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Background: The CREST trial demonstrated that for patients at standard risk of surgical complications, there was no significant difference in the primary composite outcome of periprocedural death, MI, or stroke, or late ipsilateral stroke between carotid artery stenting (CAS) and carotid endarterectomy (CEA), although CAS had a higher rate of stroke, and CEA had a higher rate of MI. The economic implications of these two strategies are unknown. Methods: We performed a prospective health economic study alongside the CREST trial. Costs were assessed from the perspective of the US health care system in 2008 dollars using a combination of resource-based and event-based methods. Costs for carotid revascularization procedures were based on measured resource use and unit costs derived from a sample of study hospitals. Non-procedural costs for these hospitalizations were estimated using hospital billing data (charges) and cost-center-specific cost-to-charge-ratios. Costs for follow-up events were estimated using national average DRG reimbursements. The primary analysis was based on a modified intention-to-treat population for which the assigned procedure was attempted (n=1212 CAS; 1193 CEA). Results: CAS was associated with higher total procedural costs (Δ=$675, see Table), driven mainly by higher costs for disposable supplies. Length of stay was shorter for CAS, with associated reductions in non-procedural hospital costs (Δ = -$436). Total cost for the index hospitalization remained slightly higher for CAS (Δ=$239) with similar differences at 1 year. Conclusions: For patients at standard risk of surgical complications, total 1-year costs are slightly higher for CAS vs. CEA, driven largely by higher initial procedural costs. Cost-utility analysis will be performed to determine whether differences in quality of life observed in CREST render CAS an economically attractive strategy. CAS (n= 1212) CEA (n=1193) Δ CAS - CEA (95% CI via bootstrap) Index Procedure Costs, $ Excluding MD Fees 6782 ± 1412 5743 ± 1370 1039 (926, 1148) MD Fees 1114 ± 240 1478 ± 108 -364 (-379, -349) Total 7896 ± 1551 7221 ± 1450 675 (555, 800) Index Hospital Stay 7159 ± 5108 7595 ± 7482 -436 (-951, -59) TOTAL Index Hosp Costs 15055 ± 5539 14816 ± 7709 239 (-302, 778) Index Hosp LOS (days) ICU LOS 0.7 ± 1.1 0.8 ± 1.4 -0.1 (-0.24, -0.04) Non-ICU LOS 1.9 ± 3.2 2.2 ± 4.1 -0.2 (-0.55, 0.06) TOTAL LOS 2.6 ± 3.3 3.0 ± 4.5 -0.4 (-0.7, -0.06) 1-year Rehosp Costs: Repeat Revasc - CAS 295 ± 2097 273 ± 2126 -21 (-142, 201) Repeat Revasc - CEA 287 ± 2088 477 ± 2510 -190 (-371, -11) Ipsilateral Stroke 556 ± 3273 384 ± 2580 172 (-75, 402) TIA 48 ± 520 33 ± 487 16 (-25, 56) Death 63 ± 846 45 ± 779 18 (-49, 85) TOTAL Rehosp costs 1321 ± 4827 1293± 4502 28 (-334, 396) TOTAL 1-year costs 16375 ± 7730 16108 ± 9030 267 (-366, 961)

