Etoposide Upregulates Survival Favoring Sphingosine-1-Phosphate in Etoposide-Resistant Retinoblastoma Cells

Abstract

Improved knowledge of retinoblastoma chemotherapy resistance is needed to raise treatment efficiency. The objective of this study was to test whether etoposide alters glucosyl-ceramide, ceramide, sphingosine, and sphingosine-1-phosphate (sphingosine-1-P) levels in parental retinoblastoma cells (WERI Rb1) or their etoposide-resistant subclones (WERI EtoR). WERI Rb1 and WERI EtoR were incubated with 400 ng/ml etoposide for 24 h. Levels of glucosyl-ceramides, ceramides, sphingosine, sphingosine-1-P were detected by Q-TOF mass spectrometry. Statistical analysis was done by ANOVA followed by Tukey post-hoc test (p < 0.05). The mRNA expression of sphingolipid pathways enzymes in WERI Rb1, WERI EtoR and four human retinoblastoma tissue samples was analyzed by quantitative real-time PCR. Pathways enzymes mRNA expression confirmed similarities of human sphingolipid metabolism in both cell lines and tissue samples, but different relative expression. Significant up-regulation of sphingosine was seen in both cell lines (p < 0.001). Only sphingosine-1-P up-regulation was significantly increased in WERI EtoR (p < 0.01), but not in WERI Rb1 (p > 0.2). Both cell lines upregulate pro-apoptotic sphingosine after etoposide incubation, but only WERI EtoR produces additional survival favorable sphingosine-1-P. These data may suggest a role of sphingosine-1-P in retinoblastoma chemotherapy resistance, although this seems not to be the only resistance mechanism.

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A Theoretical Insight into Selectivity of Inhibitors toward two Domains of Bromodomain-Containing Protein 4 Using Molecular Dynamics Simulations

Abstract

Bromodomains (BRDs) have been an attractive candidate for development of efficient inhibitors toward gene transcription. Molecular dynamics (MD) simulations followed by principal component (PC) analysis were performed to investigate binding selectivity of inhibitors RVX297, BSP, JQ1, SF2523 and CPD2 toward two domains (BD1 and BD2) of bromodomain-containing protein 4 (BRD4). The results show that inhibitor bindings exert different effect on motions of the BC-loops in BD1 and BD2. The rank of binding free energies calculated by using molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method agrees with the one determined by experiment. The results also suggest that the binding ability of RVX297, BSP and JQ1 to BD2 is stronger than that of them to BD1, while the binding ability of SF2523 to BD2 is obviously weaker than that of SF2523 to BD1. Alanine mutation calculations and the calculated inhibitor-residue interaction spectrum prove that the current five inhibitors have obvious binding selectivity toward BD1 and BD2. This study is not only helpful for further understanding the differences in internal dynamics of BD1 and BD2 caused by inhibitor bindings, but also can theoretically contribute significant guidance to designs of effective and high selective anticancer drugs targeting BD1 and BD2 in BRD4.

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According to the cross-correlation analysis and PC analysis, the inhibitor bindings generate different influence on motions of the BC-loop in the two domains BD1 and BD2 of BRD4. Alanine mutation calculations and the calculated inhibitor-residue interaction spectrum prove that the current five inhibitors have obvious binding selectivity toward BD1 and BD2.

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Allosteric Mechanism of Quinoline Inhibitors for HIV RT-associated RNase with MD simulation and Dynamics Fluctuation Network

Abstract

The human immunodeficiency virus (HIV) is a retrovirus which infects T lymphocyte of human body and causes Immunodeficiency. Reverse transcriptase inhibitors (RTIs) can inhibit some functions of RT, preventing virus synthesis (double-stranded DNA), so that HIV virus replication can be reduced. Experimental results indicate a series of benzimidazole-based inhibitors which target HIV RT-associate RNase to inhibit the reverse transcription of HIV virus. However, the allosteric mechanism is still unclear. Here, molecular dynamics simulations and dynamics fluctuation network analysis were used to reveal the binding mode between the inhibitors and RT-associate RNase. The most active molecule has more hydrophobic and electrostatic interactions than the less active inhibitor. Dynamics correlation network analysis indicates that the most active inhibitor perturbs the network of RT-associate RNase and decreases the correlation of nodes. 3D-QSAR model suggests that two robust and reliable models were constructed and validated by independent test set. 3D-QSAR model also shows that bulky negatively charged or hydrophilic substituent is favourable to bioactivity. These results reveal the allosteric mechanism of quinoline inhibitors and help to improve the bioactivity.

This article is protected by copyright. All rights reserved.

Dynamic correlation networks for bound RTNR suggest that allosteric information freely transferred within the network.

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In silico ligand-based modeling of hBACE-1 inhibitors

Abstract

Alzheimer's disease (AD) is a chronic neurodegenerative disease affecting more than 30 million people worldwide. Development of small molecule inhibitors of human β-secretase 1 (hBACE-1) is being the focus of pharmaceutical industry for the past 15-20 years. Here, we successfully applied multiple ligand-based in silico modeling techniques to understand the inhibitory activities of a diverse set of small molecule hBACE-1 inhibitors reported in the scientific literature. Strikingly, the use of only a small subset of 230 (13%) molecules allowed us to develop quality models that performed reasonably well on the validation set of 1476 (87%) inhibitors. Varying the descriptor sets and the complexity of the modeling techniques resulted in only minor improvements to the model's performance. The current results demonstrate that predictive models can be built by choosing appropriate modeling techniques in spite of using small datasets consisting of diverse chemical classes, a scenario typical in triaging of high-throughput screening (HTS) results to identify false negatives. We hope that these encouraging results will help the community to develop more predictive models that would support research efforts for the debilitating Alzheimer's disease. Additonally, the integrated diversity of the techniques employed will stimulate scientists in the field to use in silico statistical modeling techniques like these to derive better models to help advance the drug discovery projects faster.

This article is protected by copyright. All rights reserved.

Multiple ligand based QSAR approaches with increasing modeling sophistication and descriptor information content were used to develop qualitative classification and quantitative regression models using experimental IC₅₀′s reported for hBACE-1 small molecule inhibitors. The statistical modeling outcome and analysis provides a framework for extending this workflow for other therapeutic targets as well as avenues to further the lead identification and lead optimization of small molecule hBACE-1 inhibitors

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Treatment utilization and outcomes in elderly patients with locally advanced esophageal carcinoma: a review of the National Cancer Database

Abstract

For elderly patients with locally advanced esophageal cancer, therapeutic approaches and outcomes in a modern cohort are not well characterized. Patients ≥70 years old with clinical stage II and III esophageal cancer diagnosed between 1998 and 2012 were identified from the National Cancer Database and stratified based on treatment type. Variables associated with treatment utilization were evaluated using logistic regression and survival evaluated using Cox proportional hazards analysis. Propensity matching (1:1) was performed to help account for selection bias. A total of 21,593 patients were identified. Median and maximum ages were 77 and 90, respectively. Treatment included palliative therapy (24.3%), chemoradiation (37.1%), trimodality therapy (10.0%), esophagectomy alone (5.6%), or no therapy (12.9%). Age ≥80 (OR 0.73), female gender (OR 0.81), Charlson–Deyo comorbidity score ≥2 (OR 0.82), and high-volume centers (OR 0.83) were associated with a decreased likelihood of palliative therapy versus no treatment. Age ≥80 (OR 0.79) and Clinical Stage III (OR 0.33) were associated with a decreased likelihood, while adenocarcinoma histology (OR 1.33) and nonacademic cancer centers (OR 3.9), an increased likelihood of esophagectomy alone compared to definitive chemoradiation. Age ≥80 (OR 0.15), female gender (OR 0.80), and non-Caucasian race (OR 0.63) were associated with a decreased likelihood, while adenocarcinoma histology (OR 2.10) and high-volume centers (OR 2.34), an increased likelihood of trimodality therapy compared to definitive chemoradiation. Each treatment type demonstrated improved survival compared to no therapy: palliative treatment (HR 0.49) to trimodality therapy (HR 0.25) with significance between all groups. Any therapy, including palliative care, was associated with improved survival; however, subsets of elderly patients with locally advanced esophageal cancer are less likely to receive aggressive therapy. Care should be taken to not unnecessarily deprive these individuals of treatment that may improve survival.

Through analysis of the NCDB, we found both definitive and palliative therapy results in improved survival over no therapy in the elderly. Specific groups are less likely to receive aggressive therapy suggesting that nonclinical factors in treatment selection may compromise outcomes.

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MET-GRB2 Signaling-Associated Complexes Correlate with Oncogenic MET Signaling and Sensitivity to MET Kinase Inhibitors

Purpose: Targeting MET in cancer is hampered by lack of diagnostics that accurately reflect high MET signaling and dependence. We hypothesized that assays reflecting MET signaling associated protein complexes could redefine tumors dependent on MET and could add additional precision beyond genomic assessments.

Experimental Design: We used biochemical approaches, cellular viability studies, and proximity ligation assays to assess MET dependence. We examined MET signaling complexes in lung cancer patient specimens (N = 406) and patient-derived xenograft (PDX) models of solid tumors (N = 308). We evaluated response to crizotinib in a MET-amplified cohort of PDX models of lung cancer (N = 6) and provide a case report of a lung cancer patient harboring a exon14 MET splice variant.

Results: We found the interaction of MET with the adaptor protein GRB2 is necessary for oncogenic survival signaling by MET. MET-GRB2 complexes were identified only within MET-amplified PDX models and patient specimens but exhibit substantial variability. Lack of MET-GRB2 complexes was associated with lack of response to MET TKI in cell lines and PDX models. Presence of MET-GRB2 complexes can further subtype tumors with exon14 MET splice variants. Presence of these complexes correlated with response to crizotinib in one patient with exon14 MET lacking MET gene amplification.

Conclusions: Proximity assays measuring MET-GRB2 signaling complexes provide novel insights into MET-mediated signaling and could complement current clinical genomics-based assay platforms. Clin Cancer Res; 23(22); 7084–96. ©2017 AACR.

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FDA Drug Approval: Elotuzumab in Combination with Lenalidomide and Dexamethasone for the Treatment of Relapsed or Refractory Multiple Myeloma

On November 30, 2015, the FDA approved elotuzumab (Empliciti; Bristol-Myers Squibb) in combination with lenalidomide and dexamethasone for the treatment of patients with multiple myeloma who received one to three prior therapies. FDA approval was based primarily on the results of a phase III, randomized, open-label, multicenter trial, CA204004, which evaluated elotuzumab in combination with lenalidomide and dexamethasone (E-Ld) compared with lenalidomide and dexamethasone (Ld) alone in patients with relapsed or refractory multiple myeloma. Coprimary endpoints were progression-free survival (PFS) and overall response rate (ORR). The key secondary endpoint was overall survival, but these data were not mature at the time of clinical database cutoff. The trial demonstrated a statistically significant improvement in PFS, with an estimated HR of 0.70 for E-Ld over Ld [95% confidence interval (CI), 0.57–0.85; P = 0.0004). Estimated median PFS was 19.4 months in the E-Ld arm and 14.9 months in the Ld arm. ORR was 75.8% in the E-Ld arm compared with 65.5% in the Ld arm. Serious adverse reactions were reported in 65% of patients in the E-Ld arm compared with 57% in the Ld arm. The FDA approved elotuzumab with the following warnings and precautions: infusion reactions, infections, second primary malignancies, hepatotoxicity, and interference with the determination of complete response. Clin Cancer Res; 23(22); 6759–63. ©2017 AACR.

