Citations

Abstract Background: Chronic myelogenous leukemia (CML) is a BCR-ABL1 driven myeloid neoplasm, with excellent response rates to tyrosine kinase inhibitors (TKI). TKI such as imatinib (IM), dasatinib (DAS), nilotinib (NIL), bosutinib (BOS), and ponatinib (PON) currently US FDA approved, have dramatically changed survival outcomes. That being said, two main challenges exist; suboptimal responses and TKI intolerance. The reason why some patients tolerate/respond to TKI better than others is unknown, but is potentially explainable based on drug metabolism. The field of pharmacogenomics (PGX) is rapidly evolving, with commercial panel's rapidly assessing metabolic pathways such as cytochrome P450 (CYP), UGTA1A, p-glycoprotein/ABCB1. We carried out this study using a 22 gene-PGX panel to assess potential causes of TKI related intolerance and suboptimal responses in compliant patients. Methods: Twenty-nine Mayo Clinic patients with CML were prospectively recruited after informed consent. Buccal swabs were utilized for PGX testing to assess variations/polymorphisms involving CYP3A4/5, 1A2, 2C9, UGT1A1, amongst others (www.Oneome.com). Three groups of patients were recruited, those with i) TKI intolerance, ii) suboptimal responses by ELN criteria, iii) and newly diagnosed cases, to assess if PGX testing could explain TKI intolerance or resistance, or alter choice of therapy in newly diagnosed cases. A pharmacology review was obtained to interpret reports in all cases. Results: 29 patients with CML were prospectively enrolled, median age 64 years, 15 (52%) male. At last follow up (median 21 months), 3 (10%) deaths and no disease progressions were documented. 82% received first line IM, whereas 18% received first line DAS. Major metabolic pathways assessed for IM included, CYP3A4 and 5, while minor pathways included 2C19, 2C9 and 2D6. Majority (83%) of IM patients had normal major pathways with 17% being intermediate-normal. With regards to the minor pathways, 25% were rapid metabolizers for 2C19, whereas 29% were intermediate and 4% poor metabolizers for 2C9 and 50% and 16% were intermediate and poor metabolizers for 2D6. In all IM treated patients PGX testing did not completely explain intolerance or resistance and did not change clinical decision making. Front line DAS was used in 18% and the major pathway assessed was CYP 3A4, with 80% being normal and 20% being intermediate metabolizers. Once again PGX testing did not completely explain intolerance/resistance and no therapeutic changes were based on PGX analysis. Seventeen (59%) patients received second line TKI therapy (50% each for intolerance and resistance), with DAS (58%) and NIL (29%), being the two most common. Once again for DAS, 90% had normal 3A4 metabolism while 10% were intermediate and for NIL 80% were normal and 20% intermediate. Only one of 2 NIL treated patients with UGTA1A polymorphisms developed indirect hyperbilirubinemia (grade 1). In the second line setting, PGX testing did not completely explain or alter treatment decisions. BOS was used as a third line agent in 3 (>10%) patients, 2 who remained refractory and one with intolerance, and all three were normal CYP3A4 metabolizers. None of these patients developed diarrhea a common side effect of BOS. PON was used in one patient with intermediate CYP3A4 metabolism as a 5th line agent and the patient continues to have refractory disease with no thrombotic events. In all cases, no changes were made to concomitant medications based on PGX testing. Conclusion: In conclusion, while PGX testing has rapidly evolved and become commercially available, in the context of TKI therapy for CML, while it offers useful insights on minor metabolic derangements and side effects like NIL induced hyperbilirubinemia, in our study, currently limited by a small sample size, it did not completely explain TKI intolerance or resistance. Recruitment of additional patients (n=100) and correlations with plasma drug levels are currently ongoing. Table. Table. Disclosures Al-Kali: Novartis: Research Funding. Stewart:Amgen Inc., Celgene, Roche, Seattle Genetics: Research Funding; Amgen Inc., BMS, Celgene, Takeda, Roche, Seattle Genetics, Janssen, Ono: Consultancy.

