Faculty Bibliography

Dedifferentiation or phenotype switching refers to the transition from a proliferative to an invasive cellular state. We previously identified a 122-gene epigenetic gene signature that classifies primary melanomas as low- versus high-risk (denoted as Epgn1 or Epgn3). We found that the transcriptomes of the Epgn1 low-risk and Epgn3 high-risk cells are similar to the proliferative and invasive cellular states, respectively. These signatures were further validated in melanoma tumor samples. Examination of the chromatin landscape revealed differential H3K27 acetylation in the Epgn1 low-risk versus Epgn3 high-risk cell lines that corroborated with a differential super-enhancer and enhancer landscape. Melanocytic lineage genes (MITF, its targets and regulators) were associated with super-enhancers in the Epgn1 low-risk state whereas invasiveness genes were linked with Epgn3 high-risk status. We identified ITGA3 gene as marked by a super-enhancer element in the Epgn3 invasive cells. Silencing of ITGA3 enhanced invasiveness in both in vitro and in vivo systems suggesting it as a negative regulator of invasion. In conclusion, we define chromatin landscape changes associated with Epgn1/3 and phenotype switching during early steps of melanoma progression that regulate transcriptional reprogramming. This super-enhancer and enhancer-driven epigenetic regulatory mechanism resulting in major changes in the transcriptome could be important in future therapeutic targeting efforts.

In melanoma, immune cell infiltration into the tumor is associated with better patient outcomes and response to immunotherapy. T-cell non-inflamed tumors (cold tumors) are associated with tumor cell-intrinsic Wnt/β-catenin activation, and are typically resistant to anti-PD-1 alone or in combination with anti-CTLA-4 therapy. Reversal of the 'cold tumor' phenotype and identifying new effective immunotherapies are challenges. We sought to investigate the role of a newer immunotherapy agent, B7-H3, in this setting. RNA sequencing was used to identify co-targeting strategies upon B7-H3 inhibition in a well-defined preclinical melanoma model driven by β-catenin. We found that immune checkpoint molecule B7-H3 confers a suppressive tumor microenvironment by modulating antiviral signals and innate immunity. B7-H3 inhibition led to an inflamed microenvironment, up-regulation of CD47/SIRPa signaling, and together with blockade of the macrophage checkpoint CD47 resulted in additive antitumor responses. We found that the antitumor effects of the B7-H3/CD47 antibody combination were dependent on cytokine signaling pathways (CCR5/CCL5 and IL4).

Cyclic AMP (cAMP) has a key role in psoriasis pathogenesis, as indicated by the therapeutic efficacy of phosphodiesterase inhibitors that prevent the degradation of cAMP. However, whether soluble adenylate cyclase (sAC) (encoded by the ADCY10 gene), which is an important source for cAMP, is involved in Th17 cell-mediated inflammation or could be an alternative therapeutic target in psoriasis is unknown. We have utilized the imiquimod model of murine psoriasiform dermatitis to address this question. Adcy10^-/- mice had reduced erythema, scaling and swelling in the skin and reduced CD4+ IL17+ cell numbers in the draining lymph nodes, compared with wild-type mice after induction of psoriasiform dermatitis with imiquimod. Keratinocyte-specific knock out of Adcy10 had no effect on imiquimod-induced ear swelling suggesting keratinocyte sAC has no role in imiquimod-induced inflammation. During Th17 polarization in vitro, naive T cells from Adcy10^-/- mice exhibited reduced IL17 secretion and IL-17+ T-cell proliferation suggesting that differentiation into Th17 cells is suppressed without sAC activity. Interestingly, loss of sAC did not impact the expression of Th17 lineage-defining transcription factors (such as Rorc and cMaf) but rather was required for CREB-dependent gene expression, which is known to support Th17 cell gene expression. Finally, topical application of small molecule sAC inhibitors (sACi) reduced imiquimod-induced psoriasiform dermatitis and Il17 gene expression in the skin. Collectively, these findings demonstrate that sAC is important for psoriasiform dermatitis in mouse skin. sACi may provide an alternative class of topical therapeutics for Th17-mediated skin diseases.

BACKGROUND:Fractional ablative laser resurfacing has been shown to improve the final cosmetic appearance of surgical scars, but optimal timing is unknown. OBJECTIVE:To compare surgical scars treated with fractional carbon dioxide (CO 2 ) laser performed on Day 0 and Day 14. METHODS:Prospective, randomized, split-scar, physician-blinded study of 30 surgical scars on the limbs. Scars halves received fractional CO 2 laser on either Day 0 or Day 14. Scar assessment at 6 months evaluated patient preference, physician modified Manchester Scar Scale (MMSS) score, and quantitative scar analysis on histology (fractal dimension [F D ] and lacunarity [L] analysis). RESULTS:There was no significant difference in patient assessment (54% preferred Day 0 side, 46% preferred Day 14 side, p = .58) or physician assessment (mean MMSS 8.4 for Day 0 vs 8.7 for Day 14, p = .28). Fractal dimensions were similar for both interventions (mean 1.778 for Day 0 vs 1.781 for Day 14, p = .80). Lacunarity was similar for both interventions (mean 0.368 for Day 0 vs 0.345 for Day 14, p = .44). LIMITATIONS:Single-center study with wounds limited to limbs of skin Phototype I-II subjects; 4 of whom were lost to follow-up. CONCLUSION:Intraoperative CO 2 laser is noninferior to Day 14 laser resurfacing for surgical scar treatment.

Cyclic AMP (cAMP) signaling is localized to multiple spatially distinct microdomains, but the role of cAMP microdomains in cancer cell biology is poorly understood. Here, we present a tunable genetic system that allows us to activate cAMP signaling in specific microdomains. We uncover a nuclear cAMP microdomain that activates a tumor-suppressive pathway in a broad range of cancers by inhibiting YAP, a key effector protein of the Hippo pathway, inside the nucleus. We show that nuclear cAMP induces a LATS-dependent pathway leading to phosphorylation of nuclear YAP solely at serine 397 and export of YAP from the nucleus with no change in YAP protein stability. Thus, nuclear cAMP inhibition of nuclear YAP is distinct from other known mechanisms of Hippo regulation. Pharmacologic targeting of specific cAMP microdomains remains an untapped therapeutic approach for cancer; thus, drugs directed at the nuclear cAMP microdomain may provide avenues for the treatment of cancer.