Tipifarnib enhances anti-EGFR activity of cetuximab in non-HRas mutated head and neck squamous cell carcinoma cancer (HNSCC).

OBJECTIVE: To test the potential ability of tipifarnib to impair proliferation and to enhance the activity of the EGFR inhibitor cetuximab in wild-type H-Ras HNSCC, which accounts for the majority of HNSCC. MATERIALS AND METHODS: Cell growth, apoptosis and signaling changes in HNSCC cells following tipifarnib exposure in vitro were assessed by SRB, colony formation assay, annexin V staining and Western blot, respectively. A patient-derived xenograft (PDX) animal model was adopted to evaluate the efficacy of tipifarnib in vivo with and without cetuximab. RESULTS: Treatment of wild-type H-Ras HNSCC cell lines in vitro with tipifarnib reduced cell growth and increased levels of defarnesylated H-Ras in a dose-dependent manner. In a PDX mouse model, treatment with single-agent tipifarnib led to only near-significant growth inhibition. The addition of cetuximab resulted in increased anti-proliferative effect both in culture and in PDX models, which was also mirrored by Western blot and apoptosis assay results. CONCLUSION: Tipifarnib has only a moderate ability to slow tumor growth as a single agent in HNSCC with wild type H-Ras, despite specifically inhibiting the farnesyltransferase upon which the function of H-Ras depends. The combination of cetuximab and tipifarnib appears to enhance the anti-proliferative effect of single-agent tipifarnib and marginally enhance that of single agent cetuximab. These findings deserve further evaluation.

Combination of resveratrol and green tea epigallocatechin gallate induces synergistic apoptosis and inhibits tumor growth in vivo in head and neck cancer models.

Despite widespread interest in chemoprevention and therapy due to the high margin of safety of dietary natural compounds, clinical intervention with single agents has failed to yield the expected outcomes, mostly due to poor bioavailability and low potency. Combinations of natural agents with synergistic effects are gaining increasing attention. In the present study, in vitro and in vivo antitumor effects of a combination of two natural dietary agents, green tea epigallocatechin gallate (EGCG) and resveratrol were investigated. It was revealed that their combination at low doses (at which single agents induce minimal apoptosis) synergistically increased apoptosis (combination index < 1) in head and neck cancer cell lines. Synergistic apoptosis was also supported by caspase­3 and PARP cleavage. The combination also significantly inhibited growth of xenografted head and neck tumors in nude mice as supported by significant inhibition of tumor volume, tumor weight and Ki67 expression, and increase in TUNEL­positive cells. Mechanistic studies revealed that the combination inhibited AKT­mTOR signaling both in vitro and in vivo. In addition, overexpression of constitutively active AKT protected cells from apoptosis induced by the combination of EGCG and resveratrol. Collectively, the present results for the first time suggest that the combination of EGCG and resveratrol has synergistic growth inhibitory effects and provide an important rationale for future clinical development for chemoprevention and treatment of head and neck cancer.

Co-expression of fibroblast growth factor receptor 3 with mutant p53, and its association with worse outcome in oropharyngeal squamous cell carcinoma.

Fibroblast growth factor receptor 3 (FGFR3) is expressed in squamous cell carcinoma of the head and neck (SCCHN) including oropharyngeal squamous cell carcinoma (OPSCC) and is a potential therapeutic target. However, information on its correlation with other relevant cancer related proteins stratified by p16 status and its prognostic significance in OPSCC is limited. We examined FGFR3 expression and its correlation with clinical characteristics, p16 status, and mutant p53 (mp53) among 220 retrospectively collected OPSCC cases and 40 prospectively collected SCCHN cases, including a majority of OPSCC. Correlations of FGFR3 Weighted Index (WI) with p16 status and mp53 WI as well as its association with disease-free survival (DFS) and overall survival (OS) were evaluated. FGFR3 expression was detected in 61% and 70% of cases in cohorts 1 and 2, respectively. FGFR3 level was significantly higher in p16-negative tumors in both cohorts (p<0.001 and 0.006). FGFR3 expression was highly correlated with mp53 expression in both p16 + and p16- OPSCC (p<0.0001 and p = 0.0006, respectively). In cohort 1, univariate analysis showed that FGFR3 was associated with DFS but not OS. Kaplan-Meier analysis showed that higher FGFR3 and mp53 level correlated with worse DFS (p = 0.025) and OS (p = 0.009). As expected, p16 positive status was associated with improved OS and DFS (p<0.001 for both). Our results suggest that high FGFR3 expression is associated with p16 negative status and mp53 expression in OPSCC and correlates with a worse clinical outcome. The biological relationship between FGFR3 and mp53 in OPSCC deserves further investigation.