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Enhanced Cancer Immunotherapy by Chimeric Antigen Receptor-Modified T Cells Engineered to Secrete Checkpoint Inhibitors

Purpose: Despite favorable responses of chimeric antigen receptor (CAR)-engineered T-cell therapy in patients with hematologic malignancies, the outcome has been far from satisfactory in the treatment of solid tumors, partially owing to the development of an immunosuppressive tumor microenvironment. To overcome this limitation, we engineered CAR T cells secreting checkpoint inhibitors (CPI) targeting PD-1 (CAR.αPD1-T) and evaluated their efficacy in a human lung carcinoma xenograft mouse model.

Experimental Design: To evaluate the effector function and expansion capacity of CAR.αPD1-T cells in vitro, we measured the production of IFN and T-cell proliferation following antigen-specific stimulation. Furthermore, the antitumor efficacy of CAR.αPD1-T cells, CAR T cells, and CAR T cells combined with anti–PD-1 antibody was determined using a xenograft mouse model. Finally, the underlying mechanism was investigated by analyzing the expansion and functional capacity of TILs.

Results: Human anti–PD-1 CPIs secreted by CAR.αPD1-T cells efficiently bound to PD-1 and reversed the inhibitory effect of PD-1/PD-L1 interaction on T-cell function. PD-1 blockade by continuously secreted anti–PD-1 attenuated the inhibitory T-cell signaling and enhanced T-cell expansion and effector function both in vitro and in vivo. In the xenograft mouse model, we demonstrated that the secretion of anti–PD-1 enhanced the antitumor activity of CAR T cells and prolonged overall survival.

Conclusions: With constitutive anti–PD-1 secretion, CAR.αPD1-T cells are more functional and expandable, and more efficient at tumor eradication than parental CAR T cells. Collectively, our study presents an important and novel strategy that enables CAR T cells to achieve better antitumor immunity, especially in the treatment of solid tumors. Clin Cancer Res; 23(22); 6982–92. ©2017 AACR.

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Immunotherapy of Prostate Cancer: Facts and Hopes

In the last few years, immunotherapy has become an important cancer treatment modality, and although the principles of immunotherapy have evolved over many decades, the FDA approvals of sipuleucel-T and ipilimumab began a new wave in immuno-oncology. Despite the current enthusiasm, it is unlikely that any of the immunotherapeutics alone can dramatically change prostate cancer outcomes, but combination strategies are more promising and provide a reason for optimism. Several completed and ongoing studies have shown that the combination of cancer vaccines or checkpoint inhibitors with different immunotherapeutic agents, hormonal therapy (enzalutamide), radiotherapy (radium 223), DNA-damaging agents (olaparib), or chemotherapy (docetaxel) can enhance immune responses and induce more dramatic, long-lasting clinical responses without significant toxicity. The goal of prostate cancer immunotherapy does not have to be complete eradication of advanced disease but rather the return to an immunologic equilibrium with an indolent disease state. In addition to determining the optimal combination of treatment regimens, efforts are also ongoing to discover biomarkers of immune response. With such concerted efforts, the future of immunotherapy in prostate cancer looks brighter than ever. Clin Cancer Res; 23(22); 6764–70. ©2017 AACR.

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Personalized Medicine-Based Approach to Model Patterns of Chemoresistance and Tumor Recurrence Using Ovarian Cancer Stem Cell Spheroids

Purpose: Chemoresistant ovarian cancers grow in suspension within the ascites fluid. To screen the effect of chemotherapeutics and biologics on resistant ovarian cancers with a personalized basis, we developed a 3D hanging drop spheroid platform.

Experimental Design: We initiated spheroids with primary aldehyde dehydrogenase–positive (ALDH⁺) CD133⁺ ovarian cancer stem cells (OvCSC) from different patient samples and demonstrated that stem cell progeny from harvested spheroids was similar to the primary tumor. OvCSC spheroids were utilized to initiate tumors in immunodeficient mice. Drug responses to cisplatin and ALDH-targeting compound or JAK2 inhibitor determined whether the OvCSC population within the spheroids could be targeted. Cells that escaped therapy were isolated and used to initiate new spheroids and model tumor reemergence in a personalized manner.

Results: OvCSC spheroids from different patients exhibited varying and personalized responses to chemotherapeutics. Xenografts were established from OvCSC spheroids, even with a single spheroid. Distinct responses to therapy were observed in distinct primary tumor xenografts similar to those observed in spheroids. Spheroids resistant to cisplatin/ALDH inhibitor therapy had persistent, albeit lower ALDH expression and complete loss of CD133 expression, whereas those resistant to cisplatin/JAK2 inhibitor therapy were enriched for ALDH⁺ cells.

Conclusions: Our 3D hanging drop suspension platform can be used to propagate primary OvCSCs that represent individual patient tumors effectively by differentiating in vitro and initiating tumors in mice. Therefore, our platform can be used to study cancer stem cell biology and model tumor reemergence to identify new targeted therapeutics from an effective personalized medicine standpoint. Clin Cancer Res; 23(22); 6934–45. ©2017 AACR.

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Low-Dose Cyclophosphamide Induces Antitumor T-Cell Responses, which Associate with Survival in Metastatic Colorectal Cancer

Purpose: Anticancer T-cell responses can control tumors, but immunosuppressive mechanisms in vivo prevent their function. The role of regulatory T cells (Tregs) in metastatic colorectal cancer is unclear. We have previously shown depletion of Tregs enhances colorectal cancer–specific effector T-cell responses. Low-dose cyclophosphamide targets Tregs in animal models and some human studies; however, the effect of cyclophosphamide in metastatic colorectal cancer is unknown.

Experimental Design: Fifty-five patients with metastatic colorectal cancer were enrolled in a phase I/II trial and randomly assigned to receive 2-week-long courses of low-dose (50 mg twice a day) cyclophosphamide or not. The absolute number, phenotype, and antitumor function of peripheral blood–derived lymphocyte subsets were monitored throughout treatment, as well as during 18-month follow-up.

Results: Initially, cyclophosphamide reduced proliferation in all lymphocyte subsets; however, a rapid mobilization of effector T cells overcame this decrease, leading to increased absolute T-cell numbers. In contrast, a reduction in proportional and absolute Treg, B-cell, and NK-cell numbers occurred. The expansion and subsequent activation of effector T cells was focused on tumor-specific T cells, producing both granzyme B and IFN. Cyclophosphamide-treated patients demonstrating the most enhanced IFN⁺ tumor-specific T-cell responses exhibited a significant delay in tumor progression [HR = 0.29; 95% confidence interval (CI), 0.12–0.69; P = 0.0047), compared with nonresponders and no-treatment controls.

Conclusions: Cyclophosphamide-induced Treg depletion is mirrored by a striking boost in antitumor immunity. This study provides the first direct evidence of the benefit of naturally primed T cells in patients with metastatic colorectal cancer. Our results also support the concept that nonmutated self-antigens may act as useful targets for immunotherapies. Clin Cancer Res; 23(22); 6771–80. ©2017 AACR.

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Macrophages Facilitate Resistance to Anti-VEGF Therapy by Altered VEGFR Expression

Purpose: VEGF-targeted therapies have modest efficacy in cancer patients, but acquired resistance is common. The mechanisms underlying such resistance are poorly understood.

Experimental Design: To evaluate the potential role of immune cells in the development of resistance to VEGF blockade, we first established a preclinical model of adaptive resistance to anti-VEGF therapy. Additional in vitro and in vivo studies were carried out to characterize the role of macrophages in such resistance.

Results: Using murine cancer models of adaptive resistance to anti-VEGF antibody (AVA), we found a previously unrecognized role of macrophages in such resistance. Macrophages were actively recruited to the tumor microenvironment and were responsible for the emergence of AVA resistance. Depletion of macrophages following emergence of resistance halted tumor growth and prolonged survival of tumor-bearing mice. In a macrophage-deficient mouse model, resistance to AVA failed to develop, but could be induced by injection of macrophages. Downregulation of macrophage VEGFR-1 and VEGFR-3 expression accompanied upregulation of alternative angiogenic pathways, facilitating escape from anti-VEGF therapy.

Conclusions: These findings provide a new understanding of the mechanisms underlying the modest efficacy of current antiangiogenesis therapies and identify new opportunities for combination approaches for ovarian and other cancers. Clin Cancer Res; 23(22); 7034–46. ©2017 AACR.

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Genomic Characterization of Vulvar (Pre)cancers Identifies Distinct Molecular Subtypes with Prognostic Significance

Purpose: Vulvar cancer (VC) can be subclassified by human papillomavirus (HPV) status. HPV-negative VCs frequently harbor TP53 mutations; however, in-depth analysis of other potential molecular genetic alterations is lacking. We comprehensively assessed somatic mutations in a large series of vulvar (pre)cancers.

Experimental Design: We performed targeted next-generation sequencing (17 genes), p53 immunohistochemistry and HPV testing on 36 VC and 82 precursors (sequencing cohort). Subsequently, the prognostic significance of the three subtypes identified in the sequencing cohort was assessed in a series of 236 VC patients (follow-up cohort).

Results: Frequent recurrent mutations were identified in HPV-negative vulvar (pre)cancers in TP53 (42% and 68%), NOTCH1 (28% and 41%), and HRAS (20% and 31%). Mutation frequency in HPV-positive vulvar (pre)cancers was significantly lower (P = 0.001). Furthermore, a substantial subset of the HPV-negative precursors (35/60, 58.3%) and VC (10/29, 34.5%) were TP53 wild-type (wt), suggesting a third, not-previously described, molecular subtype. Clinical outcomes in the three different subtypes (HPV⁺, HPV^–/p53wt, HPV^–/p53abn) were evaluated in a follow-up cohort consisting of 236 VC patients. Local recurrence rate was 5.3% for HPV⁺, 16.3% for HPV^–/p53wt and 22.6% for HPV^–/p53abn tumors (P = 0.044). HPV positivity remained an independent prognostic factor for favorable outcome in the multivariable analysis (P = 0.020).

Conclusions: HPV^– and HPV⁺ vulvar (pre)cancers display striking differences in somatic mutation patterns. HPV^–/p53wt VC appear to be a distinct clinicopathologic subgroup with frequent NOTCH1 mutations. HPV⁺ VC have a significantly lower local recurrence rate, independent of clinicopathological variables, opening opportunities for reducing overtreatment in VC. Clin Cancer Res; 23(22); 6781–9. ©2017 AACR.

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Antagonism of EG-VEGF Receptors as Targeted Therapy for Choriocarcinoma Progression In Vitro and In Vivo

Purpose: Choriocarcinoma (CC) is the most malignant gestational trophoblastic disease that often develops from complete hydatidiform moles (CHM). Neither the mechanism of CC development nor its progression is yet characterized. We recently identified endocrine gland–derived vascular endothelial growth factor (EG-VEGF) as a novel key placental growth factor that controls trophoblast proliferation and invasion. EG-VEGF acts via two receptors, PROKR1 and PROKR2. Here, we demonstrate that EG-VEGF receptors can be targeted for CC therapy.

Experimental Design: Three approaches were used: (i) a clinical investigation comparing circulating EG-VEGF in control (n = 20) and in distinctive CHM (n = 38) and CC (n = 9) cohorts, (ii) an in vitro study investigating EG-VEGF effects on the CC cell line JEG3, and (iii) an in vivo study including the development of a novel CC mouse model, through a direct injection of JEG3-luciferase into the placenta of gravid SCID-mice.

Results: Both placental and circulating EG-VEGF levels were increased in CHM and CC (x5) patients. EG-VEGF increased JEG3 proliferation, migration, and invasion in two-dimensional (2D) and three-dimensional (3D) culture systems. JEG3 injection in the placenta caused CC development with large metastases compared with their injection into the uterine horn. Treatment of the animal model with EG-VEGF receptor's antagonists significantly reduced tumor development and progression and preserved pregnancy. Antibody-array and immunohistological analyses further deciphered the mechanism of the antagonist's actions.

