WO2003091698A2 - Cetp genetic markers for statin-specific changes in hdl cholesterol - Google Patents

Abstract

Genetic markers in the CETP gene associated with statin-specific changes in HDL cholesterol after statin treatment are disclosed. Compositions and methods for detecting and using these CETP genetic markers in a variety of clinical applications are disclosed. Such applications include articles of manufacture comprising a statin composition approved for treating patients having one of these CETP haplotypes, methods and kits for predicting the response of an individual to a given statin treatment based on their haplotype profile, and methods for treating individuals with hyperlipidemia based on their haplotype profile.

Description

CETP GENETIC MARKERS FOR STATIN-SPECIFIC CHANGES IN HDL CHOLESTEROL

RELATED APPLICATIONS

This application claims the benefit of US Provisional Application 60/375,791 filed April 26, 2002.

FIELD OF THE INVENTION

This invention relates to the field of genomics and pharmacogenetics. More specifically, this invention relates to variants of the gene for cholesteryl ester transfer protein (CETP) and their use as predictors of response to treatment with statins.

BACKGROUND OF THE INVENTION

Cardiovascular disease is a major health problem in the United States and worldwide (R. H. Knopp, N. Engl. J. Med. 341:498-511, 1999). The major cause of cardiovascular disease is atherosclerosis, which results from the formation of lipid-laden cellular lesions in one or more of the coronary arteries that supply the heart muscle with blood (Leff, T. and Gruber, P.J., "Cardiovascular Diseases" in: Meyers. R. Molecular Biology and Biotechnology (VCH Publishers 1995) pp. 149-153). High levels of low-density lipoprotein cholesterol ("LDLC") have long been associated with an increased risk of developing atherosclerosis (Leff and Gruber, supra). However, it is now widely accepted that high levels of plasma triglycerides ("TG") and low levels of high-density lipoprotein cholesterol ("HDLC") are associated with coronary artery disease as well (Gotto AM, American Journal of Cardiology, 87 (5 Suppl.) 13-18, 2001. Another risk factor for cardiovascular disease is high levels of LDL apolipoprotein B ("LDL Apo B"), which is the major lipoprotein associated with LDLC particles (American Heart Association National Center, New Release NR 96-4430 (Circ/apo B), August 1, 1996). Patients with one or more of the above risk factors are frequently treated with one or more lipid- modifying drugs to achieve certain target levels of LDLC and HDLC that are recommended by the current National Cholesterol Education Program guidelines for treatment of hypercholesterolemia. Usual medical practice is to direct initial drug therapy toward elevated LDLC with treatment of low HDLC a secondary endpoint that is often managed by addition, after some weeks, of a second therapeutic agent. One class of lipid-modifying drugs that are particularly useful for reducing elevated LDLC levels are statins, which inhibit the activity of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, the rate-limiting enzyme for cholesterol formation in the liver and other tissues (Vaughan et al., /. Amer. College Cardiol. 35:1-10, 2000;. Knopp, supra). In addition, in clinical trials of various statin compounds, increases in HDLC levels were observed, with a mean increase of 2% - 12%, depending upon the specific statin compound and the conditions under whichit was studied. While most of the common side effects of statin therapy are mild, transient and reversible (e.g., dyspepsia, abdominal pain and flatulence), more severe, long-term adverse reactions to statins occur and include hepatitis, peripheral neuropathy, insomnia, difficulty in concentrating, and elevation of creatine phosphokinase, which is correlated with rhabdomyolysis (Knopp, supra, Lupattelli G et al., Nucl. Med. Commun. 22(5): 575-8, 2001; Moghadasian MH et al., Expert Opin. Pharmacother. 1(4): 683-95, 2000).

