BR112017001785B1 - GENETIC MARKERS FOR PREDICTING RESPONSIVITY TO THERAPY WITH HDL-ELEVATING OR HDL-IMITATING AGENT - Google Patents

Abstract

The present invention provides genotyping methods and compositions for selecting patients with cardiovascular disease who will benefit from treatment with an HDL- or HDL-mimicking agent, in particular with a CETP inhibitor/modulator.

Description

[0001] The field of the invention relates to the treatment or prophylaxis of an individual with a cardiovascular disorder.

[0002] Twenty years ago, one-size-fits-all treatment was the approach that led to hugely successful drugs. Currently, with the sequencing of the human genome and advances in molecular profiling technologies, approaches to drug development are opting for a more stratified or personalized approach. These advances increasingly allow the classification of individuals into subpopulations that are at risk for a specific disease, respond to a specific treatment, do not respond to a specific treatment, or are at high risk of an adverse event when treated. In this way, such genetic tests can be used to inform diagnosis, prognosis and treatment selection. Numerous studies have demonstrated a relationship between genotype and response to pharmaceutical therapies. This approach has been widely adopted in recent years, particularly in oncology, where numerous personalized medicine approaches have been successfully developed and have provided great improvements in clinical outcomes.

[0003] In cardiovascular disorders, population stratification by genotype for a specific therapeutic intervention has been limited. One of the objectives of the present invention is to demonstrate that the population suffering from cardiovascular disorders can behave differently and consequently can respond differently to a specific treatment. Lowering LDL is an important therapeutic strategy in the treatment of cardiovascular disease. In fact, statins, which lower LDL, such as Crestor, Lipitor, Pravachol and Zocar, are widely used and are among the most prescribed drugs. For some time it was also generally accepted that increasing HDL could also be therapeutic in cardiovascular diseases. Several HDL-elevating drugs have been developed including: niacin and CETP inhibitors such as torcetrapib, anacetrapib, evacetrapib and dalcetrapib.

[0004] Cholesterylester transfer protein (CETP), also called plasma lipid transfer protein, is a hydrophobic glycoprotein that is synthesized in various tissues, mainly in the liver. CETP promotes the bidirectional transfer of cholesterol esters and triglycerides between all plasma lipoprotein particles. The first evidence of the effect of CETP activity on plasma lipoproteins was provided by observations in people with genetic CETP deficiencies. The first CETP mutation was identified in Japan in 1989 as a cause of markedly elevated HDL-C. Ten mutations associated with CETP deficiency have since been identified in Asians and one in Caucasians. It was found in Japan that 57% of individuals with HDL-C levels > 100 mg/dL have CETP gene mutations. Additionally, 37% of Japanese people with HDL-C levels between 75-100 mg/dL have CETP gene mutations. Subsequently, studies of animals treated with an anti-CETP antibody showed that inhibition of CETP resulted in a substantial increase in HDL-C concentration. Consistent with these observations in CETP-deficient patients and rabbits treated with an anti-CETP antibody, it has since been found that treatment of humans with CETP-inhibiting drugs increases the concentration of HDL cholesterol and apoA-I (the main apolipoprotein in HDLs). Numerous epidemiological studies have linked the effects of variations in CETP activity with the risk of coronary disease, including studies of human mutations (Hirano, KI Yamishita, S. and Matsuzawa Y. (2000) Curr. Opin. Lipido. 11(4) , 389-396).

[0005] Atherosclerosis and its clinical consequences, including coronary heart disease, stroke and peripheral vascular disease, represent an enormous burden on healthcare systems internationally. Drugs that inhibit CETP (CETP inhibitors) have been under development for some time with the expectation that they will be useful in treating or preventing atherosclerosis. Several classes of CETP-inhibiting drugs have been found to raise HDL, lower LDL in humans, and have therapeutic effects in the treatment of atherosclerosis and cardiovascular disease, including dalcetrapib, torcetrapib, anacetrapib, evacetrapib, BAY 60-5521, and others ( Table 1).

[0006] Table 1: Overview of the Main CETP Inhibiting Drugs and Clinical Status

[0007] However, there is evidence that these drugs may not be safe and effective in all patients. The clinical trial for torcetrapib was terminated in Phase III due to the incidence of mortality in patients to whom torcetrapib and atorvastatin were administered concomitantly compared to patients treated with atorvastatin alone. The clinical trial for dalcetrapib was also stopped in Phase III, in this case due to a lack of efficacy relative to statins alone. Additional CETP inhibitors are still being pursued in clinical trials and earlier stage development. Overall, treatment strategies utilizing CETP inhibitors that provide better efficacy while reducing off-target effects would be clinically beneficial. There is a need for biomarkers, methods, and approaches to predict response to CETP inhibitors and assess the risk of adverse events associated with administration of CETP inhibitors.

[0008] CETP inhibitors are useful for the treatment and/or prophylaxis of atherosclerosis, peripheral vascular disease, dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, cardiovascular disorders, angina, ischemia, cardiac ischemia, stroke, heart attack of myocardium, reperfusion injury, restenosis angioplasty, hypertension, and vascular complications of diabetes, obesity or endotoxemia.

[0009] Clinical trials have demonstrated that patient response to drug treatment is often heterogeneous. There is an urgent need to improve drug development, clinical development and the impact of therapeutic drugs for individuals or sub-populations of patients. Single nucleotide polymorphisms (SNPs) can be used to identify patients best suited to therapy with particular pharmaceutical agents (often this is called “pharmacogenomics”). Likewise, SNPs can be used to exclude patients from some type of treatment due to the likelihood of developing toxic side effects of gold and their likelihood of not responding to treatment has increased. Pharmacogenomics can also be used in pharmaceutical research to aid the drug development and selection process. Linder et al, Clinical Chemistry 43:254 (1997); Marshall, Nature Biotechnology 15: 1249 (1997); International Patent Application WO 97/40462, Spectra Biomedical; and Schafer et al, Nature Biotechnology 16:3 (1998).

[0010] The dalcetrapib mortality and morbidity experience (dal-OUTCOMES) was a multicenter, parallel group, placebo-controlled, randomized, double-blind study in stable coronary patients recently hospitalized for acute coronary syndrome (ACS). The study was undertaken to test the hypothesis that CETP inhibition will reduce the risk of recurrent cardiovascular events in patients with recent ACS by increasing HDL-C levels through CETP inhibition. Eligible patients entered a double-blind placebo run for approximately 4 to 6 weeks to allow patients to stabilize and for completion of planned revascularization procedures. At the end of the operating period, eligible patients in stable condition were randomized in a 1:1 ratio to 600 mg dalcetrapib or placebo on top of evidence-based medical care for ACS. Dalcetrapib is a cholesterol ester transfer protein (CETP) inhibitor. It has been shown to induce dose-related decreases in CETP activity and increases in HDL-C levels in several animal species and in humans. Decreasing CETP activity, through different approaches, has demonstrated anti-atherosclerotic effects in several animal models. The experiment was interrupted in May 2012 by the DSMB for reasons of futility. The dal-OUTCOMES study resulted in unexpected observations related to the progression of cardiovascular disease. Despite a marked increase in HDL-c, patients on treatment did not show a significant reduction in cardiovascular events and the study was terminated.

[0011] After completion of the dal-OUTCOMES study, it was hypothesized that a subgroup of study patients were responding differently to dalcetrapib and that dalcetrapib could have a significant therapeutic effect in a subpopulation of patients. A pharmacogenomic study of the dal-OUTCOMES study population was performed to study inter-individual variation in response to dalcetrapib and identify genetic markers to predict therapeutic response to dalcetrapib, or other CETP inhibitors, for patient stratification and treatment selection.

[0012] The present invention provides genotyping methods, reagents and compositions for selecting individuals who may benefit from treatment with an HDL-increasing or HDL-mimicking agent, in particular with a CETP inhibitor/modulator, with particularity that the individuals suffer from cardiovascular disorder. The invention also provides methods of treatment in patients with a cardiovascular disorder comprising genotyping and selecting patients who may benefit from treatment with an HDL-increasing or HDL-mimicking agent, in particular with a CETP inhibitor/modulator. Surprisingly, the pharmacogenomic study of the dal-OUTCOMES patient cohort found single nucleotide polymorphisms (SNPs), genetic markers, associated with an individual's response to dalcetrapib and useful in predicting therapeutic response to HDL boosting or HDL mimicking agent (in particular a CETP inhibitor/modulator) and in the treatment of patients with increased HDL or HDL mimicking agent (in particular CETP inhibitor/modulator).

[0013] The invention further provides methods for treating or preventing a cardiovascular disorder, which comprise administering to an individual in need thereof an amount of an HDL-increasing or HDL-mimicking agent that is effective for treating or preventing the disorder. cardiovascular, in which the subject has a polymorphism at Rs11647778 in the subject's ADCY9 gene. In some embodiments, the individual has a CC genotype at rs11647778. In certain embodiments, the agent that minimizes HDL or mimics HDL is comprised of niacin, fibrates, glitazone, dalcetrapib, anacetrapib, evacetrapib, DEZ-001, ATH-03, DRL-17822 (Dr. Reddy). , DLBS 1449, RVX 208, CSL-112, CER-001 or ApoA1-Milnano.

[0014] The genetic markers detected in the genotyping methods of the invention include: 20 SNPs that occur in the Adenylate Cyclase Type 9 (ADCY9) gene on chromosome 16, rs11647778 and optionally rs1967309, rs12595857, rs2239310, rs11647828, rs804945 2, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs2531971, rs2238448, rs12599911, rs12920508, or rs1333 7675, in particular rs11647778 or rs1967309, which are both in strong joint linkage disequilibrium (r2 = 0.79) and strongly associated with the response to an HDL-raising agent or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0015] Other gene markers of the invention include a SNP in the ADCY9 gene that are in linkage disequilibrium with rs11647778 or rs1967309 or provided a signal of association with clinical events with a P statistic <0.05 and may provide useful surrogate biomarkers of rs11647778 or rs1967309. According to one embodiment, a surrogate biomarker consisting of an inherited SNP in linkage disequilibrium with rs11647778 or rs1967309 is detected, and the genotype of rs11647778 or rs1967309 is inferred.

[0016] The present invention relates to methods of genotyping patients and/or treating patients with an HDL-elevating drug, in particular a CETP inhibitor. According to particular embodiments, the methods comprise evaluating the genotype of a patient at rs11647778. Three genotypes at rs11647778 are predictors of an individual's response to an HDL-raising drug, in particular a CETP inhibitor: CC, CG, and GG. Of these, the CC genotype is associated with an improved therapeutic response in patients treated with an HDL-raising drug, the CG genotype is associated with a partial response, and the GG genotype is associated with a lack of response (non-response). For the purposes of the present invention: patients who carry the CC genotype may benefit from treatment with an HDL-elevating drug; patients who carry the CG genotype can benefit from treatment with an HDL-raising drug and patients who carry the GG genotype cannot benefit from treatment with an HDL-raising drug. Two genotypes at rs11647778, CC and CG indicate a therapeutic response to a CETP inhibitor, in particular dalcetrapib, in patients with cardiovascular disorder. In particular, the CC genotype for rs11647778 is indicative of a greater therapeutic response to a CETP inhibitor, in particular dalcetrapib, in patients with cardiovascular disorders.

[0017] The present invention relates to nucleic acid molecules that contain polymorphisms or genetic variants, protein variants encoded by these nucleic acid molecules, reagents for the detection of polymorphic nucleic acid molecules, and methods of using the nucleic acid molecules. nucleic acid and proteins as well as methods of using reagents for their detection (for example, preparators and probes for use in the genotyping methods of the invention).

[0018] According to one embodiment the invention provides: methods of detecting genetic variants of the invention and detection reagents, such as probes or preparators, for use in these methods.

[0019] Specifically, the invention provides genetic markers associated with therapeutic response to an HDL-increasing agent or HDL mimic, in particular a CETP inhibitor/modulator, and synthetic nucleic acid molecules (including DNA and RNA molecules) that contain the gene variants of the invention. The invention further includes variant proteins encoded by nucleic acid molecules containing such genetic variants, antibodies to the encoded variant proteins, computer-based data storage systems containing the new gene variant or SNP information, methods for detecting these SNPs. in a test sample, methods of identifying individuals who respond therapeutically when administered an HDL-raising agent or HDL mimic, in particular a CETP inhibitor/modulator, based on the presence or absence of one or more of the variants of gene of the invention or in detecting one or more encoded variant products (e.g., variant mRNA transcripts or variant proteins) and methods of treating individuals who are carriers of a cardiovascular disease that possess one or more of the gene variants of the invention.

[0020] Exemplary embodiments of the present invention further provide methods for selecting or formulating a treatment regimen (e.g., methods for determining whether or not to administer an HDL-elevating or HDL-mimicking agent, in particular inhibitory treatment / CETP modulator to an individual).

[0021] Various embodiments of the present invention also provide methods for selecting individuals to whom an HDL-raising agent or HDL mimic (in particular CETP inhibitor/modulator) can be administered therapeutically based on the individual's genotype and methods for selecting individuals for participation in a clinical trial of an HDL or HDL mimicking agent (in particular a CETP inhibitor/modulator) based on subjects' genotypes (e.g., selection of subjects to participate in the trial who are most likely to respond positively and/or exclude subjects from the experiment who are unlikely to respond positively to treatment based on their genotype(s), in particular their genotype is CC at rs11647778, TT at rs2238448, and/or AA at rs1967309, or by selecting subjects who are unlikely to respond positively to participation in a clinical trial of an alternative drug that may benefit them.

[0022] The nucleic acid molecules of the invention can be inserted into an expression vector to produce a variant protein in a host cell. Thus, the present invention also provides a vector comprising a SNP-containing nucleic acid molecule of the invention, genetically modified host cells containing the vector, and methods for expressing a recombinant variant protein using such host cells. According to another specific embodiment, host cells, nucleic acid molecules and/or variant SNP-containing proteins can be used as targets in a method for screening or identifying therapeutic agents that are HDL-increasing agents or HDL mimics. (in particular the CETP inhibitor/modulator).

[0023] Examples of ADCY9 SNPs that can be determined/evaluated in the method as provided in this context for identifying an improved response to dalcetrapib or for treating or preventing a cardiovascular disease in a patient are those in which the mutation results in a change in nucleotide sequence at position 4,062,592, 4,065, 583, 4,059,439 and 4,051,380 (Genomic set GRCh37.p5) also known as single nucleotide polymorphisms with identifiers rs12595857, rs1967309, rs2238448 and rs1164 7778, respectively, as shown in SEQ. ID. NO.2 1, 19 and 21.

[0024] The present invention is based on the identification of genetic polymorphisms that are predictive of an increased similarity of this treatment with an HD-elevating agent - or HDL-mimicking agent, in particular the CETP inhibitor/modulator may benefit patients with cardiovascular disorders.

