Chemoradiation treatment patterns among United States Veteran Health Administration patients with unresectable stage III non-small cell lung cancer.

BACKGROUND: The Veterans Health Administration (VHA) is the largest integrated health care system in the United States (US). Among VHA patients, the rate of use of concurrent chemoradiation therapy (CCRT) among those with unresectable, stage III non-small cell lung cancer (NSCLC) is unknown. The objective was to report recent CCRT treatment patterns in VHA patients and identify characteristics associated with receipt of CCRT. METHODS: Using Department of Veteran Affairs (VA) Cancer Registry System data linked to VA electronic medical records, we determined rates of CCRT, sequential CRT (SCRT), radiation therapy (RT) only, chemotherapy (CT) only, and neither treatment. RESULTS: Among 4054 VHA patients who met study criteria, CCRT rates slightly increased from 44 to 50% between 2013 and 2017. Factors associated with decreased odds of CCRT receipt compared to any other treatment included increasing age (adjusted odds ratio [aOR] per 10 years = 0.67; 95% CI: 0.60-0.76) and Charlson-Deyo comorbidity score (aOR = 0.94; 95% CI: 0.91-0.97). White race was associated with increased odds of CCRT receipt (aOR = 1.24; 95% CI: 1.004-1.53). In a chart review sample of 200 patients, less than half (n = 85) had a documented reason for not receiving CCRT. Among these, 29% declined treatment, and 71% did not receive CCRT due to "not being a candidate" for reasons related to frailty or lung nodules being too far apart for radiation therapy. CONCLUSIONS: CCRT rates among VHA patients with unresectable, stage III NSCLC slightly increased from 2013 to 2017; however in 2017, only half were receiving CCRT. Older patients and those with multiple comorbidities were less likely to receive CCRT and even when controlling for these factors, non-white patients were less likely to receive CCRT.

EGFR mutation testing and TKI treatment patterns among veterans with stage III and IV non-small cell lung cancer.

BACKGROUND: Epidermal growth factor receptor (EGFR) mutation testing is recommended in metastatic non-small cell lung cancer (NSCLC). The objective of this study was to assess changes in EGFR mutation testing patterns and tyrosine kinase inhibitor (TKI) use in US veterans with stage III-IV NSCLC between 2013 and 2017. PATIENTS AND METHODS: Retrospective study using linked data from Department of Veterans Affairs (VA) Cancer Registry System, Corporate Data Warehouse, commercial laboratories, and clinical notes. Generalized linear mixed models accounting for clustering by VA facility were used to determine factors associated with EGFR mutation testing. RESULTS: From 2013 to 2017, EGFR mutation testing increased from 29.5% to 38.4% among veterans with stage III-IV NSCLC and from 47.0% to 57.4% among veterans with stage IV non-squamous disease. Factors associated with increased odds of testing included being married, Medicare enrollment, and adenocarcinoma histology. Factors associated with decreased odds of testing included Medicaid eligibility, stage III disease, increasing age, being a current or former smoker, increasing Charlson-Deyo comorbidity score, and receiving cancer care in the South. Appropriate use of a TKI rose from 2013 to 2017 (17.2% to 74.1%). CONCLUSION: EGFR mutation testing rates increased to almost 60% in the stage IV non-squamous NSCLC population in 2017, with residual opportunity for further increase. Several sociodemographic characteristics, comorbidities, and geographic regions were associated with EGFR mutation testing suggestive of inequitable testing decisions. Appropriate use of TKI improved drastically from 2013 to 2017 demonstrating rapidly changing practice patterns through the adoption phase of new treatment options.

Clinical Impact of 21-Gene Recurrence Score Test Within the Veterans Health Administration: Utilization and Receipt of Guideline-Concordant Care.

