A comprehensive and longitudinal evaluation of the different populations of lymphoid and myeloid cells in the peripheral blood of patients treated with chemoradiotherapy for head and neck cancer

BACKGROUND Immunotherapy provided significant survival benefits for recurrent and metastatic patients with head and neck cancer. These improvements could not be reproduced in patients treated with curative-intent chemoradiotherapy (CRT) and the optimal radio-immunotherapy (RIT) concepts have yet to be designed. Exploration and analysis of the pre-therapeutic immune status of these patients and the changes occurring during the treatment course could be crucial in rationally designing future combined treatments. METHODS Blood samples were collected from a cohort of 25 head and neck cancer patients treated with curative-intended (C)-RT prior to therapy, after the first week of treatment, and three months after treatment completion. Peripheral blood mononuclear cells (PBMCs) or all nucleated blood cells were isolated and analyzed via flow cytometry. RESULTS At baseline, patients showed reduced monocyte and lymphocyte counts compared to healthy individuals. Although overall CD8$^{+}$ T-cell frequencies were reduced, the proportion of memory subsets were increased in patients. Radiotherapy (RT) treatment led to a further increase in CD8$^{+}$ effector memory T-cells. Among myeloid populations, tumor-promoting subsets became less abundant after RT, in favor of pro-inflammatory cells. CONCLUSION The present study prospectively demonstrated a complex interplay and distinct longitudinal changes in the composition of lymphocytic and myeloid populations during curative (C)-RT of head and neck cancer. Further validation of this method in a larger cohort could allow for better treatment guidance and tailored incorporation of immunotherapies (IT) in the future

Neoadjuvant Chemoradiotherapy for Oral Cavity Cancer: Predictive Factors for Response and Interim Analysis of the Prospective INVERT-Trial

Background To study neoadjuvant chemoradiotherapy (nCRT) and potential predictive factors for response in locally advanced oral cavity cancer (LA-OCC). Methods The INVERT trial is an ongoing single-center, prospective phase 2, proof-of-principle trial. Operable patients with stage III-IVA squamous cell carcinomas of the oral cavity were eligible and received nCRT consisting of 60 Gy with concomitant cisplatin and 5-fluorouracil. Surgery was scheduled 6-8 weeks after completion of nCRT. Explorative, multiplex immunohistochemistry (IHC) was performed on pretreatment tumor specimen, and diffusion-weighted magnetic resonance imaging (DW-MRI) was conducted prior to, during nCRT (day 15), and before surgery to identify potential predictive biomarkers and imaging features. Primary endpoint was the pathological complete response (pCR) rate. Results Seventeen patients with stage IVA OCC were included in this interim analysis. All patients completed nCRT. One patient died from pneumonia 10 weeks after nCRT before surgery. Complete tumor resection (R0) was achieved in 16/17 patients, of whom 7 (41%, 95% CI: 18-67%) showed pCR. According to the Clavien-Dindo classification, grade 3a and 3b complications were found in 4 (25%) and 5 (31%) patients, respectively; grade 4-5 complications did not occur. Increased changes in the apparent diffusion coefficient signal intensities between MRI at day 15 of nCRT and before surgery were associated with better response (p=0.022). Higher abundances of programmed cell death protein 1 (PD1) positive cytotoxic T-cells (p=0.012), PD1+ macrophages (p=0.046), and cancer-associated fibroblasts (CAFs, p=0.036) were associated with incomplete response to nCRT. Conclusion nCRT for LA-OCC followed by radical surgery is feasible and shows high response rates. Larger patient cohorts from randomized trials are needed to further investigate nCRT and predictive biomarkers such as changes in DW-MRI signal intensities, tumor infiltrating immune cells, and CAFs

Association between spatial distribution of leukocyte subsets and clinical presentation of head and neck squamous cell carcinoma

