Inflammatory Gene Expression Profile Research Articles

Effect of Low Versus High Tidal-Volume Total Liquid Ventilation on Pulmonary Inflammation.

Animal experiments suggest that total liquid ventilation (TLV) induces less ventilator-induced lung injury (VILI) than conventional mechanical gas ventilation. However, TLV parameters that optimally minimize VILI in newborns remain unknown. Our objective was to compare lung inflammation between low (L-VT) and high (H-VT) liquid tidal volume and evaluate impacts on the weaning process. Sixteen anesthetized and paralyzed newborn lambs were randomized in an L-VT group (initial tidal volume of 10 mL/kg at 10/min) and an H-VT group (initial tidal volume of 20 mL/kg at 5/min). Five unventilated newborn lambs served as controls. After 4 h of TLV in the supine position, the lambs were weaned in the prone position for another 4 h. The levels of respiratory support needed during the 4 h post-TLV were compared. The anterior and posterior lung regions were assessed by a histological score and real-time quantitative PCR for IL1B, IL6, and TNF plus 12 other exploratory VILI-associated genes. All but one lamb were successfully extubated within 2 h post-TLV (72 ± 26 min vs. 63 ± 25 min, p = 0.5) with similar FiO2 at 4 h post-TLV (27 ± 6% vs. 33 ± 7%, p = 0.3) between the L-VT and H-VT lambs. No significant differences were measured in histological inflammation scores between L-VT and H-VT lambs, although lambs in both groups exhibited slightly higher scores than the control lambs. The L-VT group displayed higher IL1B mRNA expression than the H-VT group in both anterior (2.8 ± 1.5-fold increase vs. 1.3 ± 0.4-fold increase, p = 0.02) and posterior lung regions (3.0 ± 1.0-fold change increase vs. 1.1 ± 0.3-fold increase, p = 0.002), respectively. No significant differences were found in IL6 and TNF expression levels. Gene expression changes overall indicated that L-VT was associated with a qualitatively distinct inflammatory gene expression profiles compared to H-VT, which may indicate different clinical effects. In light of these findings, further mechanistic studies are warranted. In conclusion, we found no advantage of lower tidal volume use, which was in fact associated with a slightly unfavorable pattern of inflammatory gene expression.

Impact of PD-L1 and T-cell inflamed gene expression profile on survival in advanced ovarian cancer

ObjectiveProgrammed death ligand 1 (PD-L1) expression affects tumor evasion of immune surveillance. The prognostic value and relationship of PD-L1 expression to T-cell–inflamed immune signatures in ovarian cancer are unclear. The...

Immune- and tumor-intrinsic gene expression profiles of response or resistance to tislelizumab as monotherapy or in combination with chemotherapy in non-small cell lung cancer (NSCLC).

e15258 Background: Tislelizumab, an anti-PD-1 monoclonal antibody, showed clinical benefit for patients (pts) with NSCLC alone (NCT02407990, CTR20160872) and in combination with chemotherapy (NCT03432598). Gene expression profiles (GEP) associated with response and resistance to tislelizumab in these studies were assessed. Methods: The GEP of baseline tumor samples from 59 nonsquamous (NSQ) and 42 squamous (SQ) NSCLC pts treated with tislelizumab monotherapy (mono) as ≥1L treatment, and 16 NSQ and 21 SQ pts treated with tislelizumab plus chemotherapy (combo) as 1L treatment were assessed using the 1392-gene HTG GEP EdgeSeq panel. NSQ and SQ cohorts were analyzed separately due to distinct GEP features shown by PCA and t-SNE clustering. Results: Tislelizumab mono and combo showed antitumor activity in NSCLC (Table). In 80 biomarker-evaluable samples, inflamed tumor signatures (inflammatory GEP; antigen presentation GEP) were associated with longer overall survival (log-rank test, NSQ mono: P=0.04, 0.003; NSQ combo: P=0.05, 0.02; SQ combo: P=0.06, 0.06). Monotherapy non-responders (NRs) were clustered into 2 subgroups (NR1, NR2) with distinct GEPs. Compared with responders, NR1 had proliferation signatures (elevated cell cycle [CC] and DNA repair) in both NSQ ( P=0.2, 0.02) and SQ ( P=0.03, 0.4) cohorts, trending toward low inflamed tumor signatures. In NR pts receiving combo, CC and DNA repair signatures were not enriched, and high CC and DNA repair scores were observed in some SQ combo responders versus NRs ( P=0.2, 0.02). NR2 had higher M2 macrophage and Treg cell signatures versus responders in both NSQ and SQ mono, despite high inflamed tumor and low proliferation signatures. NR2 also had increased expression of genes related to immune regulation and angiogenesis, including PIK3CD, CCR2, CD244, IRAK3, and MAP4K1 ( P<0.05) in NSQ, and PIK3CD, CCR2, CD40, CD163, MMP12, VEGFC, and TEK ( P<0.05) in SQ. Conclusions: Clinical benefit in pts with NSCLC receiving tislelizumab (mono or combo) was associated with high inflamed tumor signatures, while elevated immune suppressive cell signatures may indicate resistance. High proliferation signatures were associated with resistance to monotherapy, but not to combination therapy. Both immune- and tumor-intrinsic factors may be considered for validation in future clinical trials. [Table: see text]

Association of gene expression with clinical outcomes in patients with renal cell carcinoma treated with pembrolizumab in KEYNOTE-427.

