Biomarkers associated with pulmonary exacerbations in a randomized trial of nintedanib for radiation pneumonitis.

BACKGROUND AND PURPOSE: Radiation pneumonitis (RP) is a common side effect of thoracic radiotherapy and often has a long course characterized by acute exacerbations and progression to permanent lung fibrosis. There are no validated biomarkers of prognosis in patients diagnosed with RP. MATERIALS AND METHODS: We analyzed a time course of serum chemokines, cytokines, and other proteins from patients with grade 2+ RP in a randomized clinical trial of a steroid taper plus nintedanib, a multiple tyrosine kinase inhibitor, versus placebo plus a steroid taper for the treatment of RP. Weighted gene correlation network analysis (WGCNA) and univariable zero inflated Poisson models were used to identify groups of correlated analytes and their associations with clinical outcomes. RESULTS: Thirty enrolled patients had biomarker data available, and 17 patients had enough analytes tested for network analysis. WGNCA identified ten analytes, including transforming growth factor beta-1 (TGF-ß1), monocyte chemoattractant protein-1 (MCP-1), and platelet-derived growth factor (PDGF), that in aggregate were correlated with the occurrence of pulmonary exacerbations (p = 0.008), the total number of acute pulmonary exacerbations (p = 0.002), and treatment arm (p = 0.036). By univariable analysis, an increase in rate of change of two components of the RP module were associated with an increased incidence rate of pulmonary exacerbations: interleukin 5 (IL-5, incidence rate ratio (IRR) 1.02, 95% CI 1.01-1.04, p = 0.002), and tumor necrosis factor superfamily 12 (TNFSF12, IRR 1.06, CI 1-1.11, p = 0.036). An increased slope of epidermal growth factor (EGF) was associated with a decreased incidence rate of exacerbations (IRR 0.94, CI 0.89-1, p = 0.036). CONCLUSION: We identified a panel of serum biomarkers that showed association with nintedanib treatment and acute pulmonary exacerbations in patients with RP. A confirmatory study will be needed to validate this panel for use as a prognostic tool in patients with RP.

Dosimetry and Biochemical Comparison of Early Radiation-Induced Lung Toxicity in Breast Cancer Patients Treated with 3D-CRT and IMRT: the Role of Serum Interleukin-6 and Pulmonary Surfactant Protein-D.

BACKGROUND: Radiation-induced lung disease is a potentially fatal, dose-limiting toxicity commonly seen after radiotherapy of thoracic malignancies, including breast cancer. AIM: To evaluate and compare the early lung toxicity induced by 3D-CRT and IMRT radiotherapy treatment modalities in breast cancer female patients using biochemical, dosimetry and clinical data. SUBJECTS AND METHODS: this study included 15 normal healthy controls, 15 breast cancer patients treated with IMRT, and 15 breast cancer patients treated with 3D-CRT. One blood sample was obtained from the control group and 3 blood samples were withdrawn from cases before RT, after RT and after 3 months of RT. RESULT: IMRT delivered higher radiation dose to the breast tumor and lower doses to the lung as an organ at risk. There was a non-significant increase in the serum levels of IL-6 before IMRT and 3D-CRT compared with its levels in the control group. There were significant increases in serum levels of IL-6 after RT (IMRT and 3DCRT) compared with its levels before RT. There was a non-significant decrease in the serum levels of IL-6 after 3 months of RT (IMRT and 3D-CRT) compared with its serum levels immediately after RT. There was a non-significant increase in the serum levels of SP-D before RT (IMRT and 3D-CRT) compared with its levels in the control group. There were significant-increases in serum levels of SP-D after RT (IMRT and 3D-CRT) compared with its levels before RT. There was a non-significant decrease in the serum levels of SP-D after 3 months of radiotherapy (IMRT and 3D-CRT) compared with its serum levels immediately after RT. CONCLUSION: serum of levels IL-6 and SP-D can be used to diagnose the occurrence of early lung toxicity due to radiotherapy and the rate of recovery from radiation pneumonitis is apparent in case of IMRT than 3D-CRT.

Acute Radiation-Induced Hematopoietic Depletion Does Not Alter the Onset or Severity of Pneumonitis in Mice.

