Intratumoral heterogeneity analysis reveals hidden associations between protein expression losses and patient survival in clear cell renal cell carcinoma.

Intratumoral heterogeneity (ITH) is a prominent feature of kidney cancer. It is not known whether it has utility in finding associations between protein expression and clinical parameters. We used ITH that is detected by immunohistochemistry (IHC) to aid the association analysis between the loss of SWI/SNF components and clinical parameters.160 ccRCC tumors (40 per tumor stage) were used to generate tissue microarray (TMA). Four foci from different regions of each tumor were selected. IHC was performed against PBRM1, ARID1A, SETD2, SMARCA4, and SMARCA2. Statistical analyses were performed to correlate biomarker losses with patho-clinical parameters. Categorical variables were compared between groups using Fisher\u27s exact tests. Univariate and multivariable analyses were used to correlate biomarker changes and patient survivals. Multivariable analyses were performed by constructing decision trees using the classification and regression trees (CART) methodology. IHC detected widespread ITH in ccRCC tumors. The statistical analysis of the Truncal loss (root loss) found additional correlations between biomarker losses and tumor stages than the traditional Loss in tumor (total) . Losses of SMARCA4 or SMARCA2 significantly improved prognosis for overall survival (OS). Losses of PBRM1, ARID1A or SETD2 had the opposite effect. Thus Truncal Loss analysis revealed hidden links between protein losses and patient survival in ccRCC

Nitrilase 1 modulates lung tumor progression in vitro and in vivo.

Uncovering novel growth modulators for non-small cell lung cancer (NSCLC) may lead to new therapies for these patients. Previous studies suggest Nit1 suppresses chemically induced carcinogenesis of the foregut in a mouse model. In this study we aimed to determine the role of Nit1 in a transgenic mouse lung cancer model driven by a G12D Kras mutation. Nit1 knockout mice (Nit1-/-) were crossed with KrasG12D/+ mice to investigate whether a G12D Kras mutation and Nit1 inactivation interact to promote or inhibit the development of NSCLC. We found that lung tumorigenesis was suppressed in the Nit1-null background (Nit1-/-:KrasG12D/+). Micro-CT scans and gross tumor measurements demonstrated a 5-fold reduction in total tumor volumes compared to Nit1+/+KrasG12D/+ (

From the Lung to the Finger: An Unusual Presentation of Adenosquamous Cell Carcinoma

Learning Objective To recognize unusual presentations of metastatic lung disease.https://jdc.jefferson.edu/medoncposters/1019/thumbnail.jp

Targeting fibroblast activation protein in tumor stroma with chimeric antigen receptor T cells can inhibit tumor growth and augment host immunity without severe toxicity.

The majority of chimeric antigen receptor (CAR) T-cell research has focused on attacking cancer cells. Here, we show that targeting the tumor-promoting, nontransformed stromal cells using CAR T cells may offer several advantages. We developed a retroviral CAR construct specific for the mouse fibroblast activation protein (FAP), comprising a single-chain Fv FAP [monoclonal antibody (mAb) 73.3] with the CD8α hinge and transmembrane regions, and the human CD3ζ and 4-1BB activation domains. The transduced muFAP-CAR mouse T cells secreted IFN-γ and killed FAP-expressing 3T3 target cells specifically. Adoptively transferred 73.3-FAP-CAR mouse T cells selectively reduced FAP(hi) stromal cells and inhibited the growth of multiple types of subcutaneously transplanted tumors in wild-type, but not FAP-null immune-competent syngeneic mice. The antitumor effects could be augmented by multiple injections of the CAR T cells, by using CAR T cells with a deficiency in diacylglycerol kinase, or by combination with a vaccine. A major mechanism of action of the muFAP-CAR T cells was the augmentation of the endogenous CD8(+) T-cell antitumor responses. Off-tumor toxicity in our models was minimal following muFAP-CAR T-cell therapy. In summary, inhibiting tumor growth by targeting tumor stroma with adoptively transferred CAR T cells directed to FAP can be safe and effective, suggesting that further clinical development of anti-human FAP-CAR is warranted