5024 Background: We assessed the association of baseline RNA-sequencing–based gene expression signatures and DNA alterations with response or resistance to pembrolizumab in patients with advanced renal cell carcinoma in cohorts A (clear cell; n = 110) and B (non-clear cell; n = 165) of the phase 2 KEYNOTE-427 study (NCT02853344). Methods: Using RNA-sequencing, we analyzed the association of gene expression signatures (18-gene T-cell–inflamed gene expression profile [GEP]; 10 non–T-cell–inflamed GEP canonical signatures [angiogenesis, gMDSC, glycolysis, hypoxia, mMDSC, MYC, proliferation, RAS, stromal/EMT/TGFβ, WNT]) quantifying tumor microenvironment elements (TME) with objective response rate (ORR) and progression-free survival (PFS). Canonical signatures were derived from 2 databases (TCGA, Moffit) using an algorithm that included genes based on their correlation to reference signatures in the literature. Signature definitions were finalized before linking to the clinical data, and significance was prespecified at 0.10 given the potential for limited power. Canonical signatures were analyzed through regression testing of response for association with consensus signatures after adjusting for T-cell–inflamed GEP and International Metastatic RCC Database Consortium scores in the model. P values were adjusted for multiplicity. Using whole exome sequencing, we also summarized the association of renal cell carcinoma driver gene mutations with ORR. Clinical data cutoff: Jan 30, 2019. Results: Patient characteristics for this analysis were comparable to the overall population. In cohort A, T-cell–inflamed GEP (n = 78) was statistically significantly associated with a better ORR ( P = 0.021; AUROC = 0.65) but not PFS ( P = 0.116). No other TME canonical signatures showed a correlation with ORR or PFS. ORR was estimated for mutations (Table). Conclusions: RNA-sequencing–based, T-cell–inflamed GEP was associated with ORR in patients with clear cell renal cell carcinoma receiving first-line pembrolizumab. Precision was limited by sample size for estimating ORR by specific gene mutation status. Evaluation of tissue-based biomarkers in larger studies are planned. Biomarker analyses from patients in cohort B will also be presented. Clinical trial information: NCT02853344 . [Table: see text]

YAP1 positive small-cell lung cancer subtype is associated with the T-cell inflamed gene expression profile and confers good prognosis and long term survival.

9019 Background: The dominant expression of transcription factors ASCL1, NeuroD1, YAP1 or POU2F3 characteristically defines four small cell lung cancer (SCLC) subtypes (SCLC-A, SCLC-N, SCLC-Y and SCLC-P). The clinical validation and biological relevance of these emerging SCLC subtypes is currently lacking. Methods: Using the Illumina TruSeq RNA Exome Kit, we generated RNA-Seq data from 61 cases of SCLC and pulmonary carcinoid to interrogate gene expression differences in SCLC subtypes as well as in survival outliers (top and bottom decile) matched for clinically relevant prognostic factors and treatment. We also assessed YAP1 protein expression in a blinded fashion by immunohistochemistry in 130 SCLC cases. Results: We successfully classified 68% of SCLC into one of the four SCLC subtypes whereas 81.5% of carcinoids did not fit into any of these categories. GSEA for differentially expressed genes between outlier subgroups showed significant upregulation of interferon gamma and interferon alpha response genes in late survivors. Moreover, a previously validated 18-gene T-cell inflamed gene expression profile was upregulated in late survivors and in the SCLC-Y subtype. Furthermore, the SCLC-Y subtype and late survivors showed higher expression of HLA gene family and reduced expression of cancer testis antigens. The median (95%CI) OS was 14 (4.3, 28.8), 16.7 (0.9, NA), 8.1 (2, 9.7) and 20.1 (0.6, 39.5) months respectively, for SCLC-A, N, P and Y subtypes. YAP-1 protein expression was positive in 17 of 130 (13%) SCLC cases. The majority of cases with positive YAP1 expression by immunohistochemistry, 12 of 17 cases (70.6%), were limited stage SCLC at the time of original diagnosis. Conclusions: SCLC subtypes have clinical implication as predictive and prognostic biomarker. SCLC-Y subtype is enriched for T-cell inflamed phenotype and long term survival, and may predict for clinical benefit of immunotherapy.

Pembrolizumab (pembro) plus olaparib in patients (pts) with docetaxel-pretreated metastatic castration-resistant prostate cancer (mCRPC): KEYNOTE-365 cohort A efficacy, safety, and biomarker results.

