Fluorescence Imaging-Guided Identification of Thymic Masses Using Low-Dose Indocyanine Green.

BACKGROUND: Indocyanine green (ICG) fluorescence imaging has been used to detect many types of tumors during surgery; however, there are few studies on thymic masses and the dose and time of ICG injection have not been optimized. OBJECTIVE: We aimed to evaluate the optimal ICG injection dose and timing for detecting thymic masses during surgery. METHOD: Forty-nine consecutive patients diagnosed with thymic masses on preoperative computed tomography (CT) and scheduled to undergo thymic cystectomy or thymectomy were included. Patients were administered 1, 2, or 5 mg/kg of ICG at different times. Thymic masses were observed during and after surgery using a near-infrared fluorescence imaging system, and the fluorescence signal tumor-to-normal ratio (TNR) was analyzed. RESULTS: Among the 49 patients, 14 patients with thymic cysts showed negative fluorescence signals, 33 patients with thymoma or thymic carcinoma showed positive fluorescence signals, and 2 patients showed insufficient fluorescence signals. The diagnosis of thymic masses based on CT was correct in 32 (65%) of 49 cases; however, the differential diagnosis of thymic masses based on NIR signals was correct in 47 of 49 cases (96%), including 14 cases of thymic cysts (100%) and 33 cases of thymomas or thymic carcinomas (94%). In addition, TNR was not affected by the time or dose of ICG injection, histological type, stage, or tumor size. CONCLUSIONS: Low-dose intravenous injection of ICG at flexible time can detect thymic tumors. In addition, thymic cysts can be distinguished from thymomas or thymic carcinomas during surgery by the absence of ICG fluorescence signals.

GCC2 as a New Early Diagnostic Biomarker for Non-Small Cell Lung Cancer.

No specific markers have been identified to detect non-small cell lung cancer (NSCLC) cell-derived exosomes circulating in the blood. Here, we report a new biomarker that distinguishes between cancer and non-cancer cell-derived exosomes. Exosomes isolated from patient plasmas at various pathological stages of NSCLC, NSCLC cell lines, and human pulmonary alveolar epithelial cells isolated using size exclusion chromatography were characterized. The GRIP and coiled-coil domain-containing 2 (GCC2) protein, involved in endosome-to-Golgi transport, was identified by proteomics analysis of NSCLC cell line-derived exosomes. GCC2 protein levels in the exosomes derived from early-stage NSCLC patients were higher than those from healthy controls. Receiver operating characteristic curve analysis revealed the diagnostic sensitivity and specificity of exosomal GCC2 to be 90% and 75%, respectively. A high area under the curve, 0.844, confirmed that GCC2 levels could effectively distinguish between the exosomes. These results demonstrate GCC2 as a promising early diagnostic biomarker for NSCLC.

Design and Testing of Augmented Reality-Based Fluorescence Imaging Goggle for Intraoperative Imaging-Guided Surgery.

The different pathways between the position of a near-infrared camera and the user's eye limit the use of existing near-infrared fluorescence imaging systems for tumor margin assessments. By utilizing an optical system that precisely matches the near-infrared fluorescence image and the optical path of visible light, we developed an augmented reality (AR)-based fluorescence imaging system that provides users with a fluorescence image that matches the real-field, without requiring any additional algorithms. Commercial smart glasses, dichroic beam splitters, mirrors, and custom near-infrared cameras were employed to develop the proposed system, and each mount was designed and utilized. After its performance was assessed in the laboratory, preclinical experiments involving tumor detection and lung lobectomy in mice and rabbits by using indocyanine green (ICG) were conducted. The results showed that the proposed system provided a stable image of fluorescence that matched the actual site. In addition, preclinical experiments confirmed that the proposed system could be used to detect tumors using ICG and evaluate lung lobectomies. The AR-based intraoperative smart goggle system could detect fluorescence images for tumor margin assessments in animal models, without disrupting the surgical workflow in an operating room. Additionally, it was confirmed that, even when the system itself was distorted when worn, the fluorescence image consistently matched the actual site.

Fluorescent and Iodized Emulsion for Preoperative Localization of Pulmonary Nodules.

