Analysis of microRNA Expression in Liquid-Based Cytology Samples may be Useful for Primary Lung Cancer Diagnosis

Yusuke Araki, MD; Koji Arihiro, MD, PhD; Kakuhiro Yamaguchi, MD, PhD; Shinjiro Sakamoto, MD, PhD; Yasushi Horimasu, MD, PhD; Takeshi Masuda, MD, PhD; Shintaro Miyamoto, MD, PhD; Taku Nakashima, MD, PhD; Hiroshi Iwamoto, MD, PhD; Kazunori Fujitaka, MD, PhD; Hironobu Hamada, MD, PhD; Noboru Hattori, MD, PhD

Disclosures

Am J Clin Pathol. 2021;156(4):644-652. 

In This Article

Results

Primary Lung Cancer and Noncancerous Lung Tissues

We analyzed the expression of 4 miRs (miR-21, miR-31, miR-182, and miR-183) in 18 pairs of primary lung cancer tissue and noncancerous lung tissue. As shown in Figure 2, the relative expression levels of the 4 miRs in lung cancer were significantly higher than in adjacent noncancerous tissues.

Figure 2.

Relative expression of miR-21 (A), miR-31 (B), miR-182 (C), and miR-183 (D) in lung cancer tissues and adjacent noncancerous tissues. Four microRNAs were significantly upregulated in cancer tissues. *P < .05; **P < .01. Ct, cycle threshold; N, cases diagnosed as noncancerous; T, cases diagnosed as cancerous.

Cytologic Samples

We analyzed the expression of these 4 miRs in 125 cytology samples. Eighty-two cases of cytologic samples were judged to be benign or indeterminate, and 40 were finally diagnosed as cancerous after histologic examination (Table 3). All cases initially judged as suspicious or malignant were finally diagnosed as cancerous. As shown in Figure 1, the number of samples that could be positively amplified was different for each marker; miR-21 was detected in 125 samples, miR-31 in 123 samples, miR-182 in 109 samples, and miR-183 in 102 samples. For cytologic samples, the relative expression of each miR in cancer cases was significantly higher than those of noncancer cases Figure 3. There was no significant correlation between these 4 miRs and clinical stage (Supplemental Figure 2; all supplemental material can be found at American Journal of Clinical Pathology online). Considering the expression of each tumor subtype, there was no significant difference between each subtype (Supplemental Figure 3). Notably, even for patients whose lesions were cytologically judged as benign or indeterminate, the relative expression of each miR in cancer cases was significantly higher than those of noncancer cases Figure 4.

Figure 3.

Relative expression of miR-21 (A), miR-31 (B), miR-182 (C), and miR-183 (D) in cytologic samples. Four microRNAs were significantly upregulated in samples obtained from cases finally diagnosed as cancerous. *P < .05; **P < .01. Ct, cycle threshold; N, cases diagnosed as noncancerous; T, cases diagnosed as cancerous.

Figure 4.

Relative expression of miR-21 (A), miR-31 (B), miR-182 (C), and miR-183 (D) in cytologic samples judged as benign or indeterminate. Four microRNAs were significantly upregulated in samples obtained from cases histologically diagnosed as cancerous in comparison with samples obtained from cases histologically and diagnosed as noncancerous. *P < .01. N, cases diagnosed as noncancerous; T, cases diagnosed as cancerous.

Of the 96 cases in which all 4 miRs were detected, 54 (56%) were judged as benign and 24 were finally diagnosed as cancerous. In these 54 cases, receiver operating characteristic analysis was conducted to evaluate the diagnostic value of miR expression in samples diagnosed as benign Figure 5. The diagnostic values of miR-21, miR-31, miR-182, and miR-183 were 0.676, 0.683, 0.688, and 0.772, respectively. The combined diagnostic value of expression levels of the 4 miRs was 0.810—better than for each miRNA individually for benign samples.

Figure 5.

Received operating characteristic (ROC) curve analysis of diagnostic value in cytologic samples diagnosed as benign. ROC curve with corresponding the area under the ROC curve (AUC) for miR-21 (AUC = 0.673) (A), miR-31 (AUC = 0.683) (B), miR-182 (AUC = 0.696) (C), and miR-183 (AUC = 0.769) (D) in liquid-biopsy cytology (LBC) from cancer cases vs noncancer cases. ROC curve with corresponding AUC for 4 combined miRNA expression in LBC from cancer cases vs noncancer cases (AUC = 0.810) (E). The diagnostic value of 4 combined miRNAs was better than each individual miRNA.

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