Melanoma is the sixth most common cancer in the US. Immunotherapies, such as anti‐CTLA‐4 and anti‐PD‐1, have transformed the treatment of advanced melanoma; however, a subset of patients fail to respond to immunotherapy. Thus, improved predictors of the anti‐melanoma immune response and response to immunotherapy are needed. We have previously shown that 1) gamma‐interferon‐inducible lysosomal thiol reductase (GILT) is critical for MHC class II‐restricted presentation of a melanoma antigen, 2) there is variable GILT protein expression in melanoma cells from primary and cutaneous metastatic melanoma specimens, and 3) high GILT mRNA expression in melanoma specimens is associated with improved overall survival. Here, we created a database of n = 43 metastatic melanoma patients (stage III or IV) annotated with clinical outcome. Formalin‐fixed, paraffin‐embedded sections of pre‐treatment metastatic melanoma specimens were tested for GILT protein expression using immunohistochemistry and scored by a dermatopathologist and dermatologist. GILT expression in melanoma cells was detected in 65% of lymph node and distant organ metastases, consistent with our prior finding of GILT expression in 58% of cutaneous metastases. The frequency of GILT‐staining melanoma cells was determined by the ratio of the area of tumor cells with positive GILT staining to the total area of tumor cells. The frequency of GILT‐staining melanoma cells among specimens with positive GILT staining was 15.2% ± 24.7% (mean ± SD). The median frequency of GILT staining was used to divide the patients into no/low and high GILT expression groups. The median overall survival was 52.8 months in the no/low GILT expression group, while the median overall survival was not reached in the high GILT expression group (log‐rank, p = 0.67). For the n = 34 patients who were treated with immunotherapy targeting CTLA‐4 and/or PD‐1, the median overall survival was 38.2 months in the no/low GILT expression group, while the median overall survival was not reached in the high GILT expression group (log‐rank, p = 0.83). These data suggest that high GILT expression in melanoma cells may be associated with improved overall survival, although a statistically significant association of GILT with survival was not observed in the whole group or the subset of patients treated with immunotherapy. A limitation of the current analysis is that 65% of patients have incomplete clinical outcome data. This study demonstrates variable GILT protein expression in tumor cells in metastatic melanoma specimens. Updated clinical outcome data will allow us to better evaluate the association of GILT protein expression in melanoma cells with overall survival and response to immunotherapy in metastatic melanoma patients.

Abstract Cold agglutinin disease (CAD) is an autoimmune hemolytic anemia characterized by the presence of autoantibodies (cold agglutinins) that bind to red blood cells (RBC) at low temperatures. Cold agglutinin binding to RBCs results in antibody-mediated classical complement activation with deposition of complement C4 and C3 fragments onto the RBC surface. Complement-opsonized RBCs are removed from the circulation by macrophage-driven phagocytosis in the liver or spleen and via intravascular hemolysis following assembly of the terminal lytic complex (C5b - C9), together resulting in clinical anemia. C1q is the initiating molecule for cold agglutinin-mediated complement activation on the surface of human RBCs. We hypothesized that directly blocking C1q recruitment onto cold agglutinin-sensitized RBCs will prevent complement activation and opsonization and reduce hemolysis. To this end, we have developed a humanized monoclonal antibody (ANX005) that binds with high-affinity (~10 pM) to C1q and blocks classical complement activation & hemolysis in an in vitro sheep RBC assay. We evaluated the impact of ANX005 on hemolysis and complement deposition on human RBCs that were pre-sensitized with sera from CAD subjects. ANX005 showed a dose-dependent reduction in hemolysis using both individual and pooled CAD sera as the source of cold agglutinin. We further demonstrated that C1q blockade led to a robust reduction in C4 and C3 fragment deposition onto human RBCs. These results demonstrate that C1q inhibition is an effective way to impede C4 and C3 activation and downstream assembly of the lytic complex in sera from CAD patients, and support the clinical development of ANX005 in CAD and other antibody-mediated diseases. Disclosures Gertz: Prothena Therapeutics: Research Funding; Sandoz Inc: Honoraria; NCI Frederick: Honoraria; Celgene: Honoraria; Med Learning Group: Honoraria, Speakers Bureau; Research to Practice: Honoraria, Speakers Bureau; Alnylam Pharmaceuticals: Research Funding; Novartis: Research Funding; Ionis: Research Funding; Annexon Biosciences: Research Funding; GSK: Honoraria. Qiu:Annexon Biosciences: Employment, Equity Ownership. Kendall:Annexon Biosciences: Employment, Equity Ownership. Saltarelli:Mallinckrodt: Equity Ownership; Abbvie: Equity Ownership; Annexon Biosciences: Employment, Equity Ownership, Patents & Royalties. Yednock:Annexon, Inc: Employment, Equity Ownership. Sankaranarayanan:Annexon Biosciences: Employment, Equity Ownership.