Combinatorial approaches targeting the EGFR family and c-Met in SCCHN.

OBJECTIVE: We aimed to develop novel combinations of inhibitors targeting EGFR family members and c-Met for the treatment of recurrent SCCHN. MATERIALS AND METHODS: Three different c-Met inhibitors in combination with a pan-HER inhibitor (crizotinib/afatinib, tivantinib/afatinib and cabozantinib/afatinib) were investigated for their anti-tumor effects on SCCHN cell lines in vitro. In vivo activity of the combinations was tested in SCCHN cell line xenografts and patient-derived xenograft (PDX) animal models generated from patients with recurrent SCCHN. RESULTS: Western blot assay indicated that activation of EGFR, HER2, HER3, and c-Met was blocked by all three combinations and the downstream PI3K/AKT and ERK signaling pathways were inhibited. Sulforhodamine B colorimetric assay revealed SCCHN cell growth was more effectively inhibited by the combinations than by single agents, particularly in cell lines with high c-Met expression. Furthermore, the combinations were more potent in inducing apoptosis than each of the single agents. In the PDX models, the combination treatments exhibited significantly better efficacy in tumor growth inhibition compared to the respective single agents. CONCLUSION: In conclusion, we demonstrated that the simultaneous targeting of EGFR, HER2, and c-Met is more effective than the individual inhibition of these targets in vitro and in SCCHN cell line xenograft and PDX models. Our findings pave the way for further clinical investigation of such combinations in SCCHN.

Phase Ib Study of Chemoprevention with Green Tea Polyphenon E and Erlotinib in Patients with Advanced Premalignant Lesions (APL) of the Head and Neck.

PURPOSE: On the basis of synergistic effects between green tea polyphenon E (PPE) and EGFR-tyrosine kinase inhibitor in preclinical studies, we conducted a phase Ib study of the PPE and erlotinib combination in patients with advanced premalignant lesions (APL) of the oral cavity and larynx. PATIENTS AND METHODS: Patients were treated with a fixed dose of PPE (200 mg three times a day) and dose escalation of erlotinib (50, 75, 100 mg daily) for 6 months with tissue biopsy at baseline and 6 months. Primary endpoints were safety and toxicity; secondary endpoints were evaluation of pathologic response, cancer-free survival (CFS), overall survival (OS), and biomarker modulation. RESULTS: Among 21 enrolled patients, 19 began treatment and 17 completed 6 months of treatment with PPE and erlotinib. Main characteristics of treated patients: 15 severe dysplasia or carcinoma in situ and 17 oral cavity. Only skin rash was associated with dose-limiting toxicity and MTD. Recommended doses for phase II studies are PPE 600 mg daily plus erlotinib 100 mg daily for 6 months. Pathologic responses in 17 evaluable patients: pathologic complete response (47%) and pathologic partial response (18%). The 5-year CFS and OS were 66.3% and 93%, respectively. Among tested biomarkers, only phosphorylated ERK was correlated with response to treatment. CONCLUSIONS: Treatment with PPE and erlotinib combination was well tolerated in patients with APLs of the head and neck, and showed a high rate of pathologic response with excellent CFS. This combination deserves further investigation for the chemoprevention and/or prevention of second primary tumors in early-stage head and neck cancer.

Interleukin-6/STAT3 Signaling is Prominent and Associated with Reduced Overall Survival in p16 Negative Oropharyngeal Squamous Cell Carcinoma.