Conclusions: Our work describes a novel preclinical animal model of CC and presents evidence that EG-VEGF receptors can be targeted for CC therapy. This may provide safe and less toxic therapeutic options compared with the currently used multi-agent chemotherapies. Clin Cancer Res; 23(22); 7130–40. ©2017 AACR.

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Single Dose of the CXCR4 Antagonist BL-8040 Induces Rapid Mobilization for the Collection of Human CD34+ Cells in Healthy Volunteers

Purpose: The potential of the high-affinity CXCR4 antagonist BL-8040 as a monotherapy-mobilizing agent and its derived graft composition and quality were evaluated in a phase I clinical study in healthy volunteers (NCT02073019).

Experimental Design: The first part of the study was a randomized, double-blind, placebo-controlled dose escalation phase. The second part of the study was an open-label phase, in which 8 subjects received a single injection of BL-8040 (1 mg/kg) and approximately 4 hours later underwent a standard leukapheresis procedure. The engraftment potential of the purified mobilized CD34⁺ cells was further evaluated by transplanting the cells into NSG immunodeficient mice.

Results: BL-8040 was found safe and well tolerated at all doses tested (0.5–1 mg/kg). The main treatment-related adverse events were mild to moderate. Transient injection site and systemic reactions were mitigated by methylprednisolone, paracetamol, and promethazine pretreatment. In the first part of the study, BL-8040 triggered rapid and substantial mobilization of WBCs and CD34⁺ cells in all tested doses. Four hours postdose, the count rose to a mean of 8, 37, 31, and 35 cells/μL (placebo, 0.5, 0.75, and 1 mg/kg, respectively). FACS analysis revealed substantial mobilization of immature dendritic, T, B, and NK cells. In the second part, the mean CD34⁺ cells/kg collected were 11.6 x 10⁶ cells/kg. The graft composition was rich in immune cells.

Conclusions: The current data demonstrate that BL-8040 is a safe and effective monotherapy strategy for the collection of large amounts of CD34⁺ cells and immune cells in a one-day procedure for allogeneic HSPC transplantation. Clin Cancer Res; 23(22); 6790–801. ©2017 AACR.

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Combined CDK4/6 and mTOR Inhibition Is Synergistic against Glioblastoma via Multiple Mechanisms

Purpose: Glioblastoma (GBM) is a deadly brain tumor marked by dysregulated signaling and aberrant cell-cycle control. Molecular analyses have identified that the CDK4/6-Rb-E2F axis is dysregulated in about 80% of GBMs. Single-agent CDK4/6 inhibitors have failed to provide durable responses in GBM, suggesting a need to combine them with other agents. We investigate the efficacy of the combination of CDK4/6 inhibition and mTOR inhibition against GBM.

Experimental Design: Preclinical in vitro and in vivo assays using primary GBM cell lines were performed.

Results: We show that the CDK4/6 inhibitor palbociclib suppresses the activity of downstream mediators of the mTOR pathway, leading to rebound mTOR activation that can be blocked by the mTOR inhibitor everolimus. We further show that mTOR inhibition with everolimus leads to activation of the Ras mediator Erk that is reversible with palbociclib. The combined treatment strongly disrupts GBM metabolism, resulting in significant apoptosis. Further increasing the utility of the combination for brain cancers, everolimus significantly increases the brain concentration of palbociclib.

Conclusions: Our findings demonstrate that the combination of CDK4/6 and mTOR inhibition has therapeutic potential against GBM and suggest it should be evaluated in a clinical trial. Clin Cancer Res; 23(22); 6958–68. ©2017 AACR.

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Impact of Therapy on Genomics and Transcriptomics in High-Risk Prostate Cancer Treated with Neoadjuvant Docetaxel and Androgen Deprivation Therapy

Purpose: The combination of docetaxel chemotherapy and androgen deprivation therapy (ADT) has become a standard treatment for patients with metastatic prostate cancer. The recently accrued phase III CALGB 90203 trial was designed to investigate the clinical effectiveness of this treatment approach earlier in the disease. Specimens from this trial offer a unique opportunity to interrogate the acute molecular response to docetaxel and ADT and identify potential biomarkers.

Experimental Design: We evaluated baseline clinical data, needle biopsies, and radical prostatectomy (RP) specimens from 52 (of 788) patients enrolled on CALGB 90203 at one high volume center. Pathology review, tumor and germline-targeted DNA sequencing (n = 72 genes), and expression profiling using NanoString platform (n = 163 genes) were performed to explore changes in critical prostate cancer pathways linked to aggression and resistance.

Results: Three of 52 patients had only microfocal residual cancer at prostatectomy. The most common alterations included TMPRSS2-ERG fusion (n = 32), TP53 mutation or deletion (n = 11), PTEN deletion (n = 6), FOXA1 (n = 6), and SPOP (n = 4) mutation, with no significant enrichment in posttreated specimens. We did not observe AR amplification or mutations. The degree of AR signaling suppression varied among treated tumors and there was upregulation of both AR and AR-V7 expression as well as a subset of neuroendocrine and plasticity genes.

Conclusions: These data support the feasibility of targeted and temporal genomic and transcriptome profiling of neoadjuvant-treated prostate cancer with limited formalin-fixed paraffin embedded tissue requirement. Characterization of the heterogeneity of treatment response and molecular outliers that arise posttreatment provides new insight into potential early markers of resistance. Clin Cancer Res; 23(22); 6802–11. ©2017 AACR.

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Defective Cyclin B1 Induction in Trastuzumab-emtansine (T-DM1) Acquired Resistance in HER2-positive Breast Cancer

Purpose: Trastuzumab-emtansine (T-DM1) is a standard treatment in advanced HER2-positive breast cancer. However, resistance inevitably occurs. We aimed to identify mechanisms of acquired T-DM1 resistance.

Experimental Design: HER2-positive breast cancer cells (HCC1954, HCC1419, SKBR3, and BT474) were treated in a pulse-fashion with T-DM1 to induce a resistant phenotype. Cellular and molecular effects of T-DM1 in parental versus resistant cells were compared. CDK1 kinase activity and cyclin B1 expression were assayed under various conditions. Genetic modifications to up- or downregulate cyclin B1 were conducted. Effects of T-DM1 on cyclin B1 levels, proliferation, and apoptosis were assayed in human HER2-positive breast cancer explants.

Results: We obtained three cell lines with different levels of acquired T-DM1 resistance (HCC1954/TDR, HCC1419/TDR, and SKBR3/TDR cells). HER2 remained amplified in the resistant cells. Binding to HER2 and intracellular uptake of T-DM1 were maintained in resistant cells. T-DM1 induced cyclin B1 accumulation in sensitive but not resistant cells. Cyclin B1 knockdown by siRNA in parental cells induced T-DM1 resistance, while increased levels of cyclin B1 by silencing cdc20 partially sensitized resistant cells. In a series of 18 HER2-positive breast cancer fresh explants, T-DM1 effects on proliferation and apoptosis paralleled cyclin B1 accumulation.

Conclusions: Defective cyclin B1 induction by T-DM1 mediates acquired resistance in HER2-positive breast cancer cells. These results support the testing of cyclin B1 induction upon T-DM1 treatment as a pharmacodynamic predictor in HER2-positive breast cancer. Clin Cancer Res; 23(22); 7006–19. ©2017 AACR.

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Expression of PD-L1 in Hormone-naïve and Treated Prostate Cancer Patients Receiving Neoadjuvant Abiraterone Acetate plus Prednisone and Leuprolide

Purpose: Programmed cell death ligand-1 (PD-L1)/programmed cell death-1 (PD-1) blockade has been unsuccessful in prostate cancer, with poor immunogenicity and subsequent low PD-L1 expression in prostate cancer being proposed as an explanation. However, recent studies indicate that a subset of prostate cancer may express significant levels of PD-L1. Furthermore, the androgen antagonist enzalutamide has been shown to upregulate PD-L1 expression in prostate cancer preclinical models. In this study, we evaluated the effect of neoadjuvant androgen deprivation therapy with abiraterone acetate plus prednisone and leuprolide (Neo-AAPL) on PD-L1 expression in prostate cancer.

Experimental Design: Radical prostatectomy (RP) tissues were collected from 44 patients with intermediate- to high-risk prostate cancer who underwent RP after Neo-AAPL treatment. Untreated prostate cancer tissues were collected from 130 patients, including 44 matched controls for the Neo-AAPL cases. Tumor PD-L1 expression was detected by IHC using validated anti-PD-L1 antibodies. Tumor-infiltrating CD8⁺ cells were analyzed in trial cases and matched controls. Expression of DNA mismatch repair genes was examined in PD-L1–positive tumors.

Results: Neo-AAPL–treated tumors showed a trend toward decreased PD-L1 positivity compared with matched controls (7% vs. 21% having ≥1% positive tumor cells; P = 0.062). Treated tumors also harbored significantly fewer tumor-infiltrating CD8⁺ cells (P = 0.029). In 130 untreated prostate cancers, African American ethnicity, elevated serum PSA, and small prostate independently predicted tumor PD-L1 positivity. Loss of MSH2 expression was observed in 1 of 21 PD-L1–positive tumors.

Conclusions: A subset of prostate cancer expresses PD-L1, which is not increased by Neo-AAPL treatment, indicating that combining Neo-AAPL treatment with PD-L1/PD-1 blockade may not be synergistic. Clin Cancer Res; 23(22); 6812–22. ©2017 AACR.

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Macropinocytosis of Bevacizumab by Glioblastoma Cells in the Perivascular Niche Affects their Survival

Purpose: Bevacizumab, a humanized monoclonal antibody to VEGF, is used routinely in the treatment of patients with recurrent glioblastoma (GBM). However, very little is known regarding the effects of bevacizumab on the cells in the perivascular space in tumors.

Experimental Design: Established orthotopic xenograft and syngeneic models of GBM were used to determine entry of monoclonal anti-VEGF-A into, and uptake by cells in, the perivascular space. Based on the results, we examined CD133⁺ cells derived from GBM tumors in vitro. Bevacizumab internalization, trafficking, and effects on cell survival were analyzed using multilabel confocal microscopy, immunoblotting, and cytotoxicity assays in the presence/absence of inhibitors.

Results: In the GBM mouse models, administered anti-mouse-VEGF-A entered the perivascular tumor niche and was internalized by Sox2⁺/CD44⁺ tumor cells. In the perivascular tumor cells, bevacizumab was detected in the recycling compartment or the lysosomes, and increased autophagy was found. Bevacizumab was internalized rapidly by CD133⁺/Sox2⁺-GBM cells in vitro through macropinocytosis with a fraction being trafficked to a recycling compartment, independent of FcRn, and a fraction to lysosomes. Bevacizumab treatment of CD133⁺ GBM cells depleted VEGF-A and induced autophagy thereby improving cell survival. An inhibitor of lysosomal acidification decreased bevacizumab-induced autophagy and increased cell death. Inhibition of macropinocytosis increased cell death, suggesting macropinocytosis of bevacizumab promotes CD133⁺ cell survival.

Conclusions: We demonstrate that bevacizumab is internalized by Sox2⁺/CD44⁺-GBM tumor cells residing in the perivascular tumor niche. Macropinocytosis of bevacizumab and trafficking to the lysosomes promotes CD133⁺ cell survival, as does the autophagy induced by bevacizumab depletion of VEGF-A. Clin Cancer Res; 23(22); 7059–71. ©2017 AACR.