BackgroundInteractions between tumor cells and cells in the microenvironment contribute to tumor development and metastasis. The spatial arrangement of individual cells in relation to each other influences the likelihood of whether and how these cells interact with each other.MethodsThis study investigated the effect of spatial distribution on the function of leukocyte subsets in the microenvironment of human head and neck squamous cell carcinoma (HNSCC) using multiplex immunohistochemistry (IHC). Leukocyte subsets were further classified based on analysis of two previously published HNSCC single-cell RNA datasets and flow cytometry (FC).ResultsIHC revealed distinct distribution patterns of leukocytes differentiated by CD68 and CD163. While CD68hiCD163lo and CD68hiCD163hi cells accumulated near tumor sites, CD68loCD163hi cells were more evenly distributed in the tumor stroma. PD-L1hi and PD-1hi cells accumulated predominantly around tumor sites. High cell density of PD-L1hi CD68hiCD163hi cells or PD-1hi T cells near the tumor site correlated with improved survival. FC and single cell RNA revealed high variability within the CD68/CD163 subsets. CD68hiCD163lo and CD68hiCD163hi cells were predominantly macrophages (MΦ), whereas CD68loCD163hi cells appeared to be predominantly dendritic cells (DCs). Differentiation based on CD64, CD80, CD163, and CD206 revealed that TAM in HNSCC occupy a broad spectrum within the classical M1/M2 polarization. Notably, the MΦ subsets expressed predominantly CD206 and little CD80. The opposite was observed in the DC subsets.ConclusionThe distribution patterns and their distinct interactions via the PD-L1/PD-1 pathway suggest divergent roles of CD68/CD163 subsets in the HNSCC microenvironment. PD-L1/PD-1 interactions appear to occur primarily between specific cell types close to the tumor site. Whether PD-L1/PD-1 interactions have a positive or negative impact on patient survival appears to depend on both the spatial localization and the entity of the interacting cells. Co-expression of other markers, particularly CD80 and CD206, supports the hypothesis that CD68/CD163 IHC subsets have distinct functions. These results highlight the association between spatial leukocyte distribution patterns and the clinical presentation of HNSCC

Association between spatial distribution of leukocyte subsets and clinical presentation of head and neck squamous cell carcinoma

Background: Interactions between tumor cells and cells in the microenvironment contribute to tumor development and metastasis. The spatial arrangement of individual cells in relation to each other influences the likelihood of whether and how these cells interact with each other.MethodsThis study investigated the effect of spatial distribution on the function of leukocyte subsets in the microenvironment of human head and neck squamous cell carcinoma (HNSCC) using multiplex immunohistochemistry (IHC). Leukocyte subsets were further classified based on analysis of two previously published HNSCC single-cell RNA datasets and flow cytometry (FC). Results: IHC revealed distinct distribution patterns of leukocytes differentiated by CD68 and CD163. While CD68hiCD163$^{lo}$ and CD68$^{hi}$CD163$^{hi}$ cells accumulated near tumor sites, CD68$^{lo}$CD163$^{hi}$ cells were more evenly distributed in the tumor stroma. PD-L1$^{hi}$ and PD-1$^{hi}$ cells accumulated predominantly around tumor sites. High cell density of PD-L1$^{hi}$ CD68$^{hi}$CD163$^{hi}$ cells or PD-1$^{hi}$ T cells near the tumor site correlated with improved survival. FC and single cell RNA revealed high variability within the CD68/CD163 subsets. CD68$^{hi}$CD163$^{lo}$ and CD68$^{hi}$CD163$^{hi}$ cells were predominantly macrophages (MΦ), whereas CD68$^{lo}$CD163$^{hi}$ cells appeared to be predominantly dendritic cells (DCs). Differentiation based on CD64, CD80, CD163, and CD206 revealed that TAM in HNSCC occupy a broad spectrum within the classical M1/M2 polarization. Notably, the MΦ subsets expressed predominantly CD206 and little CD80. The opposite was observed in the DC subsets. Conclusion: The distribution patterns and their distinct interactions via the PD-L1/PD-1 pathway suggest divergent roles of CD68/CD163 subsets in the HNSCC microenvironment. PD-L1/PD-1 interactions appear to occur primarily between specific cell types close to the tumor site. Whether PD-L1/PD-1 interactions have a positive or negative impact on patient survival appears to depend on both the spatial localization and the entity of the interacting cells. Co-expression of other markers, particularly CD80 and CD206, supports the hypothesis that CD68/CD163 IHC subsets have distinct functions. These results highlight the association between spatial leukocyte distribution patterns and the clinical presentation of HNSCC

Enabling ultra-high dose rate electron beams at a clinical linear accelerator for isocentric treatments

BACKGROUND AND PURPOSE Radiotherapy delivery with ultra-high dose rates (UHDR) has consistently produced normal tissue sparing while maintaining efficacy for tumour control in preclinical studies, known as the FLASH effect. Modified clinical electron linacs have been used for pre-clinical studies at reduced source-surface distance (SSD) and novel intra-operative devices are becoming available. In this context, we modified a clinical linac to deliver 16 MeV UHDR electron beams with an isocentric setup. MATERIALS AND METHODS The first Varian TrueBeam (SN 1001) was clinically operative between 2009-2022, it was then decommissioned and converted into a research platform. The 18 MeV electron beam was converted into the experimental 16 MeV UHDR. Modifications were performed by Varian and included a software patch, thinner scattering foil and beam tuning. The dose rate, beam characteristics and reproducibility were measured with electron applicators at SSD = 100 cm. RESULTS The dose per pulse at isocenter was up to 1.28 Gy/pulse, corresponding to average and instantaneous dose rates up to 256 Gy/s and 3⋅10$^{5}$ Gy/s, respectively. Beam characteristics were equivalent between 16 MeV UHDR and conventional for field sizes up to 10x10cm$^{2}$ and an overall beam reproducibility within ± 2.5% was measured. CONCLUSIONS We report on the first technical conversion of a Varian TrueBeam to produce 16 MeV UHDR electron beams. This research platform will allow isocenter experiments and deliveries with conventional setups up to field sizes of 10x10 cm$^{2}$ within a hospital environment, reducing the gap between preclinical and clinical electron FLASH investigations