INTRODUCTION: Ensuring guideline-concordant cancer care is a Department of Veterans Affairs (VA) priority, especially as the number of breast cancer patients at VA medical centers (VAMCs) grows. We assessed the utilization and clinical impact of the 21-gene Recurrence Score test, which predicts 10-year risk of breast cancer recurrence and the likelihood of chemotherapy benefit, on veterans newly diagnosed with breast cancer. PATIENTS AND METHODS: We conducted a retrospective cohort study using 2011-2012 VA Central Cancer Registry, chart review, and laboratory test data. Independent variables assessed included patient and site-of-care characteristics. The outcome of interest was whether newly diagnosed, eligible (node negative, hormone-receptor positive, human epidermal growth factor receptor 2 [HER2] negative) veterans underwent the 21-gene test. We performed descriptive statistics on all patients and multivariate logistic regression to determine associations. We correlated treatments received with test results. RESULTS: Among 328 eligible veterans, 82 (25%) had the 21-gene test; 100 eligible veterans (30%) sought care at a VAMC where no tests were ordered. Receiving care at a VAMC that had women's health services (odds ratio [OR], 1.84, 95% confidence interval [CI], 1.05-3.22) and having tumor characteristics meeting the National Comprehensive Cancer Network 2010 test criteria (OR, 3.06, 95% CI, 1.69-5.57) were positive predictors of testing; increasing age (OR, 0.93, 95% CI, 0.91-0.96 per year) and fee-based care (OR, 0.46, 95% CI, 0.26-0.82) were negative predictors. The majority of tested patients received guideline-concordant care. CONCLUSION: Site of care and tumor characteristics were important predictors of test uptake. Facilitating delivery of guideline-concordant cancer care requires improved laboratory informatics and clinical decision support.

In vivo proton-electron double-resonance imaging of extracellular tumor pH using an advanced nitroxide probe.

A variable radio frequency proton-electron double-resonance imaging (VRF PEDRI) approach for pH mapping of aqueous samples has been recently developed (Efimova et al. J. Magn. Reson. 2011, 209, 227-232). A pH map is extracted from two PEDRI acquisitions performed at electron paramagnetic resonance (EPR) frequencies of protonated and unprotonated forms of a pH-sensitive probe. To translate VRF PEDRI to an in vivo setting, an advanced pH probe was synthesized. Probe deuteration resulted in a narrow spectral line of 1.2 G compared to a nondeuterated analogue line width of 2.1 G allowing for an increase of Overhauser enhancements and reduction in rf power deposition. Binding of the probe to the cell-impermeable tripeptide, glutathione (GSH), allows for targeting to extracellular tissue space for monitoring extracellular tumor acidosis, a prognostic factor in tumor pathophysiology. The probe demonstrated pH sensitivity in the 5.8-7.8 range, optimum for measurement of acidic extracellular tumor pH (pH(e)). In vivo VRF PEDRI was performed on Met-1 tumor-bearing mice. Compared to normal mammary glands with a neutral mean pH(e) (7.1 ± 0.1), we observed broader pH distribution with acidic mean pH(e) (6.8 ± 0.1) in tumor tissue. In summary, VRF PEDRI in combination with a newly developed pH probe provides an analytical approach for spatially resolved noninvasive pHe monitoring, in vivo.

Nicotinic receptor alpha7 expression identifies a novel hematopoietic progenitor lineage.

How inflammatory responses are mechanistically modulated by nicotinic acetylcholine receptors (nAChR), especially by receptors composed of alpha7 (α7) subunits, is poorly defined. This includes a precise definition of cells that express α7 and how these impact on innate inflammatory responses. To this aim we used mice generated through homologous recombination that express an Ires-Cre-recombinase bi-cistronic extension of the endogenous α7 gene that when crossed with a reporter mouse expressing Rosa26-LoxP (yellow fluorescent protein (YFP)) marks in the offspring those cells of the α7 cell lineage (α7(lin+)). In the adult, on average 20-25 percent of the total CD45(+) myeloid and lymphoid cells of the bone marrow (BM), blood, spleen, lymph nodes, and Peyers patches are α7(lin+), although variability between litter mates in this value is observed. This hematopoietic α7(lin+) subpopulation is also found in Sca1(+)cKit(+) BM cells suggesting the α7 lineage is established early during hematopoiesis and the ratio remains stable in the individual thereafter as measured for at least 18 months. Both α7(lin+) and α7(lin-) BM cells can reconstitute the immune system of naïve irradiated recipient mice and the α7(lin+):α7(lin-) beginning ratio is stable in the recipient after reconstitution. Functionally the α7(lin+):α7(lin-) lineages differ in response to LPS challenge. Most notable is the response to LPS as demonstrated by an enhanced production of IL-12/23(p40) by the α7(lin+) cells. These studies demonstrate that α7(lin+) identifies a novel subpopulation of bone marrow cells that include hematopoietic progenitor cells that can re-populate an animal's inflammatory/immune system. These findings suggest that α7 exhibits a pleiotropic role in the hematopoietic system that includes both the direct modulation of pro-inflammatory cell composition and later in the adult the role of modulating pro-inflammatory responses that would impact upon an individual's lifelong response to inflammation and infection.

