Proximal Biomarkers Research Articles

Multiomics Analysis Confirms Effective Target Engagement for RVU120 - a First-in-Class CDK8/19 Kinase Inhibitor in AML and MR-MDS Patients and Reveals the Mechanism of Action

RVU120 is a first-in-class CDK8/19 kinase inhibitor. The primary objective of an ongoing Phase Ib study of RVU120 in AML and HR-MDS patients is to determine the preliminary safety profile and the recommended phase 2 dose of RVU120 as a single agent. Characterization of pharmacodynamic (PD) effects is a secondary objective of the trial. PD analysis supports the determination of the recommended phase 2 dose, further clinical development and ultimately positioning of this novel class of targeted agents in the treatment strategies for AML and MDS patients. A battery of tests has been used to establish the relationship between pharmacokinetics (PK) and PD activity, confirm the on-target activity of RVU120, and reveal a mechanism of action of CDK8/19 inhibitors in preclinical models. Specific protein binding of RVU120 in leukemic and normal blood cells has been verified by capturing CDK8/19 kinases after a competition with ATP acyl-phosphate biotin-labeled probes. Changes in CDK8- dependent phosphorylation of STAT5 were used as proximal biomarkers and were measured by flow cytometry. Transcriptome changes were used as downstream biomarkers and were determined by the integrated analysis of bulk RNAseq, ChIPseq and ATACseq. The molecular activity of RVU120 was correlated with anti-leukemic activity and surface marker changes. In patients, PD activity of RVU120 was established by measuring changes in pSTAT5 levels, directly in blast cells and ex vivo in established AML cell lines, exposed to plasma collected from patients treated with RVU120. Downstream transcriptomic effects in patients were assessed by RNAseq and phenotypic changes were tracked by flow cytometry surface marker panels. Preclinical profiling indicated effective CDK8/19 target engagement and inhibition of pSTAT5 levels at low nanomolar concentrations of RVU120, in both established cell lines and primary leukemic cells. The antileukemic activity was tightly correlated with levels of biomarker inhibition, however, in a group of super-responders enriched for NPM1 mutants and leukemic stem cells, efficacy was achieved at partial biomarker inhibition levels. Downstream profiling revealed inhibition of homeobox genes and induction of STAT5-dependent genes as two major transcriptomic programs regulated by RVU120. Homeobox genes inhibited by RVU120 are essential for the viability and self-renewal of leukemic cells, whereas many STAT5 -dependent genes are known as master regulators of erythropoiesis. Consistently, treatment of healthy and transformed CD34+ cells with RVU120 results in the commitment to erythropoiesis. Profiling of patient samples largely confirmed preclinical findings. Inhibition of pSTAT5 in CD34+ blasts from AML and MDS patients tightly correlated with achieved exposures after the first dose and at a steady state. At the time of submission pSTAT5 inhibition has already reached >50%, a threshold that based on preclinical predictions is sufficient for robust efficacy in selected groups of super-responder patients. Further increase at higher doses, is expected to result in deeper, more durable responses in broader populations of patients. Broad transcriptomic changes were observed in patients after treatment with RVU120. Notably, potent inhibition of homeobox genes was observed in NPM1 mutant blasts and induction of erythroid master regulator genes was observed in SF3B1 mutant cells with erythroid arrest. Clear hallmarks of erythroid differentiation were further identified in several patients by specific changes in surface markers. These patients achieved meaningful clinical responses such as reduction of blasts and transfusion independence, respectively.

