Supplementary Figure 3 from Hedgehog-Producing Cancer Cells Respond to and Require Autocrine Hedgehog Activity

Supplementary Figure 4 from Hedgehog-Producing Cancer Cells Respond to and Require Autocrine Hedgehog Activity

SHH and GLI1 levels are enhanced in primary human UC specimens.

Supplementary Figure 5 from Hedgehog-Producing Cancer Cells Respond to and Require Autocrine Hedgehog Activity

PDF file, 596K, Supplemental Figure S1. Validation of the shRNA knockdown efficiencies. Supplemental Figure S2. UC cell lines require HH signaling for proliferation. Supplemental Figure S3. SMO inhibition induces apoptosis in a subset of UC cell lines. Supplemental Figure S4. A 2507-gene signature predicts the HH-dependence of UC cells for proliferation. Supplemental Figure S5. Constitutive HH signaling is required for the anchorage-independent growth of UC cells. Supplemental Figure S6. UC cells maintain HH signaling activity through ligand expression. Supplemental Figure S7. SMO localizes to primary cilia in UC cell lines. Supplemental Figure S8. In situ hybridization (ISH) confirms the positive staining of SHH and GLI1 IHC in human UC. Supplemental Figure S9. Increased expression of SHH and GLI1 in bladders associated with cystitis cystica.

Supplemental Figures 1-5 with corresponding legends

Supplementary Figure from A Druggable UHRF1/DNMT1/GLI Complex Regulates Sonic Hedgehog–Dependent Tumor Growth

pLKO-based shRNA constructs used in this study.

<p>PDF file, 216K, List of 2507 differentially-expressed genes.</p>

Supplementary Table Legends 1-2, Figure Legends 1-6 from Hedgehog-Producing Cancer Cells Respond to and Require Autocrine Hedgehog Activity

Supplementary Figure 2 from Hedgehog-Producing Cancer Cells Respond to and Require Autocrine Hedgehog Activity

List of 2507 differentially-expressed genes

BACKGROUND:Mohs micrographic surgery (MMS)-associated anxiety is an important issue that remains poorly discussed. Identifying risk factors for MMS-associated anxiety will better equip physicians to manage patients' preoperative anxiety and result in improved patient satisfaction and outcomes.OBJECTIVE:This study aims in identification of risk factors that may play a role in MMS-associated anxiety among patients with nonmelanoma skin cancer.METHODS:A cross-sectional case series was conducted among MMS patients collect information on patient demographics and surgical locations and to measure perioperative anxiety and quality of life (QoL). In addition, adjusted linear and logistic regression analyses were performed to identify potential risk factors that predict MMS-associated anxiety.RESULTS:Significant increases in perioperative anxiety were associated with the eyelid area compared with the noneyelid facial area and nonfacial area ( p ≤ .05). Patients with graduate degrees exhibited less anxiety compared with ones who received less education ( p ≤ .05). Higher perioperative anxiety was associated with a greater impact on QoL ( p ≤ .05).CONCLUSION:This study identified that surgical location and the patient's educational level are 2 critical predicting factors for perioperative anxiety. Furthermore, perioperative anxiety could negatively affect a patient's quality of life and warrants further investigation into effective management.

Dysregulation of Sonic hedgehog (SHH) signaling drives the growth of distinct cancer subtypes, including medulloblastoma (MB). Such cancers have been treated in the clinic with a number of clinically relevant SHH inhibitors, the majority of which target the upstream SHH regulator, Smoothened (SMO). Despite considerable efficacy, many of these patients develop resistance to these drugs, primarily due to mutations in SMO. Therefore, it is essential to identify druggable, signaling components downstream of SMO to target in SMO inhibitor resistant cancers. We utilized an integrated functional genomics approach to identify epigenetic regulators of SHH signaling and identified a novel complex of Ubiquitin-like with PHD and RING finger domains 1 (UHRF1), DNA methyltransferase 1 (DNMT1), and GLI proteins. We show that this complex is distinct from previously described UHRF1/DNMT1 complexes, suggesting that it works in concert to regulate GLI activity in SHH driven tumors. Importantly, we show that UHRF1/DNMT1/GLI complex stability is targeted by a repurposed FDA-approved therapy, with a subsequent reduction in the growth of SHH-dependent MB ex vivo and in vivo. IMPLICATIONS:This work describes a novel, druggable UHRF1/DNMT1/GLI complex that regulates SHH-dependent tumor growth, and highlights an FDA-approved drug capable of disrupting this complex to attenuate tumor growth.

