Transcriptomic analysis-guided assessment of precision-cut tumor slices (PCTS) as an ex-vivo tool in cancer research.

With cancer immunotherapy and precision medicine dynamically evolving, there is greater need for pre-clinical models that can better replicate the intact tumor and its complex tumor microenvironment (TME). Precision-cut tumor slices (PCTS) have recently emerged as an ex vivo human tumor model, offering the opportunity to study individual patient responses to targeted therapies, including immunotherapies. However, little is known about the physiologic status of PCTS and how culture conditions alter gene expression. In this study, we generated PCTS from head and neck cancers (HNC) and mesothelioma tumors (Meso) and undertook transcriptomic analyses to understand the changes that occur in the timeframe between PCTS generation and up to 72 h (hrs) in culture. Our findings showed major changes occurring during the first 24 h culture period of PCTS, involving genes related to wound healing, extracellular matrix, hypoxia, and IFNγ-dependent pathways in both tumor types, as well as tumor-specific changes. Collectively, our data provides an insight into PCTS physiology, which should be taken into consideration when designing PCTS studies, especially in the context of immunology and immunotherapy.

Intraoperative Detection and Assessment of Lung Nodules.

Lung cancer is the most frequent cause of cancer-related death worldwide. Despite advances in systemic therapy, the 5-year survival remains humbling at 4% to 17%. For those diagnosed early, surgical therapy can yield potentially curative results. Surgical resection remains a cornerstone of medical care. Success hinges on sound oncologic resection principles. Various techniques can be used to identify pulmonary nodules. A challenge is intraoperative assessment of the surgical specimen to confirm disease localization and ensure an R0 resection. The primary tool is frozen section. Understanding the options available enhances the arsenal of thoracic surgeons and leads to better patient care.

Intraoperative Near-infrared Imaging Can Identify Neoplasms and Aid in Real-time Margin Assessment During Pancreatic Resection.

OBJECTIVE: To determine if intraoperative near-infrared (NIR) imaging carries benefit in resection of pancreatic neoplasms. BACKGROUND: Resection of pancreatic malignancies is hindered by high rates of local and distant recurrence from positive margins and unrecognized metastases. Improved tumor visualization could improve outcomes. We hypothesized that intraoperative NIR imaging with a clinically approved optical contrast agent could serve as a useful adjunct in assessing margins and extent of disease during pancreatic resections. METHODS: Twenty patients were enrolled in an open-label clinical trial from July 2016 to May 2018. Subjects received second window indocyanine green (ICG) (2.5-5 mg/kg) 24 hours prior to pancreatic resection. NIR imaging was performed during staging laparoscopy and after pancreas mobilization in situ and following resection ex vivo. Tumor fluorescence was quantified using tumor-to-background ratio (TBR). Fluorescence at the specimen margin was compared to pathology evaluation. RESULTS: Procedures included 9 pancreaticoduodenectomies, 10 distal pancreatectomies, and 1 total pancreatectomy; 21 total specimens were obtained. Three out of 8 noninvasive tumors were fluorescent (mean TBR 2.59 ±â€Š2.57). Twelve out of 13 invasive malignancies (n = 12 pancreatic adenocarcinoma, n = 1 cholangiocarcinoma) were fluorescent (mean TBR 4.42 ±â€Š2.91). Fluorescence at the transection margin correlated with final pathologic assessment in 12 of 13 patients. Following neoadjuvant therapy, 4 of 5 tumors were fluorescent; these 4 tumors showed no treatment response on pathology assessment. One tumor had a significant treatment response and showed no fluorescence. CONCLUSIONS: Second window ICG reliably accumulates in invasive pancreatic malignancies and provides real-time feedback during pancreatectomy. NIR imaging may help to assess the response to neoadjuvant therapy.

Implications of Hospital Volume on Costs Following Esophagectomy in the United States.