Abstract 1278: MAT2A inhibitor, IDE397, displays broad anti-tumor activity across a panel of MTAP-deleted patient-derived xenografts

Abstract Background: Approximately 15% of all solid tumors harbor deletions in methylthioadenosine phosphorylase (MTAP) (1). MTAP deficiencies have been reported to sensitize tumor cells to methionine adenosyltransferase 2A (MAT2A) modulation (2). MAT2A synthesizes the universal methyl donor and PRMT5 substrate S-adenosylmethionine (SAM) from methionine. PRMT5 symmetrically di-methylates arginine (SDMA) residues on proteins that are key to genomic integrity and proteostasis. We evaluated the anti-tumor activity of the small molecule MAT2A inhibitor IDE397 in multiple CDX models and in an ongoing study of 45 MTAP-deleted PDX models to inform clinical strategy for a First-in-Human clinical trial evaluating IDE397. Methods: CDX models utilized included the HCT116 isogenic pair and endogenous MTAP-deleted models. A series of dose response and time-course studies were performed to inform the efficacy-PD relationship. SAM was quantified by LC-MS and SDMA was visualized by western blot (WB) and further quantified by immunohistochemistry (IHC). PDX models with MTAP homozygous deletions were selected from bladder, esophageal, gastric, head and neck, NSCLC and pancreatic cancers. MTAP deletion was determined by WES and/or SNP 6.0 microarray and verified by absence of significant mRNA. Tumors were subcutaneously implanted into immunocompromised mice by serial transplantation of tumor fragments. Each PDX model included a vehicle control and an IDE397-treated group (n=5/group). IDE397 was administered orally once per day (QD). Tumor growth inhibition (TGI) was calculated using the formula TGI = (TV controlfinal - TV treatedfinal)/(TV controlfinal - TV controlinitial) x 100. Results: Administration of IDE397 resulted in TGI and/or tumor regression in MTAP-deleted CDX and PDX models. The HCT116 MTAP-deleted CDX model was more sensitive to IDE397 compared to the HCT116 MTAP-WT model. IDE397 produced a dose- and time-dependent modulation of the proximal and distal PD biomarkers SAM and SDMA. SAM was decreased in plasma and tumors irrespective of MTAP status. In contrast, SDMA was selectively modulated in MTAP-null models only. In a NSCLC CDX model, a dose dependent TGI was observed, with the higher doses leading to tumor regression. Anti-tumor activity is observed in MTAP-deleted PDX models, where IDE397 administration has resulted in TGI and tumor regressions. Conclusion: Xenograft studies indicate that IDE397 exhibits anti-tumor activity as a single agent in MTAP-deleted CDX models and in MTAP-deleted PDX models of NSCLC, pancreatic, bladder, head and neck, esophageal and gastric cancer. This data supports the clinical evaluation of IDE397 across a wide range of solid tumor types with MTAP-deletion.

Biochemical and anaplerotic applications of in vitro models of propionic acidemia and methylmalonic acidemia using patient-derived primary hepatocytes

Propionic acidemia (PA) and methylmalonic acidemia (MMA) are autosomal recessive disorders of propionyl-CoA (P-CoA) catabolism, which are caused by a deficiency in the enzyme propionyl-CoA carboxylase or the enzyme methylmalonyl-CoA (MM-CoA) mutase, respectively. The functional consequence of PA or MMA is the inability to catabolize P-CoA to MM-CoA or MM-CoA to succinyl-CoA, resulting in the accumulation of P-CoA and other metabolic intermediates, such as propionylcarnitine (C3), 3-hydroxypropionic acid, methylcitric acid (MCA), and methylmalonic acid (only in MMA). P-CoA and its metabolic intermediates, at high concentrations found in PA and MMA, inhibit enzymes in the first steps of the urea cycle as well as enzymes in the tricarboxylic acid (TCA) cycle, causing a reduction in mitochondrial energy production. We previously showed that metabolic defects of PA could be recapitulated using PA patient-derived primary hepatocytes in a novel organotypic system. Here, we sought to investigate whether treatment of normal human primary hepatocytes with propionate would recapitulate some of the biochemical features of PA and MMA in the same platform. We found that high levels of propionate resulted in high levels of intracellular P-CoA in normal hepatocytes. Analysis of TCA cycle intermediates by GC–MS/MS indicated that propionate may inhibit enzymes of the TCA cycle as shown in PA, but is also incorporated in the TCA cycle, which does not occur in PA. To better recapitulate the disease phenotype, we obtained hepatocytes derived from livers of PA and MMA patients. We characterized the PA and MMA donors by measuring key proximal biomarkers, including P-CoA, MM-CoA, as well as clinical biomarkers propionylcarnitine-to-acetylcarnitine ratios (C3/C2), MCA, and methylmalonic acid. Additionally, we used isotopically-labeled amino acids to investigate the contribution of relevant amino acids to production of P-CoA in models of metabolic stability or acute metabolic crisis. As observed clinically, we demonstrated that the isoleucine and valine catabolism pathways are the greatest sources of P-CoA in PA and MMA donor cells and that each donor showed differential sensitivity to isoleucine and valine. We also studied the effects of disodium citrate, an anaplerotic therapy, which resulted in a significant increase in the absolute concentration of TCA cycle intermediates, which is in agreement with the benefit observed clinically. Our human cell-based PA and MMA disease models can inform preclinical drug discovery and development where mouse models of these diseases are inaccurate, particularly in well-described species differences in branched-chain amino acid catabolism.