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A Phase II Trial of Neoadjuvant MK-2206, an AKT Inhibitor, with Anastrozole in Clinical Stage II or III PIK3CA-Mutant ER-Positive and HER2-Negative Breast Cancer

Purpose: Hyperactivation of AKT is common and associated with endocrine resistance in estrogen receptor–positive (ER⁺) breast cancer. The allosteric pan-AKT inhibitor MK-2206 induced apoptosis in PIK3CA-mutant ER⁺ breast cancer under estrogen-deprived condition in preclinical studies. This neoadjuvant phase II trial was therefore conducted to test the hypothesis that adding MK-2206 to anastrozole induces pathologic complete response (pCR) in PIK3CA mutant ER⁺ breast cancer.

Experimental Design: Potential eligible patients with clinical stage II/III ER⁺/HER2^– breast cancer were preregistered and received anastrozole (goserelin if premenopausal) for 28 days in cycle 0 pending tumor PIK3CA sequencing. Patients positive for PIK3CA mutation in the tumor were eligible to start MK-2206 (150 mg orally weekly, with prophylactic prednisone) on cycle 1 day 2 (C1D2) and to receive a maximum of four 28-day cycles of combination therapy before surgery. Serial biopsies were collected at preregistration, C1D1 and C1D17.

Results: Fifty-one patients preregistered and 16 of 22 with PIK3CA-mutant tumors received study drug. Three patients went off study due to C1D17 Ki67 >10% (n = 2) and toxicity (n = 1). Thirteen patients completed neoadjuvant therapy followed by surgery. No pCRs were observed. Rash was common. MK-2206 did not further suppress cell proliferation and did not induce apoptosis on C1D17 biopsies. Although AKT phosphorylation was reduced, PRAS40 phosphorylation at C1D17 after MK-2206 persisted. One patient acquired an ESR1 mutation at surgery.

Conclusions: MK-2206 is unlikely to add to the efficacy of anastrozole alone in PIK3CA-mutant ER⁺ breast cancer and should not be studied further in the target patient population. Clin Cancer Res; 23(22); 6823–32. ©2017 AACR.

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Downregulation of SAFB Sustains the NF-{kappa}B Pathway by Targeting TAK1 during the Progression of Colorectal Cancer

Purpose: To investigate the role and the underlying mechanism of scaffold attachment factor B (SAFB) in the progression of colorectal cancer (CRC).

Experimental Design: SAFB expression was analyzed in the Cancer Outlier Profile Analysis of Oncomine and in 175 paraffin-embedded archived CRC tissues. Gene Ontology analyses were performed to explore the mechanism of SAFB in CRC progression. Western blot, RT-PCR, luciferase assay, and chromatin immunoprecipitation (ChIP) were used to detect the regulation of transforming growth factor-β–activated kinase 1 (TAK1) and NF-B signaling by SAFB. The role of SAFB in invasion, metastasis, and angiogenesis was investigated using in vitro and in vivo assays. The relationship between SAFB and TAK1 was analyzed in CRC tissues.

Results: SAFB was downregulated in CRC tissues, and low expression of SAFB was significantly associated with an aggressive phenotype and poorer survival of CRC patients. The downregulation of SAFB activated NF-B signaling by targeting the TAK1 promoter. Ectopic expression of SAFB inhibited the development of aggressive features and metastasis of CRC cells both in vitro and in vivo. The overexpression of TAK1 could rescue the aggressive features in SAFB-overexpressed cells. Furthermore, the expression of SAFB in CRC tissues was negatively correlated with the expression of TAK1- and NF-B–related genes.

Conclusions: Our results show that SAFB regulated the activity of NF-B signaling in CRC by targeting TAK1. This novel mechanism provides a comprehensive understanding of both SAFB and the NF-B signaling pathway in the progression of CRC and indicates that the SAFB–TAK1–NF-B axis is a potential target for early therapeutic intervention in CRC progression. Clin Cancer Res; 23(22); 7108–18. ©2017 AACR.

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Phase I Study of a Poxviral TRICOM-Based Vaccine Directed Against the Transcription Factor Brachyury

Purpose: The transcription factor brachyury has been shown in preclinical studies to be a driver of the epithelial-to-mesenchymal transition (EMT) and resistance to therapy of human tumor cells. This study describes the characterization of a Modified Vaccinia Ankara (MVA) vector–based vaccine expressing the transgenes for brachyury and three human costimulatory molecules (B7.1, ICAM-1, and LFA-3, designated TRICOM) and a phase I study with this vaccine.

Experimental Design: Human dendritic cells (DC) were infected with MVA-brachyury-TRICOM to define their ability to activate brachyury-specific T cells. A dose-escalation phase I study (NCT02179515) was conducted in advanced cancer patients (n = 38) to define safety and to identify brachyury-specific T-cell responses.

Results: MVA-brachyury-TRICOM-infected human DCs activated CD8⁺ and CD4⁺ T cells specific against the self-antigen brachyury in vitro. No dose-limiting toxicities were observed due to vaccine in cancer patients at any of the three dose levels. One transient grade 3 adverse event (AE) possibly related to vaccine (diarrhea) resolved without intervention and did not recur with subsequent vaccine. All other AEs related to vaccine were transient and ≤grade 2. Brachyury-specific T-cell responses were observed at all dose levels and in most patients.

Conclusions: The MVA-brachyury-TRICOM vaccine directed against a transcription factor known to mediate EMT can be administered safely in patients with advanced cancer and can activate brachyury-specific T cells in vitro and in patients. Further studies of this vaccine in combination therapies are warranted and planned. Clin Cancer Res; 23(22); 6833–45. ©2017 AACR.

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Highlights of This Issue

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Epidermal Growth Factor Receptor Variant III (EGFRvIII) Positivity in EGFR-Amplified Glioblastomas: Prognostic Role and Comparison between Primary and Recurrent Tumors

Purpose: Approximately 40% of all glioblastomas have amplified the EGFR gene, and about half of these tumors express the EGFRvIII variant. The prognostic role of EGFRvIII in EGFR-amplified glioblastoma patients and changes in EGFRvIII expression in recurrent versus primary glioblastomas remain controversial, but such data are highly relevant for EGFRvIII-targeted therapies.

Experimental Design: EGFR-amplified glioblastomas from 106 patients were assessed for EGFRvIII positivity. Changes in EGFR amplification and EGFRvIII status from primary to recurrent glioblastomas were evaluated in 40 patients with EGFR-amplified tumors and 33 patients with EGFR–nonamplified tumors. EGFR single-nucleotide variants (SNV) were assessed in 27 patients. Data were correlated with outcome and validated in 150 glioblastoma patients from The Cancer Genome Atlas (TCGA) consortium.

Results: Sixty of 106 EGFR-amplified glioblastomas were EGFRvIII-positive (56.6%). EGFRvIII positivity was not associated with different progression-free or overall survival. EGFRvIII status was unchanged at recurrence in 35 of 40 patients with EGFR-amplified primary tumors (87.5%). Four patients lost and one patient gained EGFRvIII positivity at recurrence. None of 33 EGFR-nonamplified glioblastomas acquired EGFR amplification or EGFRvIII at recurrence. EGFR SNVs were frequent in EGFR-amplified tumors, but were not linked to survival.

Conclusions: EGFRvIII and EGFR SNVs are not prognostic in EGFR-amplified glioblastoma patients. EGFR amplification is retained in recurrent glioblastomas. Most EGFRvIII-positive glioblastomas maintain EGFRvIII positivity at recurrence. However, EGFRvIII expression may change in a subset of patients at recurrence, thus repeated biopsy with reassessment of EGFRvIII status is recommended for patients with recurrent glioblastoma to receive EGFRvIII-targeting agents. Clin Cancer Res; 23(22); 6846–55. ©2017 AACR.

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Neonatal airway anomaly: vallecular cyst

Description

Neonatal laryngeal cysts are an uncommon but recognised cause of inspiratory stridor and respiratory distress in infants. Although a small cyst may be asymptomatic, due to the anatomical location and the small size of an infant's respiratory tract, larger cysts may cause acute airway obstruction.1 Affected infants typically display symptoms within the first week of life, which include inspiratory stridor, respiratory distress or feeding difficulties. Congenital vallecular cysts, also known as mucus retention cysts, may arise from the mucosal surface of the true vocal fold, epiglottis or vallecula.2 Vallecular cysts are particularly dangerous as they may cause posterior displacement of the supraglottis, causing collapse of the airway during inspiration leading to respiratory compromise. Additionally, several reports have documented an association between laryngomalacia and vallecular cysts.3 Diagnosis is typically obtained through flexible nasopharyngolaryngoscopy, and endoscopic marsupialisation is the recommended surgical approach in...

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Pyoderma gangrenosum: combination therapy with excellent results in a patient with underlying hepatitis C

Description

Pyoderma gangrenosum (PG) is defined as a neutrophilic dermatosis, not related to any infections or gangrenous causes.1 It usually appears with an underlying systemic disease. We report a 55-year-old man who presented with painful bilateral skin lesions on legs with no associated constitutional symptoms. Patient had a history of untreated hepatitis C infection with negative cryoglobulinaemia screen. Surgical debridement was performed in the emergency department, at that time without a diagnosis and a biopsy plus culture of the skin was done which disclosed no evidence of microorganism, vasculitis findings and perivascular lymphoplasmacytic infiltrate. A repeat biopsy showed neutrophilic infiltrate. Clinical and histopathological diagnosis of PG was made and patient was started on intravenous steroids with some improvement,2 followed with clinical deterioration that prompted intravenous immunoglobulin therapy with excellent results.3 Patient continues to get weekly wound care and was being seen by infectious disease and rheumatology...

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Septic arthritis of knee joint due to Parvimonas micra

Parvimonas species are anaerobic, Gram-positive cocci that are a constituent of normal oral and gastrointestinal flora. We present a case of right knee joint septic arthritis due to Parvimonas micra in an immunocompromised patient. A 61-year-old male renal and pancreatic transplant recipient on immunosuppressive therapy was admitted to our hospital due to intense pain, joint swelling and inability to move his right knee over the past 9 months. After synovial fluid was drawn, cultures were positive for P. micra, an anaerobic pathogen that is part of the flora of the oral cavity. We report a rare causative pathogen for septic arthritis in an immunocompromised patient.

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Carcinoma en cuirasse in a young female

Description

Cancer or carcinoma en cuirasse is an infrequent clinical presentation of metastatic cutaneous carcinoma.1 In this condition, the skin of the chest wall is studded with carcinomatous indurated plaques. It is often seen after a patient of breast cancer undergoes mastectomy operation and develops local recurrence. The patient usually presents after a few months or years after the surgery2 (figure 1). This patient, a 22-year-old woman, had already undergone right modified radical mastectomy for infiltrating ductal carcinoma (IDC) Not Otherwise Specified (NOS) grade II with triple-negative receptor status, 1 year ago. She had now presented with a lump in the left breast and multiple cutaneous lesions on the chest wall on the right side and on the left breast since the past 5 months (figure 2). Cutaneous metastasis or carcinoma en cuirasse has occurred on the right side after mastectomy, whereas they are...

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Isolated azygos vein thrombosis: a rare phenomenon

Description

A 57-year-old man with diabetes mellitus and liver cirrhosis was electively admitted for endoscopic retrograde cholangiopancreatography (ERCP). He had history of obstructive jaundice secondary to choledocholithiasis in the preceding month, during which a biliary stent was inserted. During ERCP, he developed acute chest pain and shortness of breath. ECG, cardiac enzymes, coagulation profile and chest radiograph were normal. He was then suspected to have acute pulmonary embolism.