5544 Background: Pembro + olaparib has shown antitumor activity and acceptable safety in docetaxel-pretreated pts with mCRPC enrolled in cohort A of the phase I/II KEYNOTE-365 study (NCT02861573). Updated results with new biomarker data are reported. Methods: Pts with docetaxel-pretreated mCRPC who progressed within 6 mo of screening received pembro 200 mg IV Q3W + olaparib 400-mg capsule or 300-mg tablet PO BID. Pts might have received 1 other chemotherapy and ≤2 second-generation androgen-receptor targeted therapies. Primary end points: PSA response rate (decrease ≥50% from baseline, confirmed by a second value ≥3 wks later), ORR per RECIST v1.1, and safety. Key secondary end points: DCR, DOR, rPFS, and OS. Biospecimens (eg, blood, tissue) were collected for biomarker analysis (tissue PD-L1 expression, androgen receptor variant 7 [AR-v7] expression in circulating tumor cells [CTCs], and a T-cell–inflamed gene expression profile [GEP]). ctDNA was analyzed by Guardant Health 360 (GH360) and Omni (GH Omni) assays. FFPE tissue was analyzed by FoundationOne CDx (F1CDx) assay. Results: 84 of 87 enrolled pts were treated; 48/84 (57.1%) had measurable disease. Median (range) time from enrollment to data cutoff was 3.6 mo (0.0-29.2) for all pts and 26.7 mo (21.2-29.2) for 41 pts with ≥27 wks’ follow-up. Confirmed PSA response rate was 9% (95% CI, 3.5-16.8) in 82 pts with a baseline PSA assessment. Median time to PSA progression: 3.8 mo (95% CI, 2.9-4.4). In 24 pts with measurable disease and ≥27 wks’ follow-up, ORR was 8.3% (95% CI, 1.0-27.0; 2 PRs) and DCR ≥6 mo was 20.8% (95% CI, 7.1-42.2). Median (range) DOR was NR (12.0+ to 21.4+ mo); 2 pts had DOR ≥12 mo. In all pts, median rPFS was 4.3 mo (95% CI, 3.4-7.7) and median OS was 14.4 mo (95% CI, 8.1-18.5). Grade ≥3 TRAEs occurred in 29 pts (35%); 2 pts died of TRAEs (1 myocardial infarction, 1 unknown). Overall, 26% had PD-L1+ tumors (combined positive score ≥1). Of 31 pts with CTC data, 12.9% were AR-v7+. No BRCA1/2 mutation was detected by GH360 (n=42). Of 57 pts analyzed by GH Omni, 2 had BRCA2 mutations, 1 had a BRCA1 mutation, 4 had ATM mutations, 1 had a CHEK1 mutation, and 6 had CDK12 mutations. Of 49 pts analyzed by F1CDx, 4 had BRCA mutations; 1 pt had a copy number loss mutation not detected by ctDNA analysis. GEP was not associated with ORR or PSA response. Conclusions: Pembro + olaparib continued to show activity and acceptable safety in pts with docetaxel-pretreated mCRPC. A phase III study of this combination is ongoing (KEYLYNK-010, NCT03834519). Clinical trial information: NCT02861573 .

Chronic Intermittent Hypoxia Blunts Phrenic Motor Plasticity: Role of Inflammation

Acute intermittent hypoxia (AIH) elicits a form of spinal, respiratory motor plasticity known as phrenic long‐term facilitation (pLTF). Preconditioning with a single day of chronic intermittent hypoxia (8 hours, 2 min hypoxic episodes with 2 min intervals) abolishes AIH‐induced pLTF through a mechanism that requires systemic inflammation and spinal p38 MAP kinase activation (Huxtable et al., 2015). pLTF is restored by high‐dose, systemic administration of the non‐steroidal anti‐inflammatory drug, ketoprofen, or spinal p38 MAP kinase inhibition. Similarly, preconditioning with 7 days of CIH (8 hours per day, 2 min episodes, 2 min intervals) abolishes pLTF, but we observed that phospho‐p38 within phrenic motor neurons is unchanged from control rats (Gonzalez‐Rothi and colleagues, unpublished). Thus, mechanisms whereby 7 days of CIH undermines AIH‐induced pLTF are unclear. Since CIH induces time‐dependent spinal inflammatory gene expression profiles over time (Smith et al., 2013), we hypothesize that prolonged CIH (versus a single day) blunts LTF through an inflammation‐dependent mechanism independent of p‐38MAP kinase. In ongoing experiments, we are evaluating whether anti‐inflammatory drugs restore AIH‐induced pLTF in rats preconditioned with prolonged CIH (7–28 days). Preliminary analyses in rats preconditioned with CIH for 7 days indicate that pLTF is expressed when ketoprofen is given on each day of the CIH exposure (67% baseline), but not when a single ketoprofen dose is given 3 hours prior to assessment of pLTF (−6% baseline). These experiments are the first to explore shifting mechanisms whereby prolonged CIH and associated inflammation undermine phrenic motor plasticity. Such experiments are essential since CIH is a hallmark of sleep apnea, a condition highly prevalent in individuals living with neurological disorders (spinal cord injury, ALS, etc.). It is therefore important to identify and mitigate factors which may undermine the potential for therapeutic interventions (i.e. repetitive AIH) to harness endogenous plasticity and ultimately improve breathing.Support or Funding InformationSupported by: Department of Defense: SCI160123 and NIH R01 HL147554

Artificial Intelligence Applications in Dermatology: Where Do We Stand?