OBJECTIVE: This study was conducted to develop a fluorescent iodized emulsion comprising indocyanine green (ICG) solution and lipiodol (ethiodized oil) and evaluate its feasibility for use in a clinical setting. BACKGROUND: ICG use for the preoperative localization of pulmonary nodules is limited in terms of penetration depth and diffusion. METHODS: First, fluorescent microscopy was used to investigate the distribution of ICG-lipiodol emulsions prepared using different methods. The emulsions were injected in 15 lung lobes of 3 rabbits under computed tomography fluoroscopy guidance; evaluation with imaging and radiography was conducted after thoracotomy. Subsequently, the emulsions were used to preoperatively localize 29 pulmonary nodules in 24 human subjects, and wedge resections were performed using fluorescent imaging and C-arm fluoroscopy. RESULTS: The optimal emulsion of 10% ICG and 90% lipiodol mixed through 90 passages had even distribution and the highest signal intensity under fluorescent microscopy; it also had the best consistency in the rabbit lungs, which persisted for 24 hours at the injection site. In human subjects, the mean diameter of pulmonary nodules was 0.9 ±â€Š0.4 cm, and depth from the pleura was 1.2 ± 0.8 cm. All emulsion types injected were well localized around the target nodules without any side effects or procedure-related complications. Wedge resection with minimally invasive approach was successful in all pulmonary nodules with a free resection margin. CONCLUSIONS: A fluorescent iodized emulsion prepared by mixing ICG with lipiodol enabled accurate localization and resection of pulmonary nodules.

Near-infrared fluorescent imaging with indocyanine green in rabbit and patient specimens of esophageal cancer.

BACKGROUND: We aimed to assess the possibility of detecting esophageal cancer after intravenous injection of indocyanine green (ICG) in preclinical and clinical models. METHODS: Forty-five rabbits were surgically implanted with VX2 tumors into the esophageal muscular layer 2 weeks before esophagectomy. The rabbits received intravenous injection of ICG at doses of 1, 2, or 5 mg/kg at 3, 6, 12, 24, or 48 h before surgical removal of esophagus. Twelve patients scheduled to undergo esophagectomy were also enrolled, and all received 2 mg/kg of ICG intravenously at 3, 6, 12, or 24 h before surgical removal of esophagus. The fluorescence intensity was measured in all resected specimens from the rabbits and patients using a near-infrared (NIR) fluorescence imaging system after surgery. RESULTS: Esophageal tumors were successfully established in all rabbits, and fluorescent signals were detected in all animal and patient specimens. Tumor-to-normal ratio (TNR) analysis showed that higher doses resulted in a greater TNR. Injection of at least 2 mg/kg of ICG was required for clear visualization of the tumor, and the TNR was highest at 12 h after injection. The TNR in patients was also highest at 12 h (P=0.0004), with 2 mg/kg of ICG. None of the patients had major complications following ICG injection. CONCLUSIONS: NIR fluorescence imaging can be used to visualize esophageal cancer after systemic injection of ICG. ICG at 2 mg/kg at 12 h is optimal for tumor detection. However, since the clinical trials were conducted in a small number of patients, further studies are needed in larger populations.

Fluorescence Image-Based Evaluation of Gastric Tube Perfusion during Esophagogastrostomy.

During esophagectomy and esophagogastrostomy, the prediction of anastomotic leakage relies on the operating surgeon's tactile or visual diagnosis. Therefore, anastomotic leaks are relatively unpredictable, and new intraoperative evaluation methods or tools are essential. A fluorescence imaging system enables visualization over a wide region of interest, and provides intuitive information on perfusion intraoperatively. Surgeons can choose the best anastomotic site of the gastric tube based on fluorescence images in real time during surgery. This technology provides better surgical outcomes when used with an optimal injection dose and timing of indocyanine green.

Levels of Extracellular Vesicles in Pulmonary and Peripheral Blood Correlate with Stages of Lung Cancer Patients.

BACKGROUND: The extracellular vesicle (EV) concentration is known to be higher in cancer patients than in healthy individuals. Herein, we report that EV levels differ in the tumor-draining pulmonary vein blood and the peripheral blood of animal models and human subjects at different pathological stages of lung cancer. METHODS: Ten rabbits and 40 humans formed the study cohorts. Blood was collected from the peripheral vein of members of all groups. Pulmonary blood was collected intraoperatively from all groups except for the healthy human controls. Quantitative analysis of EV levels was performed using a nanoparticle tracking assay, a CD63 enzyme-linked immunosorbent assay, and western blotting. RESULTS: The EV levels in the peripheral blood of animals and patients with lung cancer were higher than those in the peripheral blood of healthy controls (p < 0.01 and p < 0.001, respectively). Moreover, for both animals and patients with lung cancer, the EV levels in the pulmonary blood were significantly higher than those in the preoperative peripheral blood (p < 0.01 and p < 0.0001, respectively). In patients, the pathological stages of lung cancer showed a higher correlation with the pulmonary EV levels than the peripheral EV levels. CONCLUSIONS: EV levels increased with increasing lung cancer grade, and this trend was more prominent in the pulmonary blood than in the peripheral blood.