Unique oxidation of imidazolidine nitroxides by potassium ferricyanide: strategy for designing paramagnetic probes with enhanced sensitivity to oxidative stress.

Potassium ferricyanide (PF), routinely employed for the oxidation of sterically-hindered hydroxylamines to nitroxides, is considered to be chemically inert towards the latter. In the present study, we report on an unexpected oxidative fragmentation of the imidazolidine nitroxides containing hydrogen atom in the 4-position of the heterocycle (HIMD) by PF resulting in the loss of the EPR signal. The mechanistic EPR, spectrophotometric, electrochemical and HPLC-MS studies support the assumption that the HIMD fragmentation is facilitated by the proton abstraction from the 4-position of the oxoammonium cation formed as a result of the initial one-electron HIMD oxidation. Increase in steric hindrance around the radical fragment by introducing ethyl substituents decreased the rate of ascorbate-induced HIMD reduction by more than 20 times, but did not affect the rate of ferricyanide-induced HIMD oxidation. This preferential sensitivity of HIMDs to oxidative processes has been used to detect peroxyl radicals in the presence of high concentration of the reducing agent, ascorbate. HIMD-based EPR probes capable to discriminate oxidative and reductive processes might find application in biomedicine and related fields for monitoring the oxidative stress and reactive radical species in biological systems.

Organ specific mapping of in vivo redox state in control and cigarette smoke-exposed mice using EPR/NMR co-imaging.

In vivo mapping of alterations in redox status is important for understanding organ specific pathology and disease. While electron paramagnetic resonance imaging (EPRI) enables spatial mapping of free radicals, it does not provide anatomic visualization of the body. Proton MRI is well suited to provide anatomical visualization. We applied EPR/NMR co-imaging instrumentation to map and monitor the redox state of living mice under normal or oxidative stress conditions induced by secondhand cigarette smoke (SHS) exposure. A hybrid co-imaging instrument, EPRI (1.2 GHz)/proton MRI (16.18 MHz), suitable for whole-body co-imaging of mice was utilized with common magnet and gradients along with dual EPR/NMR resonators that enable co-imaging without sample movement. The metabolism of the nitroxide probe, 3-carbamoyl-proxyl (3-CP), was used to map the redox state of control and SHS-exposed mice. Co-imaging allowed precise 3D mapping of radical distribution and reduction in major organs such as the heart, lungs, liver, bladder and kidneys. Reductive metabolism was markedly decreased in SHS-exposed mice and EPR/NMR co-imaging allowed quantitative assessment of this throughout the body. Thus, in vivo EPR/NMR co-imaging enables in vivo organ specific mapping of free radical metabolism and redox stress and the alterations that occur in the pathogenesis of disease.

In vivo monitoring of pH, redox status, and glutathione using L-band EPR for assessment of therapeutic effectiveness in solid tumors.

Approach for in vivo real-time assessment of tumor tissue extracellular pH (pH(e)), redox, and intracellular glutathione based on L-band EPR spectroscopy using dual function pH and redox nitroxide probe and disulfide nitroxide biradical, is described. These parameters were monitored in PyMT mice bearing breast cancer tumors during treatment with granulocyte macrophage colony-stimulating factor. It was observed that tumor pH(e) is about 0.4 pH units lower than that in normal mammary gland tissue. Treatment with granulocyte macrophage colony-stimulating factor decreased the value of pH(e) by 0.3 units compared with PBS control treatment. Tumor tissue reducing capacity and intracellular glutathione were elevated compared with normal mammary gland tissue. Granulocyte macrophage colony-stimulating factor treatment resulted in a decrease of the tumor tissue reducing capacity and intracellular glutathione content. In addition to spectroscopic studies, pH(e) mapping was performed using recently proposed variable frequency proton-electron double-resonance imaging. The pH mapping superimposed with MRI image supports probe localization in mammary gland/tumor tissue, shows high heterogeneity of tumor tissue pH(e) and a difference of about 0.4 pH units between average pH(e) values in tumor and normal mammary gland. In summary, the developed multifunctional approach allows for in vivo, noninvasive pH(e), extracellular redox, and intracellular glutathione content monitoring during investigation of various therapeutic strategies for solid tumors.