CT pulmonary angiography (CTPA) showed a central tubular filling defect within the azygos vein (AV). This filling defect has a small attachment at the roof of the vessel, forming acute angle with the vessel wall (figures 1 and 2). The rest of the pulmonary vasculature was fully opacified. Lung fields were normal.

Figure 1

Selected axial (A) and coronal (B) sections of CT pulmonary angiography showing tubular filling defect within the azygos vein (arrows).

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Umbilical artery perforation: a potentially life-threatening complication of umbilical artery catheterisation

Umbilical artery catheterisation is frequently performed in preterm or critically ill newborn infants for invasive monitoring of blood pressure and blood sampling for laboratory tests. It is associated with well-known complications like catheter-related infections, thromboembolic events and aneurysmal formation of the aorta. In this report, we present another major complication of umbilical artery catheterisation: umbilical artery perforation. This complication occurred in a prematurely born infant and resulted in severe haemorrhagic shock, subsequent renal failure and severe periventricular leukomalacia. Ultimately, the patient deceased 3 weeks after birth. Review of the literature identified only a few case reports and retrospective postmortem studies describing this complication. Neonatologists and paediatricians performing umbilical artery catheterisation should be aware of this rare, but potentially life-threatening complication.

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Orbitonasal metallic foreign body

Description

A 23-year-old male patient presented with the history of metallic wire injury following a burst of an instrument. The duration of injury was 2 hours. Clinical examination revealed a visual acuity of 20/20 in each eye, right brow showed a superficial skin laceration. Medial side of the right orbit had an impacted curved metallic wire. Extraocular motility in the right eye was within normal limits (figure 1). Anterior segment and posterior segment examination was unremarkable in both the eyes. B-scan ultrasound revealed a high amplitude spike along the side of the right globe but without any penetration; however, it showed curvilinear course along the medial orbital wall (figure 2A).

Figure 1

Front and lateral clinical profile showing a curvilinear metallic foreign body impacted along the superior and nasal aspect of the right upper eyelid. Extraocular motilities were unaffected.

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Norethisterone enanthate-induced cerebral venous sinus thrombosis (CVST)

A 23-year-old East Indian woman with no significant medical history, except a depot-norethisterone enanthate injection taken 3 weeks prior to admission, presented with a gradually worsening headache for the past 5 days. She had no fever, vomiting, neck stiffness, focal weakness or rash, and examination was unremarkable with no focal neurological deficits. Vasculitic, thrombophilia and sepsis screens were normal. A brain CT scan showed a left parietal lobe venous infarct, secondary to a venous dural sinus thrombosis, with MRI and Magnetic Resonance Venogram (MRV) confirming a signal void. She was diagnosed to have multiple cerebral venous sinus thrombosis due to norethisterone enanthate. She made a complete recovery following treatment with mannitol, dexamethasone and anticoagulants. A follow-up brain MRI done at 6 months was normal.

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Reversible brain lesion following growth hormone replacement therapy in an adolescent

A 12.6-year-old girl presented with a 2-month history of headache, recurrent vomiting and 5 kg weight loss. She had been receiving recombinant human growth hormone (rhGH) replacement therapy at a dose of 0.035 mg/kg for the past 10 months, due to short stature. Investigations before initiating rhGH, including brain MRI, had been normal. Physical examination revealed a nystagmus and a mildly elevated arterial blood pressure. Brain MRI revealed a lesion in the posterior aspect of the medulla oblongata, adjacent to the foramen of Magendie. rhGH therapy was discontinued, followed by a gradual resolution of the symptoms. At follow-up 3 months later, she was asymptomatic and physical examination was unremarkable. A subsequent repeat brain MRI showed complete resolution of the lesion, supporting the diagnosis of a variant of reversible posterior leucoencephalopathy syndrome. This is the first case report of a reversible brain lesion linked to rhGH replacement therapy.

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Peripheral arterial disease as seen on X-ray and arteriography

Description

An 80-year-old woman was admitted for a 2months history of non-healing ulcers on both feet. She is known to have type 2 diabetes, dyslipidaemia and hypertension for 30 years. She has a history of a femoral fracture on the left incurred after a fall 2 years ago for which she underwent open reduction and internal fixation.

On physical examination, there were ulcers on the lateral aspect of both feet and on the right malleolar area. Dorsalis pedis pulses were absent bilaterally.

An X-ray of both femurs was requested to check if the fracture healed properly and to rule out a new fracture since the patient remained bedridden despite surgery to correct the fracture. The X-ray revealed a metal implant on the left femur and with no new fracture. Incidentally, the femoral arteries were noted to be calcified along their entire length indicative of atherosclerosis (figure 1).

...

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Unusual Salmonella typhi periprosthetic joint infection involving bilateral knees: management options and literature review

A 70-year-old Indian woman, who had undergone primary bilateral total knee arthroplasty (TKA) for rheumatoid arthritis 10 months prior, presented with 10 days history of pain, swelling and erythema over both knees with pus discharging from the right knee. She had type 2 diabetes mellitus and was on long-term steroid, leflunomide and antitumour necrosis factor therapy for rheumatoid arthritis. Her clinical and laboratory features were suggestive of a haematogenous periprosthetic joint infection (PJI). The final diagnosis of bilateral Salmonella typhi PJI was made based on culture reports. Considering her underlying immunosuppression, a bilateral two-stage revision TKA was done with complete remission of symptoms and good functional recovery at last follow-up after 18 months. S. typhi infection of prosthetic joint has not been reported in the literature. Patients presenting with gastrointestinal complaints and PJI should alert the clinician to the possibility of infection with such atypical organisms endemic to the region.

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Antenatal Bartter syndrome presenting with vomiting and constipation mimicking subacute intestinal obstruction in a 20-day-old neonate

Antenatal Bartter syndrome is a rare condition that can present with different clinical features. These features include early onset maternal polyhydramnios, failure to thrive, prematurity and nephrocalcinosis.

We are presenting this 20-day-old girl who had an antenatal history of polyhydramnios. She developed persistent non-bilious vomiting that was associated with constipation soon after birth. She presented with failure to thrive and features suggestive of intestinal obstruction. On the initial evaluation, she was noted to have hypokalaemic, hyponatraemic metabolic alkalosis. The initial work-up was done to exclude surgical and renal causes of her presentation, and the diagnosis was confirmed by gene analysis to be type III—classic Bartter syndrome. She was closely monitored for her growth and development with the appropriate salt replacement therapy.

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Ectopic ACTH syndrome complicated by multiple opportunistic infections treated with percutaneous ablation of the adrenal glands

Ectopic adrenocorticotropic hormone (ACTH)-related Cushing's syndrome can lead to multiple complications including severe immunosuppression. If the ACTH-secreting tumour cannot be found, definitive treatment is surgical adrenalectomy, typically followed by glucocorticoid replacement. Here, we present a case of fulminant respiratory failure secondary to coinfection with Pneumocystis jirovecii and cytomegalovirus in a patient with ectopic ACTH-dependent Cushing's syndrome with occult primary. Due to significant deconditioning, she was unable to undergo definitive adrenalectomy and instead underwent percutaneous microwave ablation of the adrenal glands.

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Syntaxin1B contributes to regulation of the dopaminergic system through GABA transmission in the CNS

Abstract

In neuronal plasma membrane, two syntaxin isoforms, HPC-1/syntaxin1A (STX1A) and syntaxin1B (STX1B), are predominantly expressed as soluble N-ethylmaleimide-sensitive fusion attachment protein receptors, also known as t-SNAREs. We previously reported that glutamatergic and GABAergic synaptic transmission is impaired in Stx1b null mutant (Stx1b^−/−) mice, but is almost normal in Stx1a null mutant (Stx1a^−/−) mice. These observations suggested that STX1A and STX1B have distinct functions in fast synaptic transmission in the central nervous system (CNS). Interestingly, recent studies indicated that Stx1a^−/− or Stx1a^+/− mice exhibit disruption in the monoaminergic system in the CNS, causing unusual behavior that is similar to neuropsychological alterations observed in psychiatric patients. Here, we studied whether STX1B contributes to the regulation of monoaminergic system and if STX1B is related to neuropsychological properties in human neuropsychological disorders similar to STX1A. We found that monoamine release in vitro was normal in Stx1b^+/− mice unlike Stx1a^−/− or Stx1a^+/− mice, but the basal extracellular dopamine (DA) concentration in the ventral striatum was increased. DA secretion in the ventral striatum is regulated by GABAergic neurons, and Stx1b^+/− mice exhibited reduced GABA release both in vitro and in vivo, disrupting the DAergic system in the CNS of these mice. We also found that Stx1b^+/− mice exhibited reduced pre-pulse inhibition (PPI), which is believed to represent one of the prominent schizotypical behavioral profiles of human psychiatric patients. The reduction in PPI was rescued by DA receptor antagonists. These observations indicated that STX1B contributes to excess activity of the DAergic system through regulation of GABAergic transmission.

This article is protected by copyright. All rights reserved.

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PI-273, a Substrate-Competitive, Specific Small-Molecule Inhibitor of PI4KII{alpha}, Inhibits the Growth of Breast Cancer Cells

While phosphatidylinositol 4-kinase (PI4KIIα) has been identified as a potential target for antitumor therapy, the clinical applications of PI4KIIα are limited by a lack of specific inhibitors. Here we report the first small-molecule inhibitor (SMI) of human PI4KIIα. Docking-based and ligand-based virtual screening strategies were first employed to identify promising hits, followed by two rounds of kinase activity inhibition validation. 2-(3-(4-Chlorobenzoyl)thioureido)-4-ethyl-5-methylthiophene-3-carboxamide (PI-273) exhibited the greatest inhibitory effect on PI4KIIα kinase activity (IC50 = 0.47 μmol/L) and suppressed cell proliferation. Surface plasmon resonance and thermal shift assays indicated that PI-273 interacted directly with PI4KIIα. Kinetic analysis identified PI-273 as a reversible competitive inhibitor with respect to the substrate phosphatidylinositol (PI), which contrasted with most other PI kinase inhibitors that bind the ATP binding site. PI-273 reduced PI4P content, cell viability, and AKT signaling in wild-type MCF-7 cells, but not in PI4KIIα knockout MCF-7 cells, indicating that PI-273 is highly selective for PI4KIIα. Mutant analysis revealed a role of palmitoylation insertion in the selectivity of PI-273 for PI4KIIα. In addition, PI-273 treatment retarded cell proliferation by blocking cells in G2–M, inducing cell apoptosis and suppressing colony-forming ability. Importantly, PI-273 significantly inhibited MCF-7 cell-induced breast tumor growth without toxicity. PI-273 is the first substrate-competitive, subtype-specific inhibitor of PI4KIIα, the use of which will facilitate evaluations of PI4KIIα as a cancer therapeutic target. Cancer Res; 77(22); 6253–66. ©2017 AACR.

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Mitosis-Mediated Intravasation in a Tissue-Engineered Tumor-Microvessel Platform

Intravasation involves the migration of tumor cells across the local endothelium and escape into vessel flow. Although tumor cell invasiveness has been correlated to increased intravasation, the details of transendothelial migration and detachment into circulation are still unclear. Here, we analyzed the intravasation of invasive human breast cancer cells within a tissue-engineered microvessel model of the tumor microenvironment. Using live-cell fluorescence microscopy, we captured 2,330 hours of tumor cell interactions with functional microvessels and provide evidence for a mitosis-mediated mechanism where tumor cells located along the vessel periphery are able to disrupt the vessel endothelium through cell division and detach into circulation. This model provides a framework for understanding the physical and biological parameters of the tumor microenvironment that mediate intravasation of tumor cells across an intact endothelium. Cancer Res; 77(22); 6453–61. ©2017 AACR.