Currently, there are five statins sold in the United States: lovastatin and simvastatin (sold by Merck as Mevacor^® and Zocor^®, respectively); atorvastatin calcium (sold as Lipitor^® by the Parke Davis Division of Pfizer).; fluvastatin sodium (sold as Lescol^® by Novartis); and pravastatin sodium (sold as Pravachol^® by Bristol-Myers Squibb) (Knopp, supra). A sixth statin, cerivastatin sodium, was previously sold as Baycol^® by Bayer, but was voluntarily removed from the market in 2001 because of safety concerns. Five of these drugs are metabolized by cytochrome P-450 enzyme systems, while the sixth, pravastatin sodium, is metabolized by sulfation and possibly other mechanisms (Knopp, supra). Extensive studies have been performed with cerivastatin sodium, atorvastatin calcium, simvastatin, and pravastatin sodium to determine efficacy in the treatment of hypercholesterolemia. In three multicenter, placebo-controlled, dose-response studies of cerivastatin sodium, subjects with primary hypercholesterolemia experienced significantly reduced levels of total-cholesterol, LDLC, apolipoprotein B (apo B), triglycerides, total-cholesterol/HDLC ratio, and LDLC/HDLC ratios following treatment with cerivastatin sodium for an 8-week period. The mean decreases in LDLC and mean increases in HDLC for cerivastatin sodium administered once daily in the evening were 25% and 9% at 0.2 mg/day, 31% and 8% at 0.3 mg/day, and 34% and 7% at 0.4 mg/day (Physicians' Desk Reference, 2000, p. 675). Similarly, in two multicenter, placebo-controlled, dose-response studies, atorvastatin calcium given as a single dose over a six-week period significantly reduced total-cholesterol, LDLC, apo B, and triglycerides. Atorvastatin calcium at 10, 20, 40, and 80 mg/day, resulted in mean LDLC decreases/HDLC increases of 39%/6%, 43%/9%, 50%/6%, and 60%/5%, respectively (Physicians' Desk Reference,.2000, p. 2255). Also, a multicenter, double-blind, placebo-controlled, dose-response study of simvastatin showed a' significant decrease in total-cholesterol, LDLC, total cholesterol/HDLC ratio, and LDLC /HDLC ratios in subjects with familial or non-familial hypercholesterolemia. (Physicians' Desk Reference, 2000, p. 1917). In comparative studies of simvastatin at a low daily dose versus a high daily dose, the mean percent decreases in LDLC and mean percent mcreases in HDLC observed were 26% and 10% for 5 mg, 30% and 12% for 10 mg, 41% and 9% for 40 mg, and 47% and 8% for 80 mg. Finally, in multicenter, placebo-controlled studies of pravastatin sodium given in daily doses from 10-40 mg, subjects with primary hypercholesterolemia showed consistent and significant decreases in total-cholesterol, LDLC, triglycerides, total-cholesterol/HDLC ratio, and LDLC/HDLC ratios. The mean LDLC decreases and HDLC increases for pravastatin sodium administered once daily at bedtime were 22% and 7% at 10 mg/day, 32% and 2% at 20 mg/day, and 34% and 12% at 40 mg/day (Physicians' Desk Reference, 2000, p. 846).

Other comparative studies have suggested that statins differ in some of their clinical properties relevant to reducing the risks of atherosclerosis. A recent double-blind, randomized, parallel, 36-week dose escalation study with 826 hypercholesterolemic patients compared simvastatin and atorvastatin at 40 or 80 mg/day. As dose increased, simvastatin resulted in larger increases in HDLC than atorvastatin (Illingworth DR et al. (2001) Curr Med Res Opin 17(l):43-abstract only). Wierzbicki & Mikhailidis (2002 IntJ. Cardiol. 84(l):53-57) reviewed five studies comparing the dose-response effects of atorvastatin and simvastatin on HDLC in hypercholesterolemic patients to compare daily doses for both drugs ranging from 10 to 80 mg. HDLC was significantly and consistently increased by all doses of simvastatin. However while atorvastatin showed increases in HDLC at low dose, the pooled data from all five studies suggest a negative dose-response effect with smaller increases in HDLC with increasing atorvastatin concentration.

The studies described above report population means for changes in LDLC and HDLC that disguise substantial evidence of significant interindividual variation in response to statins. Indeed, any particular individual treated with a statin may experience a 10% to 70% reduction in LDLC (Aguilar- Salinas SA et al., Atherosclerosis 141 :203-207, 1998). In addition, physicians have observed that some patients treated with statins exhibit minimal or no increase in HDLC, which is not an optimal response for patients with low HDLC levels. However, physicians currently are unable to identify patients who are at risk for reduced efficacy of statin therapy, which can be expensive and is not without risk. Also, physicians must currently rely on trial and error to determine which statin and dose combination will produce the best LDLC or HDLC response in any particular patient. Thus it would be useful to understand the biological basis for variability of response to statins, including the apparent negative dose effect of atorvastatin on HDLC.

