Expression of DNA translesion synthesis polymerase η in head and neck squamous cell cancer predicts resistance to gemcitabine and cisplatin-based chemotherapy

Abstract

Purpose: The development of resistance against anticancer drugs has been a persistent clinical problem for the treatment of locally advanced malignancies in the head and neck mucosal derived squamous cell carcinoma (HNSCC). Recent evidence indicates that the DNA translesion synthesis (TLS) polymerase η (Pol η; hRad30a gene) reduces the effectiveness of gemcitabine/cisplatin. The goal of this study is to examine the relationship between the expression level of Pol η and the observed resistance against these chemotherapeutic agents in HNSCC, which is currently unknown.

Methods: Sixty-four mucosal derived squamous cell carcinomas of head and neck (HNSCC) from 1989 and 2007 at the City of Hope National Medical Center (Duarte, CA) were retrospectively analyzed. Pretreatment samples were immunostained with anti-Pol η antibody and the correlation between the expression level of Pol η and clinical outcomes were evaluated. Forty-nine cases treated with platinum (n=40) or gemcitabine (n=9) based chemotherapy were further examined for Pol η expression level for comparison with patient response to chemotherapy.

Results: The expression of Pol η was elevated in 67% of the head and neck tumor samples. Pol η expression level was significantly higher in grade 1 to grade 2 tumors (well to moderately differentiated). The overall benefit rate (complete response+ partial response) in patients treated with platinum and gemcitabine based chemotherapy was 79.5%, where low Pol η level was significantly associated with high complete response rate (p=0.03), although not associated with overall survival. Furthermore, no significant correlation was observed between Pol η expression level with gender, age, tobacco/alcohol history, tumor stage and metastatic status.

Conclusions: Our data suggest that Pol η expression may be a useful prediction marker for the effectiveness of platinum or gemcitabine based therapy for HNSCC.

Figure 1. Generation and validation of anti-Pol η antibody.

A) The N-terminal fragment of Pol η (highlighted in grey) used for the generation of an anti-Pol η antibody. B) The developed antibody used for the detection of Pol η in the lysates derived from GM637 or XP30RO cells. C) Lane 1. Identification of Pol η from the lysate from human HCT116 cells. Lane 2. Blocking with purified recombinant human Pol η protein. D) 1-3, GM 637 cells probed with secondary antibody alone (D2 and D3). 4-6, GM637 cells probed with anti-Pol η antibody (D5) or anti-PCNA antibody (D6). 7-9, XP30RO cells probed with anti-Pol η antibody (D8) or anti-PCNA antibody (D9).

Figure 2. The expression of Pol η protein in head and neck squamous cell cancer.

A) Nuclear staining of Pol η in normal squamous epithelium as control. B) Comparison of Pol η staining level between HNSCC and para-cancer normal squamous mucosa in oral cavity. C-E) HNSCC nuclear Pol η expression was classified as negative (C), low (D) and high, according to the frequency and intensity of the stained cells. F-G) Comparison of Pol η staining in primary cancer in soft palate (F) and metastatic cancer cells to lymph node (G). (Immunoperoxidase, original magnification: ×100 (A-B), ×200 (C-G)).

Figure 3. Platinum-based chemotherapy and Pol η protein expression in head and neck squamous cell cancer.

Upper row represents nuclear staining of Pol η in HNSCC before chemotherapy. Lower row represents Pol η staining in tissue obtained from the same areas of original tumors after chemotherapy. Case 1) Pol η staining level up-regulated post chemotherapy. Case 2) Pol η staining level down-regulated post treatment. Case 3-4) Pol η staining level remained same intensity post treatment (immunoperoxidase, original magnification x200).

Figure 4. Kaplan-Meier survival probability comparing tumors with high Pol η expression to those with low Pol η expression.

The blue curve corresponds to patients with high Pol η level and the red curve to patients with low Pol η level.