A Central Role for Foxp3+ Regulatory T Cells in K-Ras-Driven Lung Tumorigenesis

BACKGROUND: K-Ras mutations are characteristic of human lung adenocarcinomas and occur almost exclusively in smokers. In preclinical models, K-Ras mutations are necessary for tobacco carcinogen-driven lung tumorigenesis and are sufficient to cause lung adenocarcinomas in transgenic mice. Because these mutations confer resistance to commonly used cytotoxic chemotherapies and targeted agents, effective therapies that target K-Ras are needed. Inhibitors of mTOR such as rapamycin can prevent K-Ras-driven lung tumorigenesis and alter the proportion of cytotoxic and Foxp3+ regulatory T cells, suggesting that lung-associated T cells might be important for tumorigenesis. METHODS: Lung tumorigenesis was studied in three murine models that depend on mutant K-Ras; a tobacco carcinogen-driven model, a syngeneic inoculation model, and a transgenic model. Splenic and lung-associated T cells were studied using flow cytometry and immunohistochemistry. Foxp3+ cells were depleted using rapamycin, an antibody, or genetic ablation. RESULTS: Exposure of A/J mice to a tobacco carcinogen tripled lung-associated Foxp3+ cells prior to tumor development. At clinically relevant concentrations, rapamycin prevented this induction and reduced lung tumors by 90%. In A/J mice inoculated with lung adenocarcinoma cells resistant to rapamycin, antibody-mediated depletion of Foxp3+ cells reduced lung tumorigenesis by 80%. Likewise, mutant K-Ras transgenic mice lacking Foxp3+ cells developed 75% fewer lung tumors than littermates with Foxp3+ cells. CONCLUSIONS: Foxp3+ regulatory T cells are required for K-Ras-mediated lung tumorigenesis in mice. These studies support clinical testing of rapamycin or other agents that target Treg in K-Ras driven human lung cancer

Prognostic significance of clinical factors and Akt activation in patients with bronchioloalveolar carcinoma.

To access publisher full text version of this article. Please click on the hyperlink in Additional Link fieldPURPOSE: Lung cancer is the leading cause of cancer related mortality in the world. Bronchioloalveolar carcinoma (BAC) is a subset of NSCLC that has recently gained attention because of distinct biological and clinical features, increased incidence, and enhanced responsiveness to new therapies such as epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). However, prognostic features for BAC have not been well defined. Because activation of Akt is highly prevalent and a poor prognostic factor for other types of NSCLC, we assessed the prognostic significance of clinical features and Akt activation in patients with BAC. METHODS: Forty-six cases of BAC in Iceland were classified according to WHO 1999 criteria. Akt activation was assessed using two phospho-specific antibodies against Akt (S473 and T308) in immunohistochemical (IHC) analysis. Associations between ordered Akt levels and other dichotomous parameters were evaluated using an exact Cochran-Armitage test for trend. Survival was analyzed by the Kaplan-Meier method and log-rank test, with hazard ratios (HR) determined by Cox proportional hazard models. The Cox model was also used to assess the joint effect of multiple factors on survival when they are considered simultaneously. RESULTS: Age and histology (mucinous versus non-mucinous) were not associated with survival. Activation of Akt was highly prevalent in BAC, with only 2 out of 46 patients exhibiting negative staining with either antibody. Moderate to high Akt activation was observed in 63% of cases and was associated with non-mucinous histology. Akt activation was not associated with differences in survival or smoking status. In contrast, Cox model analysis revealed that male gender (HR 2.24, 95% CI, 1.07-4.71, p=0.032), advanced stage (III or IV) (HR 2.17, 95% CI, 1.004-4.71, p=0.049) and smoking status (HR 6.89, 95% CI, 1.49-31.88, p=0.013) were associated with a worse prognosis. CONCLUSIONS: Male gender, advanced stage, and especially smoking status (but not Akt activation) are potentially important prognostic features for BAC. These features should be considered in the design and interpretation of clinical trials that enroll BAC patients

Different inhibition of Gβγ-stimulated class IB phosphoinositide 3-kinase (PI3K) variants by a monoclonal antibody. Specific function of p101 as a Gβγ-dependent regulator of PI3Kγ enzymatic activity

Class IB phosphoinositide 3-kinases γ (PI3Kγ) are second-messenger-generating enzymes downstream of signalling cascades triggered by G-protein-coupled receptors (GPCRs). PI3Kγ variants have one catalytic p110γ subunit that can form two different heterodimers by binding to one of a pair of non-catalytic subunits, p87 or p101. Growing experimental data argue for a different regulation of p87–p110γ and p101–p110γ allowing integration into distinct signalling pathways. Pharmacological tools enabling distinct modulation of the two variants are missing. The ability of an anti-p110γ monoclonal antibody [mAb(A)p110γ] to block PI3Kγ enzymatic activity attracted us to characterize this tool in detail using purified proteins. In order to get insight into the antibody–p110γ interface, hydrogen–deuterium exchange coupled to MS (HDX-MS) measurements were performed demonstrating binding of the monoclonal antibody to the C2 domain in p110γ, which was accompanied by conformational changes in the helical domain harbouring the Gβγ-binding site. We then studied the modulation of phospholipid vesicles association of PI3Kγ by the antibody. p87–p110γ showed a significantly reduced Gβγ-mediated phospholipid recruitment as compared with p101–p110γ. Concomitantly, in the presence of mAb(A)p110γ, Gβγ did not bind to p87–p110γ. These data correlated with the ability of the antibody to block Gβγ-stimulated lipid kinase activity of p87–p110γ 30-fold more potently than p101–p110γ. Our data argue for differential regulatory functions of the non-catalytic subunits and a specific Gβγ-dependent regulation of p101 in PI3Kγ activation. In this scenario, we consider the antibody as a valuable tool to dissect the distinct roles of the two PI3Kγ variants downstream of GPCRs

Pneumonitis in Patients Receiving Thoracic Radiotherapy and Osimertinib: A Multi-Institutional Study

Introduction: Thoracic radiotherapy (TRT) is increasingly used in patients receiving osimertinib for advanced NSCLC, and the risk of pneumonitis is not established. We investigated the risk of pneumonitis and potential risk factors in this population. Methods: We performed a multi-institutional retrospective analysis of patients under active treatment with osimertinib who received TRT between April 2016 and July 2022 at two institutions. Clinical characteristics, including whether osimertinib was held during TRT and pneumonitis incidence and grade (Common Terminology Criteria for Adverse Events version 5.0) were documented. Logistic regression analysis was performed to identify risk factors associated with grade 2 or higher (2+) pneumonitis. Results: The median follow-up was 10.2 months (range: 1.9–53.2). Of 102 patients, 14 (13.7%) developed grade 2+ pneumonitis, with a median time to pneumonitis of 3.2 months (range: 1.5–6.3). Pneumonitis risk was not significantly increased in patients who continued osimertinib during TRT compared with patients who held osimertinib during TRT (9.1% versus 15.0%, p = 0.729). Three patients (2.9%) had grade 3 pneumonitis, none had grade 4, and two patients had grade 5 events (2.0%, diagnosed 3.2 mo and 4.4 mo post-TRT). Mean lung dose was associated with the development of grade 2+ pneumonitis in multivariate analysis (OR = 1.19, p = 0.021). Conclusions: Although the overall rate of pneumonitis in patients receiving TRT and osimertinib was relatively low, there was a small risk of severe toxicity. The mean lung dose was associated with an increased risk of developing pneumonitis. These findings inform decision-making for patients and providers