Purpose Ribonucleotide reductase subunit M2 (RRM2) plays an active role in tumor progression. Recently, we reported that depletion of RRM2 by systemic delivery of a nanoparticle carrying RRM2-specific siRNA suppresses head and neck tumor growth. The aim of this study is to clarify the underlying mechanism by which RRM2 depletion inhibits tumor growth. Methods siRNA-mediated gene silencing was performed to downregulate RRM2. Immunoblotting, RT-PCR, confocal microscopy, tissue fractionation, gene overexpression and knockdown were employed to analyze critical apoptosis signaling. Conventional immunohistochemistry (IHC) and quantum dot-based IHF were applied to detect RRM2 and Bcl2 expression and localization in tissue samples from patients and mice. Results Knockdown of RRM2 led to apoptosis through the intrinsic pathway in head and neck squamous cell carcinoma (HNSCC) and non-small cell lung cancer (NSCLC) cell lines. We demonstrated that Bcl-2 is a key determinant controlling apoptosis, both in vitro and in vivo and that RRM2 depletion significantly reduces Bcl-2 protein expression. We observed that RRM2 regulates Bcl-2 protein stability, with RRM2 suppression leading to increased Bcl-2 degradation, and identified their co-localization in HNSCC and NSCLC cells. In a total of 50 specimens each from HNSCC and NSCLC patients, we identified the co-localization of Bcl-2 and RRM2 and found a significant positive correlation between their expression in HNSCC (R=0.98, p<0.0001) and NSCLC (R=0.92, p<0.0001) tumor tissues. Conclusion Our novel findings add to the knowledge of RRM2 in regulating expression of the anti-apoptotic protein Bcl-2 and reveal a critical link between RRM2 and Bcl-2 in apoptosis signaling.

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.

Cancer prevention (chemoprevention) by using naturally occurring dietary agents has gained immense interest due to 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 towards 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 4.13μM, and that of luteolin was 6.96μM. In H292 cells, the IC50 of luteolin was 15.56μM, and Nano-Luteolin was 14.96μM. 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.

Purpose Epidermal growth factor receptor (EGFR) and cyclooxygenase 2 inhibitors synergistically inhibit head and neck squamous cell carcinoma tumorigenesis in preclinical studies. We conducted a phase I and pharmacokinetic study with the erlotinib and celecoxib combination in patients with advanced premalignant lesions. Patients and Methods 36 subjects with oral leukoplakia, mild, moderate, or severe dysplasia, or carcinoma in situ were screened for study participation; 12 consented and received therapy for a median of 5.38 months. Erlotinib was escalated following a standard 3+3 design at 50, 75, and 100mg orally daily and celecoxib was fixed at 400mg twice daily for 6 months. Biopsy of lesions and cytobrush of normal mucosa were performed at baseline, 3, 6 and 12 months. Erlotinib pharmacokinetics were analyzed in 10 subjects. Results The maximum tolerated dose of erlotinib with celecoxib 400mg BID was 50mg per day with skin rash being the main observed toxicity. Overall histologic response rate was 63% (complete response 43%, partial response 14%, stable disease 29%, disease progression 14%). With median follow-up of 36 months, mean time to progression to higher-grade dysplasia or carcinoma was 25.4 months. Downregulation of EGFR and p-ERK in follow-up biopsies correlated with response to treatment. Larger average erlotinib V/F (∼308L) and CL/F (8.3L/hr) compared to previous studies may be related to relatively large average bodyweights. Average erlotinib t1/2 was 25.6hr. Conclusion Encouraging responses to the celecoxib and erlotinib combination correlated with EGFR pathway inhibition. Although erlotinib-related rash was the main limitation to dose escalation, the intervention was well tolerated.