Michelle D. Williams

Epithelial to Mesenchymal Transition in Head and Neck Squamous Carcinoma : Association of Src Activation With E-cadherin Down-regulation, Vimentin Expression, and Aggressive Tumor Features

Epithelial–mesenchymal transformations (EMT) are critical for the invasion, progression, and metastasis of epithelial carcinogenesis. The role of EMT in head and neck squamous carcinoma (HNSC) tumorigenesis remains unexplored. In the current study, the expressions of several factors associated with the induction of EMT in HNSC cell lines and tumor specimens were investigated to define their functional and pathologic role in HNSC.

Subcellular-resolution molecular imaging within living tissue by fiber microendoscopy

Conventional histopathology involves sampling, sectioning and staining of tissue specimens prior to microscopic evaluation, and provides diagnostic information at a single location and point in time. In vivo microscopy and molecular-targeted optical labeling are two rapidly developing fields, which together have the potential to provide anatomical and functional indications of disease by staining and imaging tissue in situ. To address the need for high-resolution imaging instrumentation, we have developed a compact, robust, and inexpensive fiber-optic microendoscopy system based around wide-field LED illumination, a flexible 1 mm diameter fiber-optic bundle, and a color CCD camera. We demonstrate the sub-cellular resolution imaging capabilities of the system through a series of experiments, beginning with simultaneous imaging of three different cancer cell lines in culture, each targeted with a distinct fluorescent label. We used the narrow diameter probe to access subcutaneous tumors in an in vivo murine model, allowing direct comparison of microendoscopy images with macroscopic images and histopathology. A surgically resected tissue specimen from the human oral cavity was imaged across the clinical margin, demonstrating qualitative and quantitative distinction between normal and cancerous tissue based on sub-cellular image features. Finally, the fiber-optic microendoscope was used on topically-stained normal human oral mucosa in vivo, resolving epithelial cell nuclei and membranes in real-time fluorescence images. Our results demonstrate that this imaging system can potentially complement conventional diagnostic techniques, and support efforts to translate emerging molecular-diagnostic and therapeutic agents into clinical use.

Treatment with Tyrosine Kinase Inhibitors for Patients with Differentiated Thyroid Cancer: the M. D. Anderson Experience

OBJECTIVES Until recently, treatment options for patients with progressive, radioactive iodine-resistant differentiated thyroid cancer (DTC) have been limited. In our clinical practice, we have begun to use sorafenib and sunitinib for patients with progressive DTC who are not able or willing to participate in clinical trials. In this paper, we describe the University of Texas M. D. Anderson Cancer Centers experience with the off-label use of these tyrosine kinase inhibitors for DTC. METHODS Adult patients were included if they had a diagnosis of radioactive iodine-refractory DTC, were treated with single agent sorafenib or sunitinib, and had both baseline and at least one follow-up scan for restaging purposes. All imaging data were collected, as well as the TSH-suppressed thyroglobulin (Tg) levels corresponding to each scan date. The primary endpoints were radiographic response and progression-free survival (PFS). Secondary objectives were tissue-specific radiographic responses and correlation of Tg with overall response. RESULTS We identified 33 patients from our clinical database. Fifteen patients (nine women, six men) met inclusion criteria, with a median age of 61 yr (range, 38-83 yr). Eight patients had papillary and seven had follicular thyroid carcinoma. Sorafenib was used in 13 and sunitinib in two, including one patient who failed prior sorafenib therapy. All patients had evidence of progressive disease (PD) before start of therapy, with a median PFS of only 4 months. Best response in target lesions was: partial response (PR) in three (20%), stable disease (SD) in nine (60%), and PD in three (20%). Clinical benefit (PR+SD) was 80%. The sunitinib patient previously refractory to sorafenib had a 38% reduction in tumor size. The most noticeable organ-specific response was observed in lung (median change, -22%) compared to lymph nodes (median change, 0%). Pleural disease and nonirradiated bone metastases demonstrated PD. All histological subtypes had similar responses. The median PFS was 19 months. The median overall survival has not yet been reached, but at 2 yr of follow-up, overall survival is 67%. Log Tg correlated with radiographic response (P = 0.0005). CONCLUSIONS Sorafenib and sunitinib appear to be effective in patients with widely metastatic, progressive DTC, with most patients achieving SD or PR, despite having PD at baseline. The most noticeable responses occurred in the lungs in contrast with minimal changes in nodal metastases and PD in pleural and nonirradiated bone metastases, suggesting a tissue-specific response to therapy. Log Tg significantly correlated with response to treatment and therefore may have value as a surrogate marker of response.

