The present work demonstrates that Cy5.5 conjugated Fe3O4/SiO2 core/shell nanoparticles could allow us to control movement of human natural killer cells (NK-92MI) by an external magnetic field. Required concentration of the nanoparticles for the cell manipulation is as low as ~20 μg Fe/mL. However, the relative ratio of the nanoparticles loaded NK-92MI cells infiltrated into the target tumor site is enhanced by 17-fold by applying magnetic field and their killing activity is still maintained as same as the NK-92MI cells without the nanoparticles. This approach allows us to open alternative clinical treatment with reduced toxicity of the nanoparticles and enhanced infiltration of immunology to the target site.

ObjectivesTo explore lymphocyte infiltration as a potential mechanism behind CXCL14-mediated tumor growth suppression in oral cavity squamous cell carcinoma (OSCC).MethodsWe analyzed single cell RNA-sequencing (scRNA-seq) data from OSCC to identify expression changes among malignant cells in lymph nodes (LN) versus primary tumors. CXCL14 expression in murine OSCC cell lines was quantified using qRT-PCR. Short hairpin RNA knockdown of CXCL14 was performed in mouse oral cavity (MOC)1 cells, and CXCL14 overexpression was performed in MOC2 cells. Cells in each condition were injected into C57BL/6 mice with and without T cell depletion, and tumor volume was measured. At 30 days, tumors were dissociated and analyzed by flow cytometry for CD45+CD3+ T cells. CXCL14 expression was correlated with gene expression signatures of tumor infiltrating lymphocytes (TIL) in scRNA-seq data, as well as TCGA tumors.ResultsscRNA-seq revealed CXCL14 as the most significantly downregulated gene among malignant cells in LNs relative to primary tumor, supporting a role in preventing invasion and/or metastasis. In a murine immunocompetent model, CXCL14 expression was higher in indolent MOC1 cells than in more aggressive MOC2 cells. Tumor growth in vivo was significantly increased by CXCL14 knockdown in MOC1 cells relative to control, with a corresponding decrease in TIL. In MOC2 cells, tumor growth was significantly reduced by CXCL14 overexpression relative to control and TIL were increased. Both effects were lost with T cell depletion. In a human tumor scRNA-seq cohort, we found that only malignant cell CXCL14, but not non-malignant cell or fibroblast CXCL14, was associated with TIL. Bulk CXCL14 from the TCGA cohort had no association with TIL.ConclusionsHigher CXCL14 expression by tumor cells is associated with reduced tumor growth and increased TIL, supporting immune-mediated suppression of tumor growth in OSCC. Given that CXCL14 is downregulated in LN metastases compared with primary tumors, our data raise the possibility that CXCL14-mediated immune infiltration may discourage invasion and metastasis. In human scRNA-seq data, only malignant cell-specific CXCL14 was associated with TIL, suggesting a critical context-dependent effect of CXCL14 expression.

The 19 human aldehyde dehydrogenase (ALDH) gene products are enzymes that metabolize a wide range of reactive aldehydes, including those generated by the metabolism of drugs, alcohol, cigarette smoke and organic compounds. These toxic aldehydes are metabolized into respective non-reactive, non-toxic acids. We reasoned that expression of high levels of ALDH support cancer stem cells growth and chemoresistance by detoxifying metabolic aldehydes and aldehydic intermediates of common chemotherapy drugs. Thus, inhibitors of ALDHs (Aldis) may potentiate current treatments. Many malignant cancer cells and cancer stem cells (CSCs) are resistant to chemotherapy, which can lead to secondary relapses. Further, because head and neck cancer (HNC) stem cells express high levels of ALDHs, Aldis may be useful in increasing sensitivity of these tumors to chemotherapy. Testing several different HNC cell lines demonstrated that SCC4 cells are more resistant to cisplatin treatment and PCI13 cells are less resistant (e.g., at 100μM cisplatin, 30±2% of SCC4 cells survived vs. 15±6% of PCI13 cells; n=16). After cisplatin treatment (15μM for 8 days), ALDH activity increased in SCC4 more than in PCI13 (35±5% vs. 2±0.5% increase) as compared with non-treated cells measured. Co-treatment of cisplatin with Aldi-6 reduced ALDH activity in SCC4 by 67% as measured by Aldefluor assay. When cells were treated with Aldi-6 (an ALDH1, 2 and 3 inhibitor) together with cisplatin, Aldi-6 increased cell death by 30% in SCC4 and by 10% in PCI13 compared with cells treated with cisplatin alone (n=8 each). ALDH3A1 protein levels (but not ALDH1 or 2) increased with cisplatin treatment by ∼100 fold and 25 fold, respectively, in SCC4 and PCI13 as compared with non-treated cells measured by Western blot. Based on this data, we tested the efficacy of Aldi-6 in vivo. Tumor (SCC4)-bearing mice were divided into 4 groups and were treated for two weeks with vehicle, Aldi-6 (8mg/kg/day, delivered via an osmotic pump), cisplatin (2mg/kg/BW, ip injection, 2/week) or cisplatin+Aldi-6. Aldi-6 alone resulted in smaller tumors (average tumor volume (mm3): vehicle, 2,945±112; Aldi6, 773±481). Aldi-6 also enhanced cisplatin reduction of tumor size by more than 60% as compared with cisplatin treatment alone (average tumor volume (mm3): cisplatin, 1,560; Aldi6+cisplatin, 597±112; n=3-6). In conclusion, we found that increased ALDH3A1 levels in HNCs following cisplatin treatment in culture and in vivo may contribute to tumor growth and therefore, ALDH3A1 inhibitors may be suitable as chemotherapeutic agents alone; ALDH3A1 inhibitors may also reduce chemoresistance to cisplatin. Understanding the role of ALDH in HNC not only provides insight into the chemoresistance of this disease, but it may also provide novel strategies for therapy. Citation Format: Jeewon Kim, JuneHo Shin, Che-Hong Chen, Leslie Cruz, Lovisa Farnebo, Jieying Yang, John B. Sunwoo, Daria Mochly-Rosen. Novel aldehyde dehydrogenase inhibitors as potential anti-chemoresistance drugs for head and neck cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3749. doi:10.1158/1538-7445.AM2014-3749