Validation of ELISA and cellular uptake inhibition assays to detect and quantify antibodies raised against sebelipase alfa in lysosomal acid lipase deficiency

Propionic acidemia (PA) and methylmalonic acidemia (MMA) are autosomal recessive disorders of propionyl-CoA (P-CoA) catabolism, which are caused by a deficiency in the enzyme propionyl-CoA carboxylase or the enzyme methylmalonyl-CoA (MM-CoA) mutase, respectively. The functional consequence of PA or MMA is the inability to catabolize P-CoA to MM-CoA or MM-CoA to succinyl-CoA, resulting in the accumulation of P-CoA and other metabolic intermediates, such as propionylcarnitine (C3), 3-hydroxypropionic acid, methylcitric acid (MCA), and methylmalonic acid (only in MMA). P-CoA and its metabolic intermediates, at high concentrations found in PA and MMA, inhibit enzymes in the first steps of the urea cycle as well as enzymes in the tricarboxylic acid (TCA) cycle, causing a reduction in mitochondrial energy production. We previously showed that metabolic defects of PA could be recapitulated using PA patient-derived primary hepatocytes in a novel organotypic system. Here, we sought to investigate whether treatment of normal human primary hepatocytes with propionate would recapitulate some of the biochemical features of PA and MMA in the same platform. We found that high levels of propionate resulted in high levels of intracellular P-CoA in normal hepatocytes. Analysis of TCA cycle intermediates by GC–MS/MS indicated that propionate may inhibit enzymes of the TCA cycle as shown in PA, but is also incorporated in the TCA cycle, which does not occur in PA. To better recapitulate the disease phenotype, we obtained hepatocytes derived from livers of PA and MMA patients. We characterized the PA and MMA donors by measuring key proximal biomarkers, including P-CoA, MM-CoA, as well as clinical biomarkers propionylcarnitine-to-acetylcarnitine ratios (C3/C2), MCA, and methylmalonic acid. Additionally, we used isotopically-labeled amino acids to investigate the contribution of relevant amino acids to production of P-CoA in models of metabolic stability or acute metabolic crisis. As observed clinically, we demonstrated that the isoleucine and valine catabolism pathways are the greatest sources of P-CoA in PA and MMA donor cells and that each donor showed differential sensitivity to isoleucine and valine. We also studied the effects of disodium citrate, an anaplerotic therapy, which resulted in a significant increase in the absolute concentration of TCA cycle intermediates, which is in agreement with the benefit observed clinically. Our human cell-based PA and MMA disease models can inform preclinical drug discovery and development where mouse models of these diseases are inaccurate, particularly in well-described species differences in branched-chain amino acid catabolism.