BACKGROUND: With increasing focus on health care quality and cost containment, volume-based referral strategies have been proposed to improve value in high-cost procedures, such as esophagectomy. While the effect of hospital volume on outcomes has been demonstrated, our goal was to evaluate the economic consequences of volume-based referral practices for esophagectomy. METHODS: The nationwide inpatient sample (NIS) was queried for the years 2004-2013 for all patients undergoing esophagectomy. Patients were stratified by hospital volume quartile and substratified by preoperative risk and age. Clustered multivariable hierarchical logistic regression analysis was used to assess adjusted costs and mortality. RESULTS: In total, 9270 patients were clustered based on annual hospital volume quartiles of < 7, 7 to 22, 23 to 87, and > 87 esophagectomies. After stratification by patient variables, high-volume centers performed esophagectomies in high-risk patients at the same cost as low-volume centers without significant difference in resource utilization. Overall, mortality decreased across volume quartiles (lowest 8.9 versus highest 3.6%, p < 0.0001). The greatest volume-mortality differences were observed among patients aged between 70 and 80 years (lowest 12.2 versus highest 6.2%, p = 0.009). Patients with high preoperative risk also derived mortality benefits with increasing hospital volume (lowest 17.5 versus highest 11.8%, p < 0.0001). CONCLUSIONS: This study demonstrates that the mortality improvements for high-risk patients undergoing esophagectomy at high-volume centers do not come at increased costs. These results suggest that health systems should consider selectively referring high-risk patients to high-volume centers within their region.

Assessment of folate receptor alpha and beta expression in selection of lung and pancreatic cancer patients for receptor targeted therapies.

A number of folate receptor (FR) targeted small molecular drugs and monoclonal antibodies have been introduced into clinical trials to treat FR positive cancers. Because the therapeutic efficacy of these drugs depends prominently on the level of FR-α expression on the cancer cells, patients have been commonly selected for FR-targeted therapies based on the intensity of a folate-targeted radioimaging agent. Unfortunately, uptake of such imaging agents can be mediated by both major isoforms of the folate receptor, FR-α and FR-ß. Logically, if the FR positive cell population in a tumor mass is dominated by FR-ß positive macrophages, patients could be selected for therapy that have few FR-expressing cancer cells. Although several IHC studies have examined expression of either FR-α or FR-ß, no study to date has investigated expression of both FR-α and FR-ß in the same tumor mass. Herein, we utilize monoclonal antibodies specific for FR-α (mAb343) and FR-ß (m909) to query each isoform's expression in a range of cancers. We show that lung and pancreatic adenocarcinomas express the full spectrum of FR-α and FR-ß combinations with ~76% of lung adenocarcinomas expressing both FR-α and FR-ß while pancreatic cancers express primarily FR-ß. Thus, while folate-targeted imaging of lung cancer patients might accurately reflect the expression of FR-α on lung cancer cells, imaging of pancreatic cancer patients could mislead a physician into treating a nonresponding patient. Overall, these data suggest that an independent analysis of both FR-α and FR-ß should be obtained to predict the potential efficacy of a folate-targeted drug.

PDT dose dosimetry for Photofrin-mediated pleural photodynamic therapy (pPDT).

Photosensitizer fluorescence excited by photodynamic therapy (PDT) treatment light can be used to monitor the in vivo concentration of the photosensitizer and its photobleaching. The temporal integral of the product of in vivo photosensitizer concentration and light fluence is called PDT dose, which is an important dosimetry quantity for PDT. However, the detected photosensitizer fluorescence may be distorted by variations in the absorption and scattering of both excitation and fluorescence light in tissue. Therefore, correction of the measured fluorescence for distortion due to variable optical properties is required for absolute quantification of photosensitizer concentration. In this study, we have developed a four-channel PDT dose dosimetry system to simultaneously acquire light dosimetry and photosensitizer fluorescence data. We measured PDT dose at four sites in the pleural cavity during pleural PDT. We have determined an empirical optical property correction function using Monte Carlo simulations of fluorescence for a range of physiologically relevant tissue optical properties. Parameters of the optical property correction function for Photofrin fluorescence were determined experimentally using tissue-simulating phantoms. In vivo measurements of photosensitizer fluorescence showed negligible photobleaching of Photofrin during the PDT treatment, but large intra- and inter-patient heterogeneities of in vivo Photofrin concentration are observed. PDT doses delivered to 22 sites in the pleural cavity of 8 patients were different by 2.9 times intra-patient and 8.3 times inter-patient.

Novel Methods of Intraoperative Localization and Margin Assessment of Pulmonary Nodules.

Lung cancer screening has lead to frequent diagnosis of solitary pulmonary nodules, many of which require surgical biopsy for diagnosis and intervention. Subcentimeter and central nodules are particularly difficult to visualize or palpate during surgery, thus nodule localization can be a difficult problem for the thoracic surgeon. Although minimally invasive techniques including transthoracic computed tomography and bronchoscopic-guided biopsy may establish a diagnosis, these methods do not help locate nodules during surgery and can lead to inadequate tissue sampling. Therefore, surgical biopsy is often required for diagnosis and management of solitary pulmonary nodules. Additionally, after an excision, intraoperative margin assessment is important to prevent local recurrence. This is important for bronchial margins following lobectomy or parenchymal margins following sublobar resection. First, we examine methods of preoperative lesion marking, including wire placement, dye marking, ultrasound, fluoroscopy, and molecular imaging. Second, we describe the current state of the art in intraoperative margin assessment techniques.