This study addresses the hypothesis that IL-6/STAT3 signaling is of clinical relevance in oropharyngeal squamous cell carcinoma (OPSCC). We evaluated relationships between key components of this pathway in tumors from a unique cohort of n = 59 fully annotated, treatment-naïve patients with OPSCC. The multiplex Opal platform was utilized for immunofluorescence (IF) analysis of tissues to detect IL-6 and phosphorylated STAT3 (pSTAT3), taking into consideration its nuclear versus cytoplasmic localization. Abundant staining for both IL-6 and pSTAT3 was evident in tumor-rich regions of each specimen. IL-6 correlated with cytoplasmic pSTAT3 but not nuclear or total pSTAT3 in this cohort of OPSCC tumors, regardless of p16 status (r = 0.682, p < 0.0001). There was a significant association between increased total pSTAT3, nuclear pSTAT3, cytoplasmic pSTAT3 and IL-6 in p16 negative tumors. Our data indicate STAT3 phosphorylation was a key feature in p16-negative OPSCC tumors. When IL-6 data was stratified by median expression in tumors, there was no association with overall survival. In contrast, both total and nuclear pSTAT3 were significant predictors of poor overall and disease free survival. This strong inverse relationship with overall survival was present in p16 negative tumors for both total and nuclear pSTAT3, but not in p16 positive OPSCC tumors. Together these data indicate that activation of the STAT3 signaling pathway is a marker of p16 negative tumors and relevant to OPSCC prognosis and a potential target for treatment of this more aggressive OPSCC sub-population.

Prognostic implications of peritumoral vasculature in head and neck cancer.

BACKGROUND: There is conflicting evidence regarding the role of peritumoral lymphatic vessel density (LVD) and blood microvessel density (MVD) in the metastasis and prognosis of head and neck squamous cell carcinoma (HNSCC). Existing studies are limited to one or two head and neck subsites and/or small sample sizes. A larger study incorporating multiple sub-sites is needed to address the role of peritumoral LVD and MVD in HNSCC metastasis and prognosis. METHODS: Tissue samples from 200 HNSCC cases were stained simultaneously using immunohistochemistry (IHC) for markers of peritumoral LVD (lymphatic vessel marker D240) and MVD (blood vessel marker CD31). Of the stained slides, 166 and 167 were evaluable for LVD and MVD, respectively. The results were then correlated with clinicopathologic features and patient outcomes. RESULTS: Patients with metastatic disease were more likely to have high peritumoral MVD. Through multivariable analyses, MVD was not significantly related to DFS and OS, while low LVD was related to higher risk of disease progression and poor survival. CONCLUSIONS: Peritumoral MVD was found to be positively associated with metastasis, while LVD was found to be inversely related to both metastasis and progression of HNSCC. These findings may suggest a prognostic role of both peritumoral LVD and MVD in patients with HNSCC.

A Correlative Analysis of PD-L1, PD-1, PD-L2, EGFR, HER2, and HER3 Expression in Oropharyngeal Squamous Cell Carcinoma.

We explored potential associations of the PD-1/PD-L1/PD-L2 pathway with clinical characteristics, outcome, and expression of EGFR, HER2, HER3 in oropharyngeal squamous cell carcinoma (OPSCC) using an institutional database. Protein expression was assessed by IHC on tissue microarray sections (EGFR, HER2, HER3) or whole tissue sections (PD-1/PD-L1/PD-L2). Expression of EGFR, HER2, HER3, PD-L1, and PD-L2 was quantified on tumor cells. Maximum density of PD-1 positive lymphocytes was measured on a scale of 0 to 4 within the tumor mass and peritumoral stroma. Associations between biomarkers and patient outcomes were tested using descriptive and inferential statistics, logistic regression, and Cox proportional hazards models. We analyzed tissue samples from 97 OPSCC cases: median age 59 years, p16+ (71%), male (83.5%), never smokers (18%), stage 3 to 4 disease (77%). Twenty-five percent of cases were PD-L1 positive. The proportion of PD-L1+ tumors was higher in p16+ (29%) than p16- OPSCC (11%, P = 0.047). There was no correlation between PD-L1, PD-L2, PD-1, EGFR, HER2, or HER3 expression. Positive PD-L1 status correlated with advanced nodal disease on multivariate analysis (OR 5.53; 95% CI, 1.06-28.77; P = 0.042). Negative PD-L2 expression was associated with worse survival (HR 3.99; 95% CI, 1.37-11.58; P = 0.011) in p16- OPSCC. Lower density of PD-1 positive lymphocytes in peritumoral stroma was associated with significantly increased risk of death on multivariate analysis (HR 3.17; 95% CI, 1.03-9.78; P = 0.045) after controlling for prognostic factors such as stage and p16 status. PD-L1 expression on tumor cells correlates with p16 status and advanced nodal status in OPSCC. PD-1 positive lymphocytes in peritumoral stroma serve as an independent prognostic factor for overall survival. Mol Cancer Ther; 17(3); 710-6. ©2018 AACR.