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Correction: Breast Tumor Kinase Phosphorylates p190RhoGAP to Regulate Rho and Ras and Promote Breast Carcinoma Growth, Migration, and Invasion

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Multiscale Modeling of Inflammation-Induced Tumorigenesis Reveals Competing Oncogenic and Oncoprotective Roles for Inflammation

Chronic inflammation is a serious risk factor for cancer; however, the routes from inflammation to cancer are poorly understood. On the basis of the processes implicated by frequently mutated genes associated with inflammation and cancer in three organs (stomach, colon, and liver) extracted from the Gene Expression Omnibus, The Cancer Genome Atlas, and Gene Ontology databases, we present a multiscale model of the long-term evolutionary dynamics leading from inflammation to tumorigenesis. The model incorporates cross-talk among interactions on several scales, including responses to DNA damage, gene mutation, cell-cycle behavior, population dynamics, inflammation, and metabolism-immune balance. Model simulations revealed two stages of inflammation-induced tumorigenesis: a precancerous state and tumorigenesis. The precancerous state was mainly caused by mutations in the cell proliferation pathway; the transition from the precancerous to tumorigenic states was induced by mutations in pathways associated with apoptosis, differentiation, and metabolism-immune balance. We identified opposing effects of inflammation on tumorigenesis. Mild inflammation removed cells with DNA damage through DNA damage-induced cell death, whereas severe inflammation accelerated accumulation of mutations and hence promoted tumorigenesis. These results provide insight into the evolutionary dynamics of inflammation-induced tumorigenesis and highlight the combinatorial effects of inflammation and metabolism-immune balance. This approach establishes methods for quantifying cancer risk, for the discovery of driver pathways in inflammation-induced tumorigenesis, and has direct relevance for early detection and prevention and development of new treatment regimes. Cancer Res; 77(22); 6429–41. ©2017 AACR.

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In Silico Modeling of Immunotherapy and Stroma-Targeting Therapies in Human Colorectal Cancer

Despite the fact that the local immunological microenvironment shapes the prognosis of colorectal cancer, immunotherapy has shown no benefit for the vast majority of colorectal cancer patients. A better understanding of the complex immunological interplay within the microenvironment is required. In this study, we utilized wet lab migration experiments and quantitative histological data of human colorectal cancer tissue samples (n = 20) including tumor cells, lymphocytes, stroma, and necrosis to generate a multiagent spatial model. The resulting data accurately reflected a wide range of situations of successful and failed immune surveillance. Validation of simulated tissue outcomes on an independent set of human colorectal cancer specimens (n = 37) revealed the model recapitulated the spatial layout typically found in human tumors. Stroma slowed down tumor growth in a lymphocyte-deprived environment but promoted immune escape in a lymphocyte-enriched environment. A subgroup of tumors with less stroma and high numbers of immune cells showed high rates of tumor control. These findings were validated using data from colorectal cancer patients (n = 261). Low-density stroma and high lymphocyte levels showed increased overall survival (hazard ratio 0.322, P = 0.0219) as compared with high stroma and high lymphocyte levels. To guide immunotherapy in colorectal cancer, simulation of immunotherapy in preestablished tumors showed that a complex landscape with optimal stroma permeabilization and immune cell activation is able to markedly increase therapy response in silico. These results can help guide the rational design of complex therapeutic interventions, which target the colorectal cancer microenvironment. Cancer Res; 77(22); 6442–52. ©2017 AACR.

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Localized Synchrotron Irradiation of Mouse Skin Induces Persistent Systemic Genotoxic and Immune Responses

The importance of nontargeted (systemic) effects of ionizing radiation is attracting increasing attention. Exploiting synchrotron radiation generated by the Imaging and Medical Beamline at the Australian Synchrotron, we studied radiation-induced nontargeted effects in C57BL/6 mice. Mice were locally irradiated with a synchrotron X-ray broad beam and a multiplanar microbeam radiotherapy beam. To assess the influence of the beam configurations and variations in peak dose and irradiated area in the response of normal tissues outside the irradiated field at 1 and 4 days after irradiation, we monitored oxidatively induced clustered DNA lesions (OCDL), DNA double-strand breaks (DSB), apoptosis, and the local and systemic immune responses. All radiation settings induced pronounced persistent systemic effects in mice, which resulted from even short exposures of a small irradiated area. OCDLs were elevated in a wide variety of unirradiated normal tissues. In out-of-field duodenum, there was a trend for elevated apoptotic cell death under most irradiation conditions; however, DSBs were elevated only after exposure to lower doses. These genotoxic events were accompanied by changes in plasma concentrations of macrophage-derived cytokine, eotaxin, IL10, TIMP1, VEGF, TGFβ1, and TGFβ2, along with changes in tissues in frequencies of macrophages, neutrophils, and T lymphocytes. Overall, our findings have implications for the planning of therapeutic and diagnostic radiation treatments to reduce the risk of radiation-related adverse systemic effects. Cancer Res; 77(22); 6389–99. ©2017 AACR.

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CD155T/TIGIT Signaling Regulates CD8+ T-cell Metabolism and Promotes Tumor Progression in Human Gastric Cancer

The T-cell surface molecule TIGIT is an immune checkpoint molecule that inhibits T-cell responses, but its roles in cancer are little understood. In this study, we evaluated the role TIGIT checkpoint plays in the development and progression of gastric cancer. We show that the percentage of CD8 T cells that are TIGIT+ was increased in gastric cancer patients compared with healthy individuals. These cells showed functional exhaustion with impaired activation, proliferation, cytokine production, and metabolism, all of which were rescued by glucose. In addition, gastric cancer tissue and cell lines expressed CD155, which bound TIGIT receptors and inactivated CD8 T cells. In a T cell–gastric cancer cell coculture system, gastric cancer cells deprived CD8 T cells of glucose and impaired CD8 T-cell effector functions; these effects were neutralized by the additional glucose or by TIGIT blockade. In gastric cancer tumor cells, CD155 silencing increased T-cell metabolism and IFNγ production, whereas CD155 overexpression inhibited T-cell metabolism and IFNγ production; this inhibition was neutralized by TIGIT blockade. Targeting CD155/TIGIT enhanced CD8 T-cell reaction and improved survival in tumor-bearing mice. Combined targeting of TIGIT and PD-1 further enhanced CD8 T-cell activation and improved survival in tumor-bearing mice. Our results suggest that gastric cancer cells inhibit CD8 T-cell metabolism through CD155/TIGIT signaling, which inhibits CD8 T-cell effector functions, resulting in hyporesponsive antitumor immunity. These findings support the candidacy of CD155/TIGIT as a potential therapeutic target in gastric cancer. Cancer Res; 77(22); 6375–88. ©2017 AACR.

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Inflammatory Monocytes Promote Perineural Invasion via CCL2-Mediated Recruitment and Cathepsin B Expression

Perineural invasion (PNI) is an ominous event strongly linked to poor clinical outcome. Cells residing within peripheral nerves collaborate with cancer cells to enable PNI, but the contributing conditions within the tumor microenvironment are not well understood. Here, we show that CCR2-expressing inflammatory monocytes (IM) are preferentially recruited to sites of PNI, where they differentiate into macrophages and potentiate nerve invasion through a cathepsin B–mediated process. A series of adoptive transfer experiments with genetically engineered donors and recipients demonstrated that IM recruitment to nerves was driven by CCL2 released from Schwann cells at the site of PNI, but not CCL7, an alternate ligand for CCR2. Interruption of either CCL2–CCR2 signaling or cathepsin B function significantly impaired PNI in vivo. Correlative studies in human specimens demonstrated that cathepsin B–producing macrophages were enriched in invaded nerves, which was associated with increased local tumor recurrence. These findings deepen our understanding of PNI pathogenesis and illuminate how PNI is driven in part by corruption of a nerve repair program. Further, they support the exploration of inhibiting IM recruitment and function as a targeted therapy for PNI. Cancer Res; 77(22); 6400–14. ©2017 AACR.

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PD-1 Status in CD8+ T Cells Associates with Survival and Anti-PD-1 Therapeutic Outcomes in Head and Neck Cancer

Improved understanding of expression of immune checkpoint receptors (ICR) on tumor-infiltrating lymphocytes (TIL) may facilitate more effective immunotherapy in head and neck cancer (HNC) patients. A higher frequency of PD-1+ TIL has been reported in human papillomavirus (HPV)+ HNC patients, despite the role of PD-1 in T-cell exhaustion. This discordance led us to hypothesize that the extent of PD-1 expression more accurately defines T-cell function and prognostic impact, because PD-1high T cells may be more exhausted than PD-1low T cells and may influence clinical outcome and response to anti-PD-1 immunotherapy. In this study, PD-1 expression was indeed upregulated on HNC patient TIL, and the frequency of these PD-1+ TIL was higher in HPV+ patients (P = 0.006), who nonetheless experienced significantly better clinical outcome. However, PD-1high CD8+ TILs were more frequent in HPV− patients and represented a more dysfunctional subset with compromised IFN-γ secretion. Moreover, HNC patients with higher frequencies of PD-1high CD8+ TIL showed significantly worse disease-free survival and higher hazard ratio for recurrence (P < 0.001), while higher fractions of PD-1low T cells associated with HPV positivity and better outcome. In a murine HPV+ HNC model, anti-PD-1 mAb therapy differentially modulated PD-1high/low populations, and tumor rejection associated with loss of dysfunctional PD-1high CD8+ T cells and a significant increase in PD-1low TIL. Thus, the extent of PD-1 expression on CD8+ TIL provides a potential biomarker for anti-PD-1–based immunotherapy. Cancer Res; 77(22); 6353–64. ©2017 AACR.

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Combined CDK4/6 and PI3K{alpha} Inhibition Is Synergistic and Immunogenic in Triple-Negative Breast Cancer

New treatments for triple-negative breast cancer (TNBC) are urgently needed. Despite there being little evidence of clinical activity as single-agent therapies, we show that dual blockade of PI3Kα and CDK4/6 is synergistically effective against multiple RB1-wild-type TNBC models. Combined PI3Kα and CDK4/6 inhibition significantly increased apoptosis, cell-cycle arrest, and tumor immunogenicity and generated immunogenic cell death in human TNBC cell lines. Combination treatment also significantly improved disease control in human xenograft models compared with either monotherapy. Combined PI3Kα and CDK4/6 inhibition significantly increased tumor-infiltrating T-cell activation and cytotoxicity and decreased the frequency of immunosuppressive myeloid-derived suppressor cells in a syngeneic TNBC mouse model. Notably, combined PI3Kα and CDK4/6 inhibition, along with inhibition of immune checkpoints PD-1 and CTLA-4, induced complete and durable regressions (>1 year) of established TNBC tumors in vivo. Overall, our results illustrate convergent mechanisms of PI3Kα and CDK4/6 blockade on cell-cycle progression, DNA damage response, and immune-modulation and may provide a novel therapeutic approach for TNBC. Cancer Res; 77(22); 6340–52. ©2017 AACR.