Survival from partial-body irradiation (PBI) may be limited by the development of the late lung injury response of pneumonitis. Herein we investigated the hypothesis that acute hematopoietic depletion alters the onset and severity of lung disease in a mouse model. To establish depletion, C3H/HeJ mice received 8 Gy PBI with shielding of only the tibiae, ankles and feet. One week after irradiation, blood lymphocyte and neutrophil counts were each significantly reduced (P < 0.04) in these mice compared to levels in untreated controls or in mice receiving 16 Gy to the whole thorax only. All 8 Gy PBI mice survived to the experimental end point of 16 weeks postirradiation. To determine whether the hematopoietic depletion affects lung disease, groups of mice received 8 Gy PBI plus 8 Gy whole-thorax irradiation (total lung dose of 16 Gy) or 16 Gy whole-thorax irradiation only. The weight loss, survival to onset of respiratory distress (P = 0.17) and pneumonitis score (P = 0.96) of mice that received 8 Gy PBI plus 8 Gy whole-thorax irradiation were not significantly different from those of mice receiving 16 Gy whole-thorax irradiation only. Mice in respiratory distress from PBI plus whole-thorax irradiation had significantly reduced (P = 0.02) blood monocyte counts compared to levels in distressed, whole-thorax irradiated mice, and symptomatic pneumonitis was associated with increased blood neutrophil counts (P = 0.04) relative to measures from irradiated, non-distressed mice. In conclusion, survivable acute hematopoietic depletion by partial-body irradiation did not alter the onset or severity of lethal pneumonitis in the C3H/HeJ mouse model.

Identification of miRNA signatures associated with radiation-induced late lung injury in mice.

Acute radiation exposure of the thorax can lead to late serious, and even life-threatening, pulmonary and cardiac damage. Sporadic in nature, late complications tend to be difficult to predict, which prompted this investigation into identifying non-invasive, tissue-specific biomarkers for the early detection of late radiation injury. Levels of circulating microRNA (miRNA) were measured in C3H and C57Bl/6 mice after whole thorax irradiation at doses yielding approximately 70% mortality in 120 or 180 days, respectively (LD70/120 or 180). Within the first two weeks after exposure, weight gain slowed compared to sham treated mice along with a temporary drop in white blood cell counts. 52% of C3H (33 of 64) and 72% of C57Bl/6 (46 of 64) irradiated mice died due to late radiation injury. Lung and heart damage, as assessed by computed tomography (CT) and histology at 150 (C3H mice) and 180 (C57Bl/6 mice) days, correlated well with the appearance of a local, miRNA signature in the lung and heart tissue of irradiated animals, consistent with inherent differences in the C3H and C57Bl/6 strains in their propensity for developing radiation-induced pneumonitis or fibrosis, respectively. Radiation-induced changes in the circulating miRNA profile were most prominent within the first 30 days after exposure and included miRNA known to regulate inflammation and fibrosis. Importantly, early changes in plasma miRNA expression predicted survival with reasonable accuracy (88-92%). The miRNA signature that predicted survival in C3H mice, including miR-34a-5p, -100-5p, and -150-5p, were associated with pro-inflammatory NF-κB-mediated signaling pathways, whereas the signature identified in C57Bl/6 mice (miR-34b-3p, -96-5p, and -802-5p) was associated with TGF-ß/SMAD signaling. This study supports the hypothesis that plasma miRNA profiles could be used to identify individuals at high risk of organ-specific late radiation damage, with applications for radiation oncology clinical practice or in the context of a radiological incident.

Early differential diagnosis model for acute radiation pneumonitis based on multiple parameters.