Honokiol Radiosensitizes Squamous Cell Carcinoma of the Head and Neck by Downregulation of Survivin.

Purpose: Previous studies revealed diverging results regarding the role of survivin in squamous cell carcinoma of the head and neck (SCCHN). This study aimed to evaluate the clinical significance of survivin expression in SCCHN; the function of survivin in DNA-damage repair following ionizing radiation therapy (RT) in SCCHN cells; and the potential of honokiol to enhance RT through downregulation of survivin.Experimental Design: Expression of survivin in SCCHN patient primary tumor tissues (n = 100) was analyzed and correlated with clinical parameters. SCCHN cell lines were used to evaluate the function of survivin and the effects of honokiol on survivin expression in vitro and in vivoResults: Overexpression of survivin was significantly associated with lymph nodes' metastatic status (P = 0.025), worse overall survival (OS), and disease-free survival (DFS) in patients receiving RT (n = 65, OS: P = 0.024, DFS: P = 0.006) and in all patients with SCCHN (n = 100, OS: P = 0.002, DFS: P = 0.003). In SCCHN cells, depletion of survivin led to increased DNA damage and cell death following RT, whereas overexpression of survivin increased clonogenic survival. RT induced nuclear accumulation of survivin and its molecular interaction with γ-H2AX and DNA-PKCs. Survivin specifically bound to DNA DSB sites induced by I-SceI endonuclease. Honokiol (which downregulates survivin expression) in combination with RT significantly augmented cytotoxicity in SCCHN cells with acquired radioresistance and inhibited growth in SCCHN xenograft tumors.Conclusions: Survivin is a negative prognostic factor and is involved in DNA-damage repair induced by RT. Targeting survivin using honokiol in combination with RT may provide novel therapeutic opportunities. Clin Cancer Res; 24(4); 858-69. ©2017 AACR.

Systemic Delivery of Bc12-Targeting siRNA by DNA Nanoparticles Suppresses Cancer Cell Growth.

Short interfering RNA (siRNA) is a promising molecular tool for cancer therapy, but its clinical success is limited by the lack of robust in vivo delivery systems. Rationally designed DNA nanoparticles (DNPs) have emerged as facile delivery vehicles because their physicochemical properties can be precisely controlled. Nonetheless, few studies have used DNPs to deliver siRNAs in vivo, and none has demonstrated therapeutic efficacy. Herein, we constructed a number of DNPs of rectangular and tubular shapes with varied dimensions using the modular DNA brick method for the systemic delivery of siRNA that targets anti-apoptotic protein Bcl2. The siRNA delivered by the DNPs inhibited cell growth both in vitro and in vivo, which suppressed tumor growth in a xenograft model that specifically correlated with Bcl2 depletion. This study suggests that DNPs are effective tools for the systemic delivery of therapeutic siRNA and have great potential for further clinical translation.

Prognostic biomarkers in patients with human immunodeficiency virus-positive disease with head and neck squamous cell carcinoma.

