Browsing by Subject "Carcinoma, Squamous Cell"
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Item Assessment of Circularized HPV16 E7 RNA, GLUT1, and PD-L1 in Anal Squamous Cell Carcinoma(2020-05-01T05:00:00.000Z) Chamseddin, Bahir Hassan; Wang, Richard; Le, Lu Q.; Hammer, SuntreaBACKGROUND: Anal squamous cell carcinoma (ASCC) is a rare, deadly malignancy caused by high-risk human papillomaviruses in up to 90% of cases and continues to be treated by cytotoxic therapy established 40 years ago. There is a dearth of reliable biomarkers for ASCC. The prognostic implication of programmed death-ligand 1 (PD-L1) expression remains controversial while other biomarkers, like glucose-1-transporter (GLUT1) expression levels, have not been examined in the setting of ASCC. More recently, covalently closed circular RNAs has been (circRNA) expression has been shown to be widespread in cancers, and circRNAs have been proposed to be potential biomarkers. In previous studies, we discovered a novel circular E7 RNA expressed by HPV16 (circE7), which has not been assessed as a potential biomarker in any diseases. OBJECTIVE: We hypothesize that human papillomavirus infection status, increased GLUT-1 expression, increased PD-L1 expression, and HPV E7 RNA expression will serve as biomarkers for higher mortality in patients with anal squamous cell carcinoma. METHODS: A retrospective, translational case series was conducted on twenty-two subjects to analyze PD-L1, GLUT1, HPV-ISH, and HPV circE7 in relation to the clinical features and overall survival of patients with anal squamous cell carcinoma. To supplement understanding of the HPV circE7 biomarker, bioinformatic analyses of RNA-Seq data from the Cancer Genome Atlas was performed on 875 subjects with HPV-driven head and neck cancer and cervical cancer. RESULTS: Improved overall survival could be predicted histologically by pure basaloid architecture (p=0.013), PD-L1 expression (p=0.08), HPV-ISH positivity (p<0.001), but not GLUT1 expression. Quantitative RT-PCR of archived tumors revealed that high levels of circE7 in ASCC were predictive of improved overall survival (p=0.023). Bioinformatic analyses suggested that the presence of high amounts of circE7 correlated to improved survival in 875 subjects (p=0.074). CONCLUSION: Glut-1 overexpression was ubiquitous among all anal squamous cell carcinoma cases but was not predictive of survival. This study adds to the growing evidence of PD-L1 expression correlation to improved survival in ASCC. CircE7 levels correlate with improved survival in anal squamous cell carcinoma but larger, prospective studies are necessary to confirm the potential role of circE7 as a biomarker.Item Folate Receptor Beta Targeting for In Vivo Optical Imaging of Head and Neck Squamous Cell Carcinoma(2013-01-22) Sun, Joel; Thibodeaux, Joel; Huang, Gang; Wang, Yiguang; Gao, Jinming; Low, Philip S.; Sumer, Baran D.OBJECTIVE: The folate receptor (FR) is a high-affinity folic acid binding endocytic receptor uncommonly expressed in normal tissues. The α isoform (FR-α) is overexpressed in a variety of epithelial neoplastic cells. In contrast, functional expression of the β isoform (FR-β) is limited to activated macrophages. Importantly, in many malignancies FR serves as a target for the delivery of tumor specific drugs and imaging markers. Folic acid conjugated fluorescent dyes have been used to guide tumor resection in mouse models and humans. However, their potential utility in head and neck squamous cell carcinoma (HNSCC) is unclear due an incomplete characterization of FR expression in such tumors. We hypothesized that tumor infiltrating macrophages expressing FR-β could allow fluorescent visualization of HNSCC tumors using folate conjugated dyes even when FR expression in cancer cells is low. SUBJECTS AND METHODS: Immunohistochemistry was performed on a tissue microarray (TMA) containing primary tumor tissue and matched tumor free surgical margins from 22 patients who underwent HNSCC resection. Primary tumor sites included the oral tongue, base of tongue, tonsil, supraglottic larynx, glottic larynx and hypopharynx. We evaluated the expression of FR-α, FR-β, TGF-β, CD68 and arginase-1. To examine the use of folate targeting for image guided surgery, orthotopic xenograft HNSCC tumor models were generated from nude mice. The mice received 0.8 mg/kg intravenous injections of fluorescein isothiocyanate conjugated folate (Folate-FITC) and were imaged for fluorescent emission under 495nm light two hours later. RESULTS: No FR-α expression was observed in any TMA tumor specimen. All tumor samples demonstrated positive FR-β expression. Cellular morphology and CD68 expression identified the FR-β expressing cells as tumor infiltrating macrophages. No association was observed between FR-β staining and either TGF-β or arginase-1 staining. In tumor xenograft mouse models, tumors showed strong fluorescence in vivo after folate-FITC