[0025] Figure 1: SNP rs1967309 is strongly associated with a reduction in cardiovascular events (death from coronary heart disease, resuscitated cardiac arrest, non-fatal myocardial infarction, non-fatal ischemic stroke, unstable angina or unanticipated coronary revascularization) in patients treated with the CETP inhibitor dalcetrapib. The numbers show results from the genome-wide association study of patients from the dalcetrapib arm of the dal-OUTCOMES trial. A shows results in a Manhattan plot with a strong signal in the ADCY9 gene region on chromosome 16. Each point represents a P value for the association of SNPs with minor allele frequencies > 0.05 tested with the proportional hazards model Cox score for the occurrence of cardiovascular events during treatment and adjusted for gender and 5 main components for genetic ancestry. B shows results for single nucleotide polymorphisms in the ADCY9 region along with 6 imputed SNPs with P < 10-6. The x-axis shows the genomic location of the ADCY9 gene on chromosome 16. The left y-axis shows the negative log10 of P values for the comparison between cardiovascular events versus no events weighted for probability of imputation in a logistic regression model. The right y-axis shows the recombination rate. The degree of linkage disequilibrium is presented in a grayscale gradient according to r2 as estimated from the HapMap reference CEU samples.

[0026] Figure 2: Frequency of cardiovascular events (primary dal-OUTCOMES composite event or unanticipated coronary revascularization) by study termination in the dalcetrapib and placebo treatment arms separately and by rs1967309 genotypes in the ADCY9 gene. Event percentages are reported with 95% CI.

[0027] Figure 3: The cumulative incidence of cardiovascular events (primary composite event dal-OUTCOMES or unanticipated coronary revascularization) for the dalcetrapib treatment arm and the placebo arm separately and stratified by the three genotypes at the rs1967309 SNP in the ADCY9 gene ( GG, AG, AA).

[0028] Figure 4: Shows changes in lipid levels according to genotype during 24 months of treatment with dalcetrapib. Panel A. Mean ± SE (mg/dL) of change in lipid values from baseline to 1 month by CNS genotype groups of ADCY9 rs1967309 for the dalcetrapib arm. P values are shown for univariate statistics between change in lipids and genotypes. Panel B. Mean ± 95% CI for absolute LDL cholesterol values during the follow-up period of the dal-Outcomes trial for patients in the dalcetrapib arm. P value for the multivariate mixed regression model.

[0029] Figure 5: Multidimensional scaling (MDS) chart showing the first two dimensions (C1, C2) of 76,854 SNPs for 6297 individuals from the dal-Outcomes genetic study and 83 CEU founder, 186 JPT-CHB and 88 YRI founder a from the 1000 genome data set. There were 436 ethnic outliers removed from the dataset (shown with +), and samples of dal-Outcomes retained for analysis are displayed.

[0030] Figure 6: Plot of genomic variation explained by the first ten components of the principal component analysis of 76,854 SNPs for 6297 individuals from the dal-Outcomes genetic study and 83 CEU founder, 186 JPT-CHB and 88 YRI founder from the set 1000 genome data

[0031] Figure 7: Quantile-quantile (QQ) plot of -log10 observed (P values) against expectation under the null hypothesis for the genome-wide association of SNPs with MAF >0.05. The shaded region is the 95% concentration band formed by calculating the 2.5 and 97.5 percentages of the distribution under the null hypothesis. Points represent ranked P values from the Cox proportional hazards model for time to cardiovascular event occurrence of participants in the dalcetrapib arm and adjusted for gender and 5 principal components for genetic ancestry. The genomic inflation factor observed was 1.01.

[0032] Figure 8: The linkage disequilibrium of 27 SNPs genotyped in the ADCY9 gene showing r2 values, linkage disequilibrium blocks calculated using the confidence interval method are represented in black and the D' statistic is represented in shades in red. Linkage disequilibrium in 5686 Caucasians are from the dal-Outcomes study.

[0033] Figure 9: The linkage disequilibrium of 27 genotypic SNPs in the ADCY9 gene showing r2 values, linkage disequilibrium blocks calculated using the confidence interval method are represented in black and the D' statistic is represented in shades in red. Articulation imbalance in 386 participants in the dal-Plaque-2 study.

[0034] Figure 10: Manhattan plot of association of genetic variants with events (primary compound endpoint) in the dalcetrapib arm of dal-Outcomes, tested using imputed SNP dosage data within a logistic regression framework with software PLINK and adjusted for gender and 5 principal components for ancestral genetics.

[0035] Various features and embodiments of the present invention are set forth in this context, notwithstanding other features of the invention, modifications and equivalents will be apparent to a person skilled in the relevant art, based on the teachings provided. The described invention is not limited to the examples and embodiments provided, several alternative equivalents will be appreciated by those skilled in the art. As used herein, the singular forms "a", "one" and "the" include the plural unless the context clearly indicates otherwise. For example, cell "a" will also include "cells".

[0036] The term "about" when used in connection with a referenced numerical indication means the referenced numerical indication plus or minus up to 10% of that referenced numerical indication. For example, "about 100" means 90 to 110.

[0037] An "allele" is defined as any one or more alternative forms of a given gene. In a diploid cell or organism the members of an allelic pair (i.e. the two alleles of a given gene) occupy corresponding positions (locations) on a pair of homologous chromosomes and if these alleles are genetically identical the cell or organism is said to be "homozygous", but if it is genetically different the cell or organism is said to be "heterozygous" with respect to the particular gene.

[0038] A "gene" is an ordered sequence of nucleotides located at a particular position on a particular chromosome that encodes a specific functional product and may include untranslated and untranscribed sequences in the vicinity of coding regions. These non-coding sequences may contain regulatory sequences necessary for the transcription and translation of the sequence or introns, and others, or may have any function attributed to them beyond the occurrence of the SNP of interest.

[0039] The term “genotyping” refers to the determination of the genetic information that an individual carries in one or more positions in the genome. For example, genotyping may comprise determining which allele or alleles an individual carries for a single SNP or determining which allele or alleles an individual carries for a plurality of SNPs. For example, in rs1967309 the nucleotides may be an A in some individuals and a G in other individuals. Those individuals that have an A in the position have the A allele and those that have a G have the G allele. In a diploid organism the individual will have two copies of the sequence that contains the polymorphic position so that the individual can have an A allele and an allele G or, alternatively, two copies of the A alleles or two copies of the G allele. Those individuals who have two copies of the G allele are homozygous for the G allele, those individuals who have two copies of the A allele are homozygous for the A allele and those individuals who have one copy of each allele are heterozygous. The alleles are often referred to as the A allele, often the major allele, and the B allele, often the minor allele. Genotypes can be AA (homozygous A), BB (homozygous B) or AB (heterozygous). Genotyping methods generally provide for sample identification as AA, BB or AB.

[0040] The term "comprising" is intended to mean that the compositions and methods include the aforementioned elements, but do not exclude others.

[0041] "HDL-raising agent or HDL mimic" refers to compounds that increase HDL levels by any of the following mechanisms: CETP inhibition/modulation, PPAR agonism, LXR agonism, HM74 agonism ( niacin receptor) thyrotropin hormone receptor agonism, ApoA1 catabolism inducers of HDL, compounds that provide at least one of the athero-protective activities of HDL, such as compounds that would increase cellular lipid efflux (cholesterol and/or phospholipids), are endowed with antioxidant and anti-inflammatory activities. In particular, the HDL mimicking agent is comprised of ApoA1 and ApoA1 derivatives (such as apoA1 Milano, ApoA1 Paris) and other analogues, reconstituted HDL containing ApoA1 and/or ApoAII and the appropriate lipids such as phospholipids. ApoE, derivatives, analogues and peptidomimetics of amphipathic lipoproteins. Examples of "HDL or HDL mimicking agent" are comprised of niacin, fibrates, glitazone, dalcetrapib, anacetrapib, evacetrapib, DEZ-001 (formerly known as TA-8995) (Mitsubishi Tanabe Pharma), ATH-03 (Affris) , DRL- 17822 (Dr. Reddy's), DLBS- 1449 (Dexa Medica), RVX-208 (Resverlogix), CSL-112 (Cls Behring), CER-001 (Cerenis), ApoA1-Milnano (Medicine Company). Particular examples of "HDL-elevating agent or HDL-mimicking agent" are comprised of niacin, fibrates, glitazone, dalcetrapib, anacetrapib, evacetrapib, torcetrapib, preferably niacin, fibrates, glitazone, dalcetrapib, anacetrapib or evacetrapib. More particularly, the HDL-raising or mimicking agent is selected from a CETP inhibitor/modulator. Examples of CETP inhibitors/modulators are comprised of dalcetrapib, anacetrapib, evacetrapib, DEZ-001 (formerly known as TA-8995) (Mitsubishi Tanabe Pharma), ATH-03 (Affris), DRL-17822 (Dr. Reddy's), DLBS -1449 Dexa Medica). More particularly, examples of CETP inhibitors/modulators are comprised of dalcetrapib, anacetrapib, evacetrapib and torcetrapib, preferably dalcetrapib, anacetrapib and evacetrapib. More particularly, the HDL-raising or mimicking agent according to the invention refers to a CETP inhibitor/modulator, especially when the CETP inhibitor/modulator is comprised of dalcetrapib.

[0042] The "CETP inhibitor/modulator" refers to a compound that decreases CETP activity (assessed by conventional transfer assays) by inhibiting CETP and/or inducing conformational changes of the CETP polypeptide once bound to the polypeptide CETP. The CETP conformational changes of the CETP polypeptide allow CETP activity to advance between HDL particles and increase their recycling/turnover area by increasing the output of pre-beta nascent HDL formation. Preferably, CETP inhibitor/modulator refers to all compounds that would bind to cysteine 13 of the CETP polypeptide. Most preferably, the "CETP inhibitor/modulator" is selected from 2-(2-ethylbutyl)-cyclohexanecarboxylic Mercapto S-[2-[1-(2-ethylbutyl)cyclohexylcarbonylamino]-phenyl] -phenyl) -amide and/or bis[2-[1-(2-ethylbutyl)cyclohexylcarbonylamino]phenyl disulfide]. Most preferably, "CETP inhibitor/modulator" is comprised of methyl S-[2[1-(2-ethylbutyl)cyclohexylcarbonylamino]-phenyl]2-methylthiopropionate as a prodrug or acid 2-(2- ethylbutyl)-cyclohexanecarboxylic (2-Mercapto-phenyl)-amide as its active metabolite.

[0043] “Anacetrapib” refers to ((4S,5R)-5- [3,5-bis(trifluoromethyl) phenyl]-3-{[4'-fluoro-2'-methoxy-5'- (propan -2-yl)-4-(trifluoromethyl) [1,1'-biphenyl]-2-yl]methyl}-4-methyl -I,3-oxazolidin-2-one) also known as MK 0859, CAS 875446 -37-0 or a compound of formula Anacetrapib, as well as methods of preparation and use of the compound, are described in WO2006/014413, WO2006/014357, WO2007005572.

[0044] “Evacatrapib” refers to Trans-4-({(5S)-5-[{[3,5-bis(trifluoromethyl)phenyl]methyl}(2-methyl-2H-tetrazol-5-yl) acid )amino]-7,9-dimethyl-2,3,4,5-tetrahydro-1H-benzazepin-1-yl}methyl) carboxylic acid also known as LY2484595, CAS1186486-62-3 or a compound of formula (XB) Evacetrapib as well as methods of preparing and using the compound are described in WO2011002696.

[0045] “Torcetrapib” refers to the acid ethyl ester (2R,4S)-4-[(3,5-bistrifluoromethylbenzyl)methoxycarbonylamino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H -quinoline-1-carboxylic acid, also known as CP-529,414, CAS 262352-17-0 or a compound of formula (XC) Torcetrapib as well as methods of preparation and use of the compound are described in WO0017164 or WO0140190.

[0046] “BAY 60-5521” refers to (5S)-5- Quinolinol, 4-cyclohexyl -2-cyclopentyl -3-[(S)-fluoro [4- (trifluoro methyl) phenyl] methyl]-5 ,6,7,8-tetrahydro-7,7-dimethyl-, also known as CAS 893409-49-9 or a compound of formula (XD) BAY 60-5521 as well as methods of preparing and using the compound are described in WO2006063828.

[0047] "Treatment" refers to both therapeutic treatment and prophylactic or preventive measures. Those in need of treatment include those who already have the disorder as well as those in whom the disorder must be prevented or delayed.

[0048] The term "polymorphism" "polymorphic site" or "single nucleotide polymorphism site" (SNP site) or "single nucleotide polymorphism" refers to a location in the sequence of a gene that varies within a population . A polymorphism is understood as the occurrence of two or more forms of a gene, or position within a gene "allele", in a population, in such frequencies that the presence of the rarest of forms cannot be explained by mutation of an isolated form. Preferred polymorphic sites are endowed with at least two alleles. The implication is that polymorphic alleles confer some phenotypic variability to the host. Polymorphism can occur in both the coding and non-coding regions of genes. The polymorphism may occur at a single nucleotide site or may involve an insertion or deletion. The location of such a polymorphism can be identified by its nucleotide position in the gene, on the chromosome, or on the amino acid transcript that is altered by the nucleotide polymorphism. Individual polymorphisms are also assigned as unique identifiers ("Reference SNP", "refSNP" or "rs#") known to those skilled in the art and used, for example, in the Variation Single Nucleotide Polymorphism (dbSNP) Database. Nucleotide Sequences available on the NCBI website.

[0049] The terms "linkage disequilibrium" or "in linkage disequilibrium" or "LD" refer to the non-random association of alleles in a collection of individuals, in other words, it is the preferential segregation of a particular polymorphic form with another polymorphic form in a different chromosomal location more frequently than expected by chance. By contrast, alleles that co-occur at expected frequencies are said to be in "linkage equilibrium".

[0050] The prefix "rs" refers to a SNP in the database found in the NCB1 SNP database http://www.ncbi.nlm.nih.gov/snp/?term. The "rs" numbers are the NCBI form rsSNP ID.

[0051] The term "sample" includes any biological sample taken from a patient or individual that includes a cell, a tissue sample or a bodily fluid. For example, a sample may include a skin sample, a cheek cell sample, saliva, or blood cells. A sample may include, without limitation, a single cell, multiple cells, cell fragments, an aliquot of a body fluid, whole blood, platelets, serum, plasma, red blood cells, white blood cells, endothelial cells, tissue biopsies, fluid and lymphatic fluid. In particular, "sample" refers to blood cells.

[0052] The term "therapeutic agents" refers to agents capable of treating or preventing cardiovascular disorders. As used herein, an agent "capable of treating or preventing cardiovascular disorders" refers to a molecule that is capable of treating and/or preventing a cardiovascular disorder in humans and/or in a cellular or animal model of said disease. cardiovascular.

[0053] An "enhanced response polymorphism", "enhanced response genotype" or "response genotype" as used herein refers to an allelic variant or genotype at one or more polymorphic sites within the ADCY9 gene as described herein (e.g. rs11647778/CC) which predicts that an individual will respond therapeutically and benefit from treatment with an HDL-mimicking or HDL-mimicking agent (which can be measured by a decreased number of cardiovascular events) compared to a variant or polymer allelic or genotype or polymorphism (e.g., rs11647778/CG or rs11647778/GG) that predicts that an individual will respond less to elevation of HDL or administration of HDL-mimicking agent. "Reduced response", "partial response", "non-response" or "lack of therapeutic efficacy" can be measured by a relative increase in the number of cardiovascular events relative to individuals with an "enhanced response genotype". . Alternatively, "improved response", "response" or "therapeutic efficacy" may be measured by a relative decrease in the number of cardiovascular events relative to individuals carrying polymorphisms associated with "non-response" or " partial response" to an elevation of HDL or HDL that mimics the agent. In particular rs11647778/CC, and optionally rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/ AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG, rs8061182/AA and rs2238448/TT are improved response genotypes. More particularly, rs11647778/CC and optionally rs1967309/AA, and rs2238448/TT are comprised of an improved response genotype. More particularly, rs11647778/CC and rs1967309/AA are comprised of an improved response genotype.

