS and OS were explored by Cox regression analysis.Resultsthere were no significant correlations between FR+CTCs and patient age, sex, albumin, pre-albumin, C-reactive protein (CRP), ferritin and CRP/Albumin ratio, tumor size, grade of differentiation, lymph node metastasis, TNM stage, perineural invasion/vessel invasion (all P > 0.05). Nevertheless, preoperative FR+CTCs were an independent prognostic factor for DFS (HR 2.7; 95% CI 1.31-, P = 0.007) and OS (HR 3.37; 95% CI 1.06-, P = 0.04). DFS was significantly shorter for patients with post-operative FR+CTCs ≥ 17.42 FU/3ml compared with patients < 17.42 FU/3ml (P = 0.0012). For OS, it was shorter for patients with FR+CTCs ≥ 17.42 FU/3ml compared with patients < 17.42 FU/3ml, however, the difference did not reach statistical significance (P = 0.51).ConclusionsESCC patients with high FR+CTCs tend to have a worse prognosis. FR+CTCs may monitor the recurrence of cancers in time, accurately assess patient prognosis, and guide clinical decision-making.Trial registrationThe study was approved by the Sichuan Cancer Hospital & Institute Ethics Committee (No. SCCHEC-02-2022-050).

885P Clinical significance of folate receptor-positive circulating tumor cells detected by ligand-targeted polymerase chain reaction in nasopharyngeal carcinoma

885P Clinical significance of folate receptor-positive circulating tumor cells detected by ligand-targeted polymerase chain reaction in nasopharyngeal carcinoma

Folate receptor-positive circulating tumor cells predict survival and recurrence patterns in patients undergoing resection for pancreatic cancer.

ObjectiveTo evaluate the prognostic impact of folate receptor (FR)-positive circulating tumor cells (FR+ CTCs) for patients with pancreatic cancer (PC).BackgroundRisk stratification before surgery for PC patients remains challenging as there are no reliable prognostic markers currently. FR+ CTCs, detected by ligand-targeted polymerase chain reaction (LT-PCR), have shown excellent diagnostic value for PC in our previous study and prognostic value in a variety of cancer types.MethodsPeripheral blood samples from 44 consecutive patients diagnosed with PC were analyzed for FR+ CTCs. 25 patients underwent tumor resection and were assigned to the surgical group. 19 patients failed to undergo radical resection because of local advance or distant metastasis and were assigned to the non-surgical group. The impact of CTCs on relapse and survival were explored.ResultsFor the prognostic stratification, the optimal cut-off value of CTCs analyzed by receiver operating characteristic (ROC) curve was 14.49 folate units (FU)/3 ml. High CTC levels (> 14.49 FU/3 ml) were detected in 52.0% (13/25) of the patients in the surgical group and 63.2% (12/19) in the non-surgical group. In the surgical group, median disease-free survival (DFS) for patients with high CTC levels versus low CTC levels (< 14.49 FU/3 ml) was 8.0 versus 26.0 months (P = 0.008). In multivariable analysis, CTCs were an independent risk factor for DFS (HR: 4.589, P = 0.012). Concerning the recurrence patterns, patients with high CTC levels showed a significantly frequent rate of distant and early recurrence (P = 0.017 and P = 0.011). CTC levels remained an independent predictor for both distant (OR: 8.375, P = 0.014) and early recurrence (OR: 8.412, P = 0.013) confirmed by multivariable logistic regression. However, CTCs did not predict survival in the non-surgical group (P = 0.220).ConclusionFR+ CTCs in resected PC patients could predict impaired survival and recurrence patterns after surgery. Preoperative CTC levels detected by LT-PCR may help guide treatment strategies and further studies in a larger cohort are warranted.

The combination of folate receptor-positive circulating tumor cells and serum tumor markers suggests a histological diagnosis of lung cancer.

