PD-1/CD80+ small extracellular vesicles from immunocytes induce cold tumours featured with enhanced adaptive immunosuppression.

Only a minority of cancer patients benefit from immune checkpoint blockade therapy. Sophisticated cross-talk among different immune checkpoint pathways as well as interaction pattern of immune checkpoint molecules carried on circulating small extracellular vesicles (sEV) might contribute to the low response rate. Here we demonstrate that PD-1 and CD80 carried on immunocyte-derived sEVs (I-sEV) induce an adaptive redistribution of PD-L1 in tumour cells. The resulting decreased cell membrane PD-L1 expression and increased sEV PD-L1 secretion into the circulation contribute to systemic immunosuppression. PD-1/CD80+ I-sEVs also induce downregulation of adhesion- and antigen presentation-related molecules on tumour cells and impaired immune cell infiltration, thereby converting tumours to an immunologically cold phenotype. Moreover, synchronous analysis of multiple checkpoint molecules, including PD-1, CD80 and PD-L1, on circulating sEVs distinguishes clinical responders from those patients who poorly respond to anti-PD-1 treatment. Altogether, our study shows that sEVs carry multiple inhibitory immune checkpoints proteins, which form a potentially targetable adaptive loop to suppress antitumour immunity.

Quality assurance in a phase III, multicenter, randomized trial of POstmastectomy radioThErapy in Node posiTive breast cancer with or without Internal mAmmary nodaL irradiation (POTENTIAL): a planning benchmark case.

PURPOSE: To report the planning benchmark case results of the POTENTIAL trial-a multicenter, randomized, phase 3 trial-to evaluate the value of internal mammary nodal (IMN) irradiation for patients with high-risk breast cancer. METHODS: All participating institutions were provided the outlines of one benchmark case, and they generated radiation therapy plans per protocol. The plans were evaluated by a quality assurance team, after which the institutions resubmitted their revised plans. The information on beams arrangement, skin flash, inhomogeneity corrections, and protocol compliance was assessed in the first and final submission. RESULTS: The plans from 26 institutions were analyzed. Some major deviations were found in the first submission. The protocol compliance rates of dose coverage for the planning target volume of chest wall, supraclavicular fossa plus axilla, and IMN region (PTVim) were all significantly improved in the final submission, which were 96.2% vs. 69.2%, 100% vs. 76.9%, and 88.4% vs. 53.8%, respectively. For OARs, the compliance rates of heart Dmean, left anterior descending coronary artery V40Gy, ipsilateral lung V5Gy, and stomach V5Gy were significantly improved. In the first and final submission, the mean values of PTVim V100% were 79.9% vs. 92.7%; the mean values of heart Dmean were 11.5 Gy vs. 9.7 Gy for hypofractionated radiation therapy and 11.5 Gy vs. 11.0 Gy for conventional fractionated radiation therapy, respectively. CONCLUSION: The major deviations were corrected and protocol compliance was significantly improved after revision, which highlighted the importance of planning benchmark case to guarantee the planning quality for multicenter trials.

Characteristics and management of tumor treating fields-related dermatological complications in patients with glioblastoma.

Tumor treating fields (TTFields) is a novel approved modality for the treatment of glioblastoma (GBM) exhibiting a satisfactory effect. Although TTFields has shown considerable safety for the normal brain, dermatological adverse events (DAEs) often occur during therapy. However, studies focused on the identification and management of DAEs are rare. The clinical data and photos of skin lesions from 9 patients with GBM were retrospectively analyzed, and the types and grades of individual scalp dermatitis were evaluated based on the Common Terminology Criteria for Adverse Events version 5.0 (CTCAE v 5.0). Adherence and safety were also evaluated on the basis of the device monitoring data. Eight patients (88.9%) exhibited grade 1 or grade 2 CTCAE DAEs, all of whom were cured after interventions. The adherence was >90%, with no relevant safety events reported. Finally, a guideline for preventing DAEs in patients with GBM was proposed. The identification and management of TTFields-related DAEs is necessary and urgent in patients with GBM. Timely interventions of DAEs will help to improve the adherence and quality of life of patients, which ultimately improves prognosis. The proposed guideline for preventing DAEs in patients with GBM assists in the management of healthcare providers and may avoid dermatologic complications.