Abstract Background: Peripheral T-cell lymphomas (PTCLs) are aggressive non-Hodgkin lymphomas with marked clinical, pathological, and molecular heterogeneity. Outcomes following standard therapy generally are poor; however, few candidate therapeutic targets have been identified for precision medicine approaches. Retinoic acid receptor alpha (RARA) is a transcription factor that modulates cell growth and differentiation in response to natural or synthetic retinoids. Retinoids have been used successfully to treat acute promyelocytic leukemia and some cutaneous T-cell lymphomas (CTCLs). However, the function of RARA and the action of retinoids in PTCL have not been defined. Methods:Based on identification of a PTCL patient with a non-synonymous point mutation, RARA R394Q, identified in the Mayo Clinic Center for Individualized Medicine, we sought to characterize the role of RARA in PTCL cells. To investigate the role of wild-type and mutant RARA, we constructed expression vectors containing either wild-type RARA or RARA R394Q coding sequences, and also used siRNAs targeting RARA to study the role of native RARA expression. Cell lines derived from post-thymic T-cell malignancies were used for in vitro studies, including HuT78 and Mac-1 (both derived from circulating tumor cells from CTCL patients) and Karpas 299 (from an ALK-positive anaplastic large cell lymphoma). Following RARA overexpression or knockdown, we measured cell growth, cell cycle regulation, and sensitivity to synthetic retinoids. In addition, RNA sequencing and pathway analysis were performed to profile the transcriptomic response to retinoids in malignant T cells. Results:In two RARAlow cell lines, Karpas 299 and HuT78, overexpression of wild-type RARA or RARA R394Q significantly increased cell growth (p<0.001), with a greater increase observed from mutant versus wild-type RARA in Karpas 299 (136% of control versus 122%; p=0.04). Accordingly, knockdown of wild-type RARA in the RARAhigh cell line, Mac-1, resulted in a 22% inhibition of cell growth (p=0.0002). This inhibition specifically was associated with G1 cell cycle arrest (120% of control; p=0.004) and decreased protein expression of the G1-S-associated cyclin-dependent kinases, CDK2, CDK4, and CDK6. These kinases were up-regulated by overexpression of RARA in RARAlow HuT78 cells. The relatively RARA-specific retinoid, AM80 (tamibarotene), and the less specific retinoid, all-trans retinoic acid (ATRA), resulted in RARA protein degradation, cell growth inhibition that was both dose-dependent and proportional to baseline RARA expression, G1 arrest, and CDK protein up-regulation. Gene-set enrichment analysis (GSEA) of transcriptome data confirmed that genes down-regulated by AM80 were highly enriched for regulators of cell cycle and particularly G1-S transition. Finally, overexpressing RARA in RARAlow Karpas 299 and HuT78 cell lines significantly increased the ability of AM80 to inhibit CDK2/4/6 expression and cell growth (16% to 23% greater growth inhibition than control; p<0.05). Conclusions:RARA drives cyclin-dependent kinase expression and G1-S transition in malignant T cells, and promotes cell growth. These functions may be enhanced by specific RARA gene mutations. Synthetic retinoids inhibit these functions in a dose-dependent fashion, and are most effective in cells with high RARA expression. These data suggest RARA as a candidate therapeutic target in some PTCL patients. Disclosures Nowakowski: Celgene: Research Funding; Morphosys: Research Funding; Bayer: Consultancy, Research Funding.