Artificial intelligence (AI) has become a progressively prevalent Research Topic in medicine and is increasingly being applied to dermatology. There is a need to understand this technology's progress to help guide and shape the future for medical care providers and recipients. We reviewed the literature to evaluate the types of publications on the subject, the specific dermatological topics addressed by AI, and the most challenging barriers to its implementation. A substantial number of original articles and commentaries have been published to date and only few detailed reviews exist. Most AI applications focus on differentiating between benign and malignant skin lesions, however; others exist pertaining to ulcers, inflammatory skin diseases, allergen exposure, dermatopathology, and gene expression profiling. Applications commonly analyze and classify images, however, other tools such as risk assessment calculators are becoming increasingly available. Although many applications are technologically feasible, important implementation barriers have been identified including systematic biases, difficulty of standardization, interpretability, and acceptance by physicians and patients alike. This review provides insight into future research needs and possibilities. There is a strong need for clinical investigation in dermatology providing evidence of success overcoming the identified barriers. With these research goals in mind, an appropriate role for AI in dermatology may be achieved in not so distant future.

Specific bacteriophage of Bordetella bronchiseptica regulates B. bronchiseptica-induced microRNA expression profiles to decrease inflammation in swine nasal turbinate cells.

BackgroundRespiratory diseases in pigs are the main health concerns for swine producers. Similar to the diseases in human and other animals, respiratory diseases are primary related to morbidity and are the result of infection with bacteria, viruses, or both. B. bronchiseptica causes serious respiratory diseases in the swine airway track. However, the B. bronchiseptica-specific bacteriophage has diverse advantages such as decreasing antibiotic overuse and possible therapeutic potential against bacteria.ObjectiveThe objects of this study were to investigate the therapeutic effect of specific B. bronchiseptica bacteriophages and to identify genes related to bacteriophage signaling utilizing RNA microarrays in swine nasal turbinate cells.MethodsBor-BRP-1 phages were applied 24 h prior to B.bronchiseptica infection (1 × 107 cfu/ml) at several concentrations of bacterial infection. Cells were incubated to detect cytokines and 24 h to detect mucin production. And real-time quantitative PCR was performed to examine related genes expression. To determine the change of total gene expression based on B.bronchiseptica and Bor-BRP-1 treatment, we performed RNA sequencing experiments.ResultsThe results showed that B. bronchiseptica induced increased expression of several inflammatory genes such as IL-1β, IL-6, and Muc1 in a dose-dependent manner. However, Bor-BRP-1 induced reduction of gene expression compared to the B. bronchiseptica induction group. In addition, microarrays detected Bor-BRP-1-altered inflammatory gene expression against B. bronchiseptica, reducing B. bronchiseptica-induced airway inflammation in swine epithelial cells.ConclusionThese results suggest that the specific bacteriophage has a therapeutic potential to defend against B. bronchiseptica infection by altering inflammatory gene expression profiles.

Macrophage correlates with immunophenotype and predicts anti-PD-L1 response of urothelial cancer.

Immune-checkpoint blockades (ICBs) have been routinely implemented to treat metastatic urothelial cancer (mUC), whereas robust biomarkers are urgently warranted. Herein, we explored latent promising biomarkers based on 348 pretreatment mUC samples from IMvigor210.Methods: The genome, transcriptome, immunome, and metabolome were systemically analyzed using the external TCGA dataset for validation. Kaplan-Meier and ROC curve analyses were performed to estimate the predictive capacity of M1-macrophage infiltration. Chi-square/Spearman/Mann Whitney U test are used to determine its correlation to genetic, biochemical, and clinicopathological parameters.Results: M1 frequency is a robust biomarker for predicting the prognosis and response to ICBs, which is non-inferior to tumor mutation burden (TMB) or tumor neoantigen burden (TNB), and exceeds CD8 T cells, T cell inflamed gene expression profile (GEP), and PD-L1 expression. Moreover, M1 infiltration is associated with immune phenotypes (AUC = 0.785) and is negatively correlated with immune exclusion. Additionally, transcriptomic analysis showed immune activation in the high-M1 subgroup, whereas it showed steroid and drug metabolism reprograming in the M1-deficient subset, which characterized the limited sensitivity to ICB therapy. Notably, investigation of the corresponding intrinsic genomic profiles highlighted the significance of TP53 and FGFR alterations.Conclusions: M1 infiltration is a robust biomarker for immunotherapeutic response and immunophenotype determination in an mUC setting. Innate immunity activation involving macrophage polarization remodeling and anti-FGFR mutations may be promising strategies for synergy with anti-PD-L1 treatments and may help prolong the clinical survival of patients with mUC.

Pasteurella multocida specific bacteriophage suppresses P. multocida-induced inflammation: identification of genes related to bacteriophage signaling by Pasteurella multocida-infected swine nasal turbinate cells.