Abstract The MHC class I Ag presentation pathway in melanoma cells has a well-established role in immune-mediated destruction of tumors. However, the clinical significance of the MHC class II Ag presentation pathway in melanoma cells is less clear. In Ag-presenting cells, IFN-γ–inducible lysosomal thiol reductase (GILT) is critical for MHC class II–restricted presentation of multiple melanoma Ags. Although not expressed in benign melanocytes of nevi, GILT and MHC class II expression is induced in malignant melanocytes in a portion of melanoma specimens. Analysis of The Cancer Genome Atlas cutaneous melanoma data set showed that high GILT mRNA expression was associated with improved overall survival. Expression of IFN-γ, TNF-α, and IL-1β was positively associated with GILT expression in melanoma specimens. These cytokines were capable of inducing GILT expression in human melanoma cells in vitro. GILT protein expression in melanocytes was induced in halo nevi, which are nevi undergoing immune-mediated regression, and is consistent with the association of GILT expression with improved survival in melanoma. To explore potential mechanisms of GILT’s association with patient outcome, we investigated pathways related to GILT function and expression. In contrast to healthy skin specimens, in which the MHC class II pathway was nearly uniformly expressed and intact, there was substantial variation in the MHC class II pathway in the The Cancer Genome Atlas melanoma specimens. Both an active and intact MHC class II pathway were associated with improved overall survival in melanoma. These studies support a role for GILT and the MHC class II Ag presentation pathway in melanoma outcome.

170 Background: Pembro had antitumor activity as monotherapy in pretreated advanced mCRPC (KEYNOTE-028; KEYNOTE-199). KEYNOTE-365 (NCT02861573) is a phase 1b/2 umbrella study testing combinations in mCRPC; we report early results from cohort B combining pembro + docetaxel in mCRPC. Methods: Pts who failed or became intolerant to ≥4 weeks of abi or enza in the pre-chemotherapy mCRPC state received pembro 200 mg IV with docetaxel 75 mg/m2 IV Q3W plus prednisone 5 mg orally twice daily. Pts also progressed within 6 months prior to screening as determined by either PSA progression or radiologic progression in bone or soft tissue. Response was evaluated by PSA levels Q3W and imaging Q9W for first year and Q12W thereafter. Primary end points: safety and PSA response rate (confirmed PSA decline ≥50%). Key secondary end points: investigator-determined ORR (RECIST v1.1), disease control rate (DCR: CR+PR+SD ≥6 mo), time to PSA progression, rPFS, and OS. Results: 72 pts (median age 68 years; PD-L1+ 29%; visceral disease 36%; measurable disease 50%) initiated pembro + docetaxel. Median (95% CI) follow-up was 10 (8-12) mo. Efficacy is outlined in table below. Treatment-related AEs occurred in 69 (96%) pts; most frequent (≥30%) were alopecia (43%), fatigue (40%), and diarrhea (39%). Grade 3-5 treatment-related AEs occurred in 27 (38%) pts, including 2 deaths due to treatment-related AEs (pneumonitis). Conclusions: Combination of pembro + docetaxel/prednisone has activity in pts with mCRPC previously failing anti-hormonal therapy. Observed safety profile for the combination was consistent with known safety profile of each component. Clinical trial information: NCT02861573. [Table: see text]

