Heat shock protein 70 and tumor-infiltrating NK cells as prognostic indicators for patients with squamous cell carcinoma of the head and neck after radiochemotherapy: A multicentre retrospective study of the German Cancer Consortium Radiation Oncology Gro

Tumor cells frequently overexpress heat shock protein 70 (Hsp70) and present it on their cell surface, where it can be recognized by pre-activated NK cells. In our retrospective study the expression of Hsp70 was determined in relation to tumor-infiltrating CD56+ NK cells in formalin-fixed paraffin embedded (FFPE) tumor specimens of patients with SCCHN (N = 145) as potential indicators for survival and disease recurrence. All patients received radical surgery and postoperative cisplatin-based radiochemotherapy (RCT). In general, Hsp70 expression was stronger, but with variable intensities, in tumor compared to normal tissues. Patients with high Hsp70 expressing tumors (scores 3-4) showed significantly decreased overall survival (OS; p = 0.008), local progression-free survival (LPFS; p = 0.034) and distant metastases-free survival (DMFS; p = 0.044), compared to those with low Hsp70 expression (scores 0-2), which remained significant after adjustment for relevant prognostic variables. The adverse prognostic value of a high Hsp70 expression for OS was also observed in patient cohorts with p16- (p = 0.001), p53- (p = 0.0003) and HPV16 DNA-negative (p = 0.001) tumors. The absence or low numbers of tumor-infiltrating CD56+ NK cells also correlated with significantly decreased OS (p = 0.0001), LPFS (p = 0.0009) and DMFS (p = 0.0001). A high Hsp70 expression and low numbers of tumor-infiltrating NK cells have the highest negative predictive value (p = 0.00004). In summary, a strong Hsp70 expression and low numbers of tumor-infiltrating NK cells correlate with unfavorable outcome following surgery and RCT in patients with SCCHN, and thus serve as negative prognostic markers.

Stangl S ，Tontcheva N ，Sievert W ，Shevtsov M ，Niu M ，Schmid TE ，Pigorsch S ，Combs SE ，Haller B ，Balermpas P ，Rödel F ，Rödel C ，Fokas E ，Krause M ，Linge A ，Lohaus F ，Baumann M ，Tinhofer I ，Budach V ，Stuschke M ，Grosu AL ，Abdollahi A ，Debus J ，Belka C ，Maihöfer C ，Mönnich D ，Zips D ，Multhoff G ... -  《-》

CD8+ tumour-infiltrating lymphocytes in relation to HPV status and clinical outcome in patients with head and neck cancer after postoperative chemoradiotherapy: A multicentre study of the German cancer consortium radiation oncology group (DKTK-ROG).

We examined the prognostic value of tumour-infiltrating lymphocytes (TILs) in patients with squamous cell carcinoma of the head and neck (SCCHN) after surgery and postoperative cisplatin-based chemoradiotherapy. FFPE-tissue originating from the surgery of 161 patients treated in 8 DKTK partner sites was immunohistochemically stained for CD3 and CD8. Their expression was correlated with clinicopathological characteristics as well as overall survival (OS), local progression-free survival (LPFS) and distant metastases free-survival (DMFS), also in the context of the HPV16-DNA/p16 status. After a median follow-up of 48 months (range: 4100 months), OS at 4 years was 46.5% for the entire cohort. In multivariate analysis, high CD8 expression was confirmed as an independent prognostic parameter for OS (p = 0.002), LPFS (p = 0.004) and DMFS (p = 0.006), while CD3 expression lacked significance. In multivariate analysis HPV16 DNA positivity was associated with improved OS (p = 0.025) and LPFS (p = 0.013) and p16-positive patients showed improved DMFS (p = 0.008). Interestingly, high CD8 expression was a prognostic parameter for the clinical outcome in both HPV16 DNA-positive and HPV16 DNA-negative patients. Similar findings were observed in the multivariate analysis for the combined HPV16 DNA/p16 status. Altogether, CD8+ TILs constitute an independent prognostic marker in SCCHN patients treated with adjuvant chemoradiotherapy. These data indicate that CD8-positive TILs have antitumour activity and could be used for treatment stratification. Further validation of the prognostic value of CD8+ TILs as a biomarker and its role in the immune response in SCCHN patients after adjuvant chemoradiotherapy is warranted and will be performed in the prospective DKTK-ROG study.