Evaluation of Intraoperative Near-Infrared Fluorescence Visualization of the Lung Tumor Margin With Indocyanine Green Inhalation.

Importance: Identification of the tumor margin during surgery is important for precise minimal resection of lung tumors. Intravenous injection of indocyanine green (ICG) has several limitations when used for intraoperative visualization of lung cancer. Objectives: To describe a technique for intraoperative visualization of lung tumor margin using ICG inhalation and evaluate the clinical applicability of the technique in mouse and rabbit lung tumor models as well as lung specimens of patients with lung tumors. Design, Setting, and Participants: In lung tumor models of both mice and rabbits, the distribution of inhaled ICG in the lung tumor margin was investigated in vivo and ex vivo using a near-infrared imaging system. Lung tumor margin detection via inhalation of ICG was evaluated by comparing the results obtained with those of the intravenous injection method (n = 32, each time point for 4 mice). Based on preclinical data, use of ICG inhalation to help detect the tumor margin in patients with lung cancer was also evaluated (n = 6). This diagnostic study was conducted from May 31, 2017, to March 30, 2019. Main Outcomes and Measures: The use of tumor margin detection by inhaled ICG was evaluated by comparing the inhaled formulation with intravenous administration of ICG. Results: From 10 minutes after inhalation of ICG to 24 hours, the distribution of ICG in the lungs was significantly higher than that in other organs (signal to noise ratio in the lungs: 39 486.4; interquartile range [IQR], 36 983.74-43 592.5). Ex vivo and histologic analysis showed that, in both lung tumor models, inhaled ICG was observed throughout the healthy lung tissue but was rarely found in tumor tissue. The difference in the fluorescent signal between healthy and tumor lung tissues was associated with the mechanical airway obstruction caused by the tumor and with alveolar macrophage uptake of the inhaled ICG in healthy tissues. Inhalation at a 20-fold lower dose of ICG had a 2-fold higher efficiency for tumor margin detection than did the intravenous injection (2.9; IQR, 2.7-3.2; P < .001). Conclusions and Relevance: The results of this study suggest that lung-specific inhalation delivery of ICG is feasible and may be useful for the intraoperative visualization of lung tumor margin in clinical practice.

Simultaneous visualization of pulmonary nodules and intersegmental planes on fluorescent images in pulmonary segmentectomy.

OBJECTIVES: The technique of simultaneously visualizing pulmonary nodules and the intersegmental plane using fluorescent images was developed to measure the distance between them intraoperatively. METHODS: Patients who underwent pulmonary segmentectomy were consecutively included in this study between March 2016 and July 2019. Computed tomography or electromagnetic bronchoscopy-guided localization with indocyanine green-lipiodol emulsion was performed on the day of surgery. In the middle of the surgery, after dividing the segmental artery, vein and bronchus to a targeted segment, 0.3-0.5 mg/kg of indocyanine green was injected intravenously. RESULTS: In total, 31 patients (17 men and 14 women with a mean age of 63.2 ± 9.8 years) were included in this study. The mean size and depth of the nodules were 1.2 ± 0.5 (range 0.3-2.5) cm and 16.4 ± 9.9 (range 1.0-42.0) mm, respectively. Pulmonary nodules and intersegmental plane of all the patients were visualized using a fluorescent thoracoscope. The resection margins were more than the size of the tumour or were 2 (mean 2.4 ± 1.2) cm in size in all patients except one. The resection margin of this patient looked sufficient on the intraoperative view. However, adenocarcinoma in situ at the resection margin was identified based on the pathological report. The mean duration of the operation was 168.7 ± 53.3 min, and the chest tube was removed on an average of 4.7 ± 1.8 days after surgery in all patients. CONCLUSIONS: The dual visualization technique using indocyanine green could facilitate an easier measurement of the distance between pulmonary nodules and the intersegmental plane during pulmonary segmentectomy.