Dose-Volume Predictors for Radiation Esophagitis in Patients With Breast Cancer Undergoing Hypofractionated Regional Nodal Radiation Therapy.

PURPOSE: Our objective was to assess the incidence and dose-volume predictors of radiation esophagitis (RE) in patients with breast cancer undergoing hypofractionated regional nodal irradiation. METHODS AND MATERIALS: Eligible patients who received intensity modulated radiation therapy (RT) at the chest wall, the supraclavicular/infraclavicular fossa, level II axilla, and/or the internal mammary chain after mastectomy were included. The prescribed dose was 43.5 Gy in 15 fractions. RE was evaluated weekly during RT and at 1 and 2 weeks, followed by 3 and 6 months after RT, and was graded according to National Cancer Institute Common Toxicity Criteria for Adverse Events, version 3.0. The esophagus was contoured from the lower border level of the cricoid cartilage to the lower margin of the aortic arch. Esophageal total volume, mean dose, maximum dose, and the relative volumes (RV) and absolute volumes (AV) receiving at least 5 to 45 Gy by 5-Gy increments (RV5-RV45 and AV5-AV45) were evaluated. Univariable and multivariable logistic regression analyses were performed to determine risk factors for RE, and receiver operating characteristic curves were obtained to identify the thresholds of esophageal dosimetric parameters. RESULTS: In total, 298 patients were included between May 8, 2020, and January 5, 2022 (minimum post-RT follow-up: 6 months). Grade 2 and 3 RE incidence was 40.9% (122/298) and 0.3% (1/298), respectively. No grade 4 or 5 RE was observed. Esophageal RV20-RV40 and AV35-AV40 were significantly associated with the risk of grade ≥2 RE after adjusting for tumor laterality and internal mammary nodal irradiation. RV25 and AV35 were optimum dose-volume predictors for grade ≥2 RE at thresholds 20% for RV25 (35.9% vs 60.9%; P = .04) and 0.27 mL for AV35 (31.0% vs 54.6%; P = .04). CONCLUSIONS: RE is common in patients with breast cancer undergoing hypofractionated regional nodal irradiation. Maintaining the upper esophageal V25 at <20% and V35 at <0.27 mL may decrease the risk of RE.

HRS Regulates Small Extracellular Vesicle PD-L1 Secretion and Is Associated with Anti-PD-1 Treatment Efficacy.

PD-L1 localized to immunosuppressive small extracellular vesicles (sEV PD-L1) contributes to tumor progression and is associated with resistance to immune-checkpoint blockade (ICB) therapy. Here, by establishing a screening strategy with a combination of tissue microarray (TMA), IHC staining, and measurement of circulating sEV PD-L1, we found that the endosomal sorting complex required for transport (ESCRT) member protein hepatocyte growth factor-regulated tyrosine kinase substrate (HRS) was the key regulator of circulating sEV PD-L1 in head and neck squamous cell carcinoma (HNSCC) patients. Increased HRS expression was found in tumor tissues and positively correlated with elevated circulating sEV PD-L1 in patients with HNSCC. The expression of HRS was also negatively correlated to the infiltration of CD8+ T cells. Knockdown of HRS markedly reduced PD-L1 expression in HNSCC cell-derived sEVs, and these sEVs from HRS knockdown cells showed decreased immunosuppressive effects on CD8+ T cells. Knockout of HRS inhibited tumor growth in immunocompetent mice together with PD-1 blockade. Moreover, a higher HRS expression was associated with a lower response rate to anti-PD-1 therapy in patients with HNSCC. In summary, our study reveals HRS, the core component of ESCRT-0, regulates sEV PD-L1 secretion, and is associated with the response to ICB therapy in patients with HNSCC, suggesting HRS is a promising target to improve cancer immunotherapy.