Prospective Risk-Adjusted [18F]Fluorodeoxyglucose Positron Emission Tomography and Computed Tomography Assessment of Radiation Response in Head and Neck Cancer

PURPOSE [(18)F]Fluorodeoxyglucose positron emission tomography (FDG-PET)/computed tomography (CT) imaging may improve assessment of radiation response in patients with head and neck cancer, but it is not yet known for which patients this is most useful. We conducted a prospective trial to identify patient populations likely to benefit from the addition of functional imaging to the assessment of radiotherapy response. PATIENTS AND METHODS Ninety-eight patients with locally advanced cancer of the oropharynx, larynx, or hypopharynx were prospectively enrolled and treated with primary radiotherapy, with or without chemotherapy. Patients underwent FDG-PET/CT and contrast-enhanced CT imaging 8 weeks after completion of treatment. Functional and anatomic imaging response was correlated with clinical and pathologic response. Imaging accuracy was then compared between imaging modalities. RESULTS Although postradiation maximum standard uptake values were significantly higher in nonresponders compared with responders, the positive and negative predictive values of FDG-PET/CT scanning were similar to those for CT alone in the unselected study population. Subset analyses revealed that FDG-PET/CT outperformed CT alone in response assessment for patients at high risk for treatment failure (those with human papillomavirus [HPV] -negative disease, nonoropharyngeal primaries, or history of tobacco use). No benefit to FDG-PET/CT was seen for low-risk patients lacking these features. CONCLUSION These data do not support the broad application of FDG-PET/CT for radiation response assessment in unselected head and neck cancer patients. However, FDG-PET/CT may be the imaging modality of choice for patients with highest risk disease, particularly those with HPV-negative tumors. Optimal timing of FDG-PET/CT imaging after radiotherapy merits further investigation.

CRTC1/MAML2 fusion transcript in high grade mucoepidermoid carcinomas of salivary and thyroid glands and Warthin's tumors: Implications for histogenesis and biologic behavior

We analyzed 55 primary salivary gland tumors including 22 mucoepidermoid carcinomas (MECs) to determine the association of MECT1/TORC1/CRTC1‐MAML2 fusion transcript to tumor types, level of MEC differentiation and clinicopathologic parameters. Our primary salivary gland tumors were composed of 22 MECs, 11 Warthins tumors, 10 adenoid cystic carcinomas, two basaloid carcinomas, five salivary duct carcinomas, and five adenocarcinomas, not otherwise specified. We also included, for the first time, three primary MECs of the thyroid gland. We used nested RT‐PCR and subsequent sequencing techniques for detection and verification of the fusion transcript in fresh and archival specimens. Eighteen (81%) of the 22 primary salivary and one of the three thyroid glands with MEC were positive for the fusion transcript. The transcript was detected equally in low, intermediate and high grade as well as low and high stage MECs. Significant correlation between fusion negative tumors and distant metastasis was noted (P = 0.005). Four (36%) of the 11 Warthins tumors were also positive for the transcript. None of the 22 primary non‐MEC gland salivary carcinomas were positive for the transcript. We conclude that the CRTC1/MAML2 transcript may be detected in both low and high grade MEC, that fusion negative tumors may define a subset of biologically aggressive MECs tumors, that the fusion is present in primary MECs of the thyroid gland and is also detectable in Warthins tumor, and that a subset of MECs can be targeted for therapeutic intervention.