Self-targeted knockdown of CD44 improves cisplatin sensitivity of chemoresistant non-small cell lung cancer cells.

BACKGROUND: Chemoresistance remains a major challenge for effective chemotherapy of non-small-cell lung carcinoma (NSCLC). CD44 expression is related to the susceptibility of various cancer cell types to anticancer drugs. Here, we systematically investigated the CD44-dependent chemoresistance of NSCLC cells and developed a liposomal siRNA delivery system to overcome this chemoresistance by the self-targeted downregulation of CD44. METHODS: We confirmed the relationship between the expression of CD44 and the chemosensitivity of NSCLC cells using flow cytometry and MTT assay. We then generated and characterized cisplatin-resistant cell lines and compared the expression of CD44 in resistant cells to that in parental cells using western blotting. To evaluate whether the chemosensitivity of resistant cells depends on CD44 expression, we performed CD44 knockdown using CD44 siRNA and detected the chemosensitivity of these cells. Additionally, we prepared hyaluronic acid (HA)-coated liposomes as a targeted delivery system to selectively deliver CD44-specific siRNA to chemoresistant NSCLC cells and observed whether the chemosensitivity of these cells was improved. RESULTS: We found that CD44 expression is inversely proportional to the degree of cellular response to cisplatin chemotherapy and that CD44 is overexpressed in chemoresistant NSCLC cells. By performing CD44 knockdown using siRNA, we reconfirmed that the chemosensitivity of resistant cells depends on CD44 expression. We also observed that HA-liposome-mediated siRNA delivery prior to cisplatin chemotherapy significantly reduced CD44 expression and enhanced cisplatin sensitivity in chemoresistant NSCLC cells. CONCLUSIONS: These results suggest that self-targeted downregulation of chemoresistance-associated cell surface proteins during chemotherapy is an effective therapeutic strategy for overcoming the chemoresistance of NSCLC cells.

Fluorescent image-based evaluation of gastric conduit perfusion in a preclinical ischemia model.

BACKGROUND: This study evaluated near-infrared (NIR) fluorescent images to assess gastric conduit perfusion after an esophagectomy in a porcine model of gastric conduit ischemia. The time necessary to acquire a sufficient fluorescent signal to confirm ischemia in the gastric conduit after peripheral or central venous injection of indocyanine green (ICG) was also investigated. METHODS: A reversible gastric conduit ischemic pig model was established through ligation and release of the right gastroepiploic artery (RGEA, n=10). The esophageal reconstruction was performed to create an esophagogastric anastomosis. After ligation of the RGEA, ICG was injected into an ear vein (n=6) or the inferior vena cava (n=4). Under fluorescent imaging system guidance, the fluorescent signal-to-background ratio (SBR) in the gastric conduit or esophagus was measured during the entire procedure. We estimated the time necessary to acquire fluorescent signals in the gastric conduit using two different injection routes. RESULTS: When the RGEA was ligated, the SBR in the esophagus was significantly higher than that in the gastric conduit (P=0.02), and the SBR in the gastric conduit recovered within 180 s after release of the ligation. The time to acquire a fluorescent signal was faster with a central route than with a peripheral route (P=0.04). CONCLUSIONS: We successfully created an ischemic animal model of the gastric conduit. Using this animal model, we evaluated the sensitivity and applicability of the fluorescent imaging system for observation and identification of ischemic areas during an esophagectomy.

Endoscopic sentinel lymph node biopsy using indocyanine green-neomannosyl human serum albumin.