Variability of Target Volumes and Organs at Risk Delineation in Breast Cancer Radiation Therapy: Quality Assurance Results of the Pretrial Benchmark Case for the POTENTIAL Trial.

PURPOSE: To estimate the variations in clinical target volumes (CTVs) and organs at risk delineation within the quality assurance (QA) program of the POTENTIAL trial, which is a multicenter, randomized phase 3 trial evaluating postmastectomy radiation therapy (RT), with or without internal mammary nodal irradiation, for patients with high-risk breast cancer. METHODS AND MATERIALS: The simulating computed tomography scan data set of a benchmark case was sent to the participating centers, and the delineation of CTVs and organs at risk was required to be completed by the investigators following protocol guidelines. All submitted contours were reviewed and compared with the reference contours created by the QA team, using quantitative geometric analysis regarding volume and the Jaccard Index (JCI), Dice similarity coefficient, Geographic Miss Index, Discordance Index, and mean distance to agreement. In addition to the whole-volume analysis of all structures, the combination contour of the supraclavicular fossa and level III and II axilla (CTVsc + axIII + axII) was further analyzed on a slice-by-slice basis. RESULTS: The contours from 26 centers were reviewed and variations were observed between submission and reference. The variations of the CTV of the chest wall, contralateral breast, and heart were small, for which the mean JCI values were 0.62, 0.68, and 0.87, respectively. However, the mean JCI values of the CTV of the internal mammary nodal region, ipsilateral brachial plexus, left anterior descending coronary artery, and right coronary artery were 0.38, 0.21, 0.29, and 0.18, respectively, suggesting marked variations. In addition, marked under- and overoutlining variations were identified on 4 slices of CTVsc + axIII + axII on slice-by-slice analysis. CONCLUSIONS: There were residual contouring variations despite a detailed protocol being provided, confirming the importance of pretrial QA in RT and highlighting the need for education and consideration of a real-time central review of the target delineation before the trial participants begin RT.

Altered DNA Methylation and Gene Expression Profiles in Radiation-Induced Heart Fibrosis of Sprague-Dawley Rats.

Radiation-induced heart disease (RIHD) is a serious side effect of radiotherapy for thoracic tumors. Advanced myocardial fibrosis in the late phase of RIHD can lead to myocardial remodeling, heart function impairing and heart failure, resulting in serious clinical consequences, and its pathogenesis remains vague. DNA methylation is one of the important epigenetic mechanisms which often occurs in response to environmental stimuli and is crucial in regulating gene expression. We hypothesized DNA methylation may contribute to pathogenesis in radiation-induced heart fibrosis (RIHF) and altered DNA methylation patterns probably influenced the genes expression in RIHF. In present study, we found genome-wide differences in DNA methylation status and RNA expression were demonstrated and we screened out 44 genes whose altered expression maybe were regulated by CpG island methylation within the gene promoter in RIHF of Sprague-Dawley rat by employing gene expression arrays and human CpG island microarrays. Gene expression and CpG island methylation levels of several candidate genes were further validated. Our investigation provided a new dimension to reveal the specific mechanisms of RIHF and explore the potential therapeutic targets for it.

Patterns of Chest Wall Recurrence and Suggestions on the Clinical Target Volume of Breast Cancer: A Retrospective Analysis of 121 Postmastectomy Patients.