The application of next-generation sequencing in clinical practice is increasing as accuracy and interpretation have improved and the cost continues to decline rapidly. Cell-free DNA is a unique source for next-generation sequencing that could change routine clinical practice in gastroenterology and hepatology. Testing of cell-free DNA in blood and fecal samples is an easy, rapid, and noninvasive method to assess for premalignant, malignant, metabolic, infectious, inflammatory, and autoimmune gastrointestinal and liver diseases. In this review, we describe cell-free DNA technologies, current applications of cell-free DNA testing, and proposed cell-free DNA targets for gastrointestinal and hepatic diseases, with a specific focus on malignancy. In addition, we provide commentary on how cell-free DNA can be integrated into clinical practice and help guide diagnosis, prognosis, disease management, and therapeutic response.

Primary sclerosing cholangitis (PSC) is a pathogenically complex, chronic, fibroinflammatory disorder of the bile ducts without known etiology or effective pharmacotherapy. Emerging in vitro and in vivo evidence support fundamental pathophysiologic mechanisms in PSC centered on enterohepatic circulation. To date, no studies have specifically interrogated the chemical footprint of enterohepatic circulation in PSC. Herein, we evaluated the metabolome and lipidome of portal venous blood and bile obtained at the time of liver transplantation in patients with PSC (n = 7) as compared to individuals with noncholestatic, end-stage liver disease (viral, metabolic, etc. (disease control, DC, n = 19)) and to nondisease controls (NC, living donors, n = 12). Global metabolomic and lipidomic profiling was performed on serum derived from portal venous blood (portal serum) and bile using ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and differential mobility spectroscopy-mass spectroscopy (DMS-MS; complex lipid platform). The Mann–Whitney U test was used to identify metabolites that significantly differed between groups. Principal-component analysis (PCA) showed significant separation of both PSC and DC from NC for both portal serum and bile. Metabolite set enrichment analysis of portal serum and bile demonstrated that the liver-disease cohorts (PSC and DC) exhibited similar enrichment in several metabolite categories compared to NC. Interestingly, the bile in PSC was uniquely enriched for dipeptide and polyamine metabolites. Finally, analysis of patient-matched portal serum and biliary metabolome revealed that these biological fluids were more homogeneous in PSC than in DC or NC, suggesting aberrant bile formation and enterohepatic circulation. In summary, PSC and DC patients exhibited alterations in several metabolites in portal serum and bile, while PSC patients exhibited a unique bile metabolome. These specific alterations in PSC are amenable to hypothesis testing and, potentially, therapeutic pharmacologic manipulation.

11571 Background: Relapsed/refractory (R/R) non-Hodgkin lymphomas (NHL) have a poor prognosis with limited treatment options. Our expanding knowledge of molecular alterations seen in R/R NHL allows identification of patients that potentially may benefit from a precision medicine approach. However, experience in routine clinical implementation of precision medicine has been limited. Here, we summarize our clinical experience in molecular characterization of RR NHL targeted therapy (TT) using next-generation sequencing (NGS), and selection of targeted therapy (TT) based on molecular profile. Methods: We conducted a prospective study in RR NHL through the Center for Individualized Medicine at Mayo Clinic. Consenting patients underwent NGS using FoundationOne Heme panel from biopsies done at time of relapse. Results of NGS were discussed at the Genomic Tumor Board and recommendations for TT were given based on matching specific molecular alteration(s) with potential agent(s) predicted to be active based on NGS. The agents could include FDA-approved, off-label use and clinical trial therapies. Results: 28 cases were enrolled: 18 aggressive NHL, 10 follicular lymphoma (FL). Molecular alterations were present in all cases. In aggressive B-cell NHL, CDKN2A/B gene cluster alterations were seen in 73% (8/11), while seen in only 1/7 T-cell lymphomas (TCL), and 1/10 FL. TP53 deletions were second most common genomic alterations in DLBCL (57%) and seen in 40% FL. JAK-STAT and ERBB pathways were altered in TCL (2/7 each). IGH-BCl-2 gene rearrangement were common in FL (70%), followed by MLL gene alterations (50%). Targetable mutations were present in 86% (24/28) of cases. A TT was recommended in all 24 cases, but received by 2 patients only. Remaining patients did not due to benefit from current therapy (10/24), ineligibility or lack of clinical trial (7/24) or interim clinical deterioration (5/24). Conclusions: Targetable mutations were identified in most cases of RR NHL with TT recommended for all cases. However, access to TT limits potential clinical benefit of molecular-based matching strategy. More studies are needed to assess impact on clinical outcomes.