Radiation mitigating properties of the lignan component in flaxseed

BACKGROUND: Wholegrain flaxseed (FS), and its lignan component (FLC) consisting mainly of secoisolariciresinol diglucoside (SDG), have potent lung radioprotective properties while not abrogating the efficacy of radiotherapy. However, while the whole grain was recently shown to also have potent mitigating properties in a thoracic radiation pneumonopathy model, the bioactive component in the grain responsible for the mitigation of lung damage was never identified. Lungs may be exposed to radiation therapeutically for thoracic malignancies or incidentally following detonation of a radiological dispersion device. This could potentially lead to pulmonary inflammation, oxidative tissue injury, and fibrosis. This study aimed to evaluate the radiation mitigating effects of FLC in a mouse model of radiation pneumonopathy. METHODS: We evaluated FLC-supplemented diets containing SDG lignan levels comparable to those in 10% and 20% whole grain diets. 10% or 20% FLC diets as compared to an isocaloric control diet (0% FLC) were given to mice (C57/BL6) (n=15-30 mice/group) at 24, 48, or 72-hours after single-dose (13.5 Gy) thoracic x-ray treatment (XRT). Mice were evaluated 4 months post-XRT for blood oxygenation, lung inflammation, fibrosis, cytokine and oxidative damage levels, and survival. RESULTS: FLC significantly mitigated radiation-related animal death. Specifically, mice fed 0% FLC demonstrated 36.7% survival 4 months post-XRT compared to 60–73.3% survival in mice fed 10%-20% FLC initiated 24–72 hours post-XRT. FLC also mitigated radiation-induced lung fibrosis whereby 10% FLC initiated 24-hours post-XRT significantly decreased fibrosis as compared to mice fed control diet while the corresponding TGF-beta1 levels detected immunohistochemically were also decreased. Additionally, 10-20% FLC initiated at any time point post radiation exposure, mitigated radiation-induced lung injury evidenced by decreased bronchoalveolar lavage (BAL) protein and inflammatory cytokine/chemokine release at 16 weeks post-XRT. Importantly, neutrophilic and overall inflammatory cell infiltrate in airways and levels of nitrotyrosine and malondialdehyde (protein and lipid oxidation, respectively) were also mitigated by the lignan diet. CONCLUSIONS: Dietary FLC given early post-XRT mitigated radiation effects by decreasing inflammation, lung injury and eventual fibrosis while improving survival. FLC may be a useful agent, mitigating adverse effects of radiation in individuals exposed to incidental radiation, inhaled radioisotopes or even after the initiation of radiation therapy to treat malignancy

Aneurysmal Lesions of Patients with Abdominal Aortic Aneurysm Contain Clonally Expanded T Cells

Abdominal aortic aneurysm (AAA) is a common disease with often life-threatening consequences. This vascular disorder is responsible for 1-2% of all deaths in men aged 65 years or older. Autoimmunity may be responsible for the pathogenesis of AAA. Although it is well documented that infiltrating T cells are essentially always present in AAA lesions, little is known about their role in the initiation and/or progression of the disease. To determine whether T cells infiltrating AAA lesions contain clonally expanded populations of T cells, we amplified beta-chain TCR transcripts by the nonpalindromic adaptor-PCR/Vbeta-specific PCR and/or Vbeta-specific PCR, followed by cloning and sequencing. We report in this article that aortic abdominal aneurysmal lesions from 8 of 10 patients with AAA contained oligoclonal populations of T cells. Multiple identical copies of beta-chain TCR transcripts were identified in these patients. These clonal expansions are statistically significant. These results demonstrate that alphabeta TCR(+) T lymphocytes infiltrating aneurysmal lesions of patients with AAA have undergone proliferation and clonal expansion in vivo at the site of the aneurysmal lesion, in response to unidentified self- or nonself Ags. This evidence supports the hypothesis that AAA is a specific Ag-driven T cell disease