OBJECTIVE: The aim of this study was to determine the possibility of endoscopic sentinel lymph node biopsy of the head and neck region using indocyanine green-neomannosyl human serum albumin (ICG:MSA) and a custom-made intraoperative color-and-fluorescence-merged imaging system (ICFIS). METHODS: Using mouse and rabbit models of tongue cancer, we performed sentinel lymph node biopsy using an ICG:MSA tracer and custom imaging system equipped with an endoscope. RESULTS: ICG:MSA was localized to the cervical lymph node on the ipsilateral side for up to 30 minutes compared to ICG tracer (ICG n = 3, IGC:MSA n = 3). Detection of sentinel lymph nodes was attempted after injecting ICG:MSA in the mouse tongue cancer model (n = 3). During the operation, lymph nodes were easily identified and resected using ICFIS without the aid of other magnifiers. In the rabbit tongue cancer model (n = 3), the ICFIS equipped with 30-degree endoscopy was used to confirm the feasibility of endoscopic sentinel lymph node biopsy. The entire sentinel lymph node biopsy procedure was performed using the ICFIS with the 30-degree endoscope. We detected and dissected sentinel lymph nodes in the rabbit model without the other sentinel lymph node detection tools. CONCLUSION: We confirmed the usefulness of sentinel lymph node biopsy using a near infrared fluorescence technique and endoscopic system. Avoidance of radiation exposure and shine-through phenomena, which are problems of sentinel lymph node biopsy using conventional radioisotopes, are advantages of our surgical technique. By combining the endoscopic system with a sentinel lymph node biopsy procedure, we avoided visible neck scars, which lead to excellent cosmetic outcomes. LEVEL OF EVIDENCE: NA. Laryngoscope, 128:E135-E140, 2018.

Real-time computed tomography fluoroscopy-guided solitary lung tumor model in a rabbit.

Preclinical studies of lung cancer require suitable large-animal models to allow evaluation and development of surgical and interventional techniques. We assessed the feasibility and safety of a novel rabbit lung cancer model of solitary tumors, in which real-time computed tomography fluoroscopy is used to guide inoculation of VX2 carcinoma single-cell suspensions. Thirty-eight rabbits were divided into four groups according to the volume of the VX2 tissue or cell suspension, the volume of lipiodol, the volume of Matrigel, and the injection needle size. The mixtures were percutaneously injected into rabbit lungs under real-time computed tomography fluoroscopy guidance. Two weeks later, VX2 lung carcinomas were confirmed via positron emission tomography/computed tomography, necropsy, and histology. Real-time computed tomography fluoroscopy allowed the precise inoculation of the tumor cell suspensions containing lipiodol, while the use of Matrigel and a small needle prevented leakage of the suspensions into the lung parenchyma. Solitary lung tumors were successfully established in rabbits (n = 22) inoculated with single-cell suspensions (150 µL), lipiodol (150 µL), and Matrigel (150 µL) using a 26-gauge needle. This combination was determined to be optimal. Pneumothorax was observed in only two of the 38 rabbits (5.3%), both of which survived to the end of the study without any intervention. Real-time computed tomography fluoroscopy-guided inoculation of VX2 single-cell suspensions with lipiodol and Matrigel using a small needle is an easy and safe method to establish solitary lung tumors in rabbits.

Macrophage-Targeted Indocyanine Green-Neomannosyl Human Serum Albumin for Intraoperative Sentinel Lymph Node Mapping in Porcine Esophagus.

BACKGROUND: The sentinel lymph node (SLN) concept has been proposed to avoid unnecessary invasive LN dissection in surgery for esophageal cancer. This study evaluated a new macrophage-targeting fluorescent agent, indocyanine green-neomannosyl human serum albumin (ICG:MSA), for SLN mapping using a custom-made intraoperative color and fluorescence-merged imaging system (ICFIS) in porcine esophagus. METHODS: The LN targeting ability of ICG:MSA, indocyanine green-human serum albumin (ICG:HSA), and ICG was examined in vitro using the U937 differentiated monocyte cell line and in vivo in a mouse footpad model using fluorescence imaging. SLN identification in rabbit esophagus was then performed using ICG:MSA, ICG:HSA, and ICG. Finally, intraoperative SLN detection was conducted in porcine esophagus after esophagoscopic injection of ICG:MSA. RESULTS: The fluorescence signal of U937 cells treated by ICG:MSA was significantly higher than that of ICG or ICG:HSA (ICG: 1.0 ± 0.37; ICG:HSA: 3.4 ± 0.28, ICG:MSA: 6.8 ± 1.61; ICG to ICG:HSA, p = 0.03; ICG:HSA to ICG:MSA, p = 0.04; ICG to ICG:MSA, p = 0.0009). ICG:MSA was retained in popliteal LNs as long as 3 h, while ICG rapidly diffused through the entire mouse lymphatic system within 5 min. Esophageal SLN was detected within 15 min after injection of either ICG or ICG:MSA, but ICG:MSA provided more distinguishable images of LNss than ICG in rabbit esophagus. The SLN was also successfully detected in all porcine esophagus; the mean number of SLNs identified per esophagus was 1.6 ± 0.55. CONCLUSIONS: ICG:MSA has more specific macrophage-targeting properties, which could overcome the limitation of the low SLN retention of ICG, and could provide more precise real-time SLN detection during esophageal cancer surgery.