[0054] The expression "Cardiovascular events", as used herein, refers to cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke of ischemic origin, hospitalization for unstable angina and coronary revascularization.

[0055] As "oligonucleotides" are used in this context, they are comprised of nucleic acids of variable length or polynucleotides. These oligonucleotides can be useful as probes, preparators and in the manufacture of microarrays (arrays) for the detection and/or amplification of specific nucleic acids. Such DNA or RNA chains can be synthesized by the sequential addition (5'-3' or 3'-5') of activated monomers to a growing chain, which can be attached to an insoluble support. Numerous methods are known in the art for the synthesis of oligonucleotides for subsequent individual use or as part of the insoluble support, for example in matrices (BERNFIELD MR. and ROTTMAN FM. J. Biol. Chem. (1967) 242(18):4134 -43; SULSTON J. et al. PNAS (1968) 60(2):409- 415; GILLAM S. et al. NucleicAcidRes.(1975) 2(5):613-624; BONORA GM. et al. NucleicAcidRes. (1990) 18(11):3155-9; ALBERT TJ. et al. Nucleic Acid Res.(2003) 31(7):e35; GAO X. et al. Biopolymers (2004) 73(5):579-96; and MOORCROFT MJ. et al. Nucleic Acid Res. (2005) 33(8):e75). In general, oligonucleotides are synthesized through the stepwise addition of activated and protected monomers under a variety of conditions depending on the method being used. Subsequently, specific protecting groups can be removed to allow further elongation and subsequently, once the synthesis is complete, all protecting groups can be removed and the oligonucleotides removed from their solid supports for purification of the complete chains, if desired.

[0056] The term "genotype" refers to the genetic constitution of an organism, usually in relation to a gene or a few genes or a region of a gene relevant to a particular context (i.e., the genetic sites responsible for a particular phenotype). In particular, the specific combination of alleles at a certain position in a gene, such as, for example, the genotypes AA, AG or GG which are possible genotypes of the rs1967309 SNP. SNP.

[0057] A "phenotype" is defined as the observable characters of an organism. Tables 2, 3, 4 and 5 show a genotype correlation for ADCY9 SNP with values that represent an indication of response to treatment of cardiovascular disorders with an HDL-raising agent or HDL mimic.

[0058] As used in this context, the term “biomarker” refers to a sequence characteristic of a particular variant allele (polymorphic site, such as a SNP) or common type RO allele. Biomarker also refers to a peptide or epitope encoded by a particular variant or allo of the common type.

[0059] As used in this context, the term "surrogate marker" refers to a genetic variant, including a SNP, that is present in linkage disequilibrium with an improved response genotype of the invention, in particular rs11647778/CC .

[0060] As used in this context, the term “genetic marker” refers to variants of polymorphic sites of a particular gene that are associated with the response to an HDL-increasing agent or HDL mimic, in particular the CETP inhibitor/modulator. In particular, genetic marker as used herein refers to polymorphic site variants in the ADCY9 gene that are associated with response to an HDL-increasing agent or HDL mimic, in particular a CETP inhibitor/modulator.

[0061] Three genotypes in rs11647778 are predictive agents of an individual's response to an HDL-elevating drug, in particular a CETP inhibitor: CC, CG and GG. Of these, the CC genotype is associated with an improved therapeutic response in patients to an HDL-elevating drug, the CG genotype is associated with a partial response, and the GG genotype is associated with a lack of response (non-response). For the purposes of the present invention: patients with the CC genotype can benefit from treatment with an HDL-elevating drug; Patients who carry the CG genotype can benefit from treatment with an HDL-raising drug and patients who carry the GG genotype cannot benefit from treatment with an HDL-raising drug. The CC and CG genotypes at rs11647778 in patients with a cardiovascular disorder indicate a therapeutic response to a CETP inhibitor, in particular dalcetrapib. In particular, the CC genotype at rs11647778 in patients with a cardiovascular disease is indicative of a greater therapeutic response to a CETP inhibitor, in particular dalcetrapib.

[0062] Three genotypes at rs2238448 are predictive of an individual's response to an HDL-elevating drug, in particular a CETP inhibitor: TT, TC and CC. Of these three genotypes, the TT genotype is associated with an improved therapeutic response in patients treated with an HDL-elevating drug, whereas the CC genotype is associated with a lack of response (non-response). Patients who carry the TC genotype may benefit from treatment with an HDL-elevating drug, as this genotype is associated with a partial response. For the purposes of the present invention, patients carrying the TT or CT genotype can benefit from treatment with an HDL-elevating drug and patients carrying the CC genotype cannot benefit from treatment with an HDL-elevating drug. The TT genotype at rs2238448 in patients with cardiovascular disease indicates a greater therapeutic response to a CETP inhibitor, in particular dalcetrapib, compared to other genotypes.

[0063] According to certain methods described in this context, one or more biomarkers are used to identify or select individuals who will benefit from treatment with an HDL-increasing or HDL-mimicking agent, in particular a CETP inhibitor/modulator. A SNP biomarker for use in an invention may predict a therapeutic response (R) to treatment or non-response to treatment (NR). Table 2 shows the genotypes observed in the dal-Outcomes cohort, present at the rs1967309 polymorphic site, which can be used as a biomarker to predict response to dalcetrapib or HDL elevation or HDL mimic agent, in particular to another CETP inhibitor/modulator. . Table 4 shows the genotypes observed in the dal-Outcomes and dal-Plaque-2 cohorts, present at the rs11647778 polymorphic site, which can be used as a biomarker to predict response to dalcetrapib or elevation of HDL or HDL-mimicking agent, in particular to other CETP inhibitors/modulators. Each genotype shown in Table 2, either alone or in combination with genotypes at other polymorphic sites, can be used as a biomarker to predict response to an HDL-raising agent or HDL mimic, in particular to a CETP inhibitor/modulator. ).

[0064] Table 2: genetic markers and predicted response to treatment with HDL-elevating or HDL-mimicking A: Responsive PR: Partially Responsive NR: Not Responsive

[0065] Table 3: Genetic markers and predicted response to treatment with HDL-raising or HDL-mimicking agent A: Responsive PR: Partially Responsive NR: Not Responsive

[0066] Table 4: Genetic markers and predicted response to treatment with HDL-raising or HDL-mimicking agent A: Responsive PR: Partially Responsive NR: Not Responsive

[0067] Table 5: genetic markers and predicted response to treatment with HDL-raising or HDL-mimicking agent PR: Partially Responsive NR: Not Responsive

[0068] Rs11647778, rs1967309 and rs12595857 are located in an intronic (non-coding) region of the ADCY9 gene in a region that is concordant with regulatory activity in the expression of the ADCY9 gene.

[0069] In certain methods described in this context, individuals who will respond therapeutically to treatment with an HDL-increasing agent or HDL mimic, in particular with a CETP inhibitor/modulator, are identified and selected for treatment using the genotyping methods of the invention. In particular, patients who are carriers of rs11647778/CC and optionally one or more of the following improved response genotypes, are selected for treatment in the methods of the invention: rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828 /GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs804945 2/GG, rs8061182/AA, rs2238448/TT . More particularly, patients who are carriers of the rs11647778/CC and optionally rs12595857/GG, rs2238448/TT and/or rs1967309/AA genotypes are selected for treatment in the methods of the invention: More particularly, patients who are carriers of rs11647778/CC genotypes and rs1967309/AA are selected for treatment in the methods of the invention.

[0070] According to another embodiment the invention provides a method for identifying a patient who benefits from an HDL-raising or HDL-mimicking agent, the method comprising determining a genotype of said patient (e.g. example, genotyping) at one or more polymorphic sites in the ADCY9 gene.

[0071] According to another embodiment the invention provides a method for determining the responsiveness of an individual to an HDL or HDL mimicking agent, in particular a CETP inhibitor, wherein the method comprises determining a genotype of said patient (e.g. genotyping) at rs11647778 and optionally at one or more of rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs806 1182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13337675, rs12920508 , rs12599911, rs2531971 or rs2238448, using one or more of the preparers or probes exposed in this context.

[0072] According to another embodiment the invention provides a method for determining an individual's responsiveness to an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor, the method comprising determination of a genotype of said patient (e.g., genotyping) at rs11647778 and optionally at one or more of rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs 8061182, rs111590482, rs4786454, rs2283497, rs2531967 or rs3730119, rs13337675, using one or more of the preparations or probes exposed in this context.

[0073] According to a particular embodiment, the invention provides the method described in this context wherein the polymorphic sites comprise rs11647778 and optionally one or more of the following sites selected from the group consisting of: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13337675, rs1292 0508, rs12599911, rs2531971 or rs2238448, with particularity that the polymorphic site is selected from the group consisting of rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119 and rs13337675, with greater particularity in that the polymorphic site is comprised of rs11647778 and optionally rs1967309 or rs12595857, with greater particularity that the polymorphic site is comprised by rs11647778 and optionally rs1967309, with particularity that the corresponding genotypes comprise CC and AA respectively.

[0074] According to a particular embodiment, the invention provides methods for genotyping one or more polymorphic sites, wherein at least one polymorphic site is comprised of rs11647778 and optionally one or more polymorphic sites selected from the group consisting of: rs1967309 ,rs12595857,rs2239310,rs11647828,rs8049452,rs12935810,rs74702385,rs17136707,rs8061182,rs111590482,rs4786454,rs2283497,rs2 531967, rs3730119, rs13337675, rs12920508, rs12599911, rs2531971 or rs2238448, with particularity that the polymorphic site is optionally selected from the group consisting of rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs 3730119 and rs13337675 with greater particularity in which the optional polymorphic site is comprised of rs1967309 or, rs12595857, with greater particularity in that the optional polymorphic site is comprised of rs1967309.

[0075] According to a particular embodiment, the invention provides a method in which a patient carrying rs11647778/CC and optionally one or more of rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/ GG, rs8061182/AA, or rs2238448/TT benefits of treatment with an HDL- or HDL-mimic raising agent, with particularity that the HDL-raising or HDL-mimic agent is comprised of a CETP inhibitor/modulator, and with greater particularity that the HDL-raising agent is HDL or HDL imitation is comprised of S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbonyl]-amino}-phenyl) ester. According to a particular embodiment, the invention provides the method in which an HDL- or HDL-mimicking agent is administered to said patient.

[0076] According to a particular embodiment, the invention provides a method in which a patient carrying rs11647778/CC and optionally one or more of rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/ GG, rs8061182/AA, or rs2238448/TT is treated with an HDL-raising or HDL-mimicking agent, with particularity that the HDL-raising or HDL-mimicking agent is comprised of a CETP inhibitor/modulator, and with greater particularity that the HDL-raising agent or Mimic HDL is comprised of S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbonyl]-amino}-phenyl) ester.

[0077] According to a particular embodiment, the invention provides a method in which a patient carrying rs11647778/CC and optionally one or more of rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/ GG, rs8061182/AA, or rs2238448/TT is administered with an HDL-raising or HDL-mimicking agent, with particularity that the HDL-raising or HDL-mimicking agent is comprised of a CETP inhibitor/modulator, and with greater particularity that the HDL-raising agent or Mimic HDL is comprised of S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbonyl]-amino}-phenyl) ester.

[0078] According to a particular embodiment, the invention provides a method in which the patient is a carrier of a cardiovascular disease, with particularity in which the cardiovascular disease is selected from the group consisting of atherosclerosis, peripheral vascular disease, dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, angina, ischemia, cardiac ischemia, stroke, myocardial infarction, reperfusion injury, angioplastic restenosis, hypertension, and vascular complications of diabetes, obesity, or endotoxemia in a mammal, more particularly in that cardiovascular disorder is selected from the group consisting of cardiovascular disease, coronary heart disease, coronary artery disease, hypoalphalipoproteinemia, hyperbetalipoproteinemia, hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, hypertriglyceridemia, hyperlipidoproteinemia, peripheral vascular disease, angina, ischemia and stroke of the myocardium.

[0079] According to another embodiment, the invention provides a method of treating or preventing a cardiovascular disorder in a patient in need thereof, the method comprising: (a) selecting a patient who is endowed with an improved response genotype at rs11647778 and optionally at one or more of the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111 590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13337675, rs12920508, rs12599911, rs2531971 or rs2238448; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0080] According to another embodiment, the invention provides a method of treating or preventing a cardiovascular disorder in a patient in need thereof, the method comprising: (a) selecting a patient who is endowed with a response genotype enhanced at rs11647778 and optionally one or more of the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs11 1590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13337675; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0081] According to a particular embodiment, the invention provides a method of treating a cardiovascular disorder in a patient in need thereof, the method comprising: (a) selecting a patient who is endowed with an improved response polymorphism in rs11647778 and optionally rs1967309 and/or rs2238448, in particular wherein the patient is endowed with the CC genotype under rs11647778 AA genotype under rs1967309 and TT genotype under rs2238448; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0082] According to a particular embodiment, the invention provides a method of treating or preventing a cardiovascular disorder in a patient in need thereof, the method comprising: (a) selecting a patient who is endowed with a polymorphism of improved response at rs11647778 and optionally rs1967309, in particular where the patient is endowed with the CC genotype at rs11647778 and the AA 3m genotype at rs1967309; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0083] According to another embodiment, the invention provides a method of treating or preventing a cardiovascular disorder in a patient in need thereof, the method comprising: (a) genotyping a patient under rs11647778 and optionally in one or more from the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13337675, rs12920508, rs12599911, rs2531971 or rs2238448; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0084] According to another embodiment, the invention provides a method of treating or preventing a cardiovascular disorder in a patient in need thereof, the method comprising: (a) genotyping a patient under rs11647778 and optionally one or more of the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2 283497, rs2531967, rs3730119, rs13337675; (b) administering to said patient an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0085] The invention also provides a method of treating or preventing a patient comprising: a. Analyze a patient sample for the presence of the genetic marker rs11647778/CC and optionally one or more genetic markers selected from the group consisting of rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810 /GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG and rs806118 2/AA and b. administering to a patient who is a carrier of one or more of said genetic markers with an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0086] The invention also provides a method of treating a patient comprising: a. analyze a patient sample for the presence of the genetic marker rs11647778/CC and optionally one or more genetic markers selected from the group consisting of rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810 /GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA;, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG and rs80611 82/AA and b. treating a patient who is a carrier of one or more of said genetic markers with an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0087] According to a particular embodiment, the invention provides the method in which a genotype is determined at rs11647778 and optionally one or more sites selected from: rs1967309 and rs12595857.

[0088] According to a particular embodiment, the present invention provides a method of determining the responsiveness of individuals to an HDL-elevating drug comprising: a. Obtain a sample from the individual, wherein the sample comprises genetic material; B. Contact the sample with a reagent, such as soda or preparators, generating a complex between the reagent and a genetic marker selected from Table 10; w. Detect the complex to obtain a set of data associated with the sample and d. Analyze the data set to determine the presence or absence of a genetic marker.