376 Establishment of a novel in vivo mouse model of the IL-17 signaling pathway suitable for explorations of PK/PD relationships

The IL-17 signaling pathway is a key driver of psoriasis and a validated target for treatment of the disease. The purpose of the present study was to establish a mechanistic in vivo mouse model of IL-17 signaling, suitable for modelling of PK/PD relationships for new psoriasis therapies inhibiting this pathway. To obtain an IL-17 induced response in skin, which is the relevant tissue for psoriasis translations, an intradermal cytokine injection model was selected, with proximal biomarkers of clinical relevance as the primary endpoints. The biomarkers were measured in ear tissue lysate 0.5-24 hours after cytokine injection, analyzing both mRNA and protein levels. When IL-17A was injected intradermally in one ear of BalbC mice, a clear upregulation of several proximal biomarkers was seen, including CXCL1 and CCL20. However, the response was only evident at very high concentrations of IL-17A. To obtain a response with lower, clinically relevant IL-17 levels, TNFa - which is known to act in synergy with IL-17 - was co-injected with IL-17A. TNFa significantly potentiated the response of IL-17A, resulting in robust upregulation of proximal biomarkers at IL-17A concentrations that were within the concentration range reported in skin samples from psoriasis patients. The upregulated biomarkers included CXCL1 and CCL20, with the peak response observed 1-4 hours after the intradermal cytokine injection. To confirm that the biomarker response was dependent on IL-17A, we determined the therapeutic effect of an anti-IL17A antibody. Indeed, antibody doses ranging from 1 to 10 mg/kg inhibited the biomarker response. In conclusion, we have established a mechanistic mouse model of the IL-17 signaling axis which enables in-depth investigations of the PK/PD relationships of therapeutic interventions targeting this signaling pathway.

In Search of Biomarkers for Autism Spectrum Disorder.

Autism Spectrum Disorder (ASD) lacks validated measures of core social functions across development stages suitable for clinical trials. We assessed the concurrent validity between ASD clinical measures and putative biomarkers of core deficits, and their feasibility of implementation in human studies. Datasets from two adult ASD studies were combined (observational study [n = 19] and interventional study baseline data [n = 19]). Potential biomarkers included eye‐tracking, olfaction, and auditory and visual emotion recognition assessed via the Affective Speech Recognition test (ASR) and Reading‐the‐Mind‐in‐the‐Eyes Test (RMET). Current functioning was assessed with intelligence quotient (IQ), adaptive skill testing, and behavioral ratings. Autism severity was determined by the Autism Diagnostic Observation Scale‐2 and Social Communication Interaction Test (SCIT). Exploratory measures showed varying significant associations across ASD severity, adaptive skills, and behavior. Eye tracking endpoints showed little relationship to adaptive ability but correlated with severity and behavior. ASR scores significantly correlated with most adaptive behavior domains, as well as severity. Olfaction predicted visual and auditory emotion recognition. SCIT scores related moderately to multiple severity domains, and was the only measure not related with IQ. RMET accuracy was less related to ASD features. Eye tracking, SCIT, and ASR showed high test–retest reliability. We documented associations of proximal biomarkers of social functioning with multiple ASD dimensions. With the exception of SCIT, most correlations were modest, limiting utility as proxy measures of social communication. Feasibility and reliability were high for eye‐tracking, ASR, and SCIT. Overall, several novel experimental paradigms showed potential as social biomarkers or surrogate markers in ASD. Autism Research 2018, 11: 1567–1579. © 2018 The Authors. Autism Research published by International Society for Autism Research and Wiley Periodicals, Inc.Lay SummaryMore accurate measurements of treatment effects are needed to help the development of new drug treatments for autism spectrum disorders (ASD). This study evaluates the relationship between assessments designed to measure behaviors associated with social communication and cognition in ASD with clinical and diagnostic assessments of symptom severity as well as their implementation. The assessments including eye‐tracking, auditory and visual social stimuli recognition, and olfaction identification showed potential for use in the evaluation of treatments for social difficulties in ASD.

Demonstrating In-Cell Target Engagement Using a Pirin Protein Degradation Probe (CCT367766).

Demonstrating intracellular protein target engagement is an essential step in the development and progression of new chemical probes and potential small molecule therapeutics. However, this can be particularly challenging for poorly studied and noncatalytic proteins, as robust proximal biomarkers are rarely known. To confirm that our recently discovered chemical probe 1 (CCT251236) binds the putative transcription factor regulator pirin in living cells, we developed a heterobifunctional protein degradation probe. Focusing on linker design and physicochemical properties, we generated a highly active probe 16 (CCT367766) in only three iterations, validating our efficient strategy for degradation probe design against nonvalidated protein targets.