BACKGROUNDThe Milan System for Reporting Salivary Gland Cytopathology is a tiered classification scheme that includes 6 diagnostic categories. Neoplasm, which is 1 of the 6 proposed categories, consists of benign neoplasm and neoplasm of uncertain malignant potential (NUMP). NUMP is reserved for a salivary gland neoplasm without clear distinction between benign and malignant. The objective of this study was to assess the risk of malignancy (ROM) of NUMP.METHODSA retrospective analysis was conducted on 656 salivary gland fine‐needle aspiration specimens from 2010 to 2016. Cases that qualified as NUMP and had follow‐up surgical resections were reviewed and reclassified into basaloid neoplasm (BN) and salivary gland neoplasm with predominant oncocytic cell (SGNOC) groups. The ROM for each group was calculated. Fifty‐four salivary gland fine‐needle aspirations of NUMP that had surgical follow‐up were identified, which included 29 BNs and 25 SGNOCs.RESULTSHistologic follow‐up for the BN group identified 14 cellular pleomorphic adenomas, 5 basal cell adenomas, 2 benign cystadenomas, 3 adenoid cystic carcinomas, 3 epithelial and myoepithelial carcinomas, 1 basal cell adenocarcinoma, and 1 myoepithelial carcinoma. Histologic follow‐up for the SGNOC group revealed 7 nodular oncocytoses, 6 Warthin tumors, 5 oncocytomas, 1 sebaceous adenoma, 1 mucinous cystadenoma, 2 acinic cell carcinomas, 2 mucoepidermoid carcinomas, and 1 mammary analog secretory carcinoma. The ROM was calculated at 24.1% for the NUMP category overall (27.6% for BNs and 20.0% for SGNOCs).CONCLUSIONSThe ROM of the SGNOC group is similar to that of the BN group but lower than the ROM (35%) proposed by the Milan system. Cancer Cytopathol 2018. © 2018 American Cancer Society.

DNA damage is an important contributor to endothelial dysfunction and age-related vascular disease. Recently, we demonstrated in a DNA repair-deficient, prematurely aging mouse model (Ercc1Δ/− mice) that dietary restriction (DR) strongly increases life- and health span, including ameliorating endothelial dysfunction, by preserving genomic integrity. In this mouse mutant displaying prominent accelerated, age-dependent endothelial dysfunction we investigated the signaling pathways involved in improved endothelium-mediated vasodilation by DR, and explore the potential role of the renin-angiotensin system (RAS). Ercc1Δ/− mice showed increased blood pressure and decreased aortic relaxations to acetylcholine (ACh) in organ bath experiments. Nitric oxide (NO) signaling and phospho-Ser1177-eNOS were compromised in Ercc1Δ/−. DR improved relaxations by increasing prostaglandin-mediated responses. Increase of cyclo-oxygenase 2 and decrease of phosphodiesterase 4B were identified as potential mechanisms. DR also prevented loss of NO signaling in vascular smooth muscle cells and normalized angiotensin II (Ang II) vasoconstrictions, which were increased in Ercc1Δ/− mice. Ercc1Δ/− mutants showed a loss of Ang II type 2 receptor-mediated counter-regulation of Ang II type 1 receptor-induced vasoconstrictions. Chronic losartan treatment effectively decreased blood pressure, but did not improve endothelium-dependent relaxations. This result might relate to the aging-associated loss of treatment efficacy of RAS blockade with respect to endothelial function improvement. In summary, DR effectively prevents endothelium-dependent vasodilator dysfunction by augmenting prostaglandin-mediated responses, whereas chronic Ang II type 1 receptor blockade is ineffective.

Abstract Emerging evidence suggests that the primary tumor influences the development of supportive metastatic microenvironments, referred to as premetastatic niches, in certain distant organs before arrival of metastatic cells. However, the mechanisms underlying the contributions of the primary tumor to premetastatic niche formation are not fully understood. Here, we demonstrate that colorectal carcinoma cells secrete VEGFA, which stimulates tumor-associated macrophages to produce CXCL1 in the primary tumor. Elevation of CXCL1 in premetastatic liver tissue recruited CXCR2-positive myeloid-derived suppressor cells (MDSC) to form a premetastatic niche that ultimately promoted liver metastases. Importantly, premetastatic liver-infiltrating MDSCs induced tumor cell survival without involvement of innate or adaptive immune responses. Our study provides the first evidence that primary malignant cell-secreted VEGFA stimulates tumor-associated macrophages to produce CXCL1, which recruits CXCR2-positive MDSCs to form a premetastatic niche to promote liver metastases. Our findings not only shed light on how the tumor microenvironment contributes to premetastatic niche formation at distant sites, but they also provide comprehensive insights into how MDSCs are recruited to other organs where they contribute to metastatic spread of disease. Moreover, our work also provides a rationale for development of CXCR2 antagonists to inhibit or prevent metastatic spread of disease. Cancer Res; 77(13); 3655–65. ©2017 AACR.