Part of this biological basis may be genetic variation in proteins involved in lipid metabolism and atherogenecity (Kuivenhoven et al., supra). One protein involved in lipid metabolism is cholesteryl ester transfer protein (CETP), which promotes the transfer of cholesterol esters and triglycerides between lipoproteins (Stevenson, C. Critical Review in Clinical Laboratory Sciences 356(6): 517-546, 1998). CETP has been associated with both proatherogenecity and antiatherogenecity (Stevenson et al., supra), and the exact role of CETP in the development of atherosclerosis is still largely unknown (Kakko et al., Eur. J. Clin. Invest. 30:18-25, 2000). Some researchers have reported an inverse relationship between CETP activity levels and HDLC levels; however, a number of other studies have failed to detect this association (Gudnason V. et al, European Journal of Clinical Investigation 29: 116-128, 1999, internal citations omitted).

Cholesteryl ester transfer protein is encoded by a gene located on chromosome 16ql3. The CETP gene contains 16 exons that encode a 493 amino acid protein, including a 17 amino acid long leader peptide. A reference sequence for the CETP gene is shown in the contiguous lines of Figure 1 (Genaissance Reference No. 7650429; SEQ ID NO:l).

A number of polymorphic sites (PS) in the CETP gene have been identified. Polymorphisms at some of these PS are associated with baseline HDLC and CETP levels, and with the progression of atherosclerosis. One common variation of guanine or adenine is located in the first intron at a position corresponding to nucleotide 22,128 of Figure 1 and is often referred to in the literature as the TaqlB polymorphism (Kuivenhoven et al., supra), but is referred to herein as PS22. The Bl allele (guanine) has been associated with lower baseline HDLC and higher CETP concentrations than seen in patients who do not have this allele. Also, a polymorphism of guanine or adenine at a position corresponding to uLucieυuue Ό lot υi πgure i resuns m an ammo acid variation ol argrnme ( ) or glutamme ( ) at position 468 of the precursor polypeptide (i.e., position 451 of the mature protein). The Q468 allele is associated with higher plasma CETP activity in men and lower total cholesterol in women than seen in individuals who do not have this allele (Kakko et al., supra; Kakko et al., Atherosclerosis 136:233-40, 1998). Further, a guanine or adenine variation is located in the 3 ' UTR at a position corresponding to nucleotide 43507 of Figure 1. The adenine polymorphism is associated with low CETP activity (Tamminen M. et al., Atherosclerosis 124:237-247, 1996). Another polymorphism of guanine or cytosine at nucleotide position 40936 of Figure 1 results in an amino acid variation of alanine or proline at position 390 in the precursor polypeptide, i.e., position 373 of the mature CETP protein (NCBI SNP Database Ref. SNP ID #5887, July 15, 1999). However, any effect of this polymorphism on CETP function or concentration has not been reported.

Because of the demonstrated potential for variation in the CETP gene to affect the expression and function of the encoded protein, as well as HDLC levels, it would be useful to know whether additional polymorphisms exist in the CETP gene, as well as how such polymorphisms are combined in different haplotypes of this gene. None of the studies cited above assessed the clinical relevance of haplotypes of multiple CETP polymorphisms in affecting response to statin therapy. Thus, it would also be useful to determine whether any CETP haplotypes are associated with response to treatment with statins. Such information would assist the treating physician in developing the most appropriate therapy regimen for patients with cardiovascular disease. In particular, as noted above, the ability to identify populations of patients who are at risk for an undesirable HDLC response upon treatment with a statin would be useful. Identification of these patients would enable physicians to begin combination therapy for those individuals at the time of initiating therapy, thus saving the patient weeks of possibly inadequate therapy and potentially improving compliance.

SUMMARY OF THE INVENTION

Accordingly, the inventors herein have discovered a set of haplotypes in the CETP gene that are associated with statin-specific variation in HDLC response to treatment with atorvastatin calcium, pravastatin sodium, or simvastatin. The inventors have also discovered that the copy number of each of these CETP haplotypes affects the level of HDLC response to these statins, with zero copy of any of these haplotypes (defined as a statin response marker I) being correlated with a certain mean level of HDLC response and one or two copies of any of these haplotypes (defined as a statin response marker II) being correlated with a different mean level of response. The combinations of CETP haplotype and copy number that comprise statin response markers I and statin response markers II are shown in Table 1 below.

^J The location of each polymorphic site (PS) in Fig. 1 (SEQ ID NO: 1) is shown below the numbered PS.