BackgroundFolate-receptor alpha (FRα) is overexpressed in lung carcinoma. The FR-positive circulating tumor cell (FR+ CTC) has been established to be a non-invasive biomarker for lung cancer diagnosis. In this study, we sought to examine the value of FR+ CTC in the histological diagnosis of suspicious space-occupying pulmonary lesions.MethodsA total of 538 patients with suspicious space-occupying pulmonary lesions were enrolled in this study. FR+ CTCs were detected before treatment initiation using negative enrichment and ligand-targeted polymerase chain reaction assays. The enrolled patients concurrently received serum biomarker tests.ResultsA total of 282 lung cancer patients [163 with adenocarcinoma (ADC), 71 with squamous cell carcinoma (SCC), and 48 with small cell lung cancer (SCLC)], and 256 patients with benign disease who concurrently received FR+ CTC and serum biomarker tests were randomly assigned to a training set and a validation set. The FR+ CTC levels of patients with lung cancer were significantly higher than those of patients with benign disease (P<0.001). Compared to serum tumor biomarkers alone, the model combining FR+ CTC and serum biomarkers had the highest area under the receiver operating characteristic curve in the diagnosis of NSCLC, ADC, SCC, and SCLC.ConclusionsDiagnostic models that include both FR+ CTC and serum biomarkers could increase the efficiency of distinguishing between different histological types of lung cancer and benign space-occupying pulmonary diseases.

Preoperative Folate Receptor-Positive Circulating Tumor Cell Level Is a Prognostic Factor of Long Term Outcome in Non-Small Cell Lung Cancer Patients.

BackgroundSurgical resection is often the preferred treatment for non-small cell lung cancer (NSCLC) patients. Predictive biomarkers after surgery can help monitoring and treating patients promptly, so as to improve the clinical outcome. In this study, we evaluated one potential candidate biomarker, the folate receptor-positive circulating tumor cell (FR+CTC), by investigating its prognostic and predictive significance in NSCLC patients who underwent surgery.MethodsIn this prospective, observational study, we enrolled NSCLC patients who were eligible to receive surgery. Prior to operation, peripheral blood was collected from each patient for an FR+CTC analysis. FR+CTCs were isolated by negative enrichment using immunomagnetic beads to deplete leukocytes and then quantitatively detected by a ligand-targeted polymerase chain reaction (PCR) method. These patients were then given standard care and were actively followed up for seven years. At the end of the follow-up period, the association between the FR+CTC level and the prognosis in these patients was evaluated.ResultsOverall, preoperative FR+CTC level was not significantly different among NSCLC patients with adenocarcinoma or non-adenocarcinoma subtypes (P = 0.24). However, between patients with low- and high-risk pathological adenocarcinoma subtypes, the preoperative FR+CTC level was significantly different (P = 0.028). Further, patients with lower preoperative FR+CTC level had longer relapse-free survival (RFS) and overall survival (OS) than those with higher preoperative FR+CTC level (RFS: not reached vs. 33.3 months, P = 0.018; OS: not reached vs. 72.0 months, P = 0.13). In a multivariate COX regression analysis, FR+CTC level (HR = 4.10; 95% CI, 1.23–13.64; P=0.022) and pathological stage (HR = 3.16; 95% CI, 1.79–10.14; P = 0.0011) were independent prognostic factors of RFS. Moreover, FR+CTC level together with adenocarcinoma subtypes provided additional information on risk for disease recurrence compared with FR+CTC or adenocarcinoma subtype alone.ConclusionOur study demonstrated that the preoperative FR+CTC level was a potential predictor for the prognosis of NSCLC patients underwent surgery. Further, when preoperative FR+CTC level is considered together with primary tumor proliferation characteristics, its prognostic value supplements that of these conventional pathological features.

1332P Prognostic value of baseline and dynamic change value of CTC in advanced NSCLC patients treated with immune checkpoint inhibitor-based therapy

Both immune checkpoint inhibitors (ICIs) monotherapy and ICIs-chemo combination therapy have emerged as a treatment option for selected patients with advanced non-small-cell lung cancer (NSCLC). However, few serological biomarkers are available to monitor the treatment efficacy and predict the long-term prognosis. By quantitative determination of folate receptor-positive circulating tumor cell (FR+CTC), this study performed a dymanic monitoring study in NSCLC patients who underwent ICIs based therapy and investigated the correlation of CTC values to predict the prognosis. This study enrolled 38 advanced NSCLC patients with at least one measurable lesion and expected to receive ICIs treatment. Peripheral blood samples were collected before the treatment and after each cycle of ICIs treatment. FR+CTCs were enumerated by negative enrichment using immunomagnetic beads and ligand-targeted polymerase chain reaction methods. The correlation of FR+CTC baseline level and its dynamic changes with radiographic assessment and prognosis was evaluated. Of the 38 patients, 20 received first-line treatment, 9 received second-line treatment, and 9 received third-line treatment or above. CTCs were detected (≥8.7FU/3ml) in 78.9% of patients at baseline. There was no significant difference in baseline CTC values among first-line, second-line and third-line therapy patients. The CTC level decreased 18.65% after one or more cycles in the PR group, while increased 25.43% in the non-response (SD+PD) group. In the prognosis predicting study of first-line ICIs treatment subgroup, the patients treated harboring high baseline CTC level showed shorter median progression free survival time than those with low expression levels (cutoff CTC level=13; PFS: 6.83 vs. 16.30 months, P=0.0116). In sum, these data confirm the predictive significance of baseline CTCs in advanced NSCLC patients with first-line immune checkpoint inhibitors treatment. Further studies are needed to confirm whether the CTC value and its dynamic change correlated with radiological response.