OBJECTIVE: The present study aimed to construct a diagnosis model for the early differentiation of acute radiation pneumonitis (ARP) and infectious pneumonitis based on multiple parameters. METHODS: The present study included data of 152 patients admitted to the Department of Radiochemotherapy, Tangshan People's Hospital, who developed ARP (91 patients) or infectious pneumonia (IP; 61 patients) after radiotherapy. The radiophysical parameters, imaging characteristics, serological indicators, and other data were collected as independent variables, and ARP was considered as a dependent variable. Logistics univariate analysis and Spearman correlation analysis were used for selecting independent variables. Logistics multivariate analysis was used to fit the variables into the regression model to predict ARP. RESULTS: The univariate analysis showed that the positional relation between lesions and V20 area (PRLV), procalcitonin (PCT), C-reactive protein (CRP), mean lung dose (MLD), and lung volume receiving ≥20 Gy (V20) correlated with ARP while the planning target volume (PTV) dose marginally correlated with ARP. The multivariate analysis showed that the PRLV, PCT, white blood cell (WBC), and MLD were independent diagnostic factors. The nomogram was drawn on the basis of the logistics regression model. The area under the curve (AUC) of the model was 0.849, which was significantly better than that of a single indicator and the sensitivity and specificity of the model were high (82.4 and 82.0%, respectively). These results predicted by the model were highly consistent with the actual diagnostic results. The decision curve analysis (DCA) demonstrated a satisfactory positive net benefit of the model. CONCLUSION: The diagnosis model constructed in the present study is of certain value for the differential diagnosis of ARP and IP.

Early variations in lymphocytes and T lymphocyte subsets are associated with radiation pneumonitis in lung cancer patients and experimental mice received thoracic irradiation.

There were no ideal markers to predict the development of radiation pneumonitis (RP). We want to investigate the value of variations of lymphocytes and T lymphocyte subsets in predicting RP after radiotherapy (RT) of lung cancer based on previous clinical findings. A total of 182 lung cancer patients who received RT were retrospectively analyzed. Circulating lymphocytes and T lymphocyte subsets were measured before, during, and after RT. Patients were evaluated from the start of RT to 6 months post-RT. A mice model with acute radiation-induced lung injury was established and circulating lymphocytes were measured weekly until 8 weeks after irradiation. Univariate and multivariate analyses were adopted to identify risk factors of RP. Lymphocyte levels significantly decreased (P < .001) in patients before RP symptoms developed that also was able to be seen in the mice model and the values recovered during remission of symptoms. The decrease in lymphocyte count reflected the severity of RP. Meanwhile, CD4+  T lymphocyte count was significantly lower during the occurrence of symptoms in patients with RP than in those without RP (P < .001), and it improved along with RP recovery. Levels of lymphocytes and CD4+  T lymphocyte subsets proved as independent predictors of RP. Here we showed that lower peripheral blood levels of lymphocytes and CD4+  T lymphocyte were associated with an increased risk of RP, which was validated by this mice model, and thus are associated with differences in radiation-induced lung toxicity among individuals and help identify those who are susceptible to developing RP after RT.

The role of procalcitonin in differential diagnosis between acute radiation pneumonitis and bacterial pneumonia in lung cancer patients receiving thoracic radiotherapy.

Acute Radiation Pneumonitis (ARP) is one of the most common dose-limiting toxicities of thoracic radiotherapy. The accurate diagnosis of ARP remains a challenge because of the lack of a rapid biomarker capable of differentiating ARP from bacterial pneumo (BP). The aim of this study was to investigate the potential usefulness of procalcitonin (PCT) in the differential diagnosis of ARP and BP. Lung cancer patients who had undergone thoracic radiotherapy within 6 months and were admitted to hospital for ARP or BP were retrospectively analyzed. The serum levels of PCT, C-reactive protein (CRP) and white blood cells (WBC) were compared between the two groups. Receiver operating characteristic (ROC) curve was used to assess the diagnostic value of PCT, CRP and WBC in the differential diagnosis of ARP and BP and determine the best cut-off values. One hundred eighteen patients were included. Among them, seventy-seven patients were diagnosed with ARP, and 41 patients were diagnosed with BP. The PCT concentrations for patients diagnosed with ARP group were significantly lower than those in the BP group (P < 0.001). There were no differences in CRP and WBC between the two groups. The areas under the ROC curves (AUC) for PCT, CRP and WBC were 0.745, 0.589 and 0.578, respectively. The best cutoff values of PCT, CRP and WBC were 0.47 µg/L, 54.5 mg/L and 9.9 × 109/L, respectively. Low serum PCT levels are associated with ARP. PCT is a useful biomarker to distinguish ARP from BP.

Machine Learning to Build and Validate a Model for Radiation Pneumonitis Prediction in Patients with Non-Small Cell Lung Cancer.