BACKGROUND: We examined the prognostic value of a panel of biomarkers in patients with squamous cell carcinoma of the head and neck (SCCHN) who were human immunodeficiency virus (HIV) positive (HIV-positive head and neck cancer) and HIV negative (HIV-negative head and neck cancer). METHODS: Tissue microarrays (TMAs) were constructed using tumors from 41 disease site-matched and age-matched HIV-positive head and neck cancer cases and 44 HIV-negative head and neck cancer controls. Expression of tumor biomarkers was assessed by immunohistochemistry (IHC) and correlations examined with clinical variables. RESULTS: Expression levels of the studied oncogenic and inflammatory tumor biomarkers were not differentially regulated by HIV status. Among patients with HIV-positive head and neck cancer, laryngeal disease site (P = .003) and Clavien-Dindo classification IV (CD4) counts <200 cells/µL (P = .01) were associated with poor prognosis. Multivariate analysis showed that p16 positivity was associated with improved overall survival (OS; P < .001) whereas increased expression of transforming growth factor-beta (TGF-ß) was associated with poor clinical outcome (P = .001). CONCLUSION: Disease site has significant effect on the expression of biomarkers. Expression of tumor TGF-ß could be a valuable addition to the conventional risk stratification equation for improving head and neck cancer disease management strategies.

Phenformin enhances the therapeutic effect of selumetinib in KRAS-mutant non-small cell lung cancer irrespective of LKB1 status.

MEK inhibition is potentially valuable in targeting KRAS-mutant non-small cell lung cancer (NSCLC). Here, we analyzed whether concomitant LKB1 mutation alters sensitivity to the MEK inhibitor selumetinib, and whether the metabolism drug phenformin can enhance the therapeutic effect of selumetinib in isogenic cell lines with different LKB1 status. Isogenic pairs of KRAS-mutant NSCLC cell lines A549, H460 and H157, each with wild-type and null LKB1, as well as genetically engineered mouse-derived cell lines 634 (krasG12D/wt/p53-/-/lkb1wt/wt) and t2 (krasG12D/wt/p53-/-/lkb1-/-) were used in vitro to analyze the activities of selumetinib, phenformin and their combination. Synergy was measured and potential mechanisms investigated. The in vitro findings were then confirmed in vivo using xenograft models. The re-expression of wild type LKB1 increased phospho-ERK level, suggesting that restored dependency on MEK->ERK->MAPK signaling might have contributed to the enhanced sensitivity to selumetinib. In contrast, the loss of LKB1 sensitized cells to phenformin. At certain combination ratios, phenformin and selumetinib showed synergistic activity regardless of LKB1 status. Their combination reduced phospho-ERK and S6 levels and induced potent apoptosis, but was likely through different mechanisms in cells with different LKB1 status. Finally, in xenograft models bearing isogenic A549 cells, we confirmed that loss of LKB1 confers resistance to selumetinib, and phenformin significantly enhances the therapeutic effect of selumetinib. Irrespective of LKB1 status, phenformin may enhance the anti-tumor effect of selumetinib in KRAS-mutant NSCLC. The dual targeting of MEK and cancer metabolism may provide a useful strategy to treat this subset of lung cancer.

HER3 Targeting Sensitizes HNSCC to Cetuximab by Reducing HER3 Activity and HER2/HER3 Dimerization: Evidence from Cell Line and Patient-Derived Xenograft Models.

PURPOSE: Our previous work suggested that HER3 inhibition sensitizes head and neck squamous cell carcinoma (HNSCC) to EGFR inhibition with cetuximab. This study aimed to define the role of HER3 in cetuximab resistance and the antitumor mechanisms of EGFR/HER3 dual targeting in HNSCC. EXPERIMENTAL DESIGN: We treated cetuximab-resistant HNSCC UMSCC1-C and parental UMSCC1-P cell lines with anti-EGFR antibody cetuximab, anti-HER3 antibody MM-121, and their combination. We assessed activities of HER2, HER3, and downstream signaling pathways by Western blotting and cell growth by sulforhodamine B (SRB) and colony formation assays. HER3-specific shRNA was used to confirm the role of HER3 in cetuximab response. The combined efficacy and alterations in biomarkers were evaluated in UMSCC1-C xenograft and patient-derived xenograft (PDX) models. RESULTS: Cetuximab treatment induced HER3 activation and HER2/HER3 dimerization in HNSCC cell lines. Combined treatment with cetuximab and MM-121 blocked EGFR and HER3 activities and inhibited the PI3K/AKT and ERK signaling pathways and HNSCC cell growth more effectively than each antibody alone. HER3 knockdown reduced HER2 activation and resensitized cells to cetuximab. Cetuximab-resistant xenografts and PDX models revealed greater efficacy of dual EGFR and HER3 inhibition compared with single antibodies. In PDX tissue samples, cetuximab induced HER3 expression and MM-121 reduced AKT activity. CONCLUSIONS: Clinically relevant PDX models demonstrate that dual targeting of EGFR and HER3 is superior to EGFR targeting alone in HNSCC. Our study illustrates the upregulation of HER3 by cetuximab as one mechanism underlying resistance to EGFR inhibition in HNSCC, supporting further clinical investigations using multiple targeting strategies in patients who have failed cetuximab-based therapy. Clin Cancer Res; 23(3); 677-86. ©2016 AACR.