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PD-1 Expression in Head and Neck Squamous Cell Carcinomas Derives Primarily from Functionally Anergic CD4+ TILs in the Presence of PD-L1+ TAMs

Oral tongue squamous cell carcinoma (OTSCC) is the most common oral cavity tumor. In this study, we examined the basis for the activity of programmed cell death protein (PD-1)-based immune checkpoint therapy that is being explored widely in head and neck cancers. Using multispectral imaging, we systematically investigated the OTSCC tumor microenvironment (TME) by evaluating the frequency of PD-1 expression in CD8+, CD4+, and FoxP3+ tumor-infiltrating lymphocytes (TIL). We also defined the cellular sources of PD-1 ligand (PD-L1) to evaluate the utility of PD-1:PD-L1 blocking antibody therapy in this patient population. PD-L1 was expressed in 79% of the OTSCC specimens examined within the TME. Expression of PD-L1 was associated with moderate to high levels of CD4+ and CD8+ TILs. We found that CD4+ TILs were present in equal or greater frequencies than CD8+ TILs in 94% of OTSCC and that CD4+FOXP3neg TILs were colocalized with PD-1/PD-L1/CD68 more frequently than CD8+ TILs. Both CD4+PD1+ and CD8+PD1+ TILs were anergic in the setting of PD-L1 expression. Overall, our results highlight the importance of CD4+ TILs as pivotal regulators of PD-L1 levels and in determining the responsiveness of OTSCC to PD1-based immune checkpoint therapy. Cancer Res; 77(22); 6365–74. ©2017 AACR.

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PHGDH as a Key Enzyme for Serine Biosynthesis in HIF2{alpha}-Targeting Therapy for Renal Cell Carcinoma

Continuous activation of hypoxia-inducible factor (HIF) is important for progression of renal cell carcinoma (RCC) and acquired resistance to antiangiogenic multikinase and mTOR inhibitors. Recently, HIF2α antagonists PT2385 and PT2399 were developed and are being evaluated in a phase I clinical trial for advanced or metastatic clear cell RCC (ccRCC). However, resistance to HIF2α antagonists would be expected to develop. In this study, we identified signals activated by HIF2α deficiency as candidate mediators of resistance to the HIF2α antagonists. We established sunitinib-resistant tumor cells in vivo and created HIF2α-deficient variants of these cells using CRISPR/Cas9 technology. Mechanistic investigations revealed that a regulator of the serine biosynthesis pathway, phosphoglycerate dehydrogenase (PHGDH), was upregulated commonly in HIF2α-deficient tumor cells along with the serine biosynthesis pathway itself. Accordingly, treatment with a PHGDH inhibitor reduced the growth of HIF2α-deficient tumor cells in vivo and in vitro by inducing apoptosis. Our findings identify the serine biosynthesis pathway as a source of candidate therapeutic targets to eradicate advanced or metastatic ccRCC resistant to HIF2α antagonists. Cancer Res; 77(22); 6321–9. ©2017 AACR.

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Genome-Wide CRISPR Screen for Essential Cell Growth Mediators in Mutant KRAS Colorectal Cancers

Targeting mutant KRAS signaling pathways continues to attract attention as a therapeutic strategy for KRAS-driven tumors. In this study, we exploited the power of the CRISPR-Cas9 system to identify genes affecting the tumor xenograft growth of human mutant KRAS (KRASMUT) colorectal cancers. Using pooled lentiviral single-guide RNA libraries, we conducted a genome-wide loss-of-function genetic screen in an isogenic pair of human colorectal cancer cell lines harboring mutant or wild-type KRAS. The screen identified novel and established synthetic enhancers or synthetic lethals for KRASMUT colorectal cancer, including targetable metabolic genes. Notably, genetic disruption or pharmacologic inhibition of the metabolic enzymes NAD kinase or ketohexokinase was growth inhibitory in vivo. In addition, the chromatin remodeling protein INO80C was identified as a novel tumor suppressor in KRASMUT colorectal and pancreatic tumor xenografts. Our findings define a novel targetable set of therapeutic targets for KRASMUT tumors. Cancer Res; 77(22); 6330–9. ©2017 AACR.

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ASXL3 Is a Novel Pluripotency Factor in Human Respiratory Epithelial Cells and a Potential Therapeutic Target in Small Cell Lung Cancer

In this study, we generated induced pluripotent stem cells (iPSC) from normal human small airway epithelial cells (SAEC) to investigate epigenetic mechanisms of stemness and pluripotency in lung cancers. We documented key hallmarks of reprogramming in lung iPSCs (Lu-iPSC) that coincided with modulation of more than 15,000 genes relative to parental SAECs. Of particular novelty, we identified the PRC2-associated protein, ASXL3, which was markedly upregulated in Lu-iPSCs and small cell lung cancer (SCLC) lines and clinical specimens. ASXL3 overexpression correlated with increased genomic copy number in SCLC lines. ASXL3 silencing inhibited proliferation, clonogenicity, and teratoma formation by Lu-iPSCs, and diminished clonogenicity and malignant growth of SCLC cells in vivo. Collectively, our studies validate the utility of the Lu-iPSC model for elucidating epigenetic mechanisms contributing to pulmonary carcinogenesis and highlight ASXL3 as a novel candidate target for SCLC therapy. Cancer Res; 77(22); 6267–81. ©2017 AACR.

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Novel Selective Agents for the Degradation of Androgen Receptor Variants to Treat Castration-Resistant Prostate Cancer

Androgen receptor (AR) mediates the growth of prostate cancer throughout its course of development, including in abnormal splice variants (AR-SV)-driven advanced stage castration-resistant disease. AR stabilization by androgens makes it distinct from other steroid receptors, which are typically ubiquitinated and degraded by proteasomes after ligand binding. Thus, targeting AR in advanced prostate cancer requires the development of agents that can sustainably degrade variant isoforms for effective therapy. Here we report the discovery and characterization of potent selective AR degraders (SARD) that markedly reduce the activity of wild-type and splice variant isoforms of AR at submicromolar doses. Three SARDs (UT-69, UT-155, and (R)-UT-155) bind the amino-terminal transcriptional activation domain AF-1, which has not been targeted for degradation previously, with two of these SARD (UT-69 and UT-155) also binding the carboxy-terminal ligand binding domain. Despite different mechanisms of action, all three SARDs degraded wild-type AR and inhibited AR function, exhibiting greater inhibitory potency than the approved AR antagonists. Collectively, our results introduce a new candidate class of next-generation therapeutics to manage advanced prostate cancer. Cancer Res; 77(22); 6282–98. ©2017 AACR.

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Bone-Induced Expression of Integrin {beta}3 Enables Targeted Nanotherapy of Breast Cancer Metastases

Bone metastases occur in approximately 70% of metastatic breast cancer patients, often leading to skeletal injuries. Current treatments are mainly palliative and underscore the unmet clinical need for improved therapies. In this study, we provide preclinical evidence for an antimetastatic therapy based on targeting integrin β3 (β3), which is selectively induced on breast cancer cells in bone by the local bone microenvironment. In a preclinical model of breast cancer, β3 was strongly expressed on bone metastatic cancer cells, but not primary mammary tumors or visceral metastases. In tumor tissue from breast cancer patients, β3 was significantly elevated on bone metastases relative to primary tumors from the same patient (n = 42). Mechanistic investigations revealed that TGFβ signaling through SMAD2/SMAD3 was necessary for breast cancer induction of β3 within the bone. Using a micelle-based nanoparticle therapy that recognizes integrin αvβ3 (αvβ3-MPs of ∼12.5 nm), we demonstrated specific localization to breast cancer bone metastases in mice. Using this system for targeted delivery of the chemotherapeutic docetaxel, we showed that bone tumor burden could be reduced significantly with less bone destruction and less hepatotoxicity compared with equimolar doses of free docetaxel. Furthermore, mice treated with αvβ3-MP-docetaxel exhibited a significant decrease in bone-residing tumor cell proliferation compared with free docetaxel. Taken together, our results offer preclinical proof of concept for a method to enhance delivery of chemotherapeutics to breast cancer cells within the bone by exploiting their selective expression of integrin αvβ3 at that metastatic site. Cancer Res; 77(22); 6299–312. ©2017 AACR.

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Ceritinib compassionate use for patients with crizotinib-refractory, anaplastic lymphoma kinase-positive advanced non-small-cell lung cancer

Future Oncology, Ahead of Print.


http://ift.tt/2zKm8pG

Atezolizumab in urothelial bladder carcinoma

Future Oncology, Ahead of Print.


http://ift.tt/2iY5sD1

A medium invasiveness multi-level patient’s specific template for pedicle screw placement in the scoliosis surgery

Several methods including free-hand technique, fluoroscopic guidance, image-guided navigation, computer-assisted surgery system, robotic platform and patient's specific templates are being used for pedicle scr...

http://ift.tt/2msFoF1

The Motivated Cognitive Basis of Transphobia: The Roles of Right-Wing Ideologies and Gender Role Beliefs

Abstract

Transgender individuals challenge the traditional assumption that an individual's gender identity is permanently determined by their assigned sex at birth. Perceiving ambiguity surrounding indeterminate gender identities associated with transgender individuals may be especially disturbing for those who generally dislike ambiguity and have preference for order and predictability, that is, for people scoring higher on Need for Closure (NFC). We tested the associations between NFC and transphobia in two studies using community samples from the United Kingdom (n = 231) and Belgium (n = 175), and we examined whether right-wing ideological attitudes and traditional gender role beliefs mediated these relationships. Confirming our expectations, we found that NFC was significantly associated with transphobia through both stronger adherence to social conventions and obedience to authorities (i.e., right-wing authoritarianism) and stronger endorsements of traditional gender roles in the UK and Belgium, as well as through stronger preferences for hierarchy and social inequality (i.e., social dominance orientation) in the UK. Our results suggest that transgender individuals are more likely to be targets of prejudice by those higher in NFC at least partly due to the strong preference for preserving societal traditions and the resistance to a perceived disruption of traditional gender norms. Hence, attempts to reduce transphobia might be especially challenging among those high in NFC. Nevertheless, prejudice-reducing interventions could incorporate techniques that satisfy epistemic needs for predictability, certainty, and simple structure which may have higher chances of success among high NFC individuals.

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Migrant Mothering in Transition: A Qualitative Study of the Maternal Narratives and Practices of Two Generations of Rural-Urban Migrant Mothers in Southern China

Abstract

In the last few decades, mothering in many societies has been affected by mass migration. Although migrant mothering is acknowledged to be dynamic, diverse, and continually reproduced in specific socio-cultural contexts, its transformation over generations has been ignored. Drawing on qualitative data obtained from 61 rural-urban migrant women in southern China, I compare the narratives of two generations of migrant mothers and their daily mothering of their left-behind children, revealing how the intersection of gender, class, and the rural-urban divide in China shapes their gendered ideology and performance of childcare over time. Whereas first-generation migrant mothers perceive good mothering as helping their children to start their own families and establish careers and therefore prioritize economic support for their children's life endeavors as their main expression of maternal love and care, new-generation migrant mothers devote themselves to their children's overall needs, focus more on the quality of the mother-child relationship, and adopt various strategies to meet their children's emotional and educational needs. The present study gives a voice to migrant mothers, confronting the biased stereotype of migrant mothers as irresponsible parents and helping them to construct meanings for their maternal experiences during family separation. Documenting the transition in migrant mothering also underscores the importance of recognizing the changing needs of migrant mothers and their left-behind children over time.

http://ift.tt/2zYHpP8

A Resource for the Allele-Specific Analysis of DNA Methylation at Multiple Genomically Imprinted Loci in Mice

Genomically imprinted loci are expressed mono-allelically dependent upon the parent of origin. Their regulation not only illuminates how chromatin regulates gene expression but also how chromatin can be reprogrammed every generation. Because of their distinct parent of origin regulation, analysis of imprinted loci can be difficult. Single nucleotide polymorphisms (SNPs) are required to accurately assess these elements allele-specifically. However, publicly available SNP databases lack robust verification, making analysis of imprinting difficult. In addition, the allele-specific imprinting assays that have been developed employ different mouse strains, making it difficult to systemically analyze these loci. Here, we have generated a resource that will allow the allele-specific analysis of many significant imprinted loci in a single hybrid strain of Mus musculus. This resource includes verification of SNPs present within ten of the most widely used imprinting control regions and allele-specific DNA methylation assays for each gene in a C57BL/6J and CAST/EiJ hybrid strain background.