Ligand-targeted polymerase chain reaction for the detection of folate receptor-positive circulating tumour cells as a potential diagnostic biomarker for pancreatic cancer.

ObjectivesTo detect folate receptor (FR)‐positive circulating tumour cells (FR+ CTCs) by using ligand‐targeted polymerase chain reaction (LT‐PCR) in periampullary cancer patients and to investigate the diagnostic value of FR+ CTCs in distinguishing pancreatic cancer (PC) from benign pancreatic disease.Materials and MethodsCTCs were enriched from 3 mL of peripheral blood and portal vein blood by immunomagnetic depletion of leucocytes and were then detected by LT‐PCR. The diagnostic performance of FR+ CTCs in PC was investigated by receiver‐operating characteristic curve analysis.ResultsIn total, 57 consecutive patients, including 46 patients with PC, five patients with non‐pancreatic periampullary cancer (non‐PC) and six patients with benign pancreatic diseases, were enrolled. FR+ CTC levels were significantly higher in patients with malignant diseases (PC and non‐PC) than in patients with benign pancreatic diseases (P < .01). There was no notable difference in CTC levels between patients with PC and those with non‐PC (P > .05). The combination of FR+ CTCs with carbohydrate antigen 19‐9 (CA19‐9) had better diagnostic efficiency than each of these two markers alone, with high sensitivity (97.8%) and specificity (83.3%).ConclusionsLT‐PCR is feasible and reliable for detecting FR+ CTCs in patients with periampullary cancer. FR+ CTCs, especially when used in combination with CA19‐9, have potential as a biomarker for the diagnosis of PC.

WITHDRAWN: Diagnostic accuracy of folate receptor-positive circulating tumor cells detected by ligand-targeted polymerase chain reaction in patients with non-small-cell lung cancer: A meta-analysis

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Folate receptor-positive circulating tumor cells as a predictive biomarker for the efficacy of first-line pemetrexed-based chemotherapy in patients with non-squamous non-small cell lung cancer.

BackgroundThere is a lack of well-established biomarkers to predict the efficacy of pemetrexed-based chemotherapy. In this prospective phase II study, we investigated the correlation of folate receptor (FR)-positive circulating tumor cell (CTC) level with the clinical outcomes of patients with advanced non-squamous non-small cell lung cancer (nsNSCLC) when treated with pemetrexed-based chemotherapy.MethodsA total of 98 nsNSCLC patients were enrolled. Peripheral blood was collected from each patient prior to initiation of treatment. FR-positive CTCs were enriched by immunomagnetic leukocyte depletion and quantified using ligand-targeted polymerase chain reaction (LT-PCR) method.ResultsPatients with relatively low CTC level (11–16 FU/3 mL, n=32) showed a significantly shorter progression-free survival (PFS) and overall survival (OS) compared with those in the “high CTC level group” (>16 FU/3mL, n=28; median PFS, 133 versus 320 days, P<0.001; median OS, 632 days versus “not reached”, P=0.003). Patients in the “high CTC level group” also achieved superior objective response rate (ORR) and disease control rate (DCR) over those in the “low CTC level group” (ORR, 40.9% versus 9.5%, P=0.0339; DCR, 100% versus 81.0%, P=0.0485). The clinical outcomes of pemetrexed in the “negative-CTC group” (<11 FU/3mL, n=38) fell between the “high CTC level group” and the “low CTC level group” (median PFS, 290 days; median OS, 1,122 days; ORR: 21.2%, DCR: 93.9%). Further multivariate Cox proportional hazards regression analysis demonstrated that “high CTC level” was an independent factor that was significantly associated with better PFS [hazard ratio (HR) =0.26, 95% confidence interval (CI), 0.12–0.58, P=0.001] and OS (HR =0.23, 95% CI, 0.06–0.92, P=0.037).ConclusionsOur results implied that FR-positive CTC is a promising biomarker to predict the clinical outcome of pemetrexed-based chemotherapy in patients with advanced nsNSCLC.