[0089] According to a particular embodiment, the present invention provides a method of determining the responsiveness to an HDL-elevating drug in a patient who is endowed with one or more symptoms of a disease comprising: a. Obtain a sample from the patient, wherein the sample comprises genetic material; B. Contacting the genetic material with a reagent, such as a probe set or primer to generate a complex between the reagent and a genetic marker selected from Table 10; w. detect the complex to obtain a set of data associated with the sample; d. analyze the data set to determine the presence or absence of a genetic marker; and is. identify a patient who carries one or more genetic markers as a candidate to receive an HDL-raising drug.

[0090] The complex between the reagent (such as probes or preparators) and a genetic marker generated in the genotyping methods provided can be generated either by polymerase chain reaction (PCR) or DNA sequencing. Such methods are described in this context and are widely known to those skilled in the art.

[0091] The invention provides reagents (such as probes or preparators) for genotyping a genetic marker selected from rs11647778/CC, rs12595857/GG; rs1967309/AA;rs111590482/AG; rs111590482/GG; rs11647828/GG; rs12935810/GG; rs17136707/GG; rs2239310/GG; rs2283497/AA; rs2531967/AA; rs3730119/AA; rs4786454/AA; rs74702385/GA; rs74702385/AA; rs8049452/GG; rs11647778/CG, rs8061182/AA;rs1967309/GA, rs12595857/AG, rs13337675/AG, rs13337675/GG, rs17136707/AG, rs2239310/AG, rs2283497/CA, rs2531967/ GA, rs3730119/GA, rs4786454/GA, rs8049452/ GA, rs8061182/AG, rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs228 3497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG, rs8061182/AA, rs12935810/GA, rs12935810/AA, rs11647828/AA, rs2531967/GG, rs3730119 /GG, rs2239310/AA, rs12595857/AA, rs111590482/ AA, rs74702385/GG, rs11647778/GG, rs1967309/GG, rs2283497/CC, rs8061182/GG, rs17136707/AA, rs2238448/TT, rs2238448/TC, rs2238448/CC, rs8049452 /AA, rs4786454/GG, rs13337675/AA and rs11647828/AG, preferably rs11647778, in particular a primer or a probe.

[0092] According to a particular embodiment, the primer comprises the DNA strand that is 15 to 30 nucleotides in length and is hybridized under high stringency conditions to a region of chromosome 16 adjacent to rs11647778, rs1967309, rs12595857, rs2239310 , rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13 337675, rs12920508, rs12599911, rs2531971 or rs2238448.

[0093] According to a particular embodiment the primer comprises a strand of DNA that is 15 to 30 nucleotides in length and is hybridized under high stringency conditions to a region of chromosome 16 adjacent to rs11647778.

[0094] According to a particular embodiment, the primer comprises a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 adjacent to rs11647778, rs1967309, rs12595857, rs2239310 , rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs22 38448 or rs13337675.

[0095] According to a particular embodiment, the primer comprises a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 adjacent to rs11647778.

[0096] According to another embodiment, the reagent is comprised of a primer comprising a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778 ,rs1967309,rs12595857,rs2239310,rs11647828,rs8049452,rs12935810,rs74702385,rs17136707,rs8061182,rs111590482,rs4786454,rs2 283497, rs2531967, rs3730119, rs13337675, rs12920508, rs12599911, rs2531971 or rs2238448.

[0097] According to another embodiment, the reagent is comprised of a primer comprising a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778 .

[0098] According to another embodiment, the reagent is comprised of a primer comprising a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778 ,rs1967309,rs12595857,rs2239310,rs11647828,rs8049452,rs12935810,rs74702385,rs17136707,rs8061182,rs111590482,rs4786454,rs2 283497, rs2531967, rs3730119, rs2238448 or rs13337675.

[0099] According to another embodiment, the reagent is comprised of a primer that comprises a DNA chain that is comprised of 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778 .

[0100] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under conditions of high stringency to a region of chromosome 16 overlapping with rs11647778, rs1967309, rs12595857, rs2239310 , rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs13 337675, rs12920508, rs12599911, rs2531971 or rs2238448.

[0101] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778.

[0102] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778, rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730 119, rs2238448 or rs13337675.

[0103] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to a region of chromosome 16 overlapping with rs11647778.

[0104] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to an oligonucleotide selected from SEQ. ID. AT THE. 1 to SEQ. ID. NO 21.

[0105] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to an oligonucleotide that is provided with the SEQ sequence. ID NO 21.

[0106] According to another embodiment, the probe is comprised of a nucleic acid that is 15 to 30 nucleotides in length and that hybridizes under high stringency conditions to an oligonucleotide that is provided with the SEQ sequence. ID NO. 19.

[0107] According to a particular embodiment, the invention provides the method in which the HDL-raising or HDL-mimicking agents are comprised of a CETP inhibitor/modulator, in particular, wherein the HDL-raising or HDL-mimicking agent of HDL is comprised of S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbonyl]-amino}-phenyl) ester.

[0108] According to yet another embodiment, the invention provides a use of the HDL- or HDL-mimicking agent, in particular a CETP inhibitor/modulator, in the manufacture of a medicament for the treatment or prevention of a cardiovascular disorder. in a patient in need thereof, wherein the patient has an improved response genotype at rs11647778 and optionally one or more of the following sites: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs171367 07, rs8061182, rs111590482, rs4786454 , rs2283497, rs2531967, rs3730119, rs2238448, or rs13337675.

[0109] According to a particular embodiment, the invention provides for use as defined in this context, in which the patient is endowed with an improved response polymorphism in rs11647778.

[0110] According to another embodiment, the invention provides an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator for use in the treatment or prevention of a cardiovascular disorder in a patient in need thereof. , wherein the patient carries the genetic marker rs11647778/CC and optionally one or more genetic markers selected from: rs12595857/GG; rs1967309/AA;rs111590482/AG; rs111590482/GG; rs11647828/GG; rs12935810/GG; rs17136707/GG; rs2239310/GG; rs2283497/AA; rs2531967/AA; rs3730119/AA; rs4786454/AA; rs74702385/GA; rs74702385/AA; rs8049452/GG; rs8061182/AA;rs1967309/GA, rs12595857/AG, rs13337675/AG, rs13337675/GG, rs17136707/AG, rs2239310/AG, rs2283497/CA, rs2531967/GA, rs3730119/GA , rs4786454/GA, rs8049452/GA, rs8061182/ AG, rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs253 1967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG, rs8061182/AA, rs12935810/GA, rs12935810/AA, rs11647828/AA, rs2531967/GG, rs3730119/GG, rs2239310 /AA, rs12595857/AA, rs111590482/AA, rs74702385/ GG, rs1967309/GG, rs2283497/CC, rs8061182/GG, rs17136707/AA, rs8049452/AA, rs4786454/GG, rs13337675/AA, rs2238448/TT, and rs11647828/AG.

[0111] According to another embodiment, the invention provides methods for treating or preventing a cardiovascular disorder, which comprise administering to a patient in need thereof a therapeutically effective amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, wherein the patient is endowed with an improved response genotype at rs11647778 and optionally at one or more of the following sites: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182 , rs111590482, rs4786454, rs2283497, rs2531967, rs3730119 rs2238448, and rs13337675.

[0112] According to another embodiment, the invention provides methods for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof an amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder, in which the patient is endowed with an improved response genotype at rs11647778 and optionally at one or more of the following sites: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs2238448, and rs1333 7675.

[0113] According to another embodiment, the invention provides a method for the treatment or prevention of a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype in rs11647778 and optionally in one or more of the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs478645 4, rs2283497, rs2531967, rs3730119, rs13337675, rs2238448 a therapeutically effective amount of an HDL-raising agent or of HDL imitation, in particular a CETP inhibitor/modulator.

[0114] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype at rs11647778 and optionally at one or more of the following sites : rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061182, rs111590482, rs4786454, rs22 83497, rs2531967, rs3730119, rs13337675, rs2238448 an amount of an HDL-raising or HDL-mimicking agent, in particular an inhibitor / CETP modulator which is effective for the treatment or prevention of cardiovascular disorders.

[0115] According to a particular embodiment, the invention provides a method for treating a cardiovascular disorder, which comprises administering to a patient in need thereof a therapeutically effective amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, wherein treated patients are endowed with an improved response genotype, wherein the genotype is comprised of rs11647778/CC and optionally rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA;, rs4786454/AA, rs74702385/GA, rs7470238 5/AA, rs8049452/GG and/or rs8061182/AA .

[0116] According to a particular embodiment, the invention provides methods for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof an amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder, in which the patient carries the genotype rs11647778/CC and optionally rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828 /GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs80494 52/GG, rs2238448/TT, and/or rs8061182/AA.

[0117] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype in rs11647778/CC and optionally rs12595857/GG, rs1967309 /AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs47 86454/AA, rs74702385/GA, rs74702385/AA , rs8049452/GG and/or rs8061182/AA, a therapeutically effective amount of an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0118] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with the genotype rs11647778/CC and optionally rs12595857/GG, rs1967309/AA, rs111590482/ AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702 385/GA, rs74702385/AA, rs8049452/GG, rs2238448/TT, and/or rs8061182/AA, an amount of an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator which is effective for the treatment or prevention of cardiovascular disorder.

[0119] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof a therapeutically effective amount of an HDL-elevating or HDL-mimicking agent. , in particular a CETP inhibitor/modulator, wherein treated patients are endowed with a treated agent has an improved response genotype under rs11647778 and optionally rs1967309, rs2238448 or rs12595857.

[0120] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof an amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder, in which the patient is endowed with an improved response genotype at rs11647778 and optionally rs1967309, rs2238448, or rs12595857.

[0121] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype at rs11647778 and optionally rs1967309, rs2238448 or rs12595857, a therapeutically effective amount of an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0122] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype at rs11647778 and optionally rs1967309, rs2238448, or rs12595857, an amount of an HDL-raising agent or HDL-mimicking agent, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder.

[0123] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof a therapeutically effective amount of an HDL-elevating or HDL-mimicking agent. , in particular a CETP inhibitor/modulator, wherein treated patients are endowed with an improved response genotype at rs11647778 and optionally rs1967309.

[0124] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient in need thereof an amount of an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder, in which the patient is endowed with an improved response genotype at rs11647778 and optionally rs1967309.

[0125] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype in rs11647778 and optionally rs1967309, a therapeutically effective amount of an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator.

[0126] According to another embodiment, the invention provides a method for treating or preventing a cardiovascular disorder, which comprises administering to a patient who is endowed with an improved response genotype in rs11647778 and optionally rs1967309, an amount of an agent of HDL elevating or HDL mimicking, in particular a CETP inhibitor/modulator, which is effective for the treatment or prevention of cardiovascular disorder.

[0127] According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator for use in the treatment or prevention of a cardiovascular disorder, wherein the treated patient is carrying an improved response genotype rs11647778.

[0128] According to a particular embodiment, the invention provides an HDL-elevating or HDL-mimicking agent as defined in this context wherein the improved response genotype is comprised of CC.

[0129] According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent, in particular a CETP inhibitor/modulator, for use in treating or preventing a cardiovascular disorder in a patient, wherein the patient carries an improved response genotype rs11647778. According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent as defined herein, wherein the improved response genotype at rs11647778 is comprised of CC.

[0130] According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent as described in this context, wherein the cardiovascular disorder is selected from the group consisting of atherosclerosis, peripheral vascular disease , dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, angina, ischemia, cardiac ischemia, stroke, myocardial infarction, reperfusion injury, angioplastic restenosis, hypertension, and vascular complications of diabetes, obesity, or endotoxemia in a mammal .

[0131] According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent as described in this context, wherein the cardiovascular disorder is comprised of cardiovascular disease, coronary disease, coronary artery disease , hypoalphalipoproteinemia, hyperbetalipoproteinemia, hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, hypertriglyceridemia, hyperlipidoproteinemia, peripheral vascular disease, angina, ischemia or myocardial infarction.

[0132] According to a particular embodiment, the invention provides an HDL-raising or HDL-mimicking agent as described in this context, wherein the HDL-raising or HDL-mimicking agent is comprised of a of HDL, particularly a CETP inhibitor/modulator, more particularly it is comprised of S-(2-{[1-(2-ethyl - butyl)-cyclohexane carbonyl]-amino}-phenyl) ester.

[0133] According to another embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbonyl]-amino}-phenyl) ester for the treatment or prevention of a cardiovascular disorder in a patient in need of the same, who has the genotype rs11647778/CC and optionally rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs17136707/GG, rs 2239310/GG, rs2283497 /AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG, rs2238448/TT, or rs8061182/AA, with greater particularity in which the genotype is comprised of rs1164777 8/CC and optionally rs1967309/AA.

[0134] According to another embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester for the treatment or prevention of a patient with cardiovascular disorder, which carries an improved response genotype, in particular wherein the genotype is comprised of rs11647778/CC and optionally rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/GG, rs11647828/GG, rs12935810/GG, rs1713 6707 /GG, rs2239310/GG, rs2283497/AA, rs2531967/AA, rs3730119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG or rs8061182/AA, with greater particularity in which the genotype is understood by rs11647778 /CC and optionally rs1967309/AA.

[0135] According to a particular embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester for the treatment or prevention of a patient with a cardiovascular disorder which carries an improved response genotype, wherein the cardiovascular disorder is selected from the group consisting of atherosclerosis, peripheral vascular disease, dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, angina, ischemia, cardiac ischemia, stroke, myocardial infarction, reperfusion injury, angioplastic restenosis, hypertension, and vascular complications of diabetes, obesity, or endotoxemia in a mammal.

[0136] According to a particular embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester for the treatment or prevention of a cardiovascular disorder in a patient in need of the same who is endowed with an improved response genotype, in which the cardiovascular disorder is comprised of atherosclerosis, peripheral vascular disease, dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, angina, ischemia, cardiac ischemia , stroke, myocardial infarction, reperfusion injury, angioplastic restenosis, hypertension, and a vascular complication of diabetes, obesity, or endotoxemia.

[0137] According to a particular embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester for the treatment or prevention of a patient with a cardiovascular disorder which carries an improved response genotype, wherein the cardiovascular disorder is selected from the group consisting of cardiovascular disease, coronary heart disease, coronary artery disease, hypoalphalipoproteinemia, hyperbetalipoproteinemia, hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, hypertriglyceridemia, hyperlipidoproteinemia, peripheral vascular disease, angina, ischemia, and myocardial infarction.

[0138] According to a particular embodiment, the invention provides S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester for the treatment or prevention of a cardiovascular disorder in a patient in need thereof who is endowed with an improved response genotype, in which the cardiovascular disorder is comprised of cardiovascular disease, coronary heart disease, coronary artery disease, hypoalphalipoproteinemia, hyperbetalipoproteinemia, hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, hypertriglyceridemia, hyperlipidoproteinemia, peripheral vascular disease, angina, ischemia, or myocardial infarction.