Intraoperative pulmonary neoplasm identification using near-infrared fluorescence imaging.

OBJECTIVES: Near-infrared (NIR) fluorescence imaging provides surgeons with real-time visual information during surgery. The purpose of this pilot trial was to evaluate the safety and feasibility of the intraoperative detection of pulmonary neoplasms with NIR fluorescence imaging after low-dose indocyanine green (ICG) injection. METHODS: Eleven consecutive patients who were scheduled to undergo resection of pulmonary neoplasms were enrolled in this study. ICG (1 mg/kg) was administered intravenously 1 day before surgery, and the retrieved surgical specimens were examined for fluorescence signalling by using NIR fluorescence imaging system on a back table in the operating room. We analysed the fluorescence intensity, pathology, size, depth from the pleural surface and metabolic activity of the pulmonary neoplasms. RESULTS: Fluorescence signalling was detected in all specimens except in one from a patient with primary lung cancer. Two false-positive cases that presented no residual tumour with obstructive pneumonitis, after concurrent chemoradiation therapy for primary lung cancer before the operation, were identified, and their fluorescence intensity was 8.6 ± 0.4. The mean fluorescence intensity of the eight pulmonary tumours was 3.4 ± 1.9, and these tumours did not differ in pathology, size, depth from the pleural surface or metabolic activity. CONCLUSIONS: NIR fluorescence imaging could safely identify pulmonary neoplasms after the systemic injection of ICG. In addition, low-dose ICG is sufficient for NIR fluorescence imaging of pulmonary neoplasms. However, because the passive accumulation of ICG could not be used to discriminate tumours with inflammation, tumour-targeted fluorescence should be developed to solve this problem in the future.

Intraoperative fluorescence image-guided pulmonary segmentectomy.

BACKGROUND: The intraoperative color and fluorescence-merged imaging system (ICFIS) is a new technology that may aid the demarcation of intersegmental borders during pulmonary segmentectomy. This study was performed to validate, for the first time, image-guided segmentectomy using ICFIS and to find the optimal dosage of fluorescent dye to ensure safe and sustained imaging during surgery. METHODS: Nine rabbits were subjected to pulmonary segmentectomy. These constituted three groups of three rabbits each. After ligation of the segmental pulmonary artery supplying the targeted segment, the rabbits were injected intravenously with indocyanine green (ICG) at a concentration of 0.3, 0.6, or 3.0 mg/kg, depending on their group assignment. The optimal dose was determined from the rabbit study and then used to guide ICFIS during pulmonary segmentectomy in five pigs. RESULTS: The fluorescent signal contrast ratios of the targeted area to the normal lung using ICG concentrations of 0.3, 0.6, or 3.0 mg/kg were 1.9 ± 0.25, 2.0 ± 0.17, and 2.1 ± 0.06, respectively. The mean ICG washout times were 1, 3, and 6 min, respectively. Proceeding with an ICG concentration of 0.6 mg/kg, the mean washout time was found to be longer in pigs (15 min). This provided adequate time for successful ICFIS-guided segmentectomy in all five pigs, without the requirement for additional procedures for intersegmental plane demarcation. CONCLUSIONS: ICG image-guided segmentectomy using ICFIS enabled immediate visualization of the intersegmental planes. The washout time using the ICG dose determined in this study was long enough to ensure that visualization was sustained throughout the surgery.

Highly sensitive and selective anticancer effect by conjugated HA-cisplatin in non-small cell lung cancer overexpressed with CD44.