injection. Normal salivary glands and surrounding neck muscles did not demonstrate significant fluorescence. Histologic examination of the xenografts revealed that fluorescence within the tumors was confined to areas of inflammatory cell infiltration, consistent with our TMA data. Conclusion: HNSCC tumors contain a significant population of FR-β expressing macrophages. In contrast to many other carcinomas, the HNSCC tumor cells in our TMA did not express FR-α. By targeting tumor infiltrating macrophages, the folate linked delivery of fluorescent dyes can facilitate image guided HNSCC resection even when the tumor cells themselves do not express FR.Item Oncogene-Induced Signaling Heterogeneity in Lung Cancer(2015-07-22) Deb, Dhruba; Garcia, Christine K.; White, Michael A.; Cobb, Melanie H.; Altschuler, Steven J.; Wu, Lani; Minna, John D.Lung cancer causes the maximum number of cancer related deaths worldwide. In recent years, the cancer genome atlas (TCGA) initiative has identified 138 frequently occurring driver oncogenes and tumor suppressor genes in lung cancer. Currently, only 15 of these genes can be targeted therapeutically. Study of downstream signaling alterations of these oncogenes and tumor suppressor genes may identify novel therapeutic targets. Although studies on genetic heterogeneity in subclonal populations within one tumor using deep sequencing and multiple sectioning have gained popularity recently, the signaling heterogeneity within tumor cells with identical genetic changes remain poorly understood. Hence, I focus on TP53, KRas and C-Myc as they are among the most frequently occurring oncogenic alterations in lung adenocarcinoma. The downstream signaling changes of these genes may be different from one cell to another. Here, I develop high throughput approaches to study alterations of 6 major signaling readouts - phospho-Erk1/2, phospho-Stat3, Smad2/3, β-catenin, P65, and Foxo1 and quantitatively analyze thousands of cells with defined set of genetic changes. I ask - Can I utilize oncogene-induced signaling alterations in single cells to identify novel targetable vulnerabilities? Using single-cell image analysis I show that the genetically transformed HBECs with all 3 oncogenic changes (TP53, KRas and C-Myc) show significant signaling heterogeneity. They exhibit downregulated Smad2/3 signaling in single cells. Next, using a dominant negative construct, I confirm that this phenotype is partially reversible by the removal of C-Myc oncogenic stress. I further observe that the transformed HBECs exhibit upregulated Stat3 signaling in single cells. In addition, the Stat3 inhibitor Stattic causes more cell death in transformed HBECs. Interestingly, our single-cell image analysis suggests that Stat3 upregulation and Smad2/3 downregulation are mutually exclusive. Hence, Stattic will not be able to target the Smad2/3 downregulated cells. To target Smad2/3 downregulated cells, I identify Bcl6, a downstream target of Smad2/3, and I show that Bcl6 is a novel targetable vulnerability in transformed HBECs. I observe that C-Myc and Bcl6 gene expressions are strongly correlated in cell populations as well as in single-cell level. I further show that Bcl6 can be a targetable vulnerability in a subset of c-Myc addicted non-small cell lung cancers. I conclude that single-cell analysis of driver oncogenes and their downstream signaling can identify novel targetable vulnerabilities.Item Studies of the Molecular Features of Brd4-Nut and P300 That Contribute to Condensate Formation and Transcriptional Regulation(2023-05-01T05:00:00.000Z) Kosno, Martyna Olga; Kohler, Jennifer J.; Tu, Benjamin; Henne, W. Mike; Brekken, Rolf A.; Liszczak, Glen; Rosen, Michael K.Aberrant formation of biomolecular condensates has been proposed to play a role in several cancers. The oncogenic fusion protein Brd4-Nut drives aberrant gene expression and forms condensates in Nut Carcinoma (NC). It has not been clear how these condensates form and whether they modulate gene expression. Here, I dissected the molecular features of Brd4-Nut and a histone acetyltransferase (HAT), p300, and analyzed their contribution to condensate formation and transcriptional changes. I determined that a minimal fragment of Nut (MIN) in fusion with Brd4 is necessary and sufficient for binding to p300, and for condensate formation. A Brd4-p300 fusion protein also forms condensates and drives a transcriptional profile similar to Brd4-Nut(MIN). The intrinsically disordered regions, transcription factor - binding domains, and HAT activity of p300 all collectively contribute to condensate formation. Conversely, only HAT activity appears to be necessary to mimic the transcriptional profile of cells expressing Brd4-Nut. My results suggest that interaction of Brd4-Nut with p300 is important for aberrant condensate formation, and that multiple, yet distinct, regions of p300 contribute to condensate formation and transcriptional regulation.