Folate-receptor-positive circulating tumor cells as an efficacious biomarker for the diagnosis of small pulmonary nodules.

The objective of this study is to investigate the clinical significance of folate-receptor-positive circulating tumor cells (FR+CTC) for the diagnosis of lung cancer, especially in early-stage patients. A total of 72 lung cancer patients, including 31 with stage I diseases and two with stage 0 diseases, were enrolled in this study. Twenty-four patients with benign lung diseases and two healthy volunteers served as the control group. Three milliliters of peripheral blood were collected from each participant for FR+CTC analysis on enrollment. FR+CTC enumeration was performed using immunomagnetic leukocyte depletion and ligand-targeted polymerase chain reaction techniques. The study results revealed that using a cutoff value of 8.7 CTC Units/3 mL, the sensitivity, and specificity of FR+CTC for diagnosis of lung cancer were 81.94% and 73.08%, respectively. Such high sensitivity (74.19%) and specificity (73.08%) persisted even if only stage I lung cancer patients were retained in the analysis. In receiver operating characteristic analysis, the performance of FR+CTC (area under the curve = 0.8153) was superior to other clinical biomarkers such as carcinoembryonic antigen, neuron-specific enolase, and cytokeratin 19 fragments. In a subgroup analysis, patients with nodule size of >3 cm showed an improved sensitivity (88.46%); although, the specificity appeared to decrease (40%). All five patients with benign diseases in this subgroup had inflammatory diseases, indicating that large inflammatory nodules may also release FR -expressing cells into the circulatory system. FR+CTC is a reliable biomarker for the early diagnosis of small-sized lung cancer. Further study with larger sample size is required to assess the diagnostic efficiency of FR+CTC in patients with large nodule sizes.

Predictive and prognostic value of folate receptor-positive circulating tumor cells in small cell lung cancer patients treated with first-line chemotherapy.

To assess the predictive and prognostic significance of folate receptor (FR)-positive circulating tumor cells (CTCs) in patients with small cell lung cancer (SCLC) received first-line chemotherapy. Eligible patients with chemotherapy-naïve, unresectable SCLC were enrolled and blood samples were collected. CTCs were enumerated using ligand-targeted polymerase chain reaction (LT-PCR) at baseline, after two cycles of chemotherapy regimen and on disease progression. In total, 80 patients were enrolled and 67 (83.8%) had positive CTC count at baseline (CTCs ≥ 8.7 FU/3mL). The baseline CTC counts in patients with partial response (PR) were significantly higher than those with progression disease (PD) (P = 0.0365). An obvious reduction of CTC enumeration after two cycles of chemotherapy was significantly correlated with PR (P = 0.0380), instead of SD (P = 0.4934). Among positive CTC count group, patients with relative low CTC level had significantly longer progression-free survival (PFS) and overall survival (OS) than those with high CTC level (PFS: 9.1 vs 6.9 months, P = 0.0458; OS: 11.1 vs 8.6 months, P = 0.056). In multivariate analysis, distant metastases (HR = 1.466, P = 0.021) and relative low CTC level (HR = 0.656, P = 0.049) were the independent predictive factors for patients with SCLC received first-line chemotherapy. The present results demonstrated that baseline CTC counts could be the valuable predictive and prognostic biomarker for patients with SCLC received first-line chemotherapy. The reduction of CTC enumeration after two cycles of chemotherapy was a potential predictor of chemotherapeutic response in SCLC.

Clinical Significance of Folate Receptor-positive Circulating Tumor Cells Detected by Ligand-targeted Polymerase Chain Reaction in Lung Cancer.