PURPOSE: Radiation pneumonitis is an important adverse event in patients with non-small cell lung cancer (NSCLC) receiving thoracic radiotherapy. However, the risk of radiation pneumonitis grade ≥ 2 (RP2) has not been well predicted. This study hypothesized that inflammatory cytokines or the dynamic changes during radiotherapy can improve predictive accuracy for RP2. EXPERIMENTAL DESIGN: Levels of 30 inflammatory cytokines and clinical information in patients with stages I-III NSCLC treated with radiotherapy were from our prospective studies. Statistical analysis was used to select predictive cytokine candidates and clinical covariates for adjustment. Machine learning algorithm was used to develop the generalized linear model for predicting risk RP2. RESULTS: A total of 131 patients were eligible and 17 (13.0%) developed RP2. IL8 and CCL2 had significantly (Bonferroni) lower expression levels in patients with RP2 than without RP2. But none of the changes in cytokine levels during radiotherapy was significantly associated with RP2. The final predictive GLM model for RP2 was established, including IL8 and CCL2 at baseline level and two clinical variables. Nomogram was constructed based on the GLM model. The model's predicting ability was validated in the completely independent test set (AUC = 0.863, accuracy = 80.0%, sensitivity = 100%, specificity = 76.5%). CONCLUSIONS: By machine learning, this study has developed and validated a comprehensive model integrating inflammatory cytokines with clinical variables to predict RP2 before radiotherapy that provides an opportunity to guide clinicians.

[Correlations between Ape1/Ref-1, ICAM-1 and IL-17A Levels in Serum and Radiation Pneumonitis for Local Advanced Non-small Cell Lung Cancer Patients].

BACKGROUND: The main manifestations of radiation pneumonitis are injury of alveolar epithelial and endothelial cells, abnormal expression of cytokines, abnormal proliferation of fibroblasts and synthesis of fibrous matrix. The occurrence of radiation pneumonitis is associated with multiplecytokine level abnormality. These cytokines can also be used as bio-markers to predict the occurrence of radiation pneumonitis. This study was to evaluate the correlation between the change of apurinic/apyrimidinic endonuclease 1/redox factor-1 (Ape1/Ref-1), intercellular adhesion molecules 1 (ICAM-1) and interleukin-17A (IL-17A) before and after radiotherapy and radiation pneumonitis for local advanced non-small cell lung cancer (NSCLC) patients with concurrent chemoradiotherapy. METHODS: NSCLC patients (68 cases) were treated with concurrent radiotherapy and chemotherapy, every patient's normal tissue were controlled with a same radation dose. 68 local advanced NSCLC patients with concurrent chemoradiotherapy were detected the levels of Ape1/Ref-1, ICAM-1 and IL-17A in serum by ELISA before radiotherapy and in the 14th week after radiotherapy. Acute and advanced radiation pulmonary injury was graded according to Radiation Therapy Oncology Group/European Organization For Research and Treatment (RTOG/EORTC) diagnostic and grading criteria. Grade 2 or more radiation pneumonitis was taken as the main end point. RESULTS: Eighteen cases out of 68 developed radiation pneumonitis, 50 of 68 cases have no radiation pneumonia development. There was no significant change of Ape1/Ref-1 levels before and after radiotherapy in radiation pneumonitis group (P>0.05). There was no significant change of Ape1/Ref-1 concentration in serum after radiotherapy between radiation pneumonitis group and non-radiation pneumonitis group (P>0.05). Compared with before radiotherapy, upregulation degree of ICAM-1 levels in radiation pneumonitis group was significantly higher than that in non- radiation pneumonitis group (P<0.05). There was no significant change of IL-17A concentration before and after radiotherapy in radiation pneumonitis group, but after radiotherapy IL-17A concentration in serum were remarkably higher than that in non-radiation pneumonitis group (P<0.05). Correlation analysis found that the change of ICAM-1 before and after radiotherapy has no obvious correlation with the incidence of radiation pneumonitis, and IL-17A change has obvious correlation with the incidence of radiation pneumonitis. CONCLUSIONS: On the basis of strictly controlling radiation dose on normal tissue, IL-17A in serum could be the predictive factors of radiation pneumonitis for local advanced NSCLC patients with concurrent chemoradiotherapy.