Although Pasteurella multocida is highly prevalent pathogen in animals and plays an important role in swine respiratory diseases, only a few studies on the use of bacteriophages specific to Pasteurella multocida disease have been reported. The object of this study was to investigate the therapeutic effect of specific P. multocida bacteriophages and to identify genes related to bacteriophage signaling utilizing RNA microarrays in swine nasal turbinate cells. Pas-MUP-1 phages were applied 24h prior to P. multocida infection (1 × 107cfu/ml) at several concentrations of bacterial infection. Cells were incubated to detect cytokines and 24h to detect mucin production. And real-time quantitative PCR was performed to examine related genes expression. To determine the change of total gene expression based on P. multocida and Pas-MUP-1 treatment, we performed RNA sequencing experiments. We found that P. multocida-infected PT-K75 cells show increased gene expression of IL-1β, IL-6, and Muc1 in a dose-dependent manner. Interestingly, these genes resulted in decreased expression in P. multocida pretreated with the P. multocida-specific Pas-MUP-1 bacteriophage. RNA sequencing analysis revealed that bacteriophage administration regulated genes associated with immune and inflammatory responses, and the regulated genes were dramatically concentrated in the cytokine/chemokine-based signaling pathways. Pas-MUP-1 treatment was shown to regulate P. multocida induced gene expression in the bacteria. These results suggest the specific bacteriophage has therapeutic potential as an alternative to antibiotic treatment to defend against P. multocida infection by altering inflammatory gene expression profiles.

An Isocaloric Nordic Diet Modulates RELA and TNFRSF1A Gene Expression in Peripheral Blood Mononuclear Cells in Individuals with Metabolic Syndrome-A SYSDIET Sub-Study.

A healthy dietary pattern is associated with a lower risk of metabolic syndrome (MetS) and reduced inflammation. To explore this at the molecular level, we investigated the effect of a Nordic diet (ND) on changes in the gene expression profiles of inflammatory and lipid-related genes in peripheral blood mononuclear cells (PBMCs) of individuals with MetS. We hypothesized that the intake of an ND compared to a control diet (CD) would alter the expression of inflammatory genes and genes involved in lipid metabolism. The individuals with MetS underwent an 18/24-week randomized intervention to compare a ND with a CD. Eighty-eight participants (66% women) were included in this sub-study of the larger SYSDIET study. Fasting PBMCs were collected before and after the intervention and changes in gene expression levels were measured using TaqMan Array Micro Fluidic Cards. Forty-eight pre-determined inflammatory and lipid related gene transcripts were analyzed. The expression level of the gene tumor necrosis factor (TNF) receptor superfamily member 1A (TNFRSF1A) was down-regulated (p = 0.004), whereas the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) subunit, RELA proto-oncogene, was up-regulated (p = 0.016) in the ND group compared to the CD group. In conclusion, intake of an ND in individuals with the MetS may affect immune function.

HDAC3 Regulates Gingival Fibroblast Inflammatory Responses in Periodontitis

Histone deacetylases (HDACs) are important regulators of gene expression that are aberrantly regulated in several inflammatory and infectious diseases. HDAC inhibitors (HDACi) suppress inflammatory activation of various cell types through epigenetic and non-epigenetic mechanisms, and ameliorate pathology in a mouse model of periodontitis. Activation of gingival fibroblasts (GFs) significantly contributes to the development of periodontitis and the anaerobic bacterium Porphyromonas gingivalis plays a key role in driving chronic inflammation. Here, we analyzed the role of HDACs in inflammatory responses of GFs. Pan-HDACi suberoylanilide hydroxamic acid (SAHA) and/or ITF2357 (givinostat) significantly reduced TNFα- and P. gingivalis–inducible expression and/or production of a cluster of inflammatory mediators in healthy donor GFs (IL1B, CCL2, CCL5, CXCL10, COX2, and MMP3) without affecting cell viability. Selective inhibition of HDAC3/6, but not specific HDAC1, HDAC6, or HDAC8 inhibition, reproduced the suppressive effects of pan-HDACi on the inflammatory gene expression profile induced by TNFα and P. gingivalis, suggesting a critical role for HDAC3 in GF inflammatory activation. Consistently, silencing of HDAC3 expression with siRNA largely recapitulated the effects of HDAC3/6i on mRNA levels of inflammatory mediators in P. gingivalis–infected GFs. In contrast, P. gingivalis internalization and intracellular survival in GFs remained unaffected by HDACi. Activation of mitogen-activated protein kinases and NFκB signaling was unaffected by global or HDAC3/6-selective HDACi, and new protein synthesis was not required for gene suppression by HDACi. Finally, pan-HDACi and HDAC3/6i suppressed P. gingivalis–induced expression of IL1B, CCL2, CCL5, CXCL10, MMP1, and MMP3 in GFs from patients with periodontitis. Our results identify HDAC3 as an important regulator of inflammatory gene expression in GFs and suggest that therapeutic targeting of HDAC activity, in particular HDAC3, may be clinically beneficial in suppressing inflammation in periodontal disease.

P.124: Innate inflammatory gene expression profiling in potential brain-dead donors: Detailed investigation of the effect of common corticosteroid therapy.