Background: As the heterogeneity of CTCs is becoming increasingly better understood, it is clear that identifying particular subtypes of CTCs would be more relevant.Methods: We detected folate receptor (FR)-positive circulating tumor cells (FR+-CTCs) by a novel ligand-targeted polymerase chain reaction (LT-PCR) detection technique.Results: In the none-dynamic study, FR+-CTC levels of patients with lung cancer were significantly higher than controls (patients with benign lung diseases and healthy controls). With a threshold of 8.7 CTC units, FR+-CTC showed a sensitivity of 77.7% and specificity of 89.5% in the diagnosis of lung cancer. When compared with established clinical biomarkers including carcinoembryonic antigen (CEA), cytokeratin 19 fragment (CYFRA21-1), and neuron-specific enolase (NSE), FR+-CTC showed the highest diagnostic efficiency. Notably, the combination of FR+-CTC, CEA, NSE, and CYFRA21-1 could significantly improve the diagnostic efficacy in differentiating patients with lung cancer from benign lung disease. In our dynamic surveillance study, the CTC levels of 62 non-small cell lung cancer (NSCLC) patients decreased significantly after tumor resection.Conclusion: We established a LT-PCR-based FR+-CTC detection platform for patients with lung cancer that exhibits high sensitivity and specificity. This platform would be clinical useful in lung cancer diagnosis and treatment response assessment.

P1.05-009 Clinical Value of Circulating Tumor Cell in the Differential Diagnosis of Solitary Pulmonary Nodule

The diagnosis of lung cancer suffers from the lack of accurate, noninvasive and early diagnostic tests. Low-dose helical computed tomography (LDCT) identifies millions of solitary pulmonary nodules (SPN) annually, many of which are undiagnosed as either malignant or benign. When removed surgically, 18%-25% of the nodules are benign, which leads to unnecessary treatment procedures for surgeons, stress and panic for patients and waste of medical resources for government. Therefore, an accurate noninvasive test that can discriminate benign SPN from malignant is urgently needed. Circulating tumor cells (CTCs) are cells shed from either primary or secondary tumors that migrate into the circulatory system and exist at the early stage of cancer. In recent years, CTC has become the research hotspot because of its great significance in the early diagnosis of cancer, disease monitoring, prognosis evaluation and guiding individualized treatment. In this study, we evaluated the application value of CTC in the differential diagnosis of SPN. Peripheral blood samples were collected from 134 patients with solitary pulmonary nodule in Shanghai Chest Hospital from September 2013 to January 2015, including 80 patients with malignant nodule and 54 with benign nodule. CTC levels of the above subjects were detected by LT-PCR (ligand-targeted polymerase chain reaction, LT-PCR) assay, and serum CEA and CYFRA21-1 were detected by flow fluorescence assay. The CTC levels of malignant SPN patients were significantly higher than that of benign SPN patients (P<0.001). The area under the Receiver Operating Characteristic (ROC) curves of CTC and CEA were 0.817(95% CI: 0.743∼0.891) and 0.613(95% CI: 0.508∼0.718) respectively, while the CYFRA21-1 had no significant meaning in the differential diagnosis of SPN. The positive and negative predictive value (PPV and NPV) in differential diagnosis of SPN for CTC were 89% and 74%. Then the patients were divided into three groups according to the nodule diameter to evaluate the diagnostic value of CTC in SPN with different size. For SPN with diameter less than 8 mm, the PPV and NPV of CTC were 86% and 57%; For SPN with diameter between 8 mm and 20 mm, the PPV and NPV of CTC were 88% and 81%; For SPN with diameter greater than 20 mm, the PPV and NPV of CTC were 92% and 68%. Compared with traditional tumor marker, CTC detection could provide more clinical value in differential diagnosis of solitary pulmonary nodule.

Clinical value of circulating tumor cell in the differential diagnosis of solitary pulmonary nodule

Objective To evaluate the application value of circulating tumor cell (CTC) in the differential diagnosis of solitary pulmonary nodule (SPN). Methods Peripheral blood samples were collected from 134 patients with solitary pulmonary nodule in Shanghai Chest Hospital from September 2013 to January 2015, including 80 patients with malignant nodule and 54 with benign nodule. CTC levels of the above subjects were detected by ligand-targeted polymerase chain reaction (LT-PCR) assay, and serum carcinoembryonic antigen (CEA) and cytokeratin 19 fragment (CYFRA21-1) were detected by flow fluorescence assay. Results By Mann-Whitney U Test, the CTC levels of malignant SPN patients [11.06(8.77-14.41)units/3 ml] were significantly higher than those of benign SPN patients [6.65(4.49-7.84)units/3 ml] (Z=-6.217, P<0.001). The sensitivity and specificity of differential diagnosis of SPN for CTC were 80%(64/80)and 85%(46/54)respectively. According to the diameter of SPN, the patients were divided into three groups to evaluate the diagnostic value of CTC in SPN with different size. For SPN with diameter less than 8 mm, the sensitivity and specificity of CTC were 6/9 and 4/5 respectively. For SPN with diameter between 8 mm and 20 mm, the sensitivity and specificity of CTC were 83%(35/42) and 85%(29/34). For SPN with diameter greater than 20 mm, the sensitivity and specificity of CTC were 79% (23/29)and 13/15. Conclusion Comparing with the traditional tumor markers, CTC could provide more clinical value in the differential diagnosis of solitary pulmonary nodule. (Chin J Lab Med, 2016, 39: 941-945) Key words: Solitary pulmonary nodule; Neoplastic cells, circulating; Diagnosis, differential