Objective Detection and Delineation of Oral Neoplasia Using Autofluorescence Imaging

Although the oral cavity is easily accessible to inspection, patients with oral cancer most often present at a late stage, leading to high morbidity and mortality. Autofluorescence imaging has emerged as a promising technology to aid clinicians in screening for oral neoplasia and as an aid to resection, but current approaches rely on subjective interpretation. We present a new method to objectively delineate neoplastic oral mucosa using autofluorescence imaging. Autofluorescence images were obtained from 56 patients with oral lesions and 11 normal volunteers. From these images, 276 measurements from 159 unique regions of interest (ROI) sites corresponding to normal and confirmed neoplastic areas were identified. Data from ROIs in the first 46 subjects were used to develop a simple classification algorithm based on the ratio of red-to-green fluorescence; performance of this algorithm was then validated using data from the ROIs in the last 21 subjects. This algorithm was applied to patient images to create visual disease probability maps across the field of view. Histologic sections of resected tissue were used to validate the disease probability maps. The best discrimination between neoplastic and nonneoplastic areas was obtained at 405 nm excitation; normal tissue could be discriminated from dysplasia and invasive cancer with a 95.9% sensitivity and 96.2% specificity in the training set, and with a 100% sensitivity and 91.4% specificity in the validation set. Disease probability maps qualitatively agreed with both clinical impression and histology. Autofluorescence imaging coupled with objective image analysis provided a sensitive and noninvasive tool for the detection of oral neoplasia.

Clinical responses to vemurafenib in patients with metastatic papillary thyroid cancer harboring BRAFV600E mutation

BACKGROUND Clinical benefit from cytotoxic chemotherapy for metastatic papillary thyroid carcinoma (PTC) is disappointing, and effective therapeutic approaches for these patients are urgently needed. Because kinase-activating mutations in the BRAF proto-oncogene commonly occur in advanced PTC, and inhibition of BRAF(V600E) has shown promising clinical activity in melanoma, BRAF inhibitor therapy may be an effective strategy to treat metastatic PTC. METHODS The dose escalation portion of a first-in-human, phase I study of vemurafenib, a selective RAF inhibitor, included three patients with metastatic PTC harboring the BRAF(V600E) mutation. Vemurafenib was initially dosed at 240-360 mg twice a day, later escalated to 720 mg twice a day. Response evaluation was performed every 8 weeks per Response Evaluation Criteria in Solid Tumors (RECIST). RESULTS Among the three patients, one had a confirmed partial response with reduction of pulmonary target lesions by 31%, and the duration of response was 7.6 months before the disease progressed in the lungs and the bones. The time to progression was 11.7 months. The other two patients had stable disease, and the time to progression was 13.2 and 11.4 months, respectively. CONCLUSIONS Vemurafenib appears to have a promising clinical activity in patients with metastatic PTC, and our data suggest that the BRAF(V600E) mutant kinase is a relevant target for therapy in this patient population. Further investigation of inhibitors of mutated BRAF kinase in patients with PTC in a phase II study is warranted.

Differential expression of hormonal and growth factor receptors in salivary duct carcinomas : Biologic significance and potential role in therapeutic stratification of patients

