Description: Purpose: Basal cell carcinomas (BCCs) are tumors ignored by immune surveillance. Activated Hedgehog (Hh) signaling within primary cilia is a key driver in the pathogenesis of BCCs. We examined immune alterations during treatment with systemic Hh inhibitors. Experimental Design: We investigated biopsies from patients with BCC before (23 patients) and after 4 weeks of treatment (5 patients) with Hh signaling inhibitor. Ber-Ep4, BCL-2, Ki-67, CD4, CD8, MHC class I, HLA-DR-class II, and SOX9 were analyzed by immunohistochemistry. Primary cilia were analyzed by double immunofluorescence of acetylated tubulin and SOX9. Differential gene expression for 84 cytokines and chemokines was analyzed in 3 patients. Results: After 4 weeks of treatment, we found reduction of Ki-67, SOX9, Ber-EP4, and BCL-2 expression in tumors associated with morphologic signs of squamous differentiation. In addition, the number of cilia-positive BCC cells was significantly decreased. An upregulation of MHC I expression on the cell membranes of residual tumor cells and an influx of CD4 + , HLA-DR-class II + , and CD8 + cells with invasion into the tumor cell nests were found. Finally, qPCR arrays showed the differential expression of genes involved in modulating immune responses. Conclusions: We show that Hh pathway inhibitor–induced tumor regression is accompanied by a dynamic change of the microenvironment with a disruption of immune privilege involving an influx of cytotoxic T cells, activation of the adaptive immune functions, and a profound alteration of the local chemokine/cytokine network. Clin Cancer Res; 21(6); 1289–97. ©2015 AACR .

Description: Pharmacologic activation of the transcription factor NRF2 has been suggested to offer a strategy for cancer prevention. In this study, we present evidence from murine tumorigenesis experiments suggesting there may be limitations to this possibility, based on tumorigenic effects of Nrf2 in murine keratinocytes that have not been described previously. In this setting, Nrf2 expression conferred metabolic alterations in keratinocytes that were protumorigenic in nature, affecting enzymes involved in glutathione biosynthesis or in the oxidative pentose phosphate pathway and other NADPH-producing enzymes. Under stress conditions, coordinate increases in NADPH, purine, and glutathione levels promoted the survival of keratinocytes harboring oncogenic mutations, thereby promoting tumor development. The protumorigenic activity of Nrf2 in keratinocytes was particularly significant in a mouse model of skin tumorigenesis that did not rely upon chemical carcinogenesis. In exploring the clinical relevance of our findings, we confirm that NRF2 and protumorigenic NRF2 target genes were activated in some actinic keratoses, the major precancerous lesion in human skin. Overall, our results reveal an unexpected tumor-promoting activity of activated NRF2 during early phases of skin tumorigenesis. Cancer Res; 75(22); 4817–29. ©2015 AACR.

Description: Purpose: Cutaneous T-cell lymphomas (CTCL) are a heterogeneous group of malignancies that despite available therapies commonly relapse. The emergence of combination epigenetic therapies in other hematologic malignancies have made investigation of such combinations in CTCL a priority. Here, we explore the synergistic antiproliferative effects of romidepsin, an HDAC inhibitor, and azacitidine, a demethylating agent, combination in CTCL. Experimental Design: The growth inhibition under combination treatment and single agent was explored by the MTT cell viability assay and the Annexin V/propidium iodide (PI) apoptosis assay in different CTCL cell lines and tumor cells derived from Sézary syndrome patients. Quantitative analysis of a dose–effect relationship of romidepsin and azacitidine was done by the CompuSyn software. Investigation of mechanism of action was performed by flow cytometry, immunoblotting, qRT-PCR arrays, and chromatin immunoprecipitation. Global CpG methylation sequencing was utilized to study genome methylation alteration under the treatment modalities. Results: The combination of romidepsin and azacitidine exerts synergistic antiproliferative effects and induction of apoptosis involving activation of the caspase cascade in CTCL cell lines and tumor cells derived from Sézary syndrome patients. We identified genes that were selectively induced by the combination treatment, such as the tumor suppressor gene RhoB that is linked to enhanced histone acetylation at its promoter region in parallel with pronounced expression of p21. Global CpG methylation sequencing in a CTCL cell line and tumor cells demonstrated a subset of genes with a unique change in methylation profile in the combination treatment. Conclusions: The synergistic antiproliferative effects of romidepsin and azacitidine combination treatment justify further exploration in clinical trials for advanced CTCL. Clin Cancer Res; 22(8); 2020–31. ©2015 AACR .

Description: Purpose: Aging is a poor prognostic factor for melanoma. We have shown that melanoma cells in an aged microenvironment are more resistant to targeted therapy than identical cells in a young microenvironment. This is dependent on age-related secreted factors. Klotho is an age-related protein whose serum levels decrease dramatically by age 40. Most studies on klotho in cancer have focused on the expression of klotho in the tumor cell. We have shown that exogenous klotho inhibits internalization and signaling of Wnt5A, which drives melanoma metastasis and resistance to targeted therapy. We investigate here whether increasing klotho in the aged microenvironment could be an effective strategy for the treatment of melanoma. Experimental Design: PPAR increases klotho levels and is increased by glitazones. Using rosiglitazone, we queried the effects of rosiglitazone on Klotho/Wnt5A cross-talk, in vitro and in vivo , and the implications of that for targeted therapy in young versus aged animals. Results: We show that rosiglitazone increases klotho and decreases Wnt5A in tumor cells, reducing the burden of both BRAF inhibitor–sensitive and BRAF inhibitor–resistant tumors in aged, but not young mice. However, when used in combination with PLX4720, tumor burden was reduced in both young and aged mice, even in resistant tumors. Conclusions: Using glitazones as adjuvant therapy for melanoma may provide a new treatment strategy for older melanoma patients who have developed resistance to vemurafenib. As klotho has been shown to play a role in other cancers too, our results may have wide relevance for multiple tumor types. Clin Cancer Res; 23(12); 3181–90. ©2017 AACR .