Identification of proximal biomarkers of PKC agonism and evaluation of their role in HIV reactivation

DesignThe HIV latent CD4+ T cell reservoir is broadly recognized as a barrier to HIV cure. Induction of HIV expression using protein kinase C (PKC) agonists is one approach under investigation for reactivation of latently infected CD4+ T cells (Beans et al., 2013; Abreu et al., 2014; Jiang et al., 2014; Jiang and Dandekar, 2015). We proposed that an increased understanding of the molecular mechanisms of action of PKC agonists was necessary to inform on biological signaling and pharmacodynamic biomarkers. RNA sequencing (RNA Seq) was applied to identify genes and pathways modulated by PKC agonists. MethodsHuman CD4+ T cells were treated ex vivo with Phorbol 12-myristate 13-acetate, prostatin or ingenol-3-angelate. At 3 h and 24 h post-treatment, cells were harvested and RNA-Seq was performed on RNA isolated from cell lysates. The genes differentially expressed across the PKC agonists were validated by quantitative RT-PCR (qPCR). A subset of genes was evaluated for their role in HIV reactivation using siRNA and CRISPR approaches in the Jurkat latency cell model. ResultsTreatment of primary human CD4+ T cells with PKC agonists resulted in alterations in gene expression. qPCR of RNA Seq data confirmed upregulation of 24 genes, including CD69, Egr1, Egr2, Egr3, CSF2, DUSP5, and NR4A1. Gene knockdown of Egr1 and Egr3 resulted in reduced expression and decreased HIV reactivation in response to PKC agonist treatment, indicating a potential role for Egr family members in latency reversal. ConclusionOverall, our results offer new insights into the mechanism of action of PKC agonists, biomarkers of pathway engagement, and the potential role of EGR family in HIV reactivation.

Therapy with atorvastatin versus rosuvastatin reduces urinary podocytes, podocyte-associated molecules, and proximal tubule dysfunction biomarkers in patients with type 2 diabetes mellitus: a pilot study

Background: Diabetic nephropathy is a severe complication of Type 2 diabetes. Tubular lesions may play an important role in its early stages. The aim of our study was to determine if atorvastatin protects the podocytes and the proximal tubule in patients with Type 2 diabetes.Methods: A total of 63 patients with Type 2 diabetes completed this 6-months prospective pilot study. They were randomized to continue rosuvastatin therapy (control group) or to be administered an equipotent dose of atorvastatin (intervention group), and were assessed regarding urinary podocytes, podocyte-associated molecules, and biomarkers of proximal tubule dysfunction.Results: The patients from the intervention group presented a significant reduction in podocyturia (from 7.0 to 4.0 cells/ml, p < .05), urinary nephrin (from 1.7 to 1.3 mg/g, p < .001), urinary vascular endothelial growth factor (from 262.8 to 256.9, p < .01), urinary alpha1-microglobulin (from 10.0 to 8.3 mg/g, p < .01), urinary kidney injury molecule-1 (from 139.5 to 136.3 ng/g, p < .001), and urinary advanced glycation end-products (from 112.6 to 101.3 pg/ml, p < .001). Podocyturia correlated directly with the podocyte damage biomarkers, proximal tubule dysfunction biomarkers, albumin to creatinine ratio, and advanced glycation end-products, and inversely with the glomerular filtration rate.Conclusions: In patients with Type 2 diabetes, atorvastatin exerts favorable effects on the kidney. There is a correlation between the evolution of the podocytes and of the proximal tubule biomarkers, supporting the hypothesis that the glomerular changes parallel proximal tubule dysfunction in the early stages of diabetic nephropathy.