PURPOSE: Radiotherapy is a powerful strategy to prevent chest wall recurrence (CWR) of postmastectomy breast cancer (BC). This retrospective study aims at analyzing patterns of CWR to explore the delineation of clinical target volume. PATIENTS AND METHODS: Detailed clinicopathological information of postmastectomy BC patients with CWR was collected from our single cancer center based on clear criteria. To describe recurrent positions more accurately, the chest wall was divided into three layers: skin layer (skin and subcutaneous tissues), pectoralis layer (pectoralis major and minor), and rib layer (rib and intercostal muscle). The frequency distribution of recurrence location and its association with clinical pathological factors were analyzed. RESULTS: A total of 121 postmastectomy BC with CWR were included in this study. The percentages of breast tumor located in the upper outer quadrant, upper inner quadrant, lower inner quadrant, lower outer quadrant, overlapping quadrant, and areola area were 31.0% (35/113), 26.5% (30/113), 12.4% (14/113), 5.3% (6/113), 21.1% (25/113), and 2.7% (3/113), respectively. HER2-positive BC (51/113, 45.1%) is the most common BC subtype. Analysis on the patterns of CWR showed that recurrences locating in the skin layer, pectoralis layer, rib layer, mixed layers, and incision periphery accounted for 58.6% (68/116), 9.5% (11/116), 1.7% (2/116), 30.2% (35/116), and 60.5% (46/76), respectively. Rates of recurrences located in the skin and/or pectoralis layers for all BC patients, patients with concomitant distance metastasis, and patients without concomitant distance metastasis were 82.8% (96/116), 85.9% (49/57), and 81.0% (47/58), respectively. CONCLUSION: For BC patients receiving mastectomy, skin, subcutaneous tissues, pectoralis, and area around incision have a high risk of recurrence, which should be paid more attention in chest wall radiotherapy.

Prognostic value of mitochondrial DNA content and G10398A polymorphism in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is one of the leading causes of cancer-related mortality worldwide. Mitochondrial dysfunction has been postulated to render cancer cells resistant to apoptosis based on the Warburg hypothesis. However, few studies have investigated the prognostic value of mitochondrial DNA (mtDNA) content and G10398A polymorphism in NSCLC patients. mtDNA copy number and G10398A polymorphism in 128 NSCLC tissue samples were assessed by real-time PCR (RT-PCR) and PCR-RFLP respectively, and their relationship to prognosis were analyzed by survival analysis and Cox proportional hazards model. In vitro, an mtDNA deletion A549 ρ(0) cell model was utilized to assess the function of mtDNA on radiosensitivity. Cell cycle distribution and reactive oxygen species (ROS) were analyzed to elucidate the potential mechanisms. For the whole group, the median follow-up time and overall survival time were 22.5 and 23.4 months, respectively. Patients with high mtDNA content had a marginally longer survival time than patients with low mtDNA content (P=0.053). Moreover, patients with high mtDNA content plus 10398G had a significantly longer overall survival time compared with those having low mtDNA content plus 10398A (47 vs. 27 months, P<0.05). In addition, multivariate analysis showed that stage and low mtDNA content plus 10398A were the two most independent prognostic factors. In vitro, the A549 ρ(0) cells showed more resistance to radiation than ρ(+) cells. Following radiation, ρ(0) cells showed delayed G2 arrest and lower ROS level as compared to ρ(+) cells. In conclusion, the present study suggests that in patients with NSCLC, low mtDNA content plus 10398A could be a marker of poor prognosis which is associated with resistance to anticancer treatment caused by low mtDNA content plus 10398A polymorphism resulting in mitochondrial dysfunction.

Phase II trial of neoadjuvant docetaxel and cisplatin followed by intensity-modulated radiotherapy with concurrent cisplatin in locally advanced nasopharyngeal carcinoma.

