Schlafen family member 11 indicates favorable prognosis of patients with head and neck cancer following platinum-based chemoradiotherapy

Abstract

Recently, Schlafen family member 11 (SLFN11) has been reported to increase the sensitivity of cancer cells to DNA-damaging agents, including platinum derivatives; thus, SLFN11 may be a predictive biomarker for platinum-based chemoradiotherapy (CRT). In this study, we examined whether SLFN11 expression was associated with the therapeutic outcome of platinum-based CRT in head and neck squamous cell carcinoma (HNSCC). We performed immunohistochemical analyses for SLFN11 expression in 161 HNSCC tissues from patients who had been administered cisplatin-based CRT and examined the correlation between SLFN11 expression and progression-free survival (PFS). Additionally, SLFN11 expression was examined in 10 paired samples obtained before and after CRT in patients with local failure. Furthermore, in vitro experiments were performed using several HNSCC cell lines and isogenic SLFN11-knockout cells to assess the association between SLFN11 expression and drug sensitivity. PFS was found to be significantly better in the SLFN11-positive group than in the SLFN11-negative group among the 161 patients (5-year PFS: 78.8% vs. 52.8%, respectively, p < 0.001). Similar results were observed for the PFS at each primary site. The percentage of SLFN11 positivity was lower in tumor samples from patients with local failure after CRT than that in the corresponding primary tumors before CRT in 8 of 10 cases. Results of the in vitro assay demonstrated that SLFN11-knockout cells exhibited reduced sensitivity to DNA-damaging agents but not to the non-DNA-damaging agent docetaxel. Our findings suggest that SLFN11 may serve as a potential biomarker for predicting the response of HNSCC patients to platinum-based CRT.

Keywords: Schlafen family member; head and neck squamous cell carcinoma; immunohistochemistry; platinum-based chemoradiotherapy; prognostic marker.

Figure 1

Immunohistochemical examination of Schlafen family member 11 (SLFN11) in head and neck squamous cell carcinoma (HNSCC). (A) Representative images of immunohistochemical staining of SLFN11 in HNSCC. Formalin-fixed, paraffin-embedded tissue specimens of tumor biopsy samples before chemoradiotherapy (CRT) were subjected to immunohistochemistry using an anti-SLFN11 antibody. Left, an example of an SLFN11-positive case, in which most tumor cell nuclei were positive for SLFN11 (SLFN11: 95%). Right, an example of an SLFN11-negative case, in which no SLFN11-positive cells were observed in the tumor area (SLFN11: 0%). Stromal cell staining was not counted. (B) Dot and box-and-whisker plots showing the distribution of the percentage of SLFN11 positivity. Each dot represents the value for an individual case, whose primary site is indicated on the right. The highest and lowest boundaries of the box represent the 25^th and 75^th percentiles, respectively, and the whiskers above and below the box designate the maximum and minimum values, respectively. The line within the box indicates the median value.

Figure 2

Progression-free survival (PFS) in patients with SLFN11-positive HNSCC. PFS curves generated by the Kaplan–Meier method are shown for all 161 patients in (A) and those with tumors arising from each primary site are indicated at the top of the panel in (B–D). Intra-arterial and intravenous cisplatin were analyzed separately in (E, F). The difference between groups was calculated using the log-rank test.

Figure 3

Percentage of SLFN11 positivity in tumor cells before and after CRT. SLFN11 expression in tumors before CRT and in residual or recurrent tumors with local failure after CRT is plotted for 10 paired samples. The dashed line represents the threshold value (15%) used for defining SLFN11-positive samples. Data were analyzed using the Wilcoxon signed-rank test.

Figure 4

HSC-3 cells deficient in SLFN11 display reduced sensitivity to cisplatin, carboplatin, and olaparib. (A) SLFN11-KO cells from HSC-3 cells (HSC-3-KO) were generated using the CRISPR/Cas9 method. Cell lysates prepared from these cells were subjected to immunoblot analysis with an antibody for SLFN11 and β-actin (loading control). (B) HSC-3-KO and parental HSC-3 cells were exposed to cisplatin and carboplatin for 48 h, or olaparib and docetaxel for 72 h at the indicated concentrations. Cell viability was determined using a Cell Counting Kit-8 assay and the values were normalized to those of untreated cells. Data are presented as the mean ± standard error of the mean from three independent experiments.