OBJECTIVE Defects in the apoptotic machinery and augmented survival signals contribute to drug resistance in glioblastoma (GBM). Moreover, another complexity related to GBM treatment is the concept that GBM development and recurrence may arise from the expression of GBM stem cells (GSCs). Therefore, the use of a multifaceted approach or multitargeted agents that affect specific tumor cell characteristics will likely be necessary to successfully eradicate GBM. The objective of this study was to investigate the usefulness of sulforaphane (SFN)—a constituent of cruciferous vegetables with a multitargeted effect—as a therapeutic agent for GBM. METHODS The inhibitory effects of SFN on established cell lines, early primary cultures, CD133-positive GSCs, GSC-derived spheroids, and GBM xenografts were evaluated using various methods, including GSC isolation and the sphere-forming assay, analysis of reactive oxygen species (ROS) and apoptosis, cell growth inhibition assay, comet assays for assessing SFN-triggered DNA damage, confocal microscopy, Western blot analysis, and the determination of in vivo efficacy as assessed in human GBM xenograft models. RESULTS SFN triggered the significant inhibition of cell survival and induced apoptotic cell death, which was associated with caspase 3 and caspase 7 activation. Moreover, SFN triggered the formation of mitochondrial ROS, and SFN-triggered cell death was ROS dependent. Comet assays revealed that SFN increased single- and double-strand DNA breaks in GBM. Compared with the vehicle control cells, a significantly higher amount of γ-H2AX foci correlated with an increase in DNA double-strand breaks in the SFN-treated samples. Furthermore, SFN robustly inhibited the growth of GBM cell–induced cell death in established cell cultures and early-passage primary cultures and, most importantly, was effective in eliminating GSCs, which play a major role in drug resistance and disease recurrence. In vivo studies revealed that SFN administration at 100 mg/kg for 5-day cycles repeated for 3 weeks significantly decreased the growth of ectopic xenografts that were established from the early passage of primary cultures of GBM10. CONCLUSIONS These results suggest that SFN is a potent anti-GBM agent that targets several apoptosis and cell survival pathways and further preclinical and clinical studies may prove that SFN alone or in combination with other therapies may be potentially useful for GBM therapy.

OBJECTIVE Improvement in treatment outcome for patients with glioblastoma multiforme (GBM) requires a multifaceted approach due to dysregulation of numerous signaling pathways. The murine double minute 2 (MDM2) protein may fulfill this requirement because it is involved in the regulation of growth, survival, and invasion. The objective of this study was to investigate the impact of modulating MDM2 function in combination with front-line temozolomide (TMZ) therapy in GBM. METHODS The combination of TMZ with the MDM2 protein–protein interaction inhibitor nutlin3a was evaluated for effects on cell growth, p53 pathway activation, expression of DNA repair proteins, and invasive properties. In vivo efficacy was assessed in xenograft models of human GBM. RESULTS In combination, TMZ/nutlin3a was additive to synergistic in decreasing growth of wild-type p53 GBM cells. Pharmacodynamic studies demonstrated that inhibition of cell growth following exposure to TMZ/nutlin3a correlated with: 1) activation of the p53 pathway, 2) downregulation of DNA repair proteins, 3) persistence of DNA damage, and 4) decreased invasion. Pharmacokinetic studies indicated that nutlin3a was detected in human intracranial tumor xenografts. To assess therapeutic potential, efficacy studies were conducted in a xenograft model of intracranial GBM by using GBM cells derived from a recurrent wild-type p53 GBM that is highly TMZ resistant (GBM10). Three 5-day cycles of TMZ/nutlin3a resulted in a significant increase in the survival of mice with GBM10 intracranial tumors compared with single-agent therapy. CONCLUSIONS Modulation of MDM2/p53-associated signaling pathways is a novel approach for decreasing TMZ resistance in GBM. To the authors' knowledge, this is the first study in a humanized intracranial patient-derived xenograft model to demonstrate the efficacy of combining front-line TMZ therapy and an inhibitor of MDM2 protein–protein interactions.