Standard-based method for proton-electron double resonance imaging of oxygen.

Proton-electron double resonance imaging (PEDRI) has been utilized for indirect determination of oxygen concentrations in aqueous samples and living systems. Due to the complexity of the problem, there are seven oxygen related parameters that need to be measured to determine the distribution of oxygen. We present an improved approach in which image intensities from only two PEDRI acquisitions with different EPR irradiation powers are required to determine the distribution of a paramagnetic probe and oxygen in an analyzed sample. This is achieved using three reference samples with known concentrations of a paramagnetic probe and oxygen placed inside the resonator together with the measurement sample. An EPR-off image, which has low signal intensity at low magnetic field (0.02 T) is not required for the calculations, significantly reducing the total time of the experiments and the noise while enhancing the accuracy of these oxygen measurements. The Finland trityl radical was used as the paramagnetic probe and oxygen concentrations could be accurately measured and imaged over the physiological range from 0 to 240 µM.

Variable radio frequency proton-electron double-resonance imaging: application to pH mapping of aqueous samples.

Proton-electron double-resonance imaging (PEDRI) offers rapid image data collection and high resolution for spatial distribution of paramagnetic probes. Recently we developed the concept of variable field (VF) PEDRI which enables extracting a functional map from a limited number of images acquired at pre-selected EPR excitation fields using specific paramagnetic probes (Khramtsov et al., J. Magn. Reson. 202 (2010) 267-273). In this work, we propose and evaluate a new modality of PEDRI-based functional imaging with enhanced temporal resolution which we term variable radio frequency (VRF) PEDRI. The approach allows for functional mapping (e.g., pH mapping) using specifically designed paramagnetic probes with high quality spatial resolution and short acquisition times. This approach uses a stationary magnetic field but different EPR RFs. The ratio of Overhauser enhancements measured at each pixel at two different excitation frequencies corresponding to the resonances of protonated and deprotonated forms of a pH-sensitive nitroxide is converted to a pH map using a corresponding calibration curve. Elimination of field cycling decreased the acquisition time by exclusion periods of ramping and stabilization of the magnetic field. Improved magnetic field homogeneity and stability allowed for the fast MRI acquisition modalities such as fast spin echo. In total, about 30-fold decrease in EPR irradiation time was achieved for VRF PEDRI (2.4s) compared with VF PEDRI (70s). This is particularly important for in vivo applications enabling one to overcome the limiting stability of paramagnetic probes and sample overheating by reducing RF power deposition.

Induction of granzyme B expression in T-cell receptor/CD28-stimulated human regulatory T cells is suppressed by inhibitors of the PI3K-mTOR pathway.

BACKGROUND: Regulatory T cells (Tregs) can employ a cell contact- and granzyme B-dependent mechanism to mediate suppression of bystander T and B cells. Murine studies indicate that granzyme B is involved in the Treg-mediated suppression of anti-tumor immunity in the tumor microenvironment and in the Treg-mediated maintenance of allograft survival. In spite of its central importance, a detailed study of granzyme B expression patterns in human Tregs has not been performed. RESULTS: Our data demonstrated that natural Tregs freshly isolated from the peripheral blood of normal adults lacked granzyme B expression. Tregs subjected to prolonged TCR and CD28 triggering, in the presence of IL-2, expressed high levels of granzyme B but CD3 stimulation alone or IL-2 treatment alone failed to induce granzyme B. Treatment of Tregs with the mammalian target of rapamycin (mTOR) inhibitor, rapamycin or the PI3 kinase (PI3K) inhibitor LY294002 markedly suppressed granzyme B expression. However, neither rapamycin, as previously reported by others, nor LY294002 inhibited Treg proliferation or induced significant cell death in TCR/CD28/IL-2 stimulated cells. The proliferation rate of Tregs was markedly higher than that of CD4+ conventional T cells in the setting of rapamycin treatment. Tregs expanded by CD3/CD28/IL-2 stimulation without rapamycin demonstrated increased in vitro cytotoxic activity compared to Tregs expanded in the presence of rapamycin in both short term (6 hours) and long term (48 hours) cytotoxicity assays. CONCLUSION: TCR/CD28 mediated activation of the PI3K-mTOR pathway is important for granyzme B expression but not proliferation in regulatory T cells. These findings may indicate that suppressive mechanisms other than granzyme B are utilized by rapamycin-expanded Tregs.