P.124: Innate inflammatory gene expression profiling in potential brain-dead donors: Detailed investigation of the effect of common corticosteroid therapy.

Acute myeloid leukemia–induced T-cell suppression can be reversed by inhibition of the MAPK pathway

AbstractAcute myeloid leukemia (AML) remains difficult to treat due to mutational heterogeneity and the development of resistance to therapy. Targeted agents, such as MEK inhibitors, may be incorporated into treatment; however, the impact of MEK inhibitors on the immune microenvironment in AML is not well understood. A greater understanding of the implications of MEK inhibition on immune responses may lead to a greater understanding of immune evasion and more rational combinations with immunotherapies. This study describes the impact of trametinib on both T cells and AML blast cells by using an immunosuppressive mouse model of AML and primary patient samples. We also used a large AML database of functional drug screens to understand characteristics of trametinib-sensitive samples. In the mouse model, trametinib increased T-cell viability and restored T-cell proliferation. Importantly, we report greater proliferation in the CD8+CD44+ effector subpopulation and impaired activation of CD8+CD62L+ naive cells. Transcriptome analysis revealed that trametinib-sensitive samples have an inflammatory gene expression profile, and we also observed increased programmed cell death ligand 1 (PD-L1) expression on trametinib-sensitive samples. Finally, we found that trametinib consistently reduced PD-L1 and PD-L2 expression in a dose-dependent manner on the myeloid population. Altogether, our data present greater insight into the impact of trametinib on the immune microenvironment and characteristics of trametinib-sensitive patient samples.

1589TiP - KEYNOTE-495/KeyImPaCT: A randomized, biomarker-directed, phase II trial of pembrolizumab-based therapy for non–small cell lung cancer (NSCLC)

BackgroundKEYNOTE-495 is a randomized, open-label, phase II trial (NCT03516981) evaluating the clinical activity and safety of pembrolizumab (pembro)-based combination (combo) therapy in patients (pts) with treatment-naive, advanced NSCLC in biomarker-defined response groups. This biomarker-based approach is based on 2 validated, independent, next-generation biomarkers (T cell–inflamed gene expression profile [GEP] and tumor mutational burden [TMB]) and can help determine the differential efficacy of pembro-based combo therapy, based on the composite biomarker profile, and inform precision immunotherapy in the future. Trial designIn this group-sequential, adaptive randomized trial, pts (N∼288) receive pembro 200mg Q3W intravenously (IV) combined with MK-4280 (anti–LAG-3) 200mg Q3W IV, lenvatinib (lenv) 20mg PO QD, or MK-1308 (anti–CTLA-4) 25mg Q6W IV for 35 cycles (∼2 years); pts in the lenv arm may receive lenv monotherapy until disease progression or toxicity. After biomarker screening, pts are assigned to 1 of 4 groups—TMBlowGEPlow, TMBhighGEPlow, TMBlowGEPhigh, and TMBhighGEPhigh—then randomized to 1 of 3 pembro-based regimens; adaptive design elements are used to adjust randomization based on objective responses. Eligible pts are ≥18 years of age with histologically or cytologically confirmed treatment-naive, advanced NSCLC; documented absence of EGFR and B-Raf mutations and ALK and ROS1 gene rearrangements; measurable disease per RECIST v1.1; and ECOG PS 0-1. Response is assessed by imaging Q9W for the first year and Q12W thereafter using RECIST v1.1. Primary end point is investigator-assessed objective response rate (RECIST v1.1). Secondary end points are progression-free survival, overall survival, and safety. Multiple interim analyses may be performed given the group-sequential design. Enrollment is ongoing. Clinical trial identificationNCT03516981: Trial initiation, October 1, 2018. Editorial acknowledgementHolly C. Cappelli, PhD, and Dana Francis, PhD, of the ApotheCom pembrolizumab team (Yardley, PA, USA); funded by Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA. Legal entity responsible for the studyMerck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA. FundingMerck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA. DisclosureM. Gutierrez: Advisory / Consultancy: Eli Lilly, Foundation One, Essanex, Guardant 360; Speaker Bureau / Expert testimony: Merck, BMS. M.D. Hellmann: Advisory / Consultancy: Merck, Bristol-Myers Squibb, AstraZeneca, Genentech/Roche, Janssen, Nektar, Syndax, Mirati, and Shattuck Labs; Shareholder / Stockholder / Stock options: Shattuck Labs; Research grant / Funding (institution): BMS; Travel / Accommodation / Expenses: AstraZeneca, BMS; Non-remunerated activity/ies, A patent has been filed by MSK related to the use of tumor mutation burden to predict response to immunotherapy (PCT/US2015/062208), which has received licensing fees from PGDx.: MSK. M.A. Gubens: Advisory / Consultancy: AbbVie, ARIAD, AstraZeneca, BMS, Roche/Genentech, Heron, Mersana, Novartis, Pfizer; Research grant / Funding (institution), Travel / Accommodation / Expenses: Celgene, Merck, Novartis; Research grant / Funding (institution): OncoMed, Roche. C. Aggarwal: Advisory / Consultancy: BMS, Celgene, Roche/Genentech, AstraZeneca. D.S.W. Tan: Honoraria (self): Novartis, Bayer, Boehringer Ingelheim, AstraZeneca, BMS, Roche, Takeda, Pfizer; Research grant / Funding (self): Novartis, Bayer, AstraZeneca, GSK, Pfizer; Advisory / Consultancy: Novartis, Bayer, Boehringer Ingelheim. E. Felip: Advisory / Consultancy, Speaker Bureau / Expert testimony: AbbVie, AstraZeneca, BMS, Boehringer Ingelheim, Eli Lilly, Merck KGaA, MSD, Novartis, Pfizer, Roche, Takeda; Advisory / Consultancy: Blue Print Medicines, Celgene, Guardant Health. J.C. Yang: Honoraria (self), Advisory / Consultancy: Boehringer Ingelheim, Eli Lilly, Bayer, Roche/Genentech, Chugai, MSD, Pfizer, Novartis, BMS, Ono Pharmaceuticals ; Advisory / Consultancy: Merck Serono, Celgene, Merrimack, Yuhan Pharmaceuticals, Daiichi Sankyo, Hansoh Pharmaceuticals, Takeda Pharmaceuticals Blueprint Medicines. E.B. Garon: Research grant / Funding (self), Clinical trial funding: Merck, AstraZeneca, BMS. A. Basso: Full / Part-time employment: Merck . H. Ma: Full / Part-time employment: Merck . L. Fong: Research grant / Funding (self): Merck . A. Snyder: Shareholder / Stockholder / Stock options, Full / Part-time employment: Merck . J. Yuan: Full / Part-time employment: Merck . R.S. Herbst: Advisory / Consultancy: AbbVie, AstraZeneca, Biodesix, BMS, Eli Lilly, EMD Serano, Roche/Genentech, Heat Biologics, Junshi, Loxo Oncology Merck, Nektar, NextCure, Novartis, Pfizer, Sanofi, Seattle Genetics, Shire PLC, Spectrum, Symphogen, Tesaro ; Research grant / Funding (self): AstraZeneca, Eli Lilly, Merck; Advisory / Consultancy, Scientific Advisory Boards: Neon Therapeutics, Infinity Pharmaceuticals, NextCure; Officer / Board of Directors: Junshi Pharmaceuticals. All other authors have declared no conflicts of interest.