Background:Medulloblastoma (MB) is the most common pediatric brain tumor. Although standard-of-care treatment generally results in good prognosis, many patients exhibit treatment-associated lifelong disabilities. This outcome could be improved by employing therapies targeting the molecular drivers of this cancer. Attempts to do so in the SONIC HEDGEHOG MB subgroup (SHH-MB) have largely focused on the SHH pathway's principal activator, smoothened (SMO). While inhibitors targeting SMO have shown clinical efficacy, recurrence and resistance are frequently noted, likely resulting from mutations in or downstream of SMO. Therefore, identification of novel SHH regulators that act on the pathway's terminal effectors could be used to overcome or prevent such recurrence. We hypothesized that protein arginine methyltransferase 5 (PRMT5) is one such regulator and investigated its role and potential targeting in SHH-MB. Methods:PRMT5 expression in SHH-MB was first evaluated. Knockdown and pharmacological inhibitors of PRMT5 were used in SHH-MB sphere cultures to determine its effect on viability and SHH signaling. GLI1 arginine methylation was then characterized in primary SHH-MB tissue using LC-MS/MS. Finally, PRMT5 inhibitor efficacy was evaluated in vivo. Results:PRMT5 is overexpressed in SHH-MB tissue. Furthermore, SHH-MB viability and SHH activity is dependent on PRMT5. We found that GLI1 isolated from SHH-MB tissues is highly methylated, including three PRMT5 sites that affect SHH-MB cell viability. Importantly, tumor growth is decreased and survival increased in mice given PRMT5 inhibitor. Conclusions:PRMT5 is a requisite driver of SHH-MB that regulates tumor progression. A clinically relevant PRMT5 inhibitor represents a promising candidate drug for SHH-MB therapy.

The Hedgehog/GLI (HH/GLI) signaling pathway plays a critical role in human oncogenesis. Unfortunately, the clinical use of HH inhibitor(s) has been associated with serious adverse effects and mutation-related drug resistance. Since the efficacy of SMO (Smoothened) and GLI inhibitors is limited in clinical trials, there remains a critical need for the HH/GLI pathway inhibitors with different mechanisms of action. Here, we show that esophageal adenocarcinoma (EAC) cell lines are insensitive to vismodegib (SMO inhibitor) but respond to GANT61 (GLI1 inhibitor). Furthermore, we examine the role of GLI1 in tumorigenicity of EAC and how a selective bromodomain inhibitor IBET-151 downregulates transcriptional activity of the GLI1 transcription factor in EAC. Our study demonstrates that GLI1 plays an important role in tumorigenicity of EAC and that elevated GLI1 expression in patients' ultrasound-assisted endoscopic biopsy may predict the response to neoadjuvant chemotherapy (NAC) FOLFOX. Importantly, IBET-151 abrogates the growth of vismodegib-resistant EAC cells and downregulates HH/GLI by reducing the occupancy of BRD4 at the GLI1 locus. IBET-151 also attenuates tumor growth of EAC-PDXs and does so in an on-target manner as it reduces the expression of GLI1. We identify HH/GLI signaling as a novel druggable pathway in EAC as well as validate an ability of clinically relevant GLI inhibitor to attenuate the viability of vismodegib-resistant EAC cells. Therefore, we propose that selective bromodomain inhibitors, such as IBET-151, could be used as novel therapeutic agents for EAC patients harboring GLI-dependent tumors.

Anaplastic large-cell lymphoma (ALCL), a type of peripheral T-cell lymphoma, is a rare form of non-Hodgkin lymphoma, comprising 1% to 2% of non-Hodgkin lymphoma in adults in the United States.1 There are 2 major types of ALCL: systemic ALCL (S-ALCL) and primary cutaneous ALCL. S-ALCL is the more aggressive form, with high rates of metastasis and a poor prognosis. S-ALCL is divided into 2 subgroups based on the expression of anaplastic lymphoma kinase (ALK): ALK- and ALK+. The ALK- subtype is more likely to relapse than the ALK+ subtype, despite having a good initial response to standard chemotherapy.

Nevus of Ota is cosmetically burdensome and often prompts patients to seek treatment. Lasers are commonly used in removing these lesions; however, no systemic analysis has been conducted to support a gold standard laser. To conduct a meta-analysis of the efficacy and safety of Q-switched Nd:YAG lasers (QSNL), Q-switched ruby lasers (QSRL), Q-switched alexandrite lasers (QSAL), and picosecond alexandrite lasers (PSAL) in removing nevus of Ota. Inclusion criteria were nevus of Ota patients treated with QSNL, QSRL, QSAL, or PSAL and documentation of percent clearance and the rate of at least one adverse event. Articles in English, Chinese, or Japanese were included. The prespecified outcome measures were efficacy (percent clearance) and safety (rates of hyperpigmentation, hypopigmentation, scarring, and recurrence). The review included 57 studies and 13,417 patients. The pooled success rate was 64% for QSNL (95% CI 52–76%), 54% for QSRL (95% CI 39–69%), 58% for QSAL (95% CI 44–72%), and 100% for PSAL (95% CI 98–102%). The pooled adverse event rate was 5% for QSNL (95% CI 4–6%), 14% for QSRL (95% CI 9–19%), 9% for QSAL (95% CI 6–12%), and 44% (95% CI 31–57%) for PSAL. QSNL has the most evidence for effectively and safely treating nevus of Ota. PSAL potentially has a superior efficacy; however, further studies are needed to elucidate its side effect profile when treating nevus of Ota.