Neutrophil-lymphocyte ratio and a dosimetric factor for predicting symptomatic radiation pneumonitis in non-small-cell lung cancer patients treated with concurrent chemoradiotherapy.

OBJECTIVES: To identify the factors that predict the progression of radiological radiation pneumonitis (RP) to symptomatic RP, and to evaluate the usefulness of the neutrophil-lymphocyte ratio (NLR) as a marker of RP severity and prognosis in stage III non-small cell lung cancer (NSCLC) patients treated with definitive concurrent chemoradiotherapy (CCRT). MATERIALS AND METHODS: We retrospectively reviewed 61 patients treated between January 2010 and December 2015. Patients' demographic characteristics, clinical data, laboratory findings and treatment parameters were analyzed to determine the predictive factors associated with progression from radiological RP to symptomatic RP. RESULTS: Forty-seven patients (77%) exhibited radiological RP at a median of 78 days after radiation therapy (RT) completion, and 15 (32%) of these patients developed symptomatic RP. The interval between RT completion and radiological RP presentation was shorter in patients who progressed to symptomatic RP (P = .001); progression was highly probable if this latency period was ≤2 months (P = .002). Stage and RT technique correlated with symptomatic RP development (P = .046 and P = .046, respectively). Among dosimetric factors, a V20 (defined as the lung volume receiving ≥20 Gy) of >30% was the most significant predictor of symptomatic RP (P = .001). The NLR and C-reactive protein level at radiological RP were higher in patients who developed symptomatic RP (P = .067 and P = .012, respectively). On multivariate analysis, a V20 >30% and an NLR at radiological RP >6 were associated with symptomatic RP development. CONCLUSION: The NLR at radiological RP is a useful biomarker for predicting symptomatic RP development after CCRT in stage III NSCLC patients.

Serum Amyloid P-component Level May be a Biomarker for Lung Toxicities and Overall Survival after Thoracic Radiotherapy for Non-small Cell Lung Cancer.

BACKGROUND: Serum amyloid P-component (SAP) contributes to the clearance of apoptotic cells. As one of the main acute-phase reactants, SAP regulates key aspects of inflammation and sets a threshold for immune cell activation. This study aimed to investigate the association of SAP levels with symptomatic lung toxicities after thoracic radiotherapy (TRT) and overall survival (OS) in non-small lung cancer (NSCLC) patients. METHODS: The SAP level at diagnosis and during TRT was evaluated by ELISA in 113 clinically inoperable NSCLC patients. Data of radiation pneumonitis (RP)/lung fibrosis, and OS were recorded. RESULTS: The mean ± SEM values of SAP levels at baseline, week 2, and week 4 during TRT were 83.8 ± 6.4, 36.7 ± 4.8, and 36.5 ± 3.4 µg/mL, respectively (p < 0.0001). Using the median value (83.0 µg/mL) as a cutoff, patients with higher baseline SAP level had longer OS than those with lower SAP level (58.2 vs. 23.1 months, p = 0.044). Baseline SAP level, gender, pathology and BED10 were significantly associated with OS in univariate analysis, while only baseline SAP level, pathology, and BED10 were found to be prognostic for OS in multivariate analysis. Patients with symptomatic RP had lower SAP levels than those with no or mild RP (82.7 ± 7.3 vs. 96.8 ± 4.8 µg/mL, p = 0.024). CONCLUSIONS: The baseline SAP level can be used to predict symptomatic radiation pneumonitis and was prognostic for survival after TRT for NSCLC patients.

Delayed Administration of WP1066, an STAT3 Inhibitor, Ameliorates Radiation-Induced Lung Injury in Mice.