If an individual has a statin response marker I (zero copies of any of CETP haplotypes a to j), that individual is likely to experience a better HDLC response to simvastatin than an individual having statin response marker II (at least one copy of any of CETP haplotypes a to j). Additionally, if an individual has a statin response marker I, the individual is likely to experience a worse HDLC response to atorvastatin than an individual having a statin response marker II. An individual's response to pravastatin is not likely to be significantly affected by the presence of either statin response marker in the individual. In addition, individuals with a statin response marker I are likely to experience a significantly better HDLC response to simvastatin than to pravastatin sodium, and a better response to pravastatin sodium than to atorvastatin calcium. In contrast, individuals with a statin response marker II are likely to experience a better HDLC response to pravastatin sodium than to atorvastatin calcium, particularly at the highest dose. Additionally, individuals with a statin response marker II are likely to experience a better HDLC response to treatment with atorvastatin calcium than with simvastatin.

In addition, as described in more detail below, the inventors believe that additional statin response markers I and II may readily be identified based on linkage disequilibrium between any of the above CETP haplotypes or their component polymorphisms and other haplotypes or polymorphisms, respectively, that are located in the CETP gene or other genes. In particular, statin response markers of the invention include haplotypes that are in linkage disequilibrium with any of haplotypes a to j in Table 1, hereinafter referred to as "linked haplotypes", as well as "substitute haplotypes" for any of haplotypes a to j in which one or more of the polymorphisms in the original haplotype is substituted with a polymorphism(s) in linkage disequilibrium with the replaced polymorphism(s),.

The correlations between the different types of statin response markers and varying HDLC response to different statins suggest that testing for the presence of a statin response marker I or a statin response marker II in patients would provide valuable information that can be used by the treating physician to choose the most effective statin or combination therapy for achieving a desired effect on HDLC levels. In addition, these correlations suggest that any clinical trial of a statin should include in its design or analysis a consideration of the potential effect of statin response markers, and other CETP haplotypes, on the efficacy of statin response. Accordingly, some aspects of the invention are based on the correlations of statin response markers I and JJ with a differential HDLC response to treatment with statins.

In one aspect, the invention provides methods and kits for determining whether an individual has a statin response marker I or a statin response marker II. These methods and kits are useful for predicting the expected therapeutic response of an individual to treatment with statins, selecting an optimal statin for an individual or choosing appropriate therapy for an individual.

In one embodiment, a method for determining whether an individual has a statin response marker I or a statin response marker II comprises determining whether the individual has zero or at least one copy of a particular haplotype. The haplotype is selected from one of the CETP haplotypes shown in Table 1, a linked haplotype for any one of haplotypes (a)-(j) in Table 1; and a substitute haplotype for any of haplotypes (a)-(j). In another embodiment of the invention, a method for assigning an individual to a first or second statin response marker group comprises determining whether the individual has zero or at least one copy of the selected haplotype and assigning the individual to a statin response marker group based on the copy number of that haplotype. The individual is assigned to the first statin response marker group if the individual has zero copy of the haplotype and to the second statin response marker group if the individual has at least one copy of the haplotype. In preferred embodiments of these methods, the selected haplotype is one of haplotypes a to j and in particularly preferred embodiments, the selected haplotype comprises haplotype (a) or (b) in Table 1. One embodiment of a kit for determining whether an individual has a statin response marker I or a statin response marker II comprises a set of oligonucleotides designed for identifying at least one of the alleles present at each polymorphic site (PS) in a set of polymorphic sites. The set of polymorphic sites (PS) comprises the set of PS for any one of the CETP haplotypes shown in Table 1, the set of PS for a linked haplotype; or the set of PS for a substitute haplotype. In a further embodiment, the kit comprises a manual with instructions for perfoπning one or more reactions on a human nucleic acid sample to identify the allele(s) present in the individual at each polymorphic site in the set and deteπnining if the individual has a statin response marker I or a statin response marker II based on the identified allele(s)

Another aspect of the invention is a method of selecting a statin to provide an optimal HDL cholesterol response in a human individual in need of statin therapy. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and selecting a statin based on the results of the determining step. If the individual has a statin response marker I, then the selected statin is simvastatin or pravastatin sodium, while if the individual has a statin response marker II, then the selected statin is pravastatin sodium or atorvastatin calcium.