PURPOSE: To evaluate the feasibility and efficacy of neoadjuvant chemotherapy involving docetaxel and cisplatin followed by intensity-modulated radiotherapy (IMRT) with concurrent cisplatin in patients with newly diagnosed stage III to IVB nasopharyngeal carcinoma (NPC). METHODS: Docetaxel (75 mg/m(2) on day 1) and cisplatin (75 mg/m(2) on day 1) were administered on a 3-week cycle for 2 courses, followed by radical IMRT (72 Gy/33F/6.5-7 W) with concurrent cisplatin (75 mg/m(2), on day 1) every 3 weeks for 2 cycles. RESULTS: From June 2008 to October 2010, forty-six patients were recruited in this trial. Forty-five patients completed neoadjuvant setting, and all patients completed planned concurrent chemoradiotherapy (CCRT). The complete and partial response rates were 28.3 and 56.5 % after neoadjuvant chemotherapy, and 91.3, 8.7 % after CCRT, respectively. After median follow-up of 26 months (range 12-39 months), one patient experienced local recurrence and 4 patients developed distant metastasis. The 3-year overall survival and progression-free survival rate were 94.1 and 72.7 %, respectively. Neutropenia (37.0 %) and vomiting (28.3 %) were the most common Grade 3-4 adverse effects during neoadjuvant course, while mucositis (30.4 %), xerostomia (30.4 %) and radiodermatitis (21.7 %) were the most common Grades 3 acute toxicities during CCRT. Xerostomia (73.9 %), dysphagia (56.5 %), hear loss (30.4 %) and skin reaction (21.7 %) were the common Grade 1-2 late effects. There were no Grades 3-4 late toxicities. CONCLUSIONS: The protocol of neoadjuvant docetaxel and cisplatin followed by IMRT with concurrent cisplatin was well tolerated, with outstanding compliance and efficacy in locally advanced NPC, which deserved further follow-up.

Cancer in cohort of HIV-infected population: prevalence and clinical characteristics.

OBJECTIVE: To determine the incidence and survival time of acquired immunodeficiency syndrome-related malignancies among HIV-infected population. METHODS: A clinical database search, chart review and verification with health records were undertaken for all AIDS-defining cancers diagnosed in Zhongnan Hospital Wuhan University, Hubei Province, China. Kaplan-Meier method was used to evaluate survival time in HIV-infected patients with cancer. RESULTS: A total of 3,554 patients with 11,072 person-years of HIV follow-up care were reviewed from January 2004 to December 2008. Sixty-three cancer cases were identified. The median ages of HIV-positive cancer cases were 42.4 ± 8.8 years, CD4 count were 220.9 ± 142.3/µl. The common cancers were non-Hodgkin's lymphoma (NHL, 28.6%), cervical cancer (22.2%), liver cancer (17.5%). Statistically significantly elevated SIRs were observed in NHL (SIR in all = 34.5, 95% CI 11.7-89.9, SIR in males = 45.3, 95% CI 24.7-138.9, females = 12.2, 95% CI 3.9-38.2), invasive cervical cancer (SIR = 68.1, 95% CI 19.2-84.5), liver cancer (SIR = 6.0, 95% CI 2.6-12.2), nasopharyngeal cancer (SIR = 6.2, 95% CI 1.5-44.9), bladder cancer (SIR = 4.9, 95% CI 0.9-22.9), esophageal cancer (SIR = 3.1, 95% CI 0.7-14.3), and stomach cancer (SIR = 2.6, 95% CI 0.6-11.6). All cancers combined showed a statistically significantly elevated SIR of 4.1 (95% CI 2.5-4.6), SIR for all cancers was much higher in female (SIR = 4.8, 95% CI 3.2-7.3) than in male (SIR = 3.1, 95% CI 2.1-4.3). Among HIV-positive patients with cancer, the median survival time was 14.5 ± 3.8 months in NHL group, 28.9 ± 3.6 months in cervix group, 5.1 ± 1.1 months in liver group, and 26.7 ± 6.7 months in other groups. The median survival time in HIV-infected group (23.1 ± 3.5 months) was shorter than that in non-HIV-infected group (43.0 ± 5.1 months), (P < 0.05). CONCLUSIONS: NHL, cervical cancers and liver cancer are common cancers among HIV-infected individuals in Hubei, China. Most malignant diseases that arise in the setting of HIV infection tend to occur at a more advanced stage with shorter survival time.