Antitumor Activity of 2,9-Di-Sec-Butyl-1,10-Phenanthroline.

The anti-tumor effect of a chelating phen-based ligand 2,9-di-sec-butyl-1,10-phenanthroline (dsBPT) and its combination with cisplatin were examined in both lung and head and neck cancer cell lines and xenograft animal models in this study. The effects of this agent on cell cycle and apoptosis were investigated. Protein markers relevant to these mechanisms were also assessed. We found that the inhibitory effect of dsBPT on lung and head and neck cancer cell growth (IC50 ranged between 0.1-0.2 µM) was 10 times greater than that on normal epithelial cells. dsBPT alone induced autophagy, G1 cell cycle arrest, and apoptosis. Our in vivo studies indicated that dsBPT inhibited tumor growth in a dose-dependent manner in a head and neck cancer xenograft mouse model. The combination of dsBPT with cisplatin synergistically inhibited cancer cell growth with a combination index of 0.3. Moreover, the combination significantly reduced tumor volume as compared with the untreated control (p = 0.0017) in a head and neck cancer xenograft model. No organ related toxicities were observed in treated animals. Our data suggest that dsBPT is a novel and potent antitumor drug that warrants further preclinical and clinical development either as a single agent or in combination with known chemotherapy drugs such as cisplatin.

Human papillomavirus oncoprotein E6 upregulates c-Met through p53 downregulation.

PURPOSE: Human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OPSCC) carries a distinct clinical behaviour. c-Met oncogene is an important driver for tumour progression and its relationship with HPV in OPSCC was explored in the present study. EXPERIMENTAL DESIGN: Knockdown of HPV oncogene E6 or p53 alone and in combination was performed to examine their effects on c-Met expression by Western blot and quantitative real-time polymerase chain reaction. The effects of c-Met inhibition on cell proliferation, migration, and colony formation were examined in HPV-positive head and neck squamous cell carcinoma (HNSCC) cells. Retrospectively collected OPSCC patient specimens (N = 78) were stained for c-Met by immunohistochemistry and the staining levels were correlated with HPV status and patient outcomes. RESULTS: E6 knockdown decreased c-Met protein and mRNA expression in HPV-positive HNSCC cells, which was partially abolished by the elimination of p53. Reducing c-Met decreased cell proliferation, migration, and colony formation in HPV-positive HNSCC cells. In OPSCC patient samples, high c-Met expression was associated with HPV-positive status (OR = 4.11, 95%CI: 1.16-14.55, P = 0.028) and tumour stage (OR = 0.27, 95%CI: 0.08-0.93, P = 0.039) by multivariable analysis. In T3/T4 stage patients, high c-Met expression was associated with HPV positivity and low p53 levels, supporting an axis of E6-p53-c-Met regulation. Furthermore, high c-Met expression was marginally associated with poor disease-free survival in HPV-positive patients. CONCLUSIONS: Our results suggest that c-Met may serve as a novel target for treating HPV-associated OPSCC. The data also demonstrate that HPV E6 upregulates c-Met expression partially through p53 downregulation.

Combination of anti-HER3 antibody MM-121/SAR256212 and cetuximab inhibits tumor growth in preclinical models of head and neck squamous cell carcinoma.