In spite of severe side effects, chemotherapy is widely used as a major anticancer treatment in non-small cell lung cancer (NSCLC). In order to enhance the therapeutic properties and reduce side effects, enormous efforts have been devoted to direct anticancer agents specifically to tumor tissues by the use of nanoparticles, or cancer cell marker attached drugs. However, cell-specific chemotherapy is still in its infancy and is not applicable to all types of cancers due to the complexity of the cancer occurrence and progress. In this study, we demonstrate that hyaluronan (HA)-conjugated cisplatin has highly selective and sensitive anticancer effects in NSCLC cells that overexpress the trans-membrane receptor, CD44, as HA is a specific ligand for CD44. In proliferation experiments, HA-conjugated cisplatin showed dramatic cell viability decreases (from 100% to 42.31%) in H1299 cells, which overexpress CD44, whereas no such change was observed in A549 and HFL1, which have little to no expression of CD44. More importantly, conjugation with HA decreased the dosage concentration of cisplatin by 50-fold for both cytotoxic and anti-metastatic effects. In addition, HA-cisplatin conjugate treatment selectively decreased migration (from 100% to 7.8%) and invasiveness (from 100% to 21.4%, respectively) of H1299. Based on our experimental results, we strongly believe that HA-cisplatin conjugate is a potential anticancer chemo-agent, which target CD44 overexpression in NSCLC, with minimal side effects and high therapeutic properties.

The effects of sonic hedgehog signaling pathway components on non-small-cell lung cancer progression and clinical outcome.

BACKGROUND: Researchers in recent studies have reported that the sonic hedgehog (Shh) signaling pathway plays a crucial role during tumorigenesis, angiogenesis and cellular differentiation. We investigated the clinical and pathological significances of the Shh pathway and of its lymphangiogenic components in non-small-cell lung cancer (NSCLC), namely, Shh, glioma-associated oncogene homolog zinc finger protein 1 (Gli1), lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1) and vascular endothelial growth factor D (VEGF-D). METHODS: The expression of Shh, Gli1, LYVE-1 and VEGF-D in primary NSCLC tissue from 40 patients was examined using immunohistochemical assays, and relationships between expression and clinicopathological data, such as age, gender, histology, tumor size, nodal stage, visceral pleural invasion, lymphatic thromboembolism, recurrence and overall survival were investigated. RESULTS: Of the 40 specimens examined, 25 (62.5%), 20 (50.0%), 11 (27.5%) and 20 (50.0%) were positive for Shh, Gli1, LYVE-1 or VEGF-D expression, respectively. The expression of Gli1 and LYVE-1 were significantly associated (P = 0.011), and Shh and LYVE-1 expression was related to visceral pleural invasion and lymphatic thromboembolism, respectively (P < 0.05). Shh expression levels compared on survival curves were statistically significant in univariate logrank analysis (P = 0.020). However, other clinicopathological factors did not reveal any statistical significance in univariate and multivariate analyses. CONCLUSIONS: To our knowledge, this the first report of the relationship between components of the Shh signaling pathway and prognosis in NSCLC. The expression of Shh, Gli1 and LYVE-1 was found to be associated with clinicopathological factors and survival. Thus, the overexpression of the Shh signaling pathway could serve as a predictor of malignant behavior, including lymphangiogenesis, in NSCLC.

Thoracoscopic color and fluorescence imaging system for sentinel lymph node mapping in porcine lung using indocyanine green-neomannosyl human serum albumin: intraoperative image-guided sentinel nodes navigation.

PURPOSE: This study was performed to validate a newly developed sentinel lymph node (SLN) targeting tracer, indocyanine green-neomannosyl human serum albumin (ICG:MSA), and a thoracoscopic version of the intraoperative color and fluorescence imaging system (ICFIS) for lung cancer SLN mapping. METHODS: ICG alone or ICG:MSA (5 µg/kg) was injected into the rat thigh, and the results were compared. The fluorescence signal-to-background ratios of SLNs were recorded and evaluated over a 2-h period by using ICFIS. Additionally, a SLN biopsy was performed via video-assisted thoracoscopic surgery with the use of ICG:MSA in porcine lung by using thoracoscopic ICFIS. RESULTS: The newly developed ICG:MSA showed a significantly improved signal-to-background ratio compared with ICG alone throughout the trials. All SLNs were identified in both rats (ten SLNs in ten rat thighs) and pigs (ten SLNs in ten porcine lungs) under in vivo conditions. All SLNs were dissected successfully by using video-assisted thoracoscopic surgery with the help of thoracoscopic ICFIS. DISCUSSION: ICG:MSA accumulates in the SLN by uptake and retention through the mannose-specific receptors on macrophages. Thoracoscopic ICFIS successfully assisted SLN mapping despite low near-infrared light transmission in the commercial thoracoscope. On the basis of the results of the thoracoscopic SLN mapping, we anticipate that ICG:MSA and thoracoscopic ICFIS can be translated to clinical trials in the near future.