Patient recruitment into clinical studies of solid malignancies during the COVID-19 pandemic in a tertiary cancer center

Background and purpose: To analyze clinical trial activities and patient recruitment numbers into prospective clinical studies for solid malignancies during the COVID-19 pandemic in a tertiary cancer center. Materials and methods: Patient recruitment numbers in prospective clinical studies of solid malignancies were retrospectively analyzed for the years 2019 – 2021 at the Comprehensive Cancer Center Zurich (CCCZ). Changes in recruitment numbers were tested for association with organ-specific subunits, as well as organizational and treatment-related trial characteristics. To assess differences between categorical variables, Chi-squared test was used. For uni- and multivariate analysis, Cox proportional hazards were calculated. Results: In 2019, there were a total of 107 studies (registry trials, clinical phase I-III trials, and translational studies) recruiting 304 patients at the CCCZ. During the COVID-19 pandemic in 2020 and 2021, there were 120 and 125 active trials with a total recruitment of 355 and 666 patients, respectively. No significant differences between the subunits and study characteristics in changes of patient recruitment in clinical phase I-III trials were identified when the year prior to the COVID-19 pandemic (2019) was compared to the first year of the pandemic (2020) and to 2020-2021. Conclusions: Despite healthcare systems around the world have experienced significant disruption due to the COVID-19 pandemic, data from our tertiary cancer center showed that clinical trial activities were maintained at a high level during the pandemic

Potential of ChatGPT in facilitating research in radiation oncology?

PURPOSE To evaluate the potential of the artificial intelligence (AI) chatbot ChatGPT in supporting young clinical scientists with scientific tasks in radio oncological research. MATERIALS AND METHODS Seven scientific tasks were to be completed in 3 hours by 8 radiation oncologists with different scientific experience working at a university hospital: creation of a scientific synopsis, creation of a research question and corresponding clinical trial hypotheses, writing of the first paragraph of a manuscript introduction, clinical trial sample size calculation, and clinical data analyses (multivariate analysis, boxplot and survival curve). No participant had prior experience with an AI chatbot. All participants were instructed in ChatGPT v3.5 and its use was provided for all tasks. Answers were scored independently by two blinded experts. The subjective value of ChatGPT was rated by each participant. Data were analyzed with regression-, t-test and Spearman correlation (p<0.05). RESULTS Participants completed tasks 1-3 with an average score of 50% and 4-7 with 56%. Scientific experience, number of original publications and of first/last authorships showed a positive correlation with overall scoring (p=0.01-0.04). Participants with little to moderate scientific experience scored ChatGPT to be more helpful in solving tasks 4-7 compared to more experienced participants (p=0.04), with simultaneously presenting lower scorings (p=0.03). CONCLUSIONS ChatGPT did not compensate for differences in scientific experience of young clinical scientists, with less experienced researchers believing false AI-generated scientific results

Definition and validation of a radiomics signature for loco-regional tumour control in patients with locally advanced head and neck squamous cell carcinoma

Purpose: To develop and validate a CT-based radiomics signature for the prognosis of loco-regional tumour control (LRC) in patients with locally advanced head and neck squamous cell carcinoma (HNSCC) treated by primary radiochemotherapy (RCTx) based on retrospective data from 6 partner sites of the German Cancer Consortium - Radiation Oncology Group (DKTK-ROG). Material and methods: Pre-treatment CT images of 318 patients with locally advanced HNSCC were col-lected. Four-hundred forty-six features were extracted from each primary tumour volume and then fil-tered through stability analysis and clustering. First, a baseline signature was developed from demographic and tumour-associated clinical parameters. This signature was then supplemented by CT imaging features. A final signature was derived using repeated 3-fold cross-validation on the discovery cohort. Performance in external validation was assessed by the concordance index (C-Index). Furthermore, calibration and patient stratification in groups with low and high risk for loco-regional recurrence were analysed. Results: For the clinical baseline signature, only the primary tumour volume was selected. The final sig-nature combined the tumour volume with two independent radiomics features. It achieved moderatel