Salivary duct carcinoma (SDC), a rare malignancy, manifests remarkable morphologic and biologic resemblance to high-grade mammary ductal carcinoma. We contend that, similar to mammary ductal carcinoma, hormones and growth factors may play a role in SDCs. Our aim was to determine the incidence and clinical significance of the expression of several hormone and growth factor receptors and evaluate their potential in therapeutic stratification of SDC patients in the largest cohort studied to date. Eighty-four archived tumor tissue blocks were analyzed immunohistochemically for expression of estrogen receptor-β (ERβ), androgen receptor (AR), and proline, glutamic acid, and leucine-rich protein-1 and growth factor receptors HER-2 and epidermal growth factor receptor. The results were correlated with available pathologic, demographic, and clinical data from 59 of 84 cases. Proline, glutamic acid, and leucine-rich protein-1, ERβ, and AR were expressed individually in 94% (71/76), 73% (57/80), and 67% (56/84) of SDCs, respectively, and coexpressed in 45% (34/75). AR was expressed significantly more often in SDCs of men than in SDCs of women [79% (35/57) vs. 33% (9/27), P<0.001]. Epidermal growth factor receptor and HER-2 were overexpressed individually in 48% (40/83) and 25% (21/84), respectively, and co-overexpressed in 12% (10/83). Survival decreased significantly in patients with lymph node metastasis (P=0.002) and positive surgical margins (P=0.006). Lack of ERβ expression correlated with increased local and regional recurrence (P=0.05 and P=0.002, respectively). Together, these results indicate that (a) ERβ down-regulation is associated with adverse clinical features, (b) lymph node and surgical margin status are significant survival factors, and (c) the differential expression of these hormones and growth factor receptors may assist in triaging patients with SDC for novel therapies.

Multispectral optical imaging device for in vivo detection of oral neoplasia

A multispectral digital microscope (MDM) is designed and constructed as a tool to improve detection of oral neoplasia. The MDM acquires in vivo images of oral tissue in fluorescence, narrow-band (NB) reflectance, and orthogonal polarized reflectance (OPR) modes, to enable evaluation of lesions that may not exhibit high contrast under standard white light illumination. The device rapidly captures image sequences so that the diagnostic value of each modality can be qualitatively and quantitatively evaluated alone and in combination. As part of a pilot clinical trial, images are acquired from normal volunteers and patients with precancerous and cancerous lesions. In normal subjects, the visibility of vasculature can be enhanced by tuning the reflectance illumination wavelength and polarization. In patients with histologically confirmed neoplasia, we observe decreased blue/green autofluorescence and increased red autofluorescence in lesions, and increased visibility of vasculature using NB and OPR imaging. The perceived lesion borders change with imaging modality, suggesting that multimodal imaging has the potential to provide additional diagnostic information not available using standard white light illumination or by using a single imaging mode alone.

Landscape of DNA Virus Associations across Human Malignant Cancers: Analysis of 3,775 Cases Using RNA-Seq

ABSTRACT Elucidation of tumor-DNA virus associations in many cancer types has enhanced our knowledge of fundamental oncogenesis mechanisms and provided a basis for cancer prevention initiatives. RNA-Seq is a novel tool to comprehensively assess such associations. We interrogated RNA-Seq data from 3,775 malignant neoplasms in The Cancer Genome Atlas database for the presence of viral sequences. Viral integration sites were also detected in expressed transcripts using a novel approach. The detection capacity of RNA-Seq was compared to available clinical laboratory data. Human papillomavirus (HPV) transcripts were detected using RNA-Seq analysis in head-and-neck squamous cell carcinoma, uterine endometrioid carcinoma, and squamous cell carcinoma of the lung. Detection of HPV by RNA-Seq correlated with detection by in situ hybridization and immunohistochemistry in squamous cell carcinoma tumors of the head and neck. Hepatitis B virus and Epstein-Barr virus (EBV) were detected using RNA-Seq in hepatocellular carcinoma and gastric carcinoma tumors, respectively. Integration sites of viral genes and oncogenes were detected in cancers harboring HPV or hepatitis B virus but not in EBV-positive gastric carcinoma. Integration sites of expressed viral transcripts frequently involved known coding areas of the host genome. No DNA virus transcripts were detected in acute myeloid leukemia, cutaneous melanoma, low- and high-grade gliomas of the brain, and adenocarcinomas of the breast, colon and rectum, lung, prostate, ovary, kidney, and thyroid. In conclusion, this study provides a large-scale overview of the landscape of DNA viruses in human malignant cancers. While further validation is necessary for specific cancer types, our findings highlight the utility of RNA-Seq in detecting tumor-associated DNA viruses and identifying viral integration sites that may unravel novel mechanisms of cancer pathogenesis.