Improved methods are needed to risk stratify and predict outcomes in patients with primary sclerosing cholangitis (PSC). Therefore, we sought to derive and validate a prediction model and compare its performance to existing surrogate markers. The model was derived using 509 subjects from a multicenter North American cohort and validated in an international multicenter cohort (n = 278). Gradient boosting, a machine‐based learning technique, was used to create the model. The endpoint was hepatic decompensation (ascites, variceal hemorrhage, or encephalopathy). Subjects with advanced PSC or cholangiocarcinoma (CCA) at baseline were excluded. The PSC risk estimate tool (PREsTo) consists of nine variables: bilirubin, albumin, serum alkaline phosphatase (SAP) times the upper limit of normal (ULN), platelets, aspartate aminotransferase (AST), hemoglobin, sodium, patient age, and number of years since PSC was diagnosed. Validation in an independent cohort confirms that PREsTo accurately predicts decompensation (C‐statistic, 0.90; 95% confidence interval [CI], 0.84‐0.95) and performed well compared to Model for End‐Stage Liver Disease (MELD) score (C‐statistic, 0.72; 95% CI, 0.57‐0.84), Mayo PSC risk score (C‐statistic, 0.85; 95% CI, 0.77‐0.92), and SAP <1.5 × ULN (C‐statistic, 0.65; 95% CI, 0.55‐0.73). PREsTo continued to be accurate among individuals with a bilirubin <2.0 mg/dL (C‐statistic, 0.90; 95% CI, 0.82‐0.96) and when the score was reapplied at a later course in the disease (C‐statistic, 0.82; 95% CI, 0.64‐0.95). Conclusion: PREsTo accurately predicts hepatic decompensation (HD) in PSC and exceeds the performance among other widely available, noninvasive prognostic scoring systems.

The United Kingdom‐Primary Biliary Cholangitis (UK‐PBC) risk scores are a set of prognostic models that estimate the risk of end‐stage liver disease in patients with PBC at 5‐, 10‐ and 15‐year intervals. They have not been externally validated outside the United Kingdom. In this retrospective, external validation study, data were abstracted from outpatient charts and discrimination and calibration of the UK‐PBC risk scores were assessed. A total of 464 patients with PBC treated with ursodeoxycholic acid were included. The median diagnosis age was 52.4 years, and 88% were female patients. The cumulative incidence of events was 6%, 9%, and 15% at 5, 10, and 15 years, respectively. Concordance (c‐statistic) was 0.88, 0.85, and 0.84 using the 5‐, 10‐ and 15‐year risk scores, respectively, which was slightly lower than values observed in the United Kingdom validation cohort. Using the 5‐year risk score, more events were observed than predicted (25 versus 16.8; P = 0.046); using the 10‐year risk score, there was no difference between the observed and predicted number of events (35 versus 44.9; P = 0.14); conversely, using the 15‐year risk score, fewer events were observed than predicted (46 versus 67.5; P = 0.009). Limiting evaluation by the 15‐year UK‐PBC risk score to those with >10 years of follow‐up demonstrated no difference between observed and predicted events. Using the 5‐year risk score, patients within the highest quartile had statistically significant worse event‐free survival compared to the rest of the cohort: 82% versus 98% at 5 years, 73% versus 97% at 10 years, and 58% versus 93% at 15 years. Conclusion: In patients assessed at a North American tertiary medical center, the UK‐PBC risk score had excellent discrimination and was reasonably calibrated both in the short and long term. (Hepatology Communications 2018;2:676‐682)