The detection and characterization of circulating tumor cells (CTC) holds great promise for personalizing medicine and optimizing systemic therapy. However, low specificity, low sensitivity, and the time consuming nature of current approaches have impeded clinical adoption. Here we report a new method using surfaceenhanced Raman spectroscopy (SERS) to directly measure targeted CTCs in the presence of white blood cells. SERS nanoparticles with epidermal growth factor peptide as a targeting ligand have successfully identified CTCs in the peripheral blood of 19 patients with squamous cell carcinoma of the head and neck (SCCHN), with a range of 1 to 720 CTCs per milliliter of whole blood. Our technique may provide an important new clinical tool for management of patients with SCCHN and other cancers. Cancer Res; 71(5); 1526-32. Ó2011 AACR.

Gold nanorods (AuNRs)-assisted plasmonic photothermal therapy (AuNRs-PPTT) is a promising strategy for combating cancer in which AuNRs absorb near-infrared light and convert it into heat, causing cell death mainly by apoptosis and/or necrosis. Developing a valid PPTT that induces cancer cell apoptosis and avoids necrosis in vivo and exploring its molecular mechanism of action is of great importance. Furthermore, assessment of the long-term fate of the AuNRs after treatment is critical for clinical use. We first optimized the size, surface modification [rifampicin (RF) conjugation], and concentration (2.5 nM) of AuNRs and the PPTT laser power (2 W/cm) to achieve maximal induction of apoptosis. Second, we studied the potential mechanism of action of AuNRs-PPTT using quantitative proteomic analysis in mouse tumor tissues. Several death pathways were identified, mainly involving apoptosis and cell death by releasing neutrophil extracellular traps (NETs) (NETosis), which were more obvious upon PPTT using RF-conjugated AuNRs (AuNRs@RF) than with polyethylene glycol thiol-conjugated AuNRs. Cytochrome and p53-related apoptosis mechanisms were identified as contributing to the enhanced effect of PPTT with AuNRs@RF. Furthermore, Pin1 and IL18-related signaling contributed to the observed perturbation of the NETosis pathway by PPTT with AuNRs@RF. Third, we report a 15-month toxicity study that showed no long-term toxicity of AuNRs in vivo. Together, these data demonstrate that our AuNRs-PPTT platform is effective and safe for cancer therapy in mouse models. These findings provide a strong framework for the translation of PPTT to the clinic.

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Non-specific distribution of chemotherapeutic drugs (such as paclitaxel) is a major factor contributing to side effects and poor clinical outcomes in the treatment of human head and neck cancer. To develop novel drug delivery systems with enhanced efficacy and minimized adverse effects, we synthesized a ternary conjugate heparin-folic acid-paclitaxel (HFT), loaded with additional paclitaxel (T). The resulting nanoparticle, HFT-T, is expected to retain the antitumor activity of paclitaxel and specifically target folate receptor (FR)-expressing tumors, thereby increasing the bioavailability and efficacy of paclitaxel. In vitro experiments found that HFT-T selectively recognizes FR-positive human head and neck cancer cell line KB-3-1, displaying higher cytotoxicity compared to the free form of paclitaxel. In a subcutaneous KB-3-1 xenograft model, HFT-T administration enhanced the specific delivery of paclitaxel into tumor tissues and remarkably improved antitumor efficacy of paclitaxel. The average tumor volume in the HFT-T treatment group was 92.9±78.2mm3 vs 1670.3±286.1mm3 in the mice treated with free paclitaxel. Furthermore, paclitaxel tumors showed a resurgence of growth after several weeks of treatment, but this was not observed with HFT-T. This indicates that HFT-T could be more effective in preventing tumors from developing drug resistance. No significant acute in vivo toxicity was observed. These results indicate that specific delivery of paclitaxel with a ternary structured nanoparticle (HFT-T) targeting FR-positive tumor is a promising strategy to enhance chemotherapy efficacy and minimize adverse effects.