Changes of gene expression profiles of infants inflammatory genes after Rota virus infection.

Rotavirus is a medical important pediatric pathogen causes severe gastroenteritis in infants, leading to 450,000 deaths globally each year. RT-PCR-Array technique analysis is a powerful method to determine global profiles of gene expression in cells and tissues after viral infections. Using this analysis, the Ct values of 14 immune inflammatory genes were subjected to determine their gene expression rates compared with Ct values of control (non-infected persons). Five housekeeping genes were used as internal control genes. All mRNA transcripts of these genes were upregulated (from 2.2974 fold change for CCL3, CCR1, CCR3, CXCL2, IFN-a, IFNA-g, IL1B, IL1R and 21.1121 fold change for TNF-b) after infection with rotavirus. This results revealed that rotavirus infection increase the gene expression rate of 14 inflammatory genes and lead to high impact on key immune pathways in infected infants.

Phase 1 study of MK-4166, an anti-human glucocorticoid-induced tumor necrosis factor receptor (GITR) antibody, as monotherapy or with pembrolizumab (pembro) in patients (pts) with advanced solid tumors.

9509 Background: Ligation of GITR on immune cells decreases Treg-mediated suppression and enhances T cell proliferation. MK-4166 is a humanized IgG1 agonist monoclonal antibody targeting GITR. Data from the first-in-human phase 1 study (NCT02132754) of MK-4166 as monotherapy (mono) or in combination with pembro (combo) are presented. Methods: MK-4166 was tested alone (0.0015 mg IV-900 mg Q3W ×4 doses) or with pembro (fixed dose 200 mg IV Q3W up to 35 doses) in the absence of toxicity or progression. Study included a dose escalation/confirmation cohort (metastatic solid tumors) and an expansion cohort (treatment-naive and pretreated melanoma). A T cell–inflamed gene expression profile (GEP) was assessed using RNA from baseline tumor samples. End points – primary: safety/tolerability, maximum tolerated dose (MTD) of MK-4166; secondary: pharmacokinetics (PK), pharmacodynamics (PD); exploratory: objective response rate (ORR) per irRECIST1.1. Results: Of 113 pts, 48 received mono and 65 combo; 20 were in the melanoma expansion. Common AEs ( > 20%) were fatigue, infusion-related reaction, nausea, abdominal pain, and pruritus; 43 pts had grade ≥3 AEs (38.1%); 6 (5.3%) were treatment-related. One dose-limiting toxicity (bladder perforation in a urothelial pt with a neobladder) possibly related to study drug was observed with mono. MTD was not reached. No treatment-related deaths were observed. MK-4166 PK/PD showed target-mediated drug disposition concomitant with decreased GITR availability on T cells in blood with increasing doses. Four objective responses (4/45; ORR, 9%) were seen with combo in dose escalation. For ICI-naive melanoma pts (n = 13) in expansion, ORR was 69% (95% CI, 38-91), including 4 CRs and 5 PRs. No response was observed in 7 pts previously treated with ICI. High ORRs were observed in noninflamed and inflamed ICI-naive melanoma pts. Conclusions: MK-4166 at a dose up to 900 mg as monotherapy and in combination with pembro was well tolerated, with dose-related evidence of target engagement. Responses were observed with MK-4166 900 mg plus pembro, particularly in pts with melanoma naive to ICIs. Clinical trial information: NCT02132754.