[0139] According to another embodiment, the invention provides a method of predicting whether a patient diagnosed with or who is a carrier of one or more symptoms of a cardiovascular disease has an increased chance of benefiting from treatment with a blood-elevating agent. HDL-or HDL-mimicking, in particular a CETP inhibitor/modulator, comprising selecting a sample obtained from said patient for a genetic marker in the Adenylate Cyclase Type 9 gene (ADCY9) selected from rs11647778/CC, rs12595857/ XL 30119/AA, rs4786454/AA, rs74702385/GA, rs74702385/AA, rs8049452/GG, rs2238448/TT, or rs8061182/AA, wherein the presence of the genetic marker indicates that the patient has an increased chance of benefiting from said treatment with an HDL-raising or HDL-mimicking agent . According to a particular embodiment, the selected genetic marker is comprised of rs11647778/CC and optionally rs12595857/GG, rs2238448/TT, and/or rs1967309/AA. More particularly, the selected genetic marker is comprised of rs11647778/CC and optionally rs1967309/AA.

[0140] According to another embodiment, the invention provides a method of predicting whether a cardiovascular disorder patient has an increased likelihood of benefiting from treatment with an HDL-raising or HDL-mimicking agent, in particular an inhibitor/ CETP modulator, which comprises selecting a sample isolated from said patient for a genetic marker in the Adenylate Cyclase Type 9 gene (ADCY9) selected from rs11647778/CC, rs12595857/GG, rs1967309/AA, rs111590482/AG, rs111590482/ XL 385/AA, rs8049452/GG, rs8061182/AA, wherein the patient has an increased possibility of benefiting from said treatment with an HDL- or HDL-mimicking agent. According to a particular embodiment, the selected genetic marker is comprised of rs11647778/CC and is optionally selected from rs12595857/GG; rs1967309/AA. More particularly, the selected genetic marker is comprised of rs11647778/CC and optionally rs1967309/AA.

[0141] According to another embodiment, the invention provides a method for selecting a patient who is likely to react to an HDL- or HDL-mimicking agent, the method comprising: (a) Determining the genotype at rs11647778 in a sample from a patient diagnosed with or who has one or more symptoms of a cardiovascular disorder, and (b) selecting a patient with the CC genotype as being more likely to respond to an HDL-raising or HDL-mimicking agent.

[0142] According to yet another embodiment, the method further comprises determining the genotype at rs1967309 and/or rs2238448, and selecting a patient with AA genotype at rs1967309 and/or TT genotype at rs2238448 as likely to react to a blood-elevating agent. HDL or HDL imitation. According to certain embodiments, the method further comprises administering an HDL-raising or HDL-mimicking agent (e.g., dalcetrapib) to the selected patient.

[0143] According to another embodiment, the invention provides a method for selecting a patient with a cardiovascular disorder as likely to respond to a therapy comprising the HDL- or HDL-mimicking agent, the method comprising: ( a) detect a CC genotype at rs11647778 in a sample originating from the patient, (b) select the patient as most likely to react to a therapy comprising the HDL-raising or HDL-mimicking agent when rs11647778 with CC genotype is detected in a sample from the patient.

[0144] According to yet another embodiment, the invention provides a method of selecting a patient with a cardiovascular disorder as likely to respond to a therapy comprising an HDL-elevating or HDL-mimicking agent, the method comprising: ( a) detect a CC genotype at rs11647778 and AA genotype at rs1967309 in a sample from the patient, (b) select the patient as most likely to respond to a therapy comprising the HDL-raising or HDL-mimicking agent when it is detected rs11647778 with CC and rs1967309 with the AA genotype in a sample from the patient.

[0145] According to a particular embodiment, the invention provides the method described in this context, wherein the presence of a CC genotype at rs11647778 in a reference sample (prior to therapy with an HDL-raising or HDL-mimicking agent according to the invention) indicates that the patient is more likely to respond to therapy with an HDL-raising or HDL-mimicking agent.

[0146] According to a particular embodiment, the invention provides the method as described in this context, wherein the presence of the CC genotype at rs11647778 and AA genotype at rs1967309 in a reference sample indicates that the patient is more susceptible to reacting to therapy with an HDL-raising or HDL-mimicking agent.

[0147] According to a particular embodiment, the invention provides the method described in this context which further comprises c) selecting the therapy comprising an HDL-elevating or HDL-mimicking agent.

[0148] According to a particular embodiment, the invention provides the method described in this context wherein determining the genotype at rs11647778 in a sample from the patient comprises contacting the sample with a genotype-specific reagent (e.g., primer and/or or probe) that binds to the rs11647778 sequence, thereby forming a complex between the reagent and the rs11647778 sequence, and detecting the complex formed, thereby determining the genotype at rs11647778.

[0149] A method for determining the prognosis of a clinical response in a human patient to an HDL-raising or HDL-mimicking agent comprises determining the presence of at least one polymorphism in the patient's ADCY9 gene, wherein the site polymorphism is associated with a delayed, partial suboptimal response or lack of clinical response to said HDL-increasing or mimicking agent wherein at least one polymorphism site is comprised by rs11647778.

[0150] A method for determining the prognosis of a clinical response in a human patient to an HDL-elevating or HDL-mimicking agent comprises determining the presence of at least one polymorphism in the patient's ADCY9 gene, wherein the polymorphism is associated with a delayed, partial suboptimal response or lack of clinical response to said HDL-increasing or HDL-mimicking agent, wherein at least two polymorphism sites are comprised by rs11647778 and rs1967309.

[0151] According to another embodiment, the invention provides a method for determining the prognosis of a clinical response in a human patient to an HDL-elevating or HDL-mimicking agent, comprising determining the genotype of at least one site of polymorphism in the patient's ADCY9 gene, wherein the site of polymorphism is associated with a delayed, partial suboptimal, or no clinical response to said HDL-enhancing or HDL-mimicking agent. According to one embodiment, the polymorphism site is comprised of rs11647778 and the GG genotype is indicative of a delayed, partial suboptimal or no clinical response to said HDL-raising or HDL-mimicking agent. According to another embodiment, the polymorphism site is comprised of rs1967309 and the GG genotype is indicative of a late, partial suboptimal or no clinical response to said GDL-elevating or HDL-mimicking agent. According to another embodiment, the polymorphism site is comprised of rs2238448 and the CC genotype is indicative of a delayed, partial suboptimal or no clinical response to said HDL-raising or HDL-mimicking agent.

[0152] According to a particular embodiment, the invention provides the method described in this context, in which the polymorphism is determined by a genotyping analysis.

[0153] According to a particular embodiment, the invention provides the method described in this context, wherein the genotyping analysis comprises a microarray analysis or a mass spectrometry analysis or the use of specific primers and/or probes. polymorphism, in particular a primer extension reaction.

[0154] According to a particular embodiment, the invention provides the method described in this context, wherein the HDL- or HDL-mimic raising agent is comprised of an HDL-raising agent, in particular a CETP inhibitor/modulator. , with greater particularity S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomyl]-amino}-phenyl) ester.

[0155] According to a particular embodiment, the “CETP inhibitor/modulator” is comprised of S-(2-{[1-(2-ethyl-butyl)-cyclohexane carbomy]-amino}-phenyl) acid ester thioisobutyric acid, also known as-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl]amino)phenyl]2-methylpropane thioate, dalcetrapib or a compound of formula I

[0156] S-[2-([[1-(2-ethylbutyl)cyclohexyl] carbomyl] amino) phenyl] 2-methylpropane thioate has been shown to be an inhibitor of CETP activity in humans (de Grooth et al. , Circulation, 105, 2159-2165 (2002) ) and rabbits (Shinkai et al., J. Med. Chem., 43, 3566-3572 (2000); Kobayashi et al., Atherosclerosis, 162, 131-135 (2002) ); and Okamoto et al., Nature, 406 (13), 203-207 (2000) ). S-[2-([[1-(2-ethylbutyl)cyclohexyl]carbomyl]amino)phenyl]2-methylpropane thioate has been shown to increase plasma HDL cholesterol in humans (from Grooth et al., supra) and in rabbits (Shinkai et al., supra ; Kobayashi et al., supra ; Okamoto et al., supra). Furthermore, S-[2-([[1-(2-ethylbutyl)cyclohexyl]carbomyl]amino)phenyl]2-methylpropane thioate has been shown to lower LDL cholesterol in humans (from Grooth et al., supra) and rabbits. (Okamoto et al., supra). S-[2-([[1-(2-ethylbutyl)cyclohexyl] carbomyl] amino) phenyl] 2-methylpropane thioate, as well as the methods of preparation and use of the compound, are described in EP1020439, Shinkai et al., J. Med. Chem. 43:3566-3572 (2000) or WO 2007/051714, WO 2008/074677 or WO2011/000793.

[0157] According to a preferred embodiment the CETP inhibitor/modulator (for example, the compound of formula I) is comprised of a solid in crystalline or amorphous form, or more preferably in crystalline form. S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl]amino)phenyl]2-methylpropane thioate is in crystalline form A as described in WO2012/069087. Form A is characterized by a powder X-ray diffraction pattern with peaks at about 7.9°, 8.5°, 11.7°, 12.7°, 17.1°, 18.0°, 18.5°, 20 ,2°, 22.1°, 24.7° ± 0.2°, With particularity for an XRPD peaks observed with a 2Theta diffraction angle of 7.9°, 11.7°, 17.1°, 18, 5° (± 0.2°).

[0158] Other CETP inhibitors known in the art and which are of use in the present invention include: evacetrapib, anacetrapib and torcetrapib, particularly evacetrapib and anacetrapib.

[0159] For this reason, the invention provides a method for treating or preventing a cardiovascular disorder in a mammal, which method comprises administering to a mammal (preferably a mammal in need thereof) a therapeutically effective amount of the pharmaceutical composition . The mammal is preferably comprised of a human being (i.e., a male or female human being). The human being can be of any race (for example, Caucasian or Oriental).

[0160] In particular, cardiovascular disorder is comprised of atherosclerosis, peripheral vascular disease, dyslipidemia, hyperbetalipoproteinemia, hypoalphalipoproteinemia, hypercholesterolemia, hypertriglyceridemia, familial hypercholesterolemia, angina, ischemia, cardiac ischemia, stroke, myocardial infarction, reperfusion injury, restenosis angioplasma, hypertension and vascular complications of diabetes, obesity or endotoxemia. With greater particularity, cardiovascular disorder is understood as cardiovascular disease, coronary disease, coronary artery disease, hypoalphalipoproteinemia, hyperbetalipoproteinemia, hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, hypertriglyceridemia, hyperlipidoproteinemia, peripheral vascular disease, angina, ischemia or myocardial infarction.

[0161] According to certain embodiments of the present invention, individuals or patients are administered between 100 mg to 600 mg of S-[2- ([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl] amino) phenyl] 2- methylpropane thioate. In particular, individuals or patients are administered between 150 mg to 450 mg of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl]amino)phenyl]2-methylpropane thioate. More particularly, individuals or patients are administered between 250 mg and 350 mg of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl] amino) phenyl] 2-methylpropane thioate. In particular, individuals or patients are administered between 250 mg and 350 mg of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl] amino) phenyl] 2-methylpropanethioate.

[0162] According to another embodiment of the present invention, the patient for pediatric use is administered between 25 mg to 300 mg of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl] amino) phenyl ] 2-methylpropane thioate. Particularly for pediatric use, 75 mg to 150 mg of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl] amino) phenyl] 2-methylpropane thioate are administered.

[0163] The CETP inhibitor can be administered to the mammal at any suitable dosage (e.g., to achieve a therapeutically effective amount). For example, a suitable dose of a therapeutically effective amount of S-[2-([[1-(2-ethylbutyl)-cyclohexyl]-carbomyl]amino)phenyl]2-methylpropane thioate for administration to a patient will be between approximately 100 mg to about 1800 mg per day. A desirable dose is preferably about 300 mg to about 900 mg per day. A preferred dose is about 600 mg per day.

Genotyping methods

[0164] Identification of the particular genotype of a DNA sample can be accomplished by any of a number of methods widely known to a person skilled in the art. For example, identification of the polymorphism can be achieved by cloning the allele and sequencing using techniques widely known in the art. Alternatively, gene sequences can be amplified from genomic DNA, for example using PCR, and the product sequenced. Many methods for isolating and analyzing a patient's DNA for a particular genetic marker are known in the art, including polymerase chain reaction (PCR), ligation chain reaction (LCR) or ligation amplification, and amplification methods such as replication. self-sustained sequence. Various non-limiting methods used to analyze a patient's DNA for mutations at a given genetic locus are described below.

[0165] DNA microarray technologies can be used, for example, DNA chip devices and high-density microgrids for high-throughput screening applications and low-density microgrids. Methods for manufacturing microarrays are known in the art and include various inkjet deposition and “microjetting” technologies and processes, in situ or on-chip photolithographic oligonucleotide synthesis processes, and DNA probe targeting processes. electrolytic. DNA microarray hybridization applications have been successfully applied in the areas of gene expression analysis and genotyping for point mutations, single nucleotide polymorphisms (SNPs) and short tandem repeats (STRs). Additional methods include RNA interference microarrays and combinations of microarrays and other methods such as laser capture microdissection (LCM), comparative genomic hybridization (CGH), and chromatin immunoprecipitation (ChiP). See, for example, He et al. (2007) Adv. Exp. Med. Biol. 593: 117-133 and Heller (2002) Annu. Rev. Biomed. Eng. 4: 129-153. Other methods include PCR, xMAP, invasive assay, mass spectrometry, and pyrosequencing (Wang et al., 2007) Microarray Technology and Cancer Gene Profiling Vol. 593 of the Advances in Experimental Medicine and Biology book series, Pub Springer New York).

[0166] Another method of detection is comprised of the hybridization of specific alleles using probes that overlap the polymorphic site and having about 5, or alternatively 10, or alternatively 20, or alternatively 25, or alternatively 30 nucleotides surrounding the polymorphic region. For example, several probes capable of specifically hybridizing to the allelic variant or genetic marker of interest are attached to a solid phase support, e.g., a "chip." Oligonucleotide probes can be attached to a solid support through a variety of processes, including lithography. Mutation detection analysis using these chips comprises oligonucleotides, also called “DNA probe arrays” is described, for example, in Cronin et al. (1996) Human Mutation 7':244.

[0167] In other detection methods, it is necessary to first amplify at least a portion of the gene before identifying the allelic variant. Amplification can be carried out, for example, by means of PCR and/or LCR or by means of other methods that are widely known in the art.

[0168] In some cases, the presence of the specific allele in a patient's DNA can be demonstrated through restriction enzyme analysis. For example, the specific nucleotide polymorphism may result in a nucleotide sequence that comprises a restriction site that is absent from the nucleotide sequence of another allelic variant.

[0169] According to another embodiment, protection of cleavage agents (such as a nuclease, hydroxylamine or osmium tetroxide and with piperidine) may be used to detect incompatible bases in DNA/RNA DNA/RNA, or RNA heteroduplexes / DNA (see, for example, Myers et al. Science 230: 1242). In general, the "mismatched cleavage" technique begins by providing duplexes formed by hybridization of a probe, e.g., RNA or DNA, that is optionally labeled, comprising a nucleotide sequence of the allelic variant of the gene with a sample nucleic acid, obtained from a tissue sample. Double-stranded duplexes are treated with an agent that cleaves single-stranded regions of the duplex such as those formed from base pair mismatches between the control and sample strands. For example, RNA/DNA duplexes can be treated with RNase and DNA/DNA hybrids treated with SI nuclease to enzymatically digest the incompatible regions. Alternatively, either DNA/DNA or RNA/DNA duplexes can be treated with hydroxylamine or osmium tetroxide and with piperidine to digest incompatible regions. After digestion of the mismatched regions, the resulting material is then separated by size on denaturing polyacrylamide gels to determine whether the control and sample nucleic acids are derived from an identical nucleotide sequence or which nucleotides are different. See, for example, Pat. US 6,455,249; Cotton et al. (1988) Proc. Natl. Academic. Know. USA 85: 4397; Saleeba et al. (1992) Meth.Enzymol. 217:286295.