PURPOSE: The present study was designed to investigate the effects of WP1066, a specific inhibitor of STAT3 signaling, on radiation-induced lung injury in mice. METHODS: C57BL/6J mice were subjected to a single thoracic irradiation of 15 Gy X-ray and WP1066 was administrated through intraperitoneal injection. The early and delayed treatment groups were treated with WP1066 during the first 2 weeks and the second 2 weeks, respectively. The therapeutic effects of WP1066 were evaluated by survival analysis, histological examination, and measurement of inflammatory parameters and collagen deposition. The activation of STAT3 pathway was also estimated by immunohistochemical staining and Western blotting. RESULTS: Delayed treatment of WP1066, but not early treatment, prolonged survival time and prevented the development of radiation pneumonitis and the subsequent lung fibrosis in mice. WP1066 treatment also significantly suppressed the activation of STAT3 signaling in the irradiated lung tissues. CONCLUSIONS: The activation of STAT3 pathway might play an important part in the pathogenesis of radiation-induced lung injury. The protective effects of delayed treatment of WP1066 suggested STAT3 signaling could be a therapeutic target for radiation pneumonitis.

Correlations Between Serum IL-6 Levels and Radiation Pneumonitis in Lung Cancer Patients: A Meta-Analysis.

OBJECTIVE: Diagnostic significance of interleukin 6 (IL-6) for lung cancer patients with radiation pneumonitis (RP) was examined within various studies, but yielded conflicting results. Thus, this meta-analysis was performed to demonstrate correlations between serum IL-6 levels and RP in lung cancer patients. METHOD: Electronic databases updated to March 2014 were searched to find relevant studies. Relevant literatures were searched under the PubMed, Embase, Web of Science, Cochrane Library, CISCOM, CINAHL, Google Scholar, CBM and CNKI databases. STATA statistical software (Version 12.0, Stata Corporation, and College Station, TX) Standardized mean difference (SMD), and its corresponding 95% confidence intervals (CIs) were used for this meta-analysis. In addition, nine cohort studies met the inclusion criteria and involved a total of 137 RP patients and 295 non-RP patients. RESULTS: The results of combined SMD suggested that serum IL-6 levels in RP patients was significantly higher than in non-RP patients before radiotherapy. While, there was a significant difference in serum IL-6 levels of RP patients between before and after radiotherapy, we observed no difference in serum IL-6 levels between RP patients and non-RP patients after radiotherapy. Ethnicity-stratified analyses indicated that increased serum IL-6 levels were related to the risk of RP in lung cancer patients among Caucasians, but not detected among Asians (all P > 0.05). CONCLUSION: The main finding of our meta-analysis reveals that increased serum IL-6 levels may contribute to the incidence of RP in lung cancer patients, especially among Caucasians.

Bayesian network ensemble as a multivariate strategy to predict radiation pneumonitis risk.

PURPOSE: Prediction of radiation pneumonitis (RP) has been shown to be challenging due to the involvement of a variety of factors including dose-volume metrics and radiosensitivity biomarkers. Some of these factors are highly correlated and might affect prediction results when combined. Bayesian network (BN) provides a probabilistic framework to represent variable dependencies in a directed acyclic graph. The aim of this study is to integrate the BN framework and a systems' biology approach to detect possible interactions among RP risk factors and exploit these relationships to enhance both the understanding and prediction of RP. METHODS: The authors studied 54 nonsmall-cell lung cancer patients who received curative 3D-conformal radiotherapy. Nineteen RP events were observed (common toxicity criteria for adverse events grade 2 or higher). Serum concentration of the following four candidate biomarkers were measured at baseline and midtreatment: alpha-2-macroglobulin, angiotensin converting enzyme (ACE), transforming growth factor, interleukin-6. Dose-volumetric and clinical parameters were also included as covariates. Feature selection was performed using a Markov blanket approach based on the Koller-Sahami filter. The Markov chain Monte Carlo technique estimated the posterior distribution of BN graphs built from the observed data of the selected variables and causality constraints. RP probability was estimated using a limited number of high posterior graphs (ensemble) and was averaged for the final RP estimate using Bayes' rule. A resampling method based on bootstrapping was applied to model training and validation in order to control under- and overfit pitfalls. RESULTS: RP prediction power of the BN ensemble approach reached its optimum at a size of 200. The optimized performance of the BN model recorded an area under the receiver operating characteristic curve (AUC) of 0.83, which was significantly higher than multivariate logistic regression (0.77), mean heart dose (0.69), and a pre-to-midtreatment change in ACE (0.66). When RP prediction was made only with pretreatment information, the AUC ranged from 0.76 to 0.81 depending on the ensemble size. Bootstrap validation of graph features in the ensemble quantified confidence of association between variables in the graphs where ten interactions were statistically significant. CONCLUSIONS: The presented BN methodology provides the flexibility to model hierarchical interactions between RP covariates, which is applied to probabilistic inference on RP. The authors' preliminary results demonstrate that such framework combined with an ensemble method can possibly improve prediction of RP under real-life clinical circumstances such as missing data or treatment plan adaptation.