Biomarker-directed precision oncology of pembrolizumab-based combination therapy for non-small cell lung cancer: Phase II KEYNOTE-495/KeyImPaCT study.

TPS9117 Background: Pembrolizumab-based combination immunotherapy aims to improve clinical outcomes over pembrolizumab monotherapy. A biomarker-based therapeutic approach may be associated with improved response to different combination therapies of immune checkpoint inhibitors and may improve overall outcomes in NSCLC. The randomized, multicenter, open-label, phase 2 KEYNOTE-495 trial ( NCT03516981 ) will evaluate the clinical usefulness of biomarker-informed, pembrolizumab-based combination therapy in patients with treatment-naive, advanced NSCLC. Methods: This is a group-sequential, adaptive randomization trial. Patients will have histologically or cytologically confirmed treatment-naive, advanced NSCLC, documented absence of EGFR and B-Raf mutations and ALK and ROS1 gene rearrangements, measurable disease per RECIST v1.1, and ECOG PS 0-1. Tumor tissue from patients will be initially screened for 2 validated, independent, next-generation biomarkers: T cell–inflamed gene expression profile (GEP) and tumor mutational burden (TMB). Based on results of biomarker screening, patients will be assigned to 1 of 4 groups: TMBlowGEPlow, TMBhighGEPlow, TMBlowGEPhigh, and TMBhighGEPhigh. Within each group, patients will be randomly assigned to receive pembrolizumab 200 mg Q3W intravenously (IV) combined with either MK-4280 200 mg Q3W (anti–LAG-3) IV or lenvatinib 20 mg orally once daily, with the randomization assignment adaptively modified based on interim efficacy analyses. Response will be assessed by imaging every 9 weeks for the first year and every 12 weeks thereafter using RECIST v1.1. Treatment will continue for 35 cycles (~2 years). Patients in the pembrolizumab + lenvatinib arm who complete 35 treatments may continue with lenvatinib monotherapy until disease progression or toxicity. Treatment arms may be terminated during the interim analysis due to safety, prespecified futility criteria, or both. Primary end point is investigator-assessed objective response rate (RECIST v1.1). Secondary end points are progression-free survival, overall survival, and safety. Recruitment and screening are ongoing in more than 8 countries. Clinical trial information: NCT03516981.

SUN-084 Aged Insulin Resistant Macrophages Reveal Dysregulated Cholesterol Biosynthesis, Pro-Inflammatory Gene Expression, and Reduced Foam Cell Formation

Insulin resistance is the central defining feature of type 2 diabetes and increases with age. People with type 2 diabetes are at a significantly increased risk of developing atherosclerosis. Macrophages are phagocytotic leukocytes which play a central role in the development of type 2 diabetes and atherosclerosis. Despite their relevance however, the effect of insulin and insulin resistance on macrophages regarding their inflammatory status and role in atherosclerosis remain unclear. Our aim was to determine the inflammatory gene expression profile in aged insulin resistant macrophages and how this may impact foam cell formation - a key step in the development of atherosclerosis. Genome-wide transcript array analysis was performed on bone marrow derived macrophages from aged insulin resistant mice (n=3) gained through chronic hyper-stimulation of the PI3K arm of the insulin signalling pathway via hematopoietic SHIP2 knock-down (h-SHIP2KD). The transcriptome of aged h-SHIP2KD insulin resistant macrophages become reprogrammed, differentially expressing more than 4000 genes (≥2fold, P=≤ 0.05. Pathway analysis revealed that the most significantly altered intracellular signalling pathways include upregulation of interferon signalling pathwyas, components of the inflammasome (for example increased mRNA expression Nlrp3: 1.75-fold, P=0.05, Casp4: 4.31-fold, P=0.05) and changes in cholesterol biosynthesis genes (for example Fdps: 2.56-fold, P=0.05, Idi1: 3.13-fold, P=0.05, Sqle: 3.87-fold, P=0.05, Cyp51: 4.47-fold, P=0.05, Dhcr: 4.72-fold, P=0.05). Exposure of these h-SHIP2KD macrophages to acetylated LDL revealed a decreased capacity to form foam cells compared to WT controls (Mean staining intensity 2818 vs 4730 units respectively, P=0.03, n=3). These data support the emerging role of insulin a modulator of inflammation and that insulin resistance in macrophages is protective against foam cell formation. Our transcriptome expression data revealed novel targets which may be exploited for new therapies.