P-A3 Identification of proximal biomarkers of PKC agonist activity in HIV-1 latently infected cells

A small population of resting memory CD4+ T cells harboring transcriptionally silent, but replication competent human immunodeficiency-1 (HIV-1) proviruses is considered to be the major obstacle for HIV eradication in patients on anti-retroviral therapy (ART). Latent virus reactivation using protein kinase C (PKC) agonists is one approach the field is leveraging to flush HIV from latent reservoirs. Thus, increased understanding of the molecular mechanisms of action of this approach is necessary to inform of biological signaling as well as potential off-target effects associated with PKC agonists. We used an RNA-Seq- based approach to identify genes and pathways modulated following treatment of CD4+ T cells with PKC agonists. Primary human CD4+ T cells were treated with either PMA, Prostatin or Ingenol-3-Angelate for various time periods. At 3 and 24 hours post-treatment, cells were harvested and RNA-Seq was performed. Treatment with the three PKC agonists induced a time-dependent alteration in host cell gene expression pattern. Bioinformatic analysis revealed high similarity in the genes and pathways altered, with at least 42% of genes showing similar changes in expression between controls and the three compounds tested. Immune response pathways strongly associate with the gene expression changes observed at 3 hours post-treatment, while pathways associated with mitochondrial dysfunction and oxidative phosphorylation predominated at 24 hours. Quantitative reverse transcriptase PCR (qRT-PCR) validation of a subset of the RNA Seq data confirmed modulation of genes with higher expression levels across the 3 PKC agonists in HIV-1 latently infected Jurkat cells. Overall, our results could offer new insights into the mechanism of action of PKC agonists in latently infected cells, and therefore inform on eradication strategies for latent HIV.

Abstract B48: Identification of potent, cell active MTH1 inhibitors and their use in target validation studies

Abstract MTH1 is a protein that sanitizes oxidized dNTPs in the cell. It preferentially hydrolyzes 8-oxo-dGTP and 2-OH-dATP to their corresponding monophosphates and thereby prevents the incorporation of oxidized nucleotides into DNA or RNA. The functional result is a reduction in down-stream mutations, or DNA damage, thus preventing cell death. Recent publications suggest that MTH1 is a non-essential enzyme in normal cells, but is required for the survival of cancer cells as a consequence of being subjected to high levels of oxidative stress; hence its relevance as an oncology target. Gad et al., 2014 The interesting target rationale combined with a perceived high chemical tractability of the target resulting from availability of x-ray crystallographic information for the protein, led to a decision to undertake, in parallel, target validation, assay development and drug discovery. This effort resulted in the structure-based design of potent, cell active MTH1 small molecule inhibitors. This poster will describe the discovery and optimization of these inhibitors, and their use to evaluate the potential of MTH1 as an oncology target. This work was complemented by parallel genetic studies. Pharmacodynamic evaluation of target engagement using proximal biomarkers will be presented, as will phenotypic responses across a range of cancer cell lines. Citation Format: Elisabetta Leo, Alessia Petrocchi, Jennifer Bardenhagen, Maria Alimova, Xi Shi, Connor Parker, Naphtali Reyna, Matthew Hamilton, Edward Felix, Andrzej Mazan, Christian Dillon, Faika Mseeh, Joseph R. Marszalek, Carlo Toniatti, Giulio Draetta, Phil Jones, Richard T. Lewis. Identification of potent, cell active MTH1 inhibitors and their use in target validation studies. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B48.

Abstract LB-052: Kinase identification of proximal substrates (KIPS): A novel chemical genetics approach for kinase substrate identification