BACKGROUND Recently, endoscopic ultrasound‐guided fine‐needle aspiration (EUS‐FNA) has emerged as a diagnostic adjunct for small pancreatic lesions and abdominal and mediastinal lymph node diseases. DESIGN. During a 21‐month period, we performed 179 EUS‐FNAs in 166 consecutive patients; these data are the subject of this study. An average of 2.6 needle passes were obtained and aspiration was performed most commonly in the pancreas (162 cases, 91%). The FNA smears were reviewed using six diagnostic categories (negative for malignancy/nondiagnostic [NND], atypia, suspicious for malignancy, benign tumor/cyst, neuroendocrine neoplasm [NEN], and carcinoma). The review diagnosis was correlated with the histologic diagnosis made on resection or surgical biopsy specimens in 70 cases. Up to 17 months of clinical follow‐up were sought for the cases with a negative or inconclusive FNA diagnosis and no diagnostic tissue confirmation (81 cases). RESULTS The review FNA diagnoses were as follows: NND (49 cases), atypia (17 cases), suspicious for malignancy (12 cases), benign tumor/cyst excluding NEN (10 cases), NEN (6 cases), carcinoma (85 cases). Follow‐up methods included resection (49 cases), surgical biopsy (21 cases), repeat FNA or brushing cytology (28 cases), and clinical follow‐up only (81 cases). Of the 49 NND cases, 23 (47%) had positive follow‐up results (i.e., false‐negative diagnosis) that were confirmed by tissue diagnosis (resection/surgical biopsy in 11 cases [48%] and repeat FNA/brushing in 12 cases [52%]). These included pancreatic/ampullary adenocarcinoma in 20 cases, esophageal squamous carcinoma in 1 case, and NEN in 2 cases. Follow‐up also revealed carcinoma in all 12 suspicious cases and 13 pancreatic adenocarcinomas and 1 microcystic adenoma in 14 of the 17 atypical cases. Overall, repeat computed tomography (CT)‐guided FNA samples yielded a definite diagnosis in four atypical and seven NND cases, whereas EUS‐FNA results provided a definite diagnosis in three cases in which CT‐guided FNA failed and in two cases in which ampullary biopsy failed. No false‐positive cases were identified. The false‐negative rate due to inadequate sampling was 13.2%. Sensitivity (including cases with inadequate cellularity and nondiagnsotic aspirates) was 81.7% and specificity was 100%. None of the factors evaluated (lesion characteristics, aspiration site, and tumor type) significantly influenced diagnostic results. CONCLUSION EUS‐FNA is a valuable diagnostic and staging tool with high specificity and sensitivity. Negative or nondiagnostic cases on EUS‐FNA require further diagnostic work for a definitive diagnosis when clinical or radiographic findings do not correlate with the FNA results. Cancer (Cancer Cytopathol) 2002. © 2002 American Cancer Society.

The expression of p53 protein in primary head and neck squamous cell carcinoma was significantly predictive of shorter survival because of its association with earlier development of both tumor recurrence and second primary tumors. Thus, p53 expression may be a valuable marker for identifying individuals at high risk of developing a recurrence of primary disease and second primary tumors who may benefit from adjuvant therapy and chemoprevention after definitive local therapy.