Telomere length inversely correlates with radiosensitivity in human carcinoma cells with the same tissue background.

A relationship between telomeres and radiosensitivity has been established by several studies based on non-mammalian model systems, mouse models, and few human genetic diseases. However, the relationship has not been proven in human carcinoma cells, which have more clinical significance than these other models. The present study aims to determine whether telomere length is related to radiosensitivity in human carcinoma cells, and to examine the influence of tissue or genetic background. Two HEp-2 larynx squamous carcinoma cell lines, eight hepatocellular carcinoma cell lines, and five breast cancer cell lines were used. Telomere length was determined by terminal restriction fragment (TRF) Southern blot analysis and cell survival was measured by a colony-forming assay. Our results indicated that there was a significant negative correlation of telomere length and radiosensitivity in the same tissue-derived cell lines, with or without the same genetic background. Thus, telomere length may be used as a promising tool to predict the radiosensitivity of human carcinomas.

[Inverse correlation between Snail and E-cadherin expression in carcinoma cell lines and invasive ability in vitro].

OBJECTIVE: Malignant transformation of epithelial cell frequently coincides with loss of E-cadherin. Here we study the expression of Snail and E-cadherin and correlate their expression with cell differentiation and in vitro invasion. METHODS: The expression and localization of Snail and E-cadherin were studied by Northern blot and laser confocal microscopy in two normal cell lines (MDCK, NIH 3T3) and six carcinoma cell lines (A431, MCF-7, MDA-MB-453, HepG2, MDA-MB-435s, MDA-MB-231). Boyden chamber assay was done to detect the invasive ability of cells in vitro. RESULTS: Snail mRNA and protein were detected in fibroblasts NIH 3T3 and poorly differentiated carcinoma cell lines HepG2, MDA-MB-435s and MDA-MB-231. On the contrary, E-cadherin mRNA and protein were detected in normal epithelial cell line MDCK and well differentiated carcinoma cell lines A431 and MDA-MB-453. In MCF-7 cells, Snail and E-cadherin expressions were revealed both at mRNA and protein levels. The cells with higher expression of Snail had stronger ability of invasion than those with lower expression of Snail. CONCLUSION: There is an inverse correlation between Snail and E-cadherin expressions and their expressions are correlated with cell differentiation and tumor invasiveness.

[Effect of transcriptional factor snail on epithelial-mesenchymal transition and tumor metastasis].

BACKGROUND & OBJECTIVE: Transcription factor Snail mediates epithelial-mesenchymal transition (EMT), and is associated with tumor metastasis. This study was designed to observe the enhancive effect of Snail and the reverse effect of antisense-Snail on EMT of tumor cells, and explore the role of Snail in tumor metastasis. METHODS: Snail cDNA was transfected into canine renal epithelial cell line MDCK; antisense-Snail was transfected into human breast cancer cell line MDA-MB231. The expression of epithelial markers E-cadherin, beta-catenin and Cytokeratin 18, mesenchymal marker Fibronectin, metastasis-related marker matrix metalloproteinase-2 (MMP-2), and RhoA were detected by Western blot. The metastatic potential of tumor cells was examined by in vitro cell wound model and Boyden chamber invasion assay. RESULTS: The invasion potential of MDCK cells was enhanced after transfection of Snail. The expression of E-cadherin, beta-catenin, and Cytokeratin 18 was significantly lower in Snail-transfected MDCK cells than in control cells (P < 0.05); the expression of Fibronectin, MMP-2, and RhoA was significantly higher in Snail-transfected cells than in control cells (P < 0.05). Inhibiting the expression of Snail with antisense-Snail in MDA-MB231 cells led to opposite results. CONCLUSION: Snail promotes EMT in normal epithelial cells, and inhibiting the expression of Snail may reverse EMT and suppress tumor metastasis.