[0170] Changes in electrophoretic mobility can also be used to identify the particular allelic variant. For example, single-strand conformation polymorphism (SSCP) can be used to detect differences in electrophoretic mobility between mutant and common-type nucleic acids (Orita et al (1989) Proc Natl Acad. Sci USA 86: 2766 Cotton (1993 ) Mutat Res. 285: 125-144 and Hayashi (1992) Genet Anal. Tech App. 9: 73-79). The single-stranded DNA fragments of the sample and control nucleic acids are denatured and allowed to renature. The secondary structure of single-stranded nucleic acids varies depending on the sequence: The resulting change in electrophoretic mobility allows detection of a change of even a single base change. DNA fragments can be labeled or detected with labeled probes. The sensitivity of the assay can be increased by using RNA (rather than DNA), where the secondary structure is more sensitive to a change in sequence. In another preferred embodiment, the present method utilizes heteroduplex analysis to separate double-stranded heteroduplex molecules based on changes in electrophoretic mobility (Keen et al. (1991) Trends Genet 7:5).

[0171] The identity of the allelic variant or genetic marker can also be obtained by analyzing the movement of a nucleic acid comprising the polymorphic region in polyacrylamide gels containing a denaturant gradient, which is assayed using gradient gel electrophoresis denaturant (DGGE) (Myers et al. (1985) Nature 313: 495). When DGGE is used as an analysis method, the DNA will be modified to ensure that it does not completely denature, for example, by adding a GC clip of approximately 40 bp of high melting point GC-rich DNA by PCR. According to another embodiment, a temperature gradient is used instead of a denaturing agent gradient to identify differences in the mobility of control and sample DNA (Rosenbaum and Reissner (1987) Biophys Chem. 265: 1275).

[0172] Examples of techniques for detecting differences of at least one nucleotide between 2 nucleic acids include, but are not limited to, selective hybridization of oligonucleotides, selective amplification or selective extension of primers. For example, oligonucleotide probes can be prepared in which the known polymorphic nucleotide is placed centrally (allele-specific probes) and then hybridized to the target DNA under conditions that allow hybridization only if a perfect match is found (Saiki et al. 324: 163); Saiki et al. (1989) Proc. Natl. Academic. Know. USA 86: 6230). These allele-specific oligonucleotide hybridization techniques are used for the detection of nucleotide changes in the polymorphic region of the gene. For example, oligonucleotide probes that have the nucleotide sequence of the specific allelic variant are bound to a hybridization membrane and this membrane is then hybridized with labeled sample nucleic acid. Analysis of the hybridization signal will then reveal the identity of the nucleotides of a sample nucleic acid.

[0173] Alternatively, allele-specific amplification technology that relies on selective PCR amplification can be used in conjunction with the present invention. Oligonucleotides used as primers for specific amplification may carry the allelic variant of interest in the center of the molecule (so that amplification depends on differential hybridization) (Gibbs et al. (1989) Nucl. Acids Res. 17: 2437-2448) or at the extreme 3' end of a primer where, under appropriate conditions, mismatch can prevent or reduce extension of the polymerase (Prossner (1993) Tibtech11: 238 and Newton et al. (1989) Nucl. Acids Res. 17: 2503). This technique is also called "PROBE" for PRobeOligo Base Extension. Furthermore, it may be desirable to introduce a new restriction site into the mutation region to create cleavage-based detection (Gasparini et al (1992) Mol. Cell. Probes 6:1).

[0174] According to another embodiment, the identification of the allelic variant or genetic marker is performed using an oligonucleotide binding assay (OLA), as described, for example, in Pat. 4,998,617 and in Laridegren, U. et al. Science 241: 1077-1080 (1988). The OLA protocol utilizes bioligonucleotide probes that are designed to be capable of hybridizing to adjacent single-stranded sequences of a target. One of the oligonucleotides is linked to a separation tag, e.g. biotinylated, and the other is detectably labeled. If the precise complementary sequence is found in a target molecule, the oligonucleotides will hybridize in such a way that their termini are in contact, and create a binding substrate. Ligation then allows the labeled oligonucleotide to be recovered using avidin, or another biotin ligand. Nickerson, D. A. et al. described a nucleic acid detection assay that combines attributes of PCR and OLA (Nickerson, D.A. et al. (1990) Proc. Nat. Acad. Sci. USA 87: 8923-8927). In this method, PCR is used to achieve exponential amplification of target DNA, which is then detected using OLA. A variation of the OLUm method as described in Tobe et al. (1996) Nucleic Acids Res. 24: 3728 each of the allele-specific primers is labeled with a unique hapten, i.e., digoxigein and florein, and each OLA reaction is detected using hapten-specific antibodies labeled with reporter enzymes.

[0175] The invention provides a method for detecting a single nucleotide polymorphism (SNP) in ADCY9. Since single nucleotide polymorphisms are flanked by regions of invariable sequence, their analysis requires no more than determination of the identity of the single nucleotide variant and it is unnecessary to determine a complete genetic sequence for each patient. Several methods have been developed to facilitate SNP analysis.

[0176] The single base polymorphism can be detected using a specialized exonuclease-resistant nucleotide, as set forth, for example, in Pat. U.S. No. 4,656,127. According to the method, a primer complementary to the allelic sequence immediately 3' to the polymorphic site is allowed to hybridize with a target molecule obtained from a particular animal or human. If the polymorphic site on the target molecule contains a nucleotide that is complementary to the particular exonuclease-resistant nucleotide derivative present, then that derivative will be included at the end of the hybridized primer. Such inclusion makes the primer resistant to exonuclease and thus allows its detection. Once the identity of the exonuclease-resistant derivative of an example is known, a finding that the primer has become resistant to exonucleases reveals that the nucleotide present at the polymorphic site of the target molecule was complementary to that nucleotide derivative used in the reaction. This method has the advantage of not requiring the determination of large amounts of extraneous sequence data.

[0177] A solution-based method can also be used to determine the nucleotide identity of the polymorphic site (WO 91/02087). As previously explained, a primer is used that is complementary to the allelic sequences immediately 3' to a polymorphic site. The method determines the nucleotide identity of this site using labeled dideoxynucleotide derivatives, which, if complementary to the local polymorphic nucleotide, will be included in the primer terminus.

[0178] An alternative method is described in WO 92/15712. This method uses mixtures of labeled terminators and a primer that is complementary to the 3' sequence for a polymorphic site. The labeled terminator that is included is thus determined by, and complementary to, the nucleotide present at the polymorphic site of the target molecule being evaluated. The method is normally comprised of a heterogeneous phase assay, in which the preparation or target molecule is immobilized on a solid phase.

[0179] Many other primer-guided nucleotide inclusion procedures for assaying polymorphic sites in DNA have been described (Komher, JS et al (1989) Nucl. Acids Res. 17: 7779-7784; Sokolov, BP (1990) Nucl. AcidsRes 18: 3671, Syvanen, A.-C, et al (1990) Genomics 8: 684-692, Kuppuswamy, MN et al (1991) Proc. Nat. Acad. Sci. USA 88: 1143-1147; Prezant, TR and others (1992) Hum. Mutat 1: 159-164, Ugozzoli, L. et al (1992) GATA 9: 107-112, Nyren, P. et al (1993) Anal.Biochem. 208: 171 175). These methods all rely on the inclusion of labeled deoxy nucleotides to discriminate between bases at a polymorphic site.

[0180] Furthermore, it will be understood that any of the previously discussed methods for detecting changes in a gene or gene product or polymorphic variants can be used to monitor the course of treatment or therapy.

[0181] The methods described herein can be carried out, for example, using pre-packaged diagnostic kits, such as those described below, comprising at least one probe, preparation nucleic acid or reagent that can be conveniently used for the genotyping, for example, analyzing a genetic marker present in the ADCY9 gene to determine whether an individual has an increased likelihood of benefiting from treatment with an HDL-raising or HDL-mimicking agent including an HDL-raising or HDL-mimicking agent of HDL, in particular a CETP inhibitor/modulator. In particular, genetic markers are as described in this context.

[0182] The preparations or probes of the present invention, as reagents for genotyping genetic markers present in the ADCY9 gene, comprise a synthetic nucleotide sequence that is complementary and hybridizes with a contiguous sequence that reaches the ADCY9 gene, preferably from 12 to 30 nucleotides, adjacent to or spanning one or more SNPs selected from rs11647778, rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs806 1182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs2238448 and rs13337675, preferably rs11647778. According to other aspects, a primer comprises 100 or fewer nucleotides, in certain aspects 12 to 50 nucleotides or 12 to 30 nucleotides. The primer is at least 70% identical to the contiguous sequence or complement of the contiguous nucleotide sequence, preferably at least 80% identical, and more preferably at least 90% identical. A primer or probe of the invention is preferably 15 to 50 nucleotides in length comprising a region of 15 to 20 nucleotides that is complementary to a sequence selected from SEQ. IDENTITY. NO 1 - 21, in particular is complementary to the sequence SEQ. ID No. 1, 19 or 21. The degree of complementarity between a probe or preparer and SEQ. ID. AT THE. 1 - 21 can be 100%, 95%, 90%, 85%, 80% or 75%.

[0183] . Oligonucleotides, including probes and primers, "specific for" a genetic allele or genetic marker bind to the polymorphic region of a gene or bind to the polymorphic region of the gene. For oligonucleotides that are intended to be used as primers for amplification, the primers are adjacent if they are sufficiently close together to be used to produce a polynucleotide comprising the polymorphic region. According to one embodiment, the oligonucleotides are adjacent if they bind within about 12 kb, e.g., less than 1 kb of the polymorphism. Specific oligonucleotides are capable of hybridizing to a sequence and, under appropriate conditions, will not bind to a sequence that differs by a single nucleotide.

[0184] The oligonucleotides of the invention, whether used as probes or primers, can be detectably labeled. Tags can be detected directly, for example for fluorescent markers, or indirectly. Indirect detection may include any detection method known to a person skilled in the art, including biotin-avidin interactions, antibody binding, and the like. Fluorescently labeled oligonucleotides may also contain a quencher molecule. Oligonucleotides can be attached to a surface. In some embodiments, the surface is comprised of silica or glass. In some embodiments, the surface is comprised of a metallic electrode.

[0185] Probes can be used to directly determine the genotype of a sample or can be used simultaneously or subsequent to amplification. The term "probes" includes naturally occurring or recombinant single-stranded or double-stranded nucleic acids or chemically synthesized nucleic acids. They can be marked by nick translation, Klenow filling reaction, PCR or other methods known in the art. The probes of the present invention, their preparation and/or labeling are described in Sambrook et al. (1989) above. A probe may be comprised of a polynucleotide of any length suitable for selective hybridization to a nucleic acid containing a polymorphic region of the invention. The length of the probe used will depend, in part, on the nature of the assay used and the hybridization conditions employed.

[0186] Labeled probes can also be used in conjunction with the amplification of a polymorphism (Holland et al (1991) Proc. Nat. Acad. Sci. USA 88: 7276-7280). US Patent 5,210,015 describes fluorescence-based approaches to provide real-time measurements of amplification products during PCR. Such approaches have employed intercalating dyes (such as ethidium bromide) to indicate the amount of double-stranded DNA present, or have used probes containing fluorescence-quencher pairs (also referred to as the "TaqMan®" approach) in which the probe is cleaved during amplification to release a fluorescent molecule whose concentration is proportional to the amount of double-stranded DNA present. During amplification, the probe is digested by the nuclease activity of a polymerase when hybridized to the target sequence to cause the fluorescent molecule to be separated from the quenching molecule, thus causing the appearance of fluorescence from the reporter molecule. The TaqMan® approach utilizes a probe that contains a pair of reporter molecule quencher molecules that specifically bind to a region of a target polynucleotide that contains the polymorphism.

[0187] The probes can be attached to surfaces for use as "gene chips". These gene chips can be used to detect genetic variations through a series of techniques known to those skilled in the art. According to one technique, oligonucleotides are arranged on a genetic chip to determine the DNA sequence of one by hybridization sequencing, as described in U.S. patents 6,025,136 and 6,018,041. The probes of the invention can also be used for fluorescent detection of a genetic sequence. Such techniques have been described, for example, in U.S. Pat. Us. The probes of the invention can also be used for fluorescent detection of a genetic sequence. Such techniques have been described, for example, in U.S. patents 5,968,740 and 5,858,659. A probe can also be attached to an electrode surface for the electrochemical detection of nucleic acid sequences as described in patent 5,952,172 and by Kelley, S. O. et al. (1999) Nucl. Acids Res. 27: 4830-4837. One or more probes for detecting the SNP of the invention (Table 2, 3, 4, 5 or 10, in particular Table 4) can be attached to a chip and this device used to predict the response to the HDL-lowering or HDL-mimicking agent. HDL, in particular CETP inhibitor/modulator, and select an effective treatment for an individual cardiovascular disease. It is conceivable that probes for detecting the SNP of the invention could be included on a chip with a variety of other probes for uses other than predicting response to an HDL-raising or HDL-mimicking agent, in particular the inhibitor/ CETP modulator.

[0188] Furthermore, synthetic oligonucleotides used as probes or primers can be modified to become more stable. Examples of nucleic acid molecules that are modified include uncharged linkages, such as phosphoramidate, phosphothioate and methylphosphonate DNA analogs (see also U.S. patents 5,176,996, 5,264,564 and 5,256,775). Particles and probes of the invention may include, for example, inter-nucleotide modifications, such as dangling moieties (e.g., polypeptides), intercalators (e.g., acridine, psoralen), chelators, alkylators, and modified linkages (e.g., nucleic acids). alpha-anomeric). Also included are synthetic molecules that mimic nucleotide acid molecules in the ability to bind to a sequence called a hydrogen bond and other chemical interactions, including peptide bonds that replace phosphate bonds in the nucleotide backbone.

[0189] The invention relates to synthetic oligonucleotide molecules, primers and probes that hybridize under high stringency hybridization conditions with naturally occurring oligonucleotides described in this context, genetic markers of the ADCY9 gene. Oligonucleotides can be detected and/or isolated through specific hybridization under conditions of high stringency. "High stringency conditions" are known in the art and allow specific hybridization of a first oligonucleotide to a second oligonucleotide where there is a high degree of complementarity between the first and second oligonucleotides. For the genotyping methods presented in this context, this degree of complementarity is between 80% and 100% and preferably between 90% and 100%.

[0190] The SNPs of the invention can also be detected from pre-existing data, such as whole genomic sequence data present in a database. The invention provides a computer-implemented method of querying genomic data to determine a genotype to predict a patient's response to a CETP inhibitor and treat said patient accordingly, i.e., treating patients with a CETP inhibitor.