An experimental model-based exploration of cytokines in ablative radiation-induced lung injury in vivo and in vitro.

INTRODUCTION: Stereotactic ablative radiotherapy is a newly emerging radiotherapy treatment method that, compared with conventionally fractionated radiation therapy (CFRT), allows an ablative dose of radiation to be delivered to a confined area around a tumor. The aim of the present study was to investigate the changes of various cytokines that may be involved in ablative radiation-induced lung injury in vitro and in vivo. METHODS: In the in vivo study, ablative-dose radiation was delivered to a small volume of the left lung of C3H/HeJCr mice using a small-animal irradiator. The levels of 24 cytokines in the peripheral blood were tested at several time points after irradiation. For the in vitro study, three mouse cell types (type II pneumocytes, alveolar macrophages, and fibroblasts) known to play important roles in radiation-induced pneumonitis and lung fibrosis were analyzed using a co-culture system. RESULTS: In the in vivo study, we found obvious patterns of serum cytokine changes depending on the volume of tissue irradiated (2-mm vs. 3.5-mm collimator). Only the levels of 3 cytokines increased with the 2-mm collimator at the acute phase (1-2 weeks after irradiation), while the majority of cytokines were elevated with the 3.5-mm collimator. In the in vitro co-culture system, after the cells were given an ablative dose of irradiation, the levels of five cytokines (GM-CSF, G-CSF, IL-6, MCP-1, and KC) increased significantly in a dose-dependent manner. CONCLUSIONS: The cytokine levels in our radiation-induced lung injury model showed specific changes, both in vivo and in vitro. These results imply that biological studies related to ablative-dose small-volume irradiation should be investigated using the corresponding experimental models rather than on those simulating large-volume CFRT.

Cytokine levels as biomarkers of radiation fibrosis in patients treated with breast radiotherapy.

BACKGROUND: Radiation fibrosis is not easily measurable although clinical scores have been developed for this purpose. Biomarkers present an alternative more objective approach to quantification, and estimation in blood provides accessible samples. We investigated if blood cytokines could be used to measure established fibrosis in patients who have undergone radiotherapy for breast cancer. METHODS: We studied two cohorts treated by breast-conserving surgery and radiotherapy in the UK START Trial A, one with breast fibrosis (cases) and one with no or minimal fibrosis (controls). Two candidate cytokines, plasma connective tissue growth factor (CTGF) and serum interleukin-6 (IL6) were estimated by ELISA. Comparisons between cases and controls used the t-test or Mann-Whitney test and associations between blood concentration and clinical factors were assessed using the Spearman rank correlation coefficient. RESULTS: Seventy patients were included (26 cases, 44 controls). Mean time since radiotherapy was 9.9 years (range 8.3-12.0). No statistically significant differences between cases and controls in serum IL6 (median (IQR) 0.84 pg/ml (0.57-1.14), 0.75 pg/ml (0.41-1.43) respectively) or plasma CTGF (331.4 pg/ml (234.8-602.9), 334.5 pg/ml (270.0-452.8) were identified. There were no significant associations between blood cytokine concentration and age, fibrosis severity, breast size or time since radiotherapy. CONCLUSIONS: No significant difference in IL6 or CTGF concentrations was detected between patients with breast fibrosis and controls with minimal or no fibrosis.

A clinical score, including biohumoral parameters, is a useful pretest index to discriminate pulmonary infections from radiation damage in chemoradiation-treated lung cancer patients.

To obtain an easy and prompt differential diagnosis between lower airways infections and acute radiation pneumonitis in chemoradiation lung cancer patients. From 303 patients treated, only patients with severe pulmonary symptoms were hospitalized. Clinical and radiation scores were calculated evaluating clinical, biohumoral, dosimetric parameters. Out of 36 patients hospitalized, infections and acute radiation pneumonitis were reported in 66.7% and 33.3%, respectively. Patients with clinical score ≥ 2 had an Odds Ratio of 3.4 (1.4-8.3; p = .006) to have infectious pneumonia, while radiation score was not predictive.

