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. 2018 Nov 19;18(1):1133.
doi: 10.1186/s12885-018-5034-x.

Circulating tumor cells with karyotyping as a novel biomarker for diagnosis and treatment of nasopharyngeal carcinoma

Jing Zhang  1 , Huashan Shi  1 , Tingting Jiang  1 , Zhe Liu  1 , Peter P Lin  2 , Nianyong Chen  3
Affiliations

Circulating tumor cells with karyotyping as a novel biomarker for diagnosis and treatment of nasopharyngeal carcinoma

Jing Zhang et al. BMC Cancer. .

Abstract

Background: Circulating tumor cells (CTCs) have been considered great clinical significance in various cancers. However, it remains unknown that how is the role of CTCs in patients with nasopharyngeal carcinoma (NPC). We investigated the value of CTCs enumeration and karyotyping in NPC.

Methods: In the present study, we applied integrated subtraction enrichment and immunostaining-fluorescence in situ hybridization (SE-iFISH) automatic testing system to detect and characterize CTCs of NPC patients. Enumeration and aneuploidy of chromosome 8 in CTCs were examined in various stages of patients with NPC. The changes of CTCs number and karyotyping post to chemotherapy were investigated in NPC.

Results: CTCs were detected by SE-iFISH in 46 out of 50 pre-treatment NPC patients, and performed a positive rate of 92.0%. No significant association was found between disease staging and CTCs detection rate. CTCs number constantly increased with TNM stage rising (from stage II to stage IV) no matter in newly diagnosed patients without distant metastasis (M0) and relapsed or distant metastatic patients. The number of CTCs decreased after treatment in patients with partial response (PR), while increased in patients with progressive disease or stable disease (PD/SD). More interestingly, CTCs karyotyping indicated that aneuploidy of chromosome 8 in CTCs was dramatically related to chemotherapeutic efficacy in NPC. Positive correlation was found between CTCs count and plasma EBV DNA level of NPC patients.

Conclusions: CTCs could be detected in various stages of NPC patients using SE-iFISH. CTCs number could indicate the severity degree of disease in NPC. Dynamically monitoring the variations in CTCs number may predict chemotherapy efficacy during treatment. CTCs karyotyping is related to the sensibility of chemotherapy and drug resistance, and karyotyping of CTCs might predict therapeutic efficacy and evaluate chemo-resistance in NPC. CTCs could be used as a monitoring indicator in the fields of treatment, diagnosis and follow-up of NPC.

Keywords: Biomarker; Circulating tumor cells; Karyotyping; Nasopharyngeal carcinoma; SE-iFISH.

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Conflict of interest statement

Ethics approval and consent to participate

The ethics committee of West China Hospital of Sichuan University has approved the work and written informed consent was received from all participants prior to the study.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests when conducting the research.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
Identification of CTCs and CTM in NPC by SE-iFISH system. DAPI: blue, CEP8: orange, CD45: red, EpCAM: green. a. DAPI+/CD45-/EpCAM+/CEP8 = 2 (white arrow); b. DAPI+/CD45-/EpCAM+/CEP8 > 2 (white arrow); c DAPI+/CD45-/EpCAM-/CEP8 > 2 (white arrow), DAPI+/CD45+/EpCAM-/CEP8 = 2 (red arrow); d. DAPI+/CD45-/EpCAM-/CEP8 > 2 (white arrow); e. a cluster of CTCs (blue arrow)
Fig. 2
Fig. 2
The relationship between CTCs number and therapeutic efficacy in NPC. a. CTCs number in 12 NPC patients decreased from 8.8 ± 13.1 (before treatment) to 4.3 ± 7.2 (after treatment) following 2 to 4 cycles of chemotherapy. b. CTCs number in patients with PR decreased from 10.8 ± 14.8 (before treatment) to 4.7 ± 8.3 (after treatment). c. CTCs number in PD/SD patients increased from 2.7 ± 1.5 (before treatment) to 3.0 ± 1.7 (after treatment). * P < 0.05
Fig. 3
Fig. 3
Correlation of ploidy of chromosome 8 in CTCs and disease staging. a. The ratio of CTCs with different karyotypes were 82.8% (triploidy), 44.8% (tetraploidy) and 48.3% (multiploidy) in newly diagnosed (M0) patients. b. For newly diagnosed (M0) patients, the frequency of triploid CTCs in stage II/III/IV were 50.0, 75.0, and 93.3%, respectively. Tetraploid CTCs ratio in stage II/III/IV were 0, 50.0, 46.7%. Multiploid CTCs ratio in stage II/III/IV were 0, 58.3, 53.3%. c. The ratio of CTCs with different karyotypes were 4.8% (diploidy), 57.1% (triploidy), 47.6% (tetraploidy) and 66.7% (multiploidy) in patients with relapse or distant metastasis. d. For patients with relapse or distant metastasis, the frequency of triploid CTCs in stage II/III/IV were 50.0, 57.0 and 58.3%, respectively. Tetraploid CTCs ratio in stage II/III/IV were 100, 57.0, 33.3%. Multiploid CTCs ratio in stage II/III/IV were 0, 57.0, 83.3%. Diploid CTCs was detected in only one patient. Multiploidy contains pentaploidy and those > 5 copies of chromosome 8 CTCs. Stage for patients with relapse/distant metastasis after treatment refered to TNM stage at NPC initial diagnosis
Fig. 4
Fig. 4
Comparison of CTCs karyotyping before and after treatment. a. The ratio of triploid, tetraploid, and multiploid CTCs in 12 NPC patients decreased after chemotherapy, while diploid CTCs was remained the same. b. For PR patients, the ratio of tetraploid CTCs definitely decreased from 66.7 to 33.3%, while triploid CTCs decreased from 88.9 to 66.7%, and multiploid CTCs decreased from 44.4 to 33.3% after treatment. c. For PD/SD patients, the frequency of tetraploid CTCs decreased from 66.7 to 33.3%, while triploid CTCs increased from 66.7 to 100%, and multiploid CTCs remained unchanged (33.3% versus 33.3%) after treatment
Fig. 5
Fig. 5
The relationship between CTCs number and plasma EBV DNA level of NPC patients. a. Positive correlation between CTCs number and plasma EBV DNA level was found in the 67 tested samples including 50 patients prior to treatment and 17 patients post to treatment (R = 0.326, P = 0.007). b. Positive correlation between CTCs number and plasma EBV DNA level in 50 pre-treatment patients was found (R = 0.345, P = 0.014). R, Pearson linear correlation coefficient; P, P value
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