http://ift.tt/2hE2Bm5

Natural Variation in SER1 and ENA6 Underlie Condition-Specific Growth Defects in Saccharomyces cerevisiae

Despite their ubiquitous use in laboratory strains, naturally occurring loss-of-function mutations in genes encoding core metabolic enzymes are relatively rare in wild isolates of Saccharomyces cerevisiae. Here, we identify a naturally occurring serine auxotrophy in a sake brewing strain from Japan. Through a cross with a honey wine (white tecc) brewing strain from Ethiopia, we map the minimal medium growth defect to SER1, which encodes 3-phosphoserine aminotransferase and is orthologous to the human disease gene, PSAT1. To investigate the impact of this polymorphism under conditions of abundant external nutrients, we examine growth in rich medium alone or with additional stresses, including the drugs caffeine and rapamycin and relatively high concentrations of copper, salt, and ethanol. Consistent with studies that found widespread effects of different auxotrophies on RNA expression patterns in rich media, we find that the SER1 loss of function allele dominates the quantitative trait locus (QTL) landscape under many of these conditions, with a notable exacerbation of the effect in the presence of rapamycin and caffeine. We also identify a major effect QTL associated with growth on salt that maps to the gene encoding the sodium exporter, ENA6. We demonstrate that the salt phenotype is largely driven by variation in the ENA6 promoter, which harbors a deletion that removes binding sites for the Mig1 and Nrg1 transcriptional repressors. Thus, our results identify natural variation associated with both coding and regulatory regions of the genome that underlie strong growth phenotypes.

http://ift.tt/2jq323T

Targeting the MYCN-PARP-DNA Damage Response Pathway inNeuroendocrine Prostate Cancer

Purpose: We investigated MYCN-regulated molecular pathways in castration-resistant prostate cancer (CRPC) classified by morphological criteria as adenocarcinoma or neuroendocrine to extend the molecular phenotype, establish driver pathways, and identify novel approaches to combination therapy for NEPC. Results: Using comparative bioinformatics analyses of CRPC-Adeno and CRPC-Neuro RNA sequence data from public datasets and a panel of 28 PDX models we identified a MYCN-PARP-DNA damage response (DDR) pathway that is enriched in CRPC with neuroendocrine differentiation (NED) and CRPC-Neuro. ChIP-PCR assay revealed that N-MYC transcriptionally activates PARP1, PARP2, BRCA1, RMI2, and TOPBP1 through binding to the promoters of these genes. MYCN or PARP1 gene knockdown significantly reduced the expression of MYCN-PARP-DDR pathway genes and NED markers, and inhibition with MYCNsi and/or PARPsi, BRCA1si, or RMI2si significantly suppressed malignant activities including cell viability, colony formation and cell migration in C4-2b4 and NCI-H660 cells. Targeting this pathway with AURKA inhibitor PHA739358 and PARP inhibitor olaparib generated similar therapeutic effects to gene knockdown in vitro and significantly suppressed tumor growth in both C4-2b4 and MDA PDX144-13C subcutaneous models in vivo. Conclusions: Our results identify a novel MYCN-PARP-DDR pathway that is driven by N-MYC in a subset of CRPC-Adeno and in NEPC. Targeting this pathway using in vitro and in vivo CRPC-Adeno and CRPC-Neuro models demonstrated a novel therapeutic strategy for NEPC. Further investigation of N-MYC-regulated DDR gene targets and the biological and clinical significance of MYCN-PARP-DDR signaling will more fully elucidate the importance of the MYCN-PARP-DDR signaling pathway in the development and maintenance of NEPC.

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A Phase I Clinical Trial of the Poly(ADP-ribose) Polymerase Inhibitor Veliparib and Weekly Topotecan in Patients with Solid Tumors

Purpose: To determine the dose limiting toxicities (DLTs), maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D) of veliparib in combination with weekly topotecan in patients with solid tumors. Correlative studies were included to assess the impact of topotecan and veliparib on poly(ADP-ribose) levels in peripheral blood mononuclear cells, serum pharmacokinetics of both agents, and potential association of germline repair gene mutations with outcome. Experimental Design: Eligible patients had metastatic nonhematological malignancies with measurable disease. Using a 3+3 design, patients were treated with veliparib orally twice daily on days 1-3, 8-10 and 15-17 and topotecan intravenously on days 2, 9 and 16 every 28 days. Tumor responses were assessed by RECIST. Results: Of 58 patients enrolled, 51 were evaluable for the primary endpoint. The MTD and RP2D was veliparib 300 mg twice daily on days 1-3, 8-10 and 15-17 along with topotecan 3 mg/m2 on days 2, 9 and 16 of a 28-day cycle. DLTs were grade 4 neutropenia lasting >5 days. The median number of cycles was 2 (1-26). The objective response rate was 10%, with 1 complete and 4 partial responses. 22 patients (42%) had stable disease ranging from 4 to 26 cycles. Patients with germline BRCA1, BRCA2 or RAD51D mutations remained on study longer than those without homologous recombination repair (HRR) gene mutations (median 4 vs. 2 cycles). Conclusions: Weekly topotecan in combination with veliparib has a manageable safety profile and appears to warrant further investigation.

http://ift.tt/2AKppoO

Novel predictors of breast cancer survival derived from miRNA activity analysis

Purpose: Breast cancer is among the leading causes of cancer death; discovery of novel prognostic markers is needed to improve outcomes. Combining systems biology and epidemiology, we investigated microRNA-associated genes and breast cancer survival in a well-characterized population-based study. Experimental Design: A recently developed algorithm, ActMiR, was used to identify key microRNAs "activities" which were predictive of breast cancer mortality in published databases. We profiled microRNA-associated genes in tumors from our well-characterized population-based cohort of 606 women with first primary breast cancer. Cox proportional hazards models were used to estimate hazard ratios (HR) and 95% confidence intervals (CI), after 15+ years of follow-up with 119 breast cancer-specific deaths. Results: miR-500a activity was identified as a key microRNA for estrogen receptor positive breast cancer mortality using public databases. From a panel of 161 miR-500a associated genes profiled, 73 were significantly associated with breast cancer-specific mortality (FDR<0.05) in our population, among which two clusters were observed to have opposing directions of association. For example, high level of SUSD3 was associated with reduced breast cancer-specific mortality [HR 0.3, 95% CI: 0.2-0.4], while the opposite was observed for TPX2 [HR 2.7, 95% CI: 1.8-3.9]. Most importantly, we identified set of genes for which associations with breast cancer-specific mortality were independent of known prognostic factors including hormone receptor status and PAM-50 derived Risk of Recurrence scores. These results are validated in independent datasets. Conclusions: We identified novel markers that may improve prognostic efficiency while shedding light on molecular mechanisms of breast cancer progression.

http://ift.tt/2yBHtA5

Phase I Study of Chimeric Antigen Receptor Modified T Cells in Patients with EGFR-positive Advanced Biliary Tract Cancers

Purpose: This study is an expanded and parallel clinical trial of epidermal growth factor receptor (EGFR)-specific chimeric antigen receptor-engineered autologous T (CART) cell immunotherapy (NCT01869166) to assess the safety and activity of CART-EGFR cell therapy in EGFR-positive advanced unresectable, relapsed/metastatic biliary tract cancers (BTCs). Patients and Methods: Patients with EGFR-positive (>50%) advanced unresectable, relapsed/metastatic BTCs were enrolled. Well produced CART-EGFR cells were infused in a manner of dose escalation after the conditioning treatment with nab-paclitaxel (100-250 mg/m2) and cyclophosphamide (15-35 mg/kg). Results: A total of 19 patients (14 cholangiocarcinomas and 5 gallbladder carcinomas) received 1- to 3-cycles of CART-EGFR cell infusion (Median CART cell dose, 2.65 x 106kg, range, 0.8 to 4.1 x 106/kg) within 6 months. The CART-EGFR cell infusion was tolerated, but 3 patients suffered grade ≥ 3 acute fever/chill. Grade 1/2 target-mediated toxicities including mucosal/cutaneous toxicities and acute pulmonary edema and grade ≥ 3 lymphopenia and thrombocytopenia related to the conditioning treatment were observed. Of 17 evaluable patients, 1 achieved complete response and 10 achieved stable disease. The median progression-free survival was 4 months (range, 2.5 months to 22 months) from the first cycle of treatment. Analysis of data indicated the enrichment of central memory T cells (Tcm) in the infused CART-EGFR cells improved the clinical outcome. Conclusion: The CART-EGFR cell immunotherapy was a safe and active strategy for EGFR-positive advanced BTCs. The enrichment of Tcm in the infused CART-EGFR cells could predict clinical response.

http://ift.tt/2AJGJKT

Immunotherapy of Hepatocellular Carcinoma: Facts and Hopes

Treatment of patients with hepatocellular carcinoma in the advanced stage remains a great challenge, with very few drugs approved. After decades of failures of immune therapies, immune checkpoint inhibitors have emerged as potentially effective treatments for patients with hepatocellular carcinoma in the advanced stage. Immune checkpoints, including human cancer, cytotoxic T-lymphocyte protein 4 (CTLA-4) and programmed cell death protein 1 (PD-1), are surface proteins expressed in a variety of immune cells, and mostly provide immunosuppressive signals. Monoclonal antibodies able to block these molecules have shown antitumor activity against a wide spectrum of human cancers. Clinical experience with checkpoint inhibitors in hepatocellular carcinoma includes early trials with the anti-CTLA-4 agent tremelimumab, and a large phase 2 trial with the anti-PD-1 agent nivolumab. The latter has shown strong activity particularly as second-line therapy, both in terms of tumor response and patient survival. At least three topics should be the focus of future research. The search for activity in patients at less advanced stages, including the adjuvant treatment of patients with resectable or ablatable tumors. The enhanced efficacy of combination therapies, including particularly the combination with those targeted and locoregional therapies that may have a synergistic effect or act upon mechanisms of primary or acquired resistance to checkpoint inhibitors. And the identification of clinical features and serum or tissue biomarkers that would allow a better patient selection for individual treatments. Hopefully, ongoing trials will help design better treatments in the next future.

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Gastrointestinal Cancers: Timing Is Everything

In this edition of Oncology Scan, we have some transitions to announce. First, Stanley Liauw will be rotating off the editorial board. Dr Liauw has been an Associate Editor since 2013, during which time he has been an integral member of the editorial team, providing thoughtful reviews and carefully rendered, well-balanced decisions. We thank him for his service these past 4 years. Additionally, I will be stepping down as Senior Editor of the gastrointestinal section, and we are very pleased to announce that Salma Jabbour will assume the role as the new Senior Editor.

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The Second Stain: A Viral Whodunnit

This patient (1) would be presented in our multidisciplinary head and neck tumor board. Clinical/radiographic evidence of extranodal extension with infiltration of the sternocleidomastoid would influence our recommendation for primary radiation therapy.

http://ift.tt/2zFq5OS

Chemoradiation-Induced Alteration of Programmed Death-Ligand 1 and CD8+ Tumor-Infiltrating Lymphocytes Identified Patients With Poor Prognosis in Rectal Cancer: A Matched Comparison Analysis

To evaluate chemoradiotherapy (CRT)-induced changes in the expression levels of programmed death-ligand 1 (PD-L1) and CD8+ tumor-infiltrating lymphocytes (TILs) and prognostic associations in rectal cancer.

http://ift.tt/2zZGM8n