The EGFR monoclonal antibody cetuximab is the only approved targeted agent for treating head and neck squamous cell carcinoma (HNSCC). Yet resistance to cetuximab has hindered its activity in this disease. Intrinsic or compensatory HER3 signaling may contribute to cetuximab resistance. To investigate the therapeutic benefit of combining MM-121/SAR256212, an anti-HER3 monoclonal antibody, with cetuximab in HNSCC, we initially screened 12 HNSCC cell lines for total and phosphorylated levels of the four HER receptors. We also investigated the combination of MM-121 with cetuximab in preclinical models of HNSCC. Our results revealed that HER3 is widely expressed and activated in HNSCC cell lines. MM-121 strongly inhibited phosphorylation of HER3 and AKT. When combined with cetuximab, MM-121 exerted a more potent antitumor activity through simultaneously inhibiting the activation of HER3 and EGFR and consequently the downstream PI3K/AKT and ERK pathways in vitro. Both high and low doses of MM-121 in combination with cetuximab significantly suppressed tumor growth in xenograft models and inhibited activations of HER3, EGFR, AKT, and ERK in vivo. Our work is the first report on this new combination in HNSCC and supports the concept that HER3 inhibition may play an important role in future therapy of HNSCC. Our results open the door for further mechanistic studies to better understand the role of HER3 in resistance to EGFR inhibitors in HNSCC.

Honokiol enhances paclitaxel efficacy in multi-drug resistant human cancer model through the induction of apoptosis.

Resistance to chemotherapy remains a major obstacle in cancer therapy. This study aimed to evaluate the molecular mechanism and efficacy of honokiol in inducing apoptosis and enhancing paclitaxel chemotherapy in pre-clinical multi-drug resistant (MDR) cancer models, including lineage-derived human MDR (KB-8-5, KB-C1, KB-V1) and their parental drug sensitive KB-3-1 cancer cell lines. In vitro analyses demonstrated that honokiol effectively inhibited proliferation in KB-3-1 cells and the MDR derivatives (IC50 ranging 3.35 ± 0.13 µg/ml to 2.77 ± 0.22 µg/ml), despite their significant differences in response to paclitaxel (IC50 ranging 1.66 ± 0.09 ng/ml to 6560.9 ± 439.52 ng/ml). Honokiol induced mitochondria-dependent and death receptor-mediated apoptosis in MDR KB cells, which was associated with inhibition of EGFR-STAT3 signaling and downregulation of STAT3 target genes. Combined treatment with honokiol and paclitaxel synergistically augmented cytotoxicity in MDR KB cells, compared with treatment with either agent alone in vitro. Importantly, the combined treatment significantly inhibited in vivo growth of KB-8-5 tumors in a subcutaneous model. Tumor tissues from the combination group displayed a significant inhibition of Ki-67 expression and an increase in TUNEL-positive cells compared with the control group. These results suggest that targeting multidrug resistance using honokiol in combination with chemotherapy drugs may provide novel therapeutic opportunities.

Luteolin nanoparticle in chemoprevention: in vitro and in vivo anticancer activity.

Cancer prevention (chemoprevention) by using naturally occurring dietary agents has gained immense interest because of the broad safety window of these compounds. However, many of these compounds are hydrophobic and poorly soluble in water. They frequently display low bioavailability, poor systemic delivery, and low efficacy. To circumvent this problem, we explored a novel approach toward chemoprevention using nanotechnology to deliver luteolin, a natural compound present in green vegetables. We formulated water-soluble polymer-encapsulated Nano-Luteolin from hydrophobic luteolin, and studied its anticancer activity against lung cancer and head and neck cancer. In vitro studies demonstrated that, like luteolin, Nano-Luteolin inhibited the growth of lung cancer cells (H292 cell line) and squamous cell carcinoma of head and neck (SCCHN) cells (Tu212 cell line). In Tu212 cells, the IC50 value of Nano-Luteolin was 4.13 µmol/L, and that of luteolin was 6.96 µmol/L. In H292 cells, the IC50 of luteolin was 15.56 µmol/L, and Nano-Luteolin was 14.96 µmol/L. In vivo studies using a tumor xenograft mouse model demonstrated that Nano-Luteolin has a significant inhibitory effect on the tumor growth of SCCHN in comparison to luteolin. Our results suggest that nanoparticle delivery of naturally occurring dietary agents like luteolin has many advantages and may have potential application in chemoprevention in clinical settings.