[0191] The nucleic acid sample for use in genotyping methods, treatment selection or treatment methods can be obtained from any type of cell or tissue from a patient. For example, a patient's bodily fluid, a sample (e.g., blood) can be obtained using known techniques. Alternatively, nucleic acid tests can be performed on dry samples (e.g., hair or skin). More particularly, a sample nucleic acid for genotyping methods, treatment selection or treatment methods will be obtained from the blood cell type.

[0192] The invention described in this context refers to methods and reagents that are useful for determining and identifying the allele present in the ADCY9 gene at rs11647778 and optionally rs1967309, rs2238448 or rs12595857, or any other genetic variant in linkage disequilibrium with these SNPs as shown in Figures 8 and 9, ranging from position chr16:4049365 to chr16:4077178 (group GRCh37/hg19). In particular, the invention also relates to methods and reagents for determining and identifying the allele present in the ADCY9 gene at rs11647778 and optionally rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs171367 07, rs8061182, rs111590482, rs4786454, rs2283497 , Rs2531967, rs3730119, rs13337675, rs2238448, more particularly at rs11647778 and optionally rs1967309 and/or rs12595857, and more particularly at rs11647778 and optionally rs1967309.

[0193] As set out in this context, the invention also provides treatment selection methods that comprise the detection of one or more genetic markers present in the ADCY9 gene. According to some embodiments, the methods utilize probes or primers that comprise nucleotide sequences that are complementary to a polymorphic region of ADCY9. Accordingly, the invention provides kits comprising probes and primers for carrying out the genotyping methods of the invention.

[0194] According to some embodiments, the invention provides kits that are of use in determining whether a patient with a cardiovascular disorder has an increased likelihood of benefiting from treatment with an HDL-elevating or HDL-mimicking agent, in particular a CETP inhibitor/modulator. These kits contain one or more of the reagents, in particular preparations or probes, described in this context and instructions for their use. By way of example only, the invention also provides utility kits for determining whether a patient with a cardiovascular disorder is more likely to benefit from treatment with S-(2-{[1-(2-ethyl-butyl) thioisobutyric acid ) - cyclohexane carbomy] -amino} -phenyl) ester comprising a first oligonucleotide and a second oligonucleotide specific for a CC polymorphism in the ADCY9 rs11647778 SNP and optionally for an AA polymorphism in the ADCY9 rs1967309 SNP.

[0195] Kits may comprise at least one probe or primer that is capable of specifically hybridizing with the polymorphic region of ADCY9 and instructions for use. Kits may comprise at least one of the nucleic acids previously described in this context. Kits that are useful for amplifying at least a portion of ADCY9 generally comprise two primers, at least one of which is capable of hybridizing to the allelic variant sequence. Such kits are suitable for genotype detection by, for example, fluorescence detection, by electrochemical detection or by other types of detection.

[0196] Still other kits of the invention comprise at least one reagent useful for carrying out the assay. For example, the kit may comprise an enzyme. Alternatively, the kit may comprise a buffer or any other useful reagent.

[0197] Kits may include all or some of the positive controls, negative controls, reagents, preparators, sequencing markers, probes and antibodies described herein to determine the subject's genotype in the ADCY9 polymorphic region.

[0198] The following examples are intended merely to illustrate the practice of the present invention and are not provided by way of limitation. The present invention relates to the following nucleotide and amino acid sequences:

[0199] The sequences provided here are available in the NCBI database and can be retrieved from www.ncbi.nlm.nih.gov/sites/entrez?db=gene; These sequences also refer to annotated and modified sequences. The present invention also provides techniques and methods in which homologous sequences and variants of the concise sequences provided herein are used. Preferably, such "variants" are genetic variants. The ON NCB1 database, the nucleotide sequence that encodes Adenylate Cyclase Type 9 (ACDY9) in homo sapiens, is available. .

[0200] Homo sapiens Adenylate Cyclase Type 9 (ADCY9), RefSeqGene on chromosome 16

[0201] The accession number of NCBI Reference Sequence: NCBI is NG_011434.

[0202] Genomic configuration of Homo sapiens chromosome 16 tig, GRCh37.p10 Primary Assembly

[0203] NCBI Reference Sequence: NCBI accession number is NT_010393.16

[0204] The intronic sequences for the SNPs of the homo sapiens ACDY9 gene that provide the designation "rs", the alleles and the corresponding SEQ ID number designations are shown in Tables 6, 7 and 8. The polymorphisms are identified in bold and next bracket.

[0205] Table 6: ACDY9 SNPs and respective intronic sequenceU 1. NCBI Genome Reference Source Build 37.3

[0206] Table 7: List of genetic variants in the ADCY9 gene on chr16 that provided evidence of association (P < 0.05) with response to dalcetrapib treatment from the GWAS study with reference to the genotyping chip sequence used for the experiment (Illumina OMNI2.5S): Chr: chromosome number; P value: for association with cardiovascular events (primary composite event or unanticipated coronary revascularization) in patients treated with the CETP inhibitor dalcetrapib; 1: Reference sequence from the 1000 Genomes public database, as presented in the ILLUMINA annotation file for the OMNI 2.5S Chip HumanOmni25Exome-8v1_A.csv; 2: Reference sequence from the dbSNP public database version 131 from NCBI, as presented in the ILLUMINA annotation file for the OMNI 2.5S Chip HumanOmni25Exome-8v1_A.csv.

[0207] Table 8: List of additional genetic variants in the ADCY9 gene on chr16: References: a. rs1967309 1. Location r2 and D' values of the 1000 Genomes in public database version 137 of NCBI 2. Reference sequence & HGV Names of dbSNP in public database version 137 of NCBI. Example 1

[0208] The Dal-OUTCOMES trial (NC20971) was a double-blind, randomized, placebo-controlled, parallel-group, multi-center, phase III study to evaluate the safety and efficacy of the CETP inhibitor dalcetrapib in recently hospitalized patients due to an illness Acute Coronary Syndrome (ACS). At the time of interim analysis, the study included 15871 randomized patients assigned to two treatment groups: placebo (7933 patients) and dalcetrapib (600 mg per day, 7938 patients). The study did not show any evidence of reduced event rate in the primary efficacy endpoint in the dalcetrapib arm compared to the placebo arm. Details of the dal-OUTCOMES study can be found in G. Schwartz et al., N. Engl. J. Med.367; 22, 2012.

[0209] dal-Outcomes patients were recruited 4 to 12 weeks after hospitalization for acute coronary syndrome characterized by elevated cardiac biomarkers, with symptoms of acute myocardial ischemia, electrocardiographic ischemic abnormalities that were new or presumed to be new, or loss of viable myocardium in the image. Patients without elevated cardiac biomarkers were eligible to participate if symptoms of acute myocardial ischemia were accompanied by electrocardiographic changes that were new or presumed to be new and by additional evidence of obstructive coronary disease. (Schwartz GG et al., Am Heart J 2009; 158: 896-901 e3). Patients who had myocardial infarction associated with percutaneous coronary intervention were also eligible. All patients were following individualized programs to achieve LDL cholesterol levels of 100 mg per deciliter (2.6 mmol per liter) or lower by statin therapy and diet. No specific statin agent was specified, and patients were not excluded based on LDL cholesterol values. Patients' serum triglyceride levels of 400 mg per deciliter (4.5 mmol per liter) or more were excluded. (Schwartz GG et al., N Engl J Med 2012;367:2089-99). Participants eligible for the dal-Outcomes trial entered a single-blind placebo administration period of approximately 4 to 6 weeks to allow patients to stabilize and complete planned revascularization procedures. At the end of the hospitalization period, eligible patients in stable condition were randomized in a 1:1 ratio to 600 mg dalcetrapib or placebo in addition to evidence-based medical care for acute coronary syndrome. Cardiovascular events were adjudicated by an independent clinical physician. For the pharmacogenomics study, 6338 patients were recruited from April 2008 to July 2010 at 461 sites in 14 countries and provided written informed consent to participate in the genetic study of the dal-OUTCOMES study. DNA was extracted from whole blood. After cleaning the genomic data, samples from 5749 Caucasian patients were used for the genome-wide association study (GWAS).

GENOTYPING

[0210] The Illumina Infinium HumanOmni2.5Exome-8v1_A BeadChip including 2,567,845 genetic variants (Illumina, San Diego, California) was used for GWAS discovery of 5749 Caucasian participants for the dal-OUTCOMES study. The chromosome 16 region including 5 genes upstream and downstream of the adenylate cyclase type 9 (ADCY9) gene (CHR16: 3,400,000-4,600,000) was imputed using IMPUTE2, providing 17,764 variants for analysis with an average completion rate of 99 .76%. A Sequenom panel was used for confirmation of imputed nucleotide polymorphisms (SNPs) in the ADCY9 gene identified in dal-OUTCOMES and were tested in the dal-PLAQUE-2 study, including 20 SNPs with P values <0.05 from GWAS discovery.

[0211] DNA was extracted from 1 ml of whole blood using the QiaSymphony DNA midi kit version 1.1 (Qiagen, Garstilgweg, Switzerland) and quantified using the QuantiFluor™ dsDNA System (Promega, Madison, WI) with an average concentration of 91.5 ng/μL and average yield of 18.3 μg. 97.9% of DNA samples had concentrations above 25 ng/μL. Agarose gel electrophoresis confirmed the high quality of the DNA with no signs of degradation. DNA was normalized to 50 ng/μL.

[0212] Genome-General Association Study (GWAS) - Genomic genotyping was performed using 200 ng of genomic DNA in a GLP environment at the Beaulieu-Saucier Pharmacogenomics Center (Montreal, Canada). The Illumina Infinium HumanOmni2.5Exome-8v1_A BeadChip (Illumina, San Diego, CA) including 2,567,845 genetic markers was used and processed according to the manufacturer's specifications. This chip includes more than 200,000 exome variants identified through the Exome Chip Consortium. The remainder of the BeadChip content is a mix of rare and common SNPs from the 1000 Genome Project designed to be maximally informative for diverse world populations. Each BeadChip was scanned and analyzed using the Illumina iScan Reader. Digitized images were analyzed using GenomeStudio version 2011.1 of Illumina with the GenTrain 2.0 cluster algorithm, using a No-Call threshold of 0.15, without manual cluster adjustment, and using the cluster file Illumina HumanOmni25Exome-8v1_A from the manufacturer. Genotype data files were produced in three batches of comparable size as data became available.

[0213] Sequenom - A single Sequenom panel was designed and validated using HapMap DNA samples. The panel included 27 SNPs in the ADCY9 gene selected according to: a P value < 0.05 in GWAS discovery (n = 15), a P value < 10-6 by imputation (n = 6; Function (n = 5; 1 excluded due to low MAF), or literature (n = 4; 1 failed in production). The Sequenom MassArray Maldi-TOF system (Sequenom, La Jolla, CA) was used with Sequenom's Typer 4.0.22 Software. Each plate was grouped using autocluster and manually modified as needed. Two DNA samples from the Coriell Institute (NA11993 and NA07357) used as controls on each genotyping plate showed 100% agreement across all plate replicates and 100% genotype agreement with expectations for corresponding SNPs in the 1000 genomes and HapMap reference database. 17 SNPs genotyped in the GWAS and Sequenom panel provided an average genotype concordance of 99.95% (min: 99.41, max: 100.00%) and 11 SNPs previously imputed provided an average genotypic agreement with the most likely genotype (imputation probability > 0.80) of 99.75 (min: 98.48, maximum: 99.96).

Genomic quality checks

[0214] Discovery of dal-Outcomes GWAS. Three genotyping files in PLINK format generated by GenomeStudio were combined and transformed into binary PLINK format. GenomeStudio final report files were used to generate gender plots, LRR and BAF plots. PyGenClean (Lemieux Perreault LP et al., Bioinformatics 2013;29:1704-5) and PLINK version 1.07 were used for the quality control (QC) and genetic data cleaning process. The genotyping experiment consisted of 68 plates of DNA samples. On each plate, there were two controls, an alternating Coriell DNA from four pre-selected samples (NA11993, NA11992, NA10860 and NA12763) and an internal DNA control from four pre-selected samples. Coriell sample pair agreement ranged from 0.999807 to 0.999958. The pairwise agreement of the four internal controls ranged from 0.99976 to 0.99995. Comparison of Coriell genotypes with the expectation of data from 1000 genomes provided agreement with a variation of 0.996028 to 0.997783.

[0215] Detailed genetic data cleaning steps are presented in Table 9. Duplicate SNPs were evaluated for concordance, completion rate, allele calling, and MAF. SNPs with different alleles or different MAF were retained. Identical and concordant SNPs were merged. The genotyping completion rate for samples and SNPs was set to 98%. SNP genotyping plate bias (based on the 96-well plates used to dilute DNA samples) was marked but not removed, as the effect of genetic ancestry could not be excluded. Peer Identity by State (IBD) was used to conduct close family relationship checks. All but one pair-member of related pairs and duplicate samples (IBS2 * ratio > 0.80) were marked and removed based on a selection of uncorrelated SNPs (r2 < 0.1). The IBS pairwise matrix was used as a distance metric to identify the critical relationship between samples and isolated samples by multidimensional scaling (MDS). The first two MDS components of each individual were plotted including the HapMap data genotypes CEU, JPT-CHB, and YRI (retaining only founder individuals). Elements isolated from the main Caucasian cluster were marked and removed by the k-nearest neighbor (Figure 5). The scree plot and the accumulated explained variation were calculated using the smartpca program from the EIGENSOFT suite (version 3.0). The options used were a numoutlieriter (maximum number of outlier removal iterations) of 0 (off) and altnormstyle (normalization formula) equal to NO. (Price AL Nat Genet 2006; 38: 904-9) (Figure 6).

Imputation

[0216] The region of chromosome 16 including 5 genes upstream and downstream of ADCY9 (CHR16: 3,401,847 4,598,558; NCBI build GRCh37) was imputed using the programs IMPUTE2 (version 2.3.0) and GTOOL (version 0.7.0) on Linux with 1096 genotyped SNPs and 5749 white samples. (Howie BN et al., PLoS Genet 2009; 5: e1000529). Strand alignment was resolved automatically by casting non-A/T and C/G SNPs according to position. Ambiguous A/T and C/G SNPs were considered absent and were imputed. The Impute2 reference package ALL_1000G_fase1integrated_v3_chr16_impute was used for imputation. The inventors used a cutoff of 0.80 for the imputed genotype probabilities per genotype and per individual and a completion rate of 98% or greater. 17,764 variants remained for analysis with an average completion rate of 99.76%.

[0217] From a total of 1,223,798 common SNPs analyzed, a single region with genome-wide significance was found to be associated with cardiovascular events by Cox proportional hazards modeling in the dalcetrapib arm, identifying a SNPs in the ADCY9 gene on the chromosome 16: rs1967309 (P = 2.41 x 10-8) (Figure 1A). The genetic variant at this position has a lower random frequency of 0.41 and the additive genetic effect of one allele (A) has an HR = 0.65 (95% CI: 0.56, 0.76) for cardiovascular events in the arm of dalcetrapib (Table 12a). Neighboring SNPs in linkage disequilibrium with rs1967309 and which provided lower but supportive evidence for association were located in a common recombination block (Figure 1B). Imputation in this region identified 9 additional SNPs in the ADCY9 gene with P < 10 5, including 6 with P < 10-6 (Figure 10). None of the 835,255 rare SNPs analyzed in 36,268 SKAT gene sets passed the significance threshold for follow-up.