The invention further provides a method of treating an individual with a statin, wherein the individual needs to maintain or increase her HDLC level. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and choosing a treatment for the individual based on the results of the determining step. If the individual has a statin response marker II, then the chosen treatment is (a) prescribing pravastatin sodium, (b) prescribing no greater than 10 mg/day ol atorvastatin calcium, (c) prescribing simvastatin m combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker II, and (d) prescribing 80 mg/day of atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker II. If the individual has a statin response marker I, then the chosen treatment is selected from the group consisting of (i) prescribing simvastatin, (ii) prescribing pravastatin sodium and (iii) prescribing atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker I.

In yet another embodiment, the invention provides a method for predicting an individual's HDL cholesterol response to treatment with a statin. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and making a response prediction based on the results of the determining step. If the individual is determined to have a statin response marker I, then the response prediction is that the individual will likely experience an increase in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a decrease in HDLC if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and a negligible change in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day. If the individual is determined to have a statin response marker II, then the response prediction is that the individual will likely experience a decrease in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day or with atorvastatin calcium at 80 mg/day, an increase in HDLC if treated with atorvastatin calcium at 10 mg/day, and an increase in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day. In other aspects, the invention provides: (i) a method of seeking regulatory approval for marketing a pharmaceutical formulation comprising a statin as at least one active ingredient for treating a disease or condition in a population partially or wholly defined by having a statin response marker, (ii) an article of manufacture comprising the pharmaceutical formulation that is marketed for treating the defined population, (iii) a method of manufacturing a drug product comprising the pharmaceutical formulation, and (iv) a method of marketing the drug product for treating the defined population. In one preferred embodiment, the statin is selected from the group consisting of simvastatin, a pharmaceutically acceptable salt of simvastatin acid, lovastatin, a pharmaceutically acceptable salt of lovastatin acid, and the defining statin response marker is a statin response marker I. In another preferred embodiment, the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker II. In particularly preferred embodiments, the disease or condition is a cardiovascular or coronary artery disorder, e.g., hypercholesterolemia.

The method of seeking regulatory approval comprises conducting at least one clinical trial which comprises administering the pharmaceutical formulation to first and second treatment groups of patients having the disease or condition, wherein each patient in the first treatment group has a statin response marker I and each patient in the second treatment group has a statin response marker II, demonstrating that one of the treatment groups exhibits a mean percent change in HDLC that is better than the mean percent change in HDLC exhibited by the other treatment group, and filing with a regulatory agency an application for marketing approval of the pharmaceutical formulation with a label stating that the pnarmaceutical tormulation is indicated tor treatmg the disease or condition in patients having the statin response marker as in the treatment group exhibiting the better mean change in HDLC. In preferred embodiments, the regulatory agency is the United States Food and Drug Administration (FDA) or the European. Agency for the Evaluation of Medicinal Products (EMEA), or a future equivalent of these agencies.

In one embodiment, the article of manufacture comprises the pharmaceutical formulation and at least one indicium identifying a population for whom the pharmaceutical formulation is indicated, wherein the identified population is partially or wholly defined by having a statin response marker I or a statin response marker II, and a trial population having the defining statin response marker exhibits a better mean HDLC response to the statin than a trial population lacking the defining statin response marker. Another embodiment of the article of manufacture comprises packaging material and the pharmaceutical formulation contained within the packaging material, wherein the packaging material comprises a label approved by a regulatory agency for the pharmaceutical formulation, wherein the label states that the pharmaceutical formulation is indicated for a population partially or wholly defined by having a statin response marker I or a statin response marker II. In other embodiments, an article of manufacture according to the invention comprises a pharmaceutical formulation comprising a statin and an HDLC-modulating agent as separate active ingredients.

The method for manufacturing the drug product comprises combining in a package a pharmaceutical formulation comprising a statin as at least one active ingredient and a label which states that the drug product is indicated for treating a population defined wholly or partially by having a statin response marker I or statin response marker II, wherein a trial population having the defining statin response marker exhibits a better mean HDLC response to the statin than a trial population lacking the defining statin response marker.

The method for marketing the drug product comprises promoting to a target audience the use of the drug product for treatmg individuals who belong to the defined population.

BRIEF DESCRIPTION OF THE SEQUENCES

SEQ ID NO:l is a reference sequence for the CETP gene (Genaissance Reference No. 7650429; contiguous lines), with the two alternative allelic variants of each polymorphic site indicated by the appropriate nucleotide symbol (R= G or A, Y= T or C, M= A or C, K