Abstract Protein kinases are attractive targets for pharmacological intervention through their frequent disease associated aberration of cellular signalling networks. In this context an understanding of the myriad of substrates on which a protein kinase operates can help to reveal potential pharmacodynamic and patient stratification biomarkers. Kinase Identification of Proximal Substrates (KIPS) utilises an innovative chemical genetics approach to identify novel proximal biomarkers of kinase target response. This approach relies on the generation of a desensitized mutant protein that retains its kinase function, but loses sensitivity to small-molecule mediated inhibition. Comparative phospho-proteomics is performed on immuno-complexes of tagged wild type and mutant protein kinases in the presence and absence of inhibitor to isolate specific phospho-proteomic changes in proximal substrates. To exemplify the KIPS approach we have focused on the identification of novel substrates of the atypical protein kinase C isoform, PKCι. Through structure based design we identified two acidic amino acid residues in the PKCι nucleotide binding site that are crucial for efficacy of a previously characterized PKCι inhibitor, CRT0066854. Upon mutation to non-acidic alanine residues the PKCι protein is rendered insensitive to compound-mediated inhibition, allowing a comparison of wild type and mutant responses to PKCι inhibition to be made. Using HCT116 cells, we identified Myosin X as a putative substrate for PKCι, and demonstrated its specificity to the PKCι signalling axis through alanine-scanning peptide arrays and generation of phospho-specific antibodies. We show that the key acidic amino acids utilised in KIPS are highly conserved across the AGC and CAMK kinase super families. Furthermore, extensive compound SAR, diverse kinome profiling and structural biology have identified a tool box of inhibitor compounds with a broad range of activities across these kinase families leading to broader applicability of KIPS to enable the identification of individual kinase specific substrates and response across approximately 20% of the human kinome. Citation Format: jon roffey, Andrew Turnbull, Christian Dillon, Susan Boyd, Philippe Riou, Mark Linch, Peter Parker, Sven Kjaer, Neil McDonald. Kinase identification of proximal substrates (KIPS): A novel chemical genetics approach for kinase substrate identification. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-052. doi:10.1158/1538-7445.AM2015-LB-052

A27 Pharmacodynamic And Proximal Biomarkers For Modulation Of Huntingtin Levels: Preclincial Studies For Translation To The Clinic

HTT suppression is a key therapeutic strategy for HD. Reducing the amount of the disease-causing expanded HTT protein in the brain of patients is predicted to slow the progression of the disease. Several approaches are being employed to lower HTT including antisense oligonucleotides and siRNAs, as well as gene therapy approaches using viral delivery of miRNAs, shRNAs, and zinc-finger repressor proteins. To enable the advancement of such therapeutics to the clinic, translatable HTT lowering pharmacodynamic and proximal biomarkers are being explored in preclinical models of HD. Specifically, efforts to identify both molecular and imaging measures that are sensitive to HTT levels will be presented. One or more of these pharmacodynamics markers will be essential measures to demonstrate that the delivery of a HTT lowering therapy does, in fact, lower the amount of HTT in the brains of HD patients.

DEVELOPING BIOMARKERS IN MOOD DISORDERS RESEARCH THROUGH THE USE OF RAPID-ACTING ANTIDEPRESSANTS

An impediment to progress in mood disorders research is the lack of analytically valid and qualified diagnostic and treatment biomarkers. Consistent with the National Institute of Mental Health (NIMH)'s Research Domain Criteria (RDoC) initiative, the lack of diagnostic biomarkers has precluded us from moving away from a purely subjective (symptom-based) toward a more objective diagnostic system. In addition, treatment response biomarkers in mood disorders would facilitate drug development and move beyond trial-and-error toward more personalized treatments. As such, biomarkers identified early in the pathophysiological process are proximal biomarkers (target engagement), while those occurring later in the disease process are distal (disease pathway components). One strategy to achieve this goal in biomarker development is to increase efforts at the initial phases of biomarker development (i.e. exploration and validation) at single sites with the capability of integrating multimodal approaches across a biological systems level. Subsequently, resultant putative biomarkers could then undergo characterization and surrogacy as these latter phases require multisite collaborative efforts. We have used multimodal approaches - genetics, proteomics/metabolomics, peripheral measures, multimodal neuroimaging, neuropsychopharmacological challenge paradigms and clinical predictors - to explore potential predictor and mediator/moderator biomarkers of the rapid-acting antidepressants ketamine and scopolamine. These exploratory biomarkers may then be used for a priori stratification in larger multisite controlled studies during the validation and characterization phases with the ultimate goal of surrogacy. In sum, the combination of target engagement and well-qualified disease-related measures are crucial to improve our pathophysiological understanding, personalize treatment selection, and expand our armamentarium of novel therapeutics.