Prospective study of proton beam radiation therapy for adjuvant and definitive treatment of thymoma and thymic carcinoma: Early response and toxicity assessment.

BACKGROUND AND PURPOSE: Radiation is an important modality in treatment of thymic tumors. However, toxicity may reduce its overall benefit. We hypothesized that double-scattering proton beam therapy (DS-PT) can achieve excellent local control with limited toxicity in patients with thymic malignancies. METHODS AND MATERIALS: Patients with thymoma or thymic carcinoma treated with DS-PT between 2011 and 2015 were prospectively analyzed for toxicity and patterns of failure on an IRB-approved study. RESULTS: Twenty-seven consecutive patients were evaluated. Patients were a median of 56 years and had thymoma (85%). They were treated with definitive (22%), salvage (15%) or adjuvant (63%) DS-PT to a median of 61.2/1.8 Gy [CGE]. No patient experienced grade ⩾3 toxicity. Acute grade 2 toxicities included dermatitis (37%), fatigue (11%), esophagitis (7%), and pneumonitis (4%). Late grade 2 toxicity was limited to a single patient with chronic dyspnea. At a median follow-up of 2 years, 100% local control was achieved. Three-year regional control, distant control, and overall survival rates were 96% (95% CI 76-99%), 74% (95% CI 41-90%), and 94% (95% CI 63-99%), respectively. CONCLUSIONS: This is the first cohort and prospective series of proton therapy to treat thymic tumors, demonstrating low rates of early toxicity and excellent initial outcomes.

Assessment of folate receptor-ß expression in human neoplastic tissues.

Over-expression of folate receptor alpha on cancer cells has been frequently exploited for delivery of folate-targeted imaging and therapeutic agents to tumors. Because limited information exists on expression of the beta isoform of the folate receptor in human cancers (FR-ß), we have evaluated the immunohistochemical staining pattern of FR-ß in 992 tumor sections from 20 different human cancer types using a new anti-human FR-ß monoclonal antibody. FR-ß expression was shown to be more pronounced in cells within the stroma, primarily macrophages and macrophage-like cells than cancer cells in every cancer type studied. Moreover, FR-ß expression in both cancer and stromal cells was found to be statistically more prominent in females than males. A significant positive correlation was also observed between FR-ß expression on stromal cells and both the stage of the cancer and the presence of lymph node metastases. Based on these data we conclude FR-ß may constitute a good target for specific delivery of therapeutic agents to activated macrophages and that accumulation of FR-ß positive macrophages in the stroma could serve as a useful indicator of a tumor's metastatic potential.

Characterization of surgical models of postoperative tumor recurrence for preclinical adjuvant therapy assessment.

PURPOSE: Nearly 30% of cancer patients undergoing curative surgery succumb to distant recurrent disease. Despite large implications and known differences between primary and recurrent tumors, preclinical adjuvant therapy evaluation frequently occurs only in primary tumors and not recurrent tumors. We hypothesized that well characterized and reproducible models of postoperative systemic recurrences should be used for preclinical evaluation of adjuvant approaches. EXPERIMENTAL DESIGN: We examined traditional animal models of cancer surgery that generate systemic cancer recurrences. We also investigated models of systemic cancer recurrences that incorporate spontaneously metastatic cell lines and surgical resection. For each model, we critiqued feasibility, reproducibility and similarity to human recurrence biology. Using our novel model, we then tested the adjuvant use of a novel systemic inhibitor of TGF-ß, 1D11. RESULTS: Traditional surgical models are confounded by immunologic factors including concomitant immunity and perioperative immunosuppression. A superior preclinical model of postoperative systemic recurrences incorporates spontaneously metastatic cell lines and primary tumor excision. This approach is biologically relevant and readily feasible. Using this model, we discovered that "perioperative" TGF-ß blockade has strong anti-tumor effects in the setting of advanced disease that would not be appreciated in primary tumor cell lines or other surgical models. CONCLUSIONS: There are multiple immunologic effects that rendered previous models of postoperative cancer recurrences inadequate. Use of spontaneously metastatic cell lines followed by surgical resection eliminates these confounders, and best resembles the clinical scenario. This preclinical model provides more reliable preclinical information when evaluating new adjuvant therapies.