Association between white blood cell count following radiation therapy with radiation pneumonitis in non-small cell lung cancer.

PURPOSE: Radiation pneumonitis (RP) is an inflammatory response to radiation therapy (RT). We assessed the association between RP and white blood cell (WBC) count, an established metric of systemic inflammation, after RT for non-small cell lung cancer. METHODS AND MATERIALS: We retrospectively analyzed 366 patients with non-small cell lung cancer who received ≥60 Gy as definitive therapy. The primary endpoint was whether WBC count after RT (defined as 2 weeks through 3 months after RT completion) was associated with grade ≥3 or grade ≥2 RP. Median lung volume receiving ≥20 Gy (V20) was 31%, and post-RT WBC counts ranged from 1.7 to 21.2 × 10(3) WBCs/µL. Odds ratios (ORs) associating clinical variables and post-RT WBC counts with RP were calculated via logistic regression. A recursive-partitioning algorithm was used to define optimal post-RT WBC count cut points. RESULTS: Post-RT WBC counts were significantly higher in patients with grade ≥3 RP than without (P<.05). Optimal cut points for post-RT WBC count were found to be 7.4 and 8.0 × 10(3)/µL for grade ≥3 and ≥2 RP, respectively. Univariate analysis revealed significant associations between post-RT WBC count and grade ≥3 (n=46, OR=2.6, 95% confidence interval [CI] 1.4‒4.9, P=.003) and grade ≥2 RP (n=164, OR=2.0, 95% CI 1.2‒3.4, P=.01). This association held in a stepwise multivariate regression. Of note, V20 was found to be significantly associated with grade ≥2 RP (OR=2.2, 95% CI 1.2‒3.4, P=.01) and trended toward significance for grade ≥3 RP (OR=1.9, 95% CI 1.0-3.5, P=.06). CONCLUSIONS: Post-RT WBC counts were significantly and independently associated with RP and have potential utility as a diagnostic or predictive marker for this toxicity.

Mapping metabolic changes associated with early Radiation Induced Lung Injury post conformal radiotherapy using hyperpolarized ¹³C-pyruvate Magnetic Resonance Spectroscopic Imaging.

PURPOSE: Radiation Pneumonitis (RP) limits radiotherapy. Detection of early metabolic changes in the lungs associated with RP may provide an opportunity to adjust treatment before substantial toxicities occur. In this work, regional lactate-to-pyruvate signal ratio (lac/pyr) was quantified in rat lungs and heart following administration of hyperpolarized (13)C-pyruvate magnetic resonance imaging (MRI) at day 5, 10, 15 and 25-post conformal radiotherapy. These results were also compared to histology and blood analyses. METHODS: The lower right lungs of 12 Sprague Dawley rats were irradiated in 2 fractions with a total dose of 18.5 Gy using a modified micro-CT system. Regional lactate and pyruvate data were acquired from three irradiated and three age-matched healthy rats at each time point on days 5, 10, 15 and 25-post radiotherapy. Arterial blood was collected from each animal prior to the (13)C-pyruvate injection and was analyzed for blood lactate concentration and arterial oxygen concentration (paO2). Macrophage count was computed from the histology of all rat lungs. RESULTS: A significant increase in lac/pyr was observed in both right and left lungs of the irradiated cohort compared to the healthy cohort for all time points. No increase in lac/pyr was observed in the hearts of the irradiated cohort compared to the hearts of the healthy cohorts. Blood lactate concentration and paO2 did not show a significant change between the irradiated and the healthy cohorts. Macrophage count in both right and left lungs was elevated for the irradiated cohort compared to the healthy cohort. CONCLUSIONS: Metabolic changes associated with RP may be mapped as early as five days post conformal radiotherapy. Over the small sample size in each cohort, elevated macrophage count, consistent with early phase of inflammation was highly correlated to increases in lac/pyr in both the irradiated and unirradiated lungs. Further experiments with larger sample size may improve the confidence of this finding.