Balermpas P ，Rödel F ，Rödel C ，Krause M ，Linge A ，Lohaus F ，Baumann M ，Tinhofer I ，Budach V ，Gkika E ，Stuschke M ，Avlar M ，Grosu AL ，Abdollahi A ，Debus J ，Bayer C ，Stangl S ，Belka C ，Pigorsch S ，Multhoff G ，Combs SE ，Mönnich D ，Zips D ，Fokas E ... -  《-》

Comparison of the composition of lymphocyte subpopulations in non-relapse and relapse patients with squamous cell carcinoma of the head and neck before, during radiochemotherapy and in the follow-up period: a multicenter prospective study of the German Ca

Radiochemotherapy (RCT) has been shown to induce changes in immune cell homeostasis which might affect antitumor immune responses. In the present study, we aimed to compare the composition and kinetics of major lymphocyte subsets in the periphery of patients with non-locoregional recurrent (n = 23) and locoregional recurrent (n = 9) squamous cell carcinoma of the head and neck (SCCHN) upon primary RCT. EDTA-blood of non-locoregional recurrent SCCHN patients was collected before (t0), after application of 20-30 Gy (t1), in the follow-up period 3 (t2) and 6 months (t3) after RCT. In patients with locoregional recurrence blood samples were taken at t0, t1, t2 and at the time of recurrence (t5). EDTA-blood of age-related, healthy volunteers (n = 22) served as a control (Ctrl). Major lymphocyte subpopulations were phenotyped by multiparameter flow cytometry. Patients with non-recurrent SCCHN had significantly lower proportions of CD19+ B cells compared to healthy individuals before start of any therapy (t0) that dropped further until 3 months after RCT (t2), but reached initial levels 6 months after RCT (t3). The proportion of CD3+ T and CD3+/CD4+ T helper cells continuously decreased between t0 and t3, whereas that of CD8+ cytotoxic T cells and CD3+/CD56+ NK-like T cells (NKT) gradually increased in the same period of time in non-recurrent patients. The percentage of CD4+/CD25+/FoxP3+ regulatory T cells (Tregs) decreased directly after RCT, but increased above initial levels in the follow-up period 3 (t2) and 6 (t3) months after RCT. Patients with locoregional recurrence showed similar trends with respect to B, T cells and Tregs between t0 and t5. CD4+ T helper cells remained stably low between t0 and t5 in patients with locoregional recurrence compared to Ctrl. NKT/NK cell subsets (CD56+/CD69+, CD3-/CD56+, CD3-/CD94+, CD3-/NKG2D+, CD3-/NKp30+, CD3-/NKp46+) increased continuously up to 6 months after RCT (t0-t3) in patients without locoregional recurrence, whereas in patients with locoregional recurrence, these subsets remained stably low until time of recurrence (t5). Monitoring the kinetics of lymphocyte subpopulations especially activatory NK cells before and after RCT might provide a clue with respect to the development of an early locoregional recurrence in patients with SCCHN. However, studies with larger patient cohorts are needed. Observational Study on Biomarkers in Head and Neck Cancer (HNprädBio), NCT02059668. Registered on 11 February 2014, https://clinicaltrials.gov/ct2/show/NCT02059668 .

Niu M ，Combs SE ，Linge A ，Krause M ，Baumann M ，Lohaus F ，Ebert N ，Tinhofer I ，Budach V ，von der Grün J ，Rödel F ，Grosu AL ，Multhoff G ... -  《Radiation Oncology》

The PD-1/PD-L1 axis and human papilloma virus in patients with head and neck cancer after adjuvant chemoradiotherapy: A multicentre study of the German Cancer Consortium Radiation Oncology Group (DKTK-ROG).