Folate Receptor–Positive Circulating Tumor Cell Detected by LT-PCR–Based Method as a Diagnostic Biomarker for Non–Small-Cell Lung Cancer

To investigate the diagnostic performance of folate receptor-positive circulating tumor cells in distinguishing non-small-cell lung cancer (NSCLC) from lung benign disease by using a novel ligand-targeted polymerase chain reaction (PCR) detection technique. Circulating tumor cells were enriched from 3-ml peripheral blood by immunomagnetic depletion of leukocytes and then labeled with a conjugate of a tumor-specific ligand folic acid and a synthesized oligonucleotide. After washing off free conjugates, the stripped bound conjugates were analyzed by quantitative PCR. Seven hundred fifty-six participants (473 patients with NSCLC, 227 patients with lung benign disease, and 56 healthy donors) were randomly assigned to a training set and a test set. The circulating tumor cell (CTC) levels in patients with NSCLC were significant higher than those with lung benign disease (p < 0.001) and healthy donors (p < 0.001). Compared with carcinoembryonic antigen, neuron-specific enolase, and Cyfra21-1, CTCs displayed the highest area under the receiver operating characteristic curve (training set, 0.815; validation set, 0.813) in the diagnosis of NSCLC, with a markedly sensitivity (training set, 72.46%; validation set, 76.37%) and specificity (training set, 88.65%; validation set, 82.39%). The model combining CTCs with carcinoembryonic antigen, neuron-specific enolase, and Cyfra21-1 was more effective for the diagnosis of NSCLC than tumor makers alone (sensitivity and specificity in the training set, 84.21% and 83.91%; validation set, 88.78% and 87.36%, respectively). In addition, the CTC levels were higher in patients with stage III/IV NSCLC compared with those with stage I/II disease. Ligand-targeted PCR technique was feasible and reliable for detecting folate receptor-positive CTCs in patients with NSCLC, and CTC levels could be used as a useful biomarker for the diagnosis of NSCLC.

Quantification of Rare Circulating Tumor Cells in Non-Small Cell Lung Cancer by Ligand-Targeted PCR

BackgroundQuantification of circulating tumor cells (CTC) is valuable for evaluation of non-small cell lung cancer (NSCLC). The sensitivity of current methods constrains their use to detect rare CTCs in early stage. Here we evaluate a novel method, ligand-targeted polymerase chain reaction (LT-PCR), that can detect rare CTCs in NSCLC patients.MethodsCTCs were enriched by immunomagnetic depletion of leukocytes and then labeled by a conjugate of a tumor-specific ligand and an oligonucleotide. After washing off free conjugates, the bound conjugates were stripped from CTCs and then analyzed by qPCR. To evaluate the clinical utility, blood samples were obtained from 72 NSCLC patients (33 initially diagnosed and 39 on chemotherapy), 20 benign patients, and 24 healthy donors.ResultsExperiments with healthy blood spiked with tumor cells indicated the LT-PCR allows specific detection of CTC. The clinical study showed that the initially diagnosed patients have an average of 20.8 CTC units with metastatic diseases, 11.8 CTC units with localized diseases, and 6.0 CTC units with benign diseases. With the threshold of 8.5 CTC units, the assay can detect 80% of stage I/II, 67% of stage III, and 93% of stage IV cancer. With the benign patients and healthy donors as control group, the method can detect cancer with a sensitivity of 81.8% and a specificity of 93.2%.ConclusionThe LT-PCR would allow quantification of CTC in NSCLC patients at a more sensitive level, providing a potential tool for stratifying malignant lung diseases, especially at early stage.