Purpose: Our previous study revealed that simultaneously targeting epidermal growth factor receptor (EGFR) tyrosine kinase and cyclooxygenase-2 (COX-2) additively or synergistically inhibited growth of squamous cell carcinoma of the head and neck (SCCHN) in vitro. However, an in vivo efficacy of this combined treatment in SCCHN has not been studied. Experimental Design: Nude mice were pretreated with control (1% Tween 80), ZD1839 (50 mg/kg) alone, celecoxib (50 mg/kg) alone, or a combination of ZD1839 and celecoxib at the same dosages for 7 days before injection of a human SCCHN cell line Tu212.The animals were continuously treated with the agents 5 days a week for about 11weeks. Results: Tumor growth in the combined treatment was significantly inhibited compared with the control (P < 0.001), ZD1839 (P = 0.005), or celecoxib (P < 0.001). At the same time, a dramatic delay of tumor progression was observed in the combined treatment compared with all other three groups. Molecular analysis showed that the combined treatment significantly decreased prostaglandin E metabolite production. The cooperative effect of these two agents in combination was also associated with down-regulation of phosphorylated EGFR, phosphorylated extracellular signal-regulated kinase, and phosphorylated signal transducers and activators of transcription 3 levels and reduction of vascular endothelial growth factor and Ki-67 expression. Specifically, gene silencing of both EGFR and COX-2 by small interfering RNA further confirmed the cooperative antitumor effect. Conclusion:The current results strongly suggest that a cooperative effect of the combined treatment on tumor progression is mediated through blocking both EGFR-and COX-2-related pathways. This combination regimen may provide a promising strategy for cancer therapy and chemoprevention in SCCHN.

One of the mechanisms of the antitumor activity of green tea (2)-epigallocatechin-3-gallate (EGCG) is associated with its effect on epidermal growth factor receptor (EGFR)-mediated signaling transduction pathways. We investigated whether combining EGCG with the EGFR-tyrosine kinase inhibitor (EGFR-TKI) erlotinib may augment erlotinib-induced cell growth inhibition of squamous cell carcinoma of the head and neck (SCCHN) in a mouse xenograft model. In vitro studies with 5 head and neck cancer cell lines revealed that synergistic cell growth inhibition by the combination of EGCG and erlotinib was associated with significantly greater inhibition of pEGFR and pAKT, increased activation of caspases 9, 3 and PARP compared to the inhibition induced by EGCG or erlotinib alone. Erlotinib inhibited phosphorylation of EGFR, stabilizing EGFR at the plasma membrane, whereas EGCG induced EGFR internalization and ubiquitin-degradation, ultimately undermining EGFR signaling. The efficacy of the combination treatment was investigated with nude mice (n 5 25) orally gavaged with vehicle control, EGCG, erlotinib or the combination at the same doses for 7 days, followed by subcutaneous injection with Tu212 cells. Animals were continuously administered the agents 5 days weekly for 7 weeks. The combined treatment resulted in significantly greater inhibition of tumor growth and delayed tumor progression as a result of increased apoptosis, decreased cell proliferation and reduced pEGFR and pAKT compared to the single agent treatment groups. Our results suggest a synergistic antitumor effect of a combined treatment with EGCG and erlotinib, and provide a promising regimen for future chemoprevention and treatment of SCCHN. ' 2008 Wiley-Liss, Inc.Key words: squamous cell carcinoma of the head and neck; (2)-epigallocatechin-3-gallate; erlotinib Squamous cell carcinoma of the head and neck (SCCHN) accounts for 3% of all cancers in the United States, with approximately 45,660 new cases and more than 11,210 expected deaths in 2007. 1 Despite advances in conventional therapies, including surgery, radiation and chemotherapy, the overall survival rate for SCCHN has not significantly improved in the past 3 decades. 2 Overexpression of epidermal growth factor receptor (EGFR) and its ligands TGF-a or EGF has been observed in 80-90% of SCCHN specimens 3-6 and correlates with poor disease-free and overall survival, and increased risk of disease recurrence and metastasis. 6,7 Erlotinib (OSI-774 or Tarceva), an EGFR tyrosine kinase inhibitor (TKI), has shown strong antitumor and chemopreventive efficacies in a variety of cancer types including SCCHN through blocking EGFR-related signal transduction pathways. 5,[8][9][10] Our previous studies showed that EGFR-TKI could inhibit tumor growth through blocking EGFR-related pathways. 3,11 However, the efficacy of monotherapy is severely limited by heterogeneity of the tumor cell population and redundant growth and survival pathways.The development of preventive approaches using specific natural or synthetic che...