Clonally Expanded Alpha-Chain T-Cell Receptor (TCR) Transcripts are Present in Aneurysmal Lesions of Patients With Abdominal Aortic Aneurysm (AAA)

Abdominal aortic aneurysm (AAA) is a life-threatening immunological disease responsible for 1 to 2% of all deaths in 65 year old or older individuals. Although mononuclear cell infiltrates have been demonstrated in AAA lesions and autoimmunity may be responsible for the initiation and account for the propagation of the disease, the information available about the pathogenesis of AAA is limited. To examine whether AAA lesions from patients with AAA contain clonally expanded α-chain TCR transcripts, we amplified by the non-palindromic adaptor-PCR (NPA-PCR)/Vα-specific PCR and/or the Vα-specific PCR these α-chain TCR transcripts. The amplified transcripts were cloned and sequenced. Substantial proportions of identical α-chain TCR transcripts were identified in AAA lesions of 4 of 5 patients, demonstrating that clonally expanded T cells are present in these AAA lesions. These results were statistically significant by the bimodal distribution. Three of 5 of these patients were typed by DNA-based HLA-typing and all three expressed DRB1 alleles containing the DRβGln70 amino acid residue that has been demonstrated to be associated with AAA. All three patients exhibited clonally expanded T cells in AAA lesions. Four of the 5 patients with AAA who exhibited clonal expansions of α-chain TCR transcripts, also exhibited clonal expansions of β-chain TCR transcripts in AAA lesions, as we have demonstrated previously (J Immunol 192:4897, 2014). αβ TCR-expressing T cells infiltrating AAA lesions contain T-cell clones which have undergone proliferation and clonal expansion in vivo in response to as yet unidentified specific antigens that may be self or nonself. These results provide additional evidence supporting the hypothesis that AAA is a specific antigen-driven T-cell autoimmune disease

Non Small Cell Carcinoma of the Lung- From Morphology to Molecular Profiling

Dr. Charalambos Solomides is an Associate Professor of Pathology at the Thomas Jefferson University Hospital. At present, he holds many positions at Jefferson. He is a Staff Surgical Pathologist, Staff Cytopathologist, Director of Immunnohistochemistry Research Lab, Director of Cytopathology and the Director of Fine Needle Aspiration Services. His interests are in Non-small cell lung carcinoma, radiation induced lung injury and idiopathic pulmonary fibrosis

Correlations between Gene Amplification and Protein Expression of Topoisomerase 2A (TOP2A) in Squamous Cell Carcinoma of the Lung

Background: While DNA topoisomerase 2A (TOP2A) plays an essential role in maintaining the structural integrity of the double helix during replication and recombination, excessive expression of this enzyme may promote malignant cell transformations. In fact, increased levels of TOP2A have been observed in various cancer cell lines including squamous cell carcinoma of the lung. This study sought to identify correlations between genotypic and phenotypic evidence of TOP2A obtained via in situ hybridization (ISH) and immunohistochemistry (IHC) techniques. Methods Tissue microarrays created from 29 samples of Stage I Squamous Cell Carcinoma of the lung were stained with VENTANA BenchMark ULTRA platform with dual color ISH molecular probes TOP2A / CEP17 and antiTOP2A antibody (clone JS5B4-rabbit monoclonal antibody). Gene copy numbers were analyzed using bright field microscopy. Gene amplification was considered in cells exhibiting gene copies \u3e 3 or TOP2A:CEP17 ratios \u3e 1.82. IHC stains were quantified using Spectrum software (Apeiro technologies) using the nuclear algorithm. All levels of protein expression (+1 to +3) were considered positive. Results: A moderate Pearson Correlation (0.4) between TOP2A gene amplification and protein expression was identified. Conclusion: While gene amplification moderately correlated with protein expression of TOP2A, additional factors influencing protein expression independently of gene amplification should be identified