We examined the prognostic role of PD-1+ and CD8+ tumor infiltrating lymphocytes (TILs), and PD-L1+ cells in patients with squamous cell carcinoma of the head and neck (SCCHN) treated with surgery and postoperative chemoradiotherapy (CRT). FFPE samples from 161 patients were immunohistochemically stained for PD-1, CD8 and PD-L1. The immune marker expression was correlated with clinicopathologic characteristics, and overall survival (OS), local progression-free survival (LPFS) and distant metastases free-survival (DMFS), also in the context of HPV16 DNA/p16 status. The median follow-up was 48 months (range: 4-100). The 2-year-OS was 84.1% for the entire cohort. High PD-1 and PD-L1 expression were more common in patients with positive HPV16 DNA (p < 0.001 and p = 0.008, respectively) and high infiltration by CD8+ TILs (p < 0.001 for both markers). High PD-L1 expression correlated with superior OS (p = 0.025), LPFS (p = 0.047) and DMFS (p = 0.048) in multivariable analysis, whereas no significance could be demonstrated for PD-1. Patients with CD8high /PD-L1high expression had favorable outcome (p < 0.001 for all endpoints) compared to other groups. We validated the superior OS data on CD8high /PD-L1high using the Cancer Genome Atlas TCGA dataset (n = 518; p = 0.032). High PD-L1 expression was a favorable prognostic marker in HPV16-negative but not HPV16-positive patients. In conclusion, HPV-positive tumors showed higher expression of immune markers. PD-L1 expression constitutes an independent prognostic marker in SCCHN patients post-adjuvant CRT. In conjunction with CD8 status, these data provide an important insight on the immune contexture of SCCHN and are directly relevant for future treatment stratification with PD-1/PD-L1 immune checkpoint inhibitors to complement CRT.

Balermpas P ，Rödel F ，Krause M ，Linge A ，Lohaus F ，Baumann M ，Tinhofer I ，Budach V ，Sak A ，Stuschke M ，Gkika E ，Grosu AL ，Abdollahi A ，Debus J ，Stangl S ，Ganswindt U ，Belka C ，Pigorsch S ，Multhoff G ，Combs SE ，Welz S ，Zips D ，Lim SY ，Rödel C ，Fokas E ，DKTK-ROG ... -  《-》

HPV16 DNA status is a strong prognosticator of loco-regional control after postoperative radiochemotherapy of locally advanced oropharyngeal carcinoma: results from a multicentre explorative study of the German Cancer Consortium Radiation Oncology Group (

To investigate the impact of HPV status in patients with locally advanced head and neck squamous cell carcinoma (HNSCC), who received surgery and cisplatin-based postoperative radiochemotherapy. For 221 patients with locally advanced squamous cell carcinoma of the hypopharynx, oropharynx or oral cavity treated at the 8 partner sites of the German Cancer Consortium, the impact of HPV DNA, p16 overexpression and p53 expression on outcome were retrospectively analysed. The primary endpoint was loco-regional tumour control; secondary endpoints were distant metastases and overall survival. In the total patient population, univariate analyses revealed a significant impact of HPV16 DNA positivity, p16 overexpression, p53 positivity and tumour site on loco-regional tumour control. Multivariate analysis stratified for tumour site showed that positive HPV 16 DNA status correlated with loco-regional tumour control in patients with oropharyngeal carcinoma (p=0.02) but not in the oral cavity carcinoma group. Multivariate evaluation of the secondary endpoints in the total population revealed a significant association of HPV16 DNA positivity with overall survival (p<0.01) but not with distant metastases. HPV16 DNA status appears to be a strong prognosticator of loco-regional tumour control after postoperative cisplatin-based radiochemotherapy of locally advanced oropharyngeal carcinoma and is now being explored in a prospective validation trial.

Lohaus F ，Linge A ，Tinhofer I ，Budach V ，Gkika E ，Stuschke M ，Balermpas P ，Rödel C ，Avlar M ，Grosu AL ，Abdollahi A ，Debus J ，Bayer C ，Belka C ，Pigorsch S ，Combs SE ，Mönnich D ，Zips D ，von Neubeck C ，Baretton GB ，Löck S ，Thames HD ，Krause M ，Baumann M ，DKTK-ROG ... -  《-》