[0218] There was no detectable genetic effect for rs1967309 in the placebo group alone (Cox proportional hazards, HR = 0.92; P = 0.25) (Table 12b). The gene-by-treatment arm interaction term was indicative of a statistical interaction (P = 0.0014; beta: -0.340). Stratification by genotypes in the dalcetrapib arm shows that homozygotes for the minor allele (AA) and heterozygotes (AG) at rs1967309 have, respectively, HR = 0.40 (95% CI 0.28, 0.57) and HR = 0.68 (95% CI: 0.55; Cardiovascular when compared to the GG reference homozygotes (Figure 3). Considering only patients with the homozygous AA genotype at rs1967309 (n = 961), there was a reduction of 39% on the pre-specified composite endpoint of coronary death, resuscitated cardiac arrest, non-fatal myocardial failure, non-fatal ischemic stroke, unstable angina or unanticipated coronary revascularization with dalcetrapib compared with placebo (HR = 0.61; 95% CI: 0. 41; 0.92). Similar results were observed when coronary revascularizations were removed from the primary outcome (HR = 0.60; 95% CI: 0.35; 1.02). When considering only patients with the GG genotype Homozygous at rs1967309 (n = 1984), there was a 27% increase in cardiovascular events for dalcetrapib treatment versus placebo (HR = 1.27, 95% CI 1.02, 1.58). Heterozygous carriers showed an intermediate response (n = 2796, HR = 0.94, 95% CI 0.77, 1.16). The results were confirmed by genotyping 27 SNPs in the ADCY9 gene using Sequenom technology (Table 10).

[0219] A total of eight polymorphisms (genotyped or imputed) were associated with P values less than 10-6 in the discovery cohort of 5749 patients from the OUTCOMES study pharmacogenomics study, with the pre-specified primary endpoint of death from coronary disease, resuscitated cardiac arrest, non-fatal myocardial infarction, non-fatal ischemic stroke, unstable angina objective evidence of ischemia or coronary revascularization. In this genome-wide discovery analysis, two of these eight SNPs in the ADCY9 gene were associated with P values less than 5 x 10-8. For the polymorphic nucleotide rs1967309, patients with the AA genotype (minor or least common allele) benefited from a 39% reduction in cardiovascular events when treated with dalcetrapib compared with placebo, while those with the GG genotype (major allele ) were exposed to a 27% increased risk. Heterozygous patients with the AG genotype had an intermediate response. These results were observed only with dalcetrapib, not in the placebo group, and there were statistical interactions between the polymorphisms and study arm (dalcetrapib versus placebo) in association with clinical events, thus strengthening the link with the ADCY9 gene. Hazard ratios were similar when coronary revascularizations were removed from the analysis, which provided a direct comparison with the primary composite endpoint of the dal-OUTCOMES global trial (coronary heart disease, resuscitated cardiac arrest, nonfatal myocardial infarction, nonfatal ischemic stroke or unstable angina). All eight polymorphisms were in strong linkage disequilibrium (r2 > 0.77), which reinforces the postulate that this region of the ADCY9 gene determines clinical responses to dalcetrapib.

PLASMA LIPIDS

[0220] The genotypes of SNPs rs1967309 were significantly associated with changes over time in dalcetrapib-induced lipids in dal-OUTCOMES. Using univariate statistics, genotypes were associated with change in total cholesterol at 1 month (P = 0.0001, P = 0.004 when controlling for gender and genetic ancestry): the GG genotype had a smaller increase (10.0 ± 23.3 mg/dl) (12.9 ± 30.3) and AA (13.8 ± 24.0). This effect was not observed in the placebo arm. A similar result was observed for changes in LDL cholesterol after 1 month of treatment with dalcetrapib: The changes were -2.0 ± 20.0, -1.0 ± 20.0 and + 0.6 ± 21. 0 mg/dl in carriers of GG, AG and AA genotype of rs1967309 respectively (univariate P = 0.003, adjusted P = 0.006). During the dalcetrapib treatment period, GG carriers had slightly lower overall LDL-cholesterol levels (adjusted P = 0.048, repeated measures analysis, Figure 4).

[0221] Table 9. Summary information on genetic data cleaning procedures performed before statistical analysis

[0222] Table 10: Hazard Ratio (HR) over time to events (first occurrence of CHD death, Ml, ACS hospitalization, resuscitated cardiac arrest, atherothrombotic stroke, or unanticipated coronary revascularization) in the dalcetrapib treatment arm versus the placebo arm stratified by genotypes of SNPs identified as associated with cardiovascular events in the ADCY9 gene. Treatment effects of dalcetrapib versus placebo stratified by genotype for validation 20 SNPs genotyped in 5686 patients from the dal-Outcpmes study population genotyped by Sequenom. MAF (smallest random frequency) and r2 were assessed in all available subjects (n = 5686) NR = non-responder; PR = partially responsive; R = responsive:

[0223] Table 11. r2 value among all SNPs based on 5686 from the dal-Outcomes study. Example 2

[0224] The dal-PLAQUE-2 study (Tardif JC et al., Circulation Cardiovascular imaging 2011; 4: 319-33) was a parallel group, phase 3b multicenter, double-blind, randomized, placebo-controlled, effect of dalcetrapib on the progression of atherosclerotic disease in patients with evidence of coronary artery disease and carotid media thickness of at least 0.65 mm in the distal wall of the common carotid arteries, assessed by B-mode ultrasonography. 931 patients were randomized to receive dalcetrapib 600 mg daily or matching placebos with an intended treatment duration of 24 months. However, the trial was prematurely terminated concurrently with the termination of dal OUTCOMES (the latter due to futility). Patients continued study medication until returning for follow-up carotid imaging at 12 months. B-mode carotid ultrasound recordings at baseline, month 6, and month 12 were analyzed centrally at the Montreal Heart Institute central laboratory. Intima-media thickness (IMT) was analyzed in the far wall of common carotid arteries. A 10-mm-long segment of the common carotid artery was analyzed in serial examinations using automated edge detection software (Carotid Analyzer, Medical Imaging Applications, Coralville, Iowa). Among the 411 participants in the dal-PLAQUE-2 trial who consented to the genetic study, 386 had imaging measurements at baseline, 6 months, and 12 months (194 and 192 in the dalcetrapib and placebo groups, respectively). DNA was extracted from whole blood. Research protocols were approved by the relevant institutional review boards or ethics committees, and all human participants gave written informed consent.

[0225] Central analysis of carotid imaging studies in the dal-Plaque-2 assay

[0226] All acquired carotid ultrasound recordings were analyzed in the central laboratory of the Montreal Heart Institute by experienced technicians supervised by a physician. Images were required to meet quality and inclusion and exclusion criteria for eligibility before random assignment to treatment study. Methods for analysis of carotid intimal thickness-mean included side-by-side viewing of baseline and follow-up studies.

[0227] Carotid arterial wall segments were evaluated in a longitudinal view, perpendicular to the ultrasound beam, with a horizontal view of the arterial image to see the interfaces between the blood and vascular structures in the longest possible segment. The location of the end of the common carotid artery was used as a reference in repositioning and analyzing the comparative length of the segments in the baseline and follow-up studies. Intima-media thickness was analyzed in the distant wall of the right and left common carotid arteries to reduce measurement variability. The mean values of the intima-media thickness of the right and left common carotid arteries were used. A corresponding 10-mm-long segment of the common carotid artery was analyzed in serial examinations (baseline, 6 months, and 12 months), starting 5 mm proximal to the carotid bifurcation. Carotid Analyzer edge detection software (Medical Imaging Applications LLC, Coralville, Iowa) was used to analyze intima-media thickness using the automated option. Two sets of carotid artery borders were identified: first, the media - adventitial borders were determined and verified by the reader. When necessary, their locations were semi-automatically modified to the correct location. When these borders were approved, the lumen-intima boundaries were automatically detected using the media-adventitia boundaries for guidance. Frames from loop sequences of intima - media thickness were selected for reading, and poor quality outlier frames were removed from analysis when necessary.

[0228] Of the 27 SNPs selected for genotyping by Sequenom, 20 SNPs in the ADCY9 gene had P < 0.05 in the discovery cohort. Association with SNPs was tested in the dal-PLAQUE-2 trial to obtain supporting evidence for association, based on imaging data obtained after 6 and 12 months of treatment in the dalcetrapib arm (n = 194 ). Ten of 20 SNPs provided association with IMT measures in dal-PLAQUE-2 (P < 0.05, Table 13). Notably, marker rs2238448 (which was in linkage disequilibrium with rs1967309 (r2 = 0.80)) was associated with IMT in dal-PLAQUE-2 (P = 0.009) and with events in dal-OUTCOMES (P = 8.88 x 10-8 HR = 0.67, 95% CI: 0.58, 0.78). After 12 months of dalcetrapib treatment, changes in IMT were -0.027 ± 0.079 mm in homozygous carriers of the minor allele (TT), 0.000 ± 0.048 mm for heterozygotes, and +0.009 ± 0.038 mm for homozygous carriers of the common allele (CC). At rs2238448 (P = 0.009). This effect appeared at 6 months. All 20 SNPs showed a decrease in IMT according to the genotype group that showed reduced cardiovascular risk in the dal-OUTCOMES trial (Table 13). None of the genotyped SNPs showed association in the placebo arm (all P > 0.05). Only the gene-by-treatment interaction term of rs2531967 reached significance (P = 0.024) in dal-PLAQUE-2, likely due to the small sample size. SNP rs1967309 did not reach significance in dal-PLAQUE-2 (P = 0.114), but the reduction in IMT was of similar magnitude to that with rs2238448 and consistent with the findings in dal OUTCOMES. After 12 months of dalcetrapib treatment, changes in IMT were 0.021 ± 0.083 mm in AA homozygotes, -0.001 ± 0.048 mm for heterozygotes, and +0.005 ± 0.042 mm for GG homozygotes at rs1967309.

[0229] Table 13: Change from baseline in carotid intima-media thickness after 6 and 12 months of docetrapib treatment. 20 SNPs with P < 0.05 in dal-OUTCOMES are shown with results for the dakPLAQUE»2 study.

Statistical analyzes for both examples 1 and 2

[0230] Genome-wide association tests were performed using JMP Genomics software version 6.1. Significant results were validated in SAS software (v. 9.3) (SAS Institute Inc., Cary, NC, USA). The 1 degree of freedom additive genetic test was used when genotypes were coded as 0, 1 or 2, according to the minor allele count. In the presence of covariates, the P value for additive genetic testing is adjusted for the covariate such that where r is the probability of having the event and where the null (H0) under additive genetic testing is b1 = 0. A Cox proportional hazards regression model was used to test the gene-by-treatment interaction of according to: log (event rate) ~ genotype + treatment arm + genotype * treatment arm + gender + principal components, using samples from treatment arm and placebo. The GWAS discovery models included adjustment for gender and the 5 principal components for genetic ancestry. MAFs were calculated using samples from both treatment arms. Rare variants (MAF <0.05) were analyzed together with genes using the sequence association kernel test (SKAT) (Wu MC et al., Am J Hum Genet 2011; 89: 82-93). Including covariates and using a beta weighting function with parameters set to 1 and set to 25. Variants, positioned between two genes were analyzed as a distinct set.

[0231] For the dal-Plaque-2 population, mixed regression models were tested for repeated measures analysis in SAS software. The Plaque-2 endpoint tested was the average of the IMT measurements of the common carotid arteries at the 6-month and 12-month visits, using the baseline measurement as a covariate. Adjustment of the significance threshold for multiple testing of the 27 SNPs genotyped in the Sequenom panel was not performed, due to the highly correlated nature of the selected SNPs. As a reference, the number of independent tests (Meff) was estimated using the method of Gao et al, Genet Epidemiol 2008; 32: 361-9. For Meff = 8, using the sample population of dal-Outcomes

[0232] For the linkage disequilibrium plots (Figures 8 and 9), Haploview (version 4.2) (Barrett JC et al., Bioinformatics 2005; 21: 263-5) was used to display the r2 linkage disequilibrium values in a pairwise matrix chart with a color heat chart of D' values. Linkage disequilibrium blocks were constructed using the confidence interval method set to [0.70, 0.98], maximum upper confidence for strong recombination 0.9 and the fraction of strong LD in informative comparisons > 0. 95.

Claims (8)

1. Use of dalcetrapib characterized by the fact that it is in the manufacture of a medicinal product for the treatment or prevention of a cardiovascular disorder in an individual, wherein the individual is known to carry a CC or CG enhanced response genotype in nucleotide polymorphism unique in SEQ ID NO: 21 and optionally carries an improved response genotype, at one or more of the following locations: rs1967309, rs12595857, rs2239310, rs11647828, rs8049452, rs12935810, rs74702385, rs17136707, rs8061 182, rs111590482, rs4786454, rs2283497, rs2531967, rs3730119, rs2238448 and rs13337675. 2. Use according to claim 1, characterized by the fact that the patient has the improved response genotype CC in the single nucleotide polymorphism in SEQ ID NO: 21 and, optionally, has the improved response genotype selected from GG in the single nucleotide polymorphism in SEQ ID NO: 2 AA in the single nucleotide polymorphism in SEQ ID NO: 20; AG in the single nucleotide polymorphism in SEQ ID NO: 10; GG in the single nucleotide polymorphism in SEQ ID NO: 10; GG in the single nucleotide polymorphism in SEQ ID NO: 4; GG in the single nucleotide polymorphism in SEQ ID NO: 6; GG in the single nucleotide polymorphism in SEQ ID NO: 8; GG in the single nucleotide polymorphism in SEQ ID NO: 3; AA in the single nucleotide polymorphism in SEQ ID NO: 12; AA in the single nucleotide polymorphism in SEQ ID NO: 13; AA in the single nucleotide polymorphism in SEQ ID NO: 14; AA in the single nucleotide polymorphism in SEQ ID NO: 11; GA in the single nucleotide polymorphism in SEQ ID NO: 7; AA in the single nucleotide polymorphism in SEQ ID NO: 7; GG in the single nucleotide polymorphism in SEQ ID NO: 5; and AA in the single nucleotide polymorphism in SEQ ID NO: 9. 3. Use according to claim 1, characterized by the fact that the individual is known to carry the enhanced response genotype CC in the single nucleotide polymorphism in SEQ ID NO: 21. 4. Use according to claim 1, characterized by the fact that the individual carries an improved response genotype in rs1967309. 5. Use according to any one of claims 1, 3 and 4, characterized by the fact that the individual carries the improved response genotypes CC in the single nucleotide polymorphism in SEQ ID NO: 21 and AA in the single nucleotide polymorphism in SEQ ID NO: 1. 6. Use according to any one of claims 1, 3 and 4, characterized by the fact that the individual carries the improved response genotypes CC in the single nucleotide polymorphism in SEQ ID NO: 21 and TT in the single nucleotide polymorphism in SEQ ID NO: 19. 7. Use according to any one of claims 1, 3 or 4, characterized by the fact that the individual carries the improved response genotypes CC in the single nucleotide polymorphism in SEQ ID NO: 21, TT in the single nucleotide polymorphism in SEQ ID NO: 19 and AA in the single nucleotide polymorphism in SEQ ID NO: 1. 8. Use according to any one of claims 1 and 3 to 7, characterized by the fact that the cardiovascular disorder is acute coronary syndrome (ACS).