De-intensification of postoperative radiotherapy in head and neck cancer irrespective of human papillomavirus status—results of a prospective multicenter phase II trial (DIREKHT Trial)

BackgroundCurrent standard treatment concepts in head and neck squamous cell carcinoma (HNSCC) are based on former studies using 2D and 3D treatment plans. However, modern radiation techniques allow for a more precise and individual dose application. Therefore, in a clearly defined patient population, de-intensified risk-adapted radiation is investigated.MethodsPatients with newly diagnosed HNSCC after surgery (with resection margins ≥1 mm and cM0) with the following tumor stages (TNM 7th Edition) were eligible for the study: oral cavity, oropharynx, or larynx: pT1–3, pN0–pN2b; hypopharynx: pT1–2, pN1. The patients should have either a low risk of local recurrence [≤pT2, resection margin ≥5 mm, no peritumoral lymphangiosis (L0), and no perineural invasion] or contralateral lymph node metastasis (≤3 ipsilateral lymph node metastases, in case of well-lateralized oropharyngeal or oral cavity cancer contralateral cN0, otherwise pN0). Patients were assigned to three different treatment regimes with reduction of the treated volume, radiation dose, or both, according to tumor stage and results of surgery performed. The primary objective was to show an LRR of &lt;10% after 2 years.FindingsA total of 150 patients were enrolled. Tumor localizations were as follows: n = 53 (35.3%), oral cavity; n = 94 (62.7%), oropharynx (82% HPV-positive); n = 2 (1.3%), hypopharynx; and n = 1 (0.7%), larynx. A total of 61 patients (41.0%) were stage IVA, 81 (54.0%) were stage III, and 8 (5.3%) were stage II. Median follow-up was 36 months. Cumulative incidence of 2y-LRR was 5.6% (95% CI: 1.7%–9.2%) in the whole study population and 14.1% (95% CI: 3.8%–23.2%) in patients with oral cavity cancer. Cumulative incidence of 2y-LRR in non-irradiated or dose-reduced regions was 3.5% (95% CI: 0.4%–6.5%). After 2 years, disease-free survival was 92% (95% CI: 87%–96%) and overall survival was 94% (95% CI: 90%–98%) for the complete study cohort. Acute III° toxicity was as follows: dysphagia, 30%; xerostomia, 7%; mucositis, 19%; and dermatitis, 4%. Dysphagia and xerostomia decrease over time. After 27 months, late dysphagia III° and xerostomia II° were 1% and 9%, respectively.InterpretationThe study met its primary objective. De-intensification of postoperative radiotherapy irrespective of HPV status in a predefined patient population is associated with a favorable toxicity profile without compromising LRR. In an unplanned subgroup analysis, a significantly increased risk of LRR was observed in patients with oral cavity cancer. In these patients, de-intensified radiotherapy should be applied with caution

Merkel Cell Polyoma Viral Load and Intratumoral CD8+ Lymphocyte Infiltration Predict Overall Survival in Patients With Merkel Cell Carcinoma

Introduction: Merkel cell carcinoma (MCC) is linked to the presence of clonally integrated Merkel cell polyomavirus (MCPyV) in up to 80% of the cases. The aim of the study was to determine the prognostic value of baseline MCPyV viral load and lymphocytic infiltration.Methods: MCPyV DNA prevalence, integration status and viral load were determined by specific quantitative real-time PCR in surgical specimens obtained from 49 patients with MCC treated with (n = 22, 45%) or without postoperative radiotherapy (RT). CD8+ tumor infiltrating lymphocytes (TILs) and programmed death ligand 1 (PD-L1) status were assessed using immunohistochemistry. MCPyV characteristics and immune marker expression were correlated with clinicopathological factors and overall survival (OS).Results: Median age at diagnosis was 74 (range, 42–100); 51% of the patients were female. One-, three, and five-year OS rates were 83.8, 58.6, and 47.1%, respectively. A positive MCPyV status was associated with female gender (p = 0.042). Tumor localization (head/arms vs. trunk) positively correlated with PD-L1 status (p = 0.011) and combined CD8/PD-L1 expression (p = 0.038). Overall CD8+ infiltration was inversely associated with N-stage (p = 0.048). Stromal TILs correlated significantly with both PD-L1 expression (p = 0.010) and N-stage (p = 0.037). A high viral load (&gt;median) was significantly associated with worse OS (p = 0.029) and high intratumoral CD8+ infiltration with improved OS for the entire cohort (p = 0.045).Conclusion: These data provide important insight on the role of MCPy DNA viral load and TILs in the context of PD-L1 in patients with Merkel cell carcinoma. Future clinical studies should aim to explore the effect of PD-1/PD-L1 immune-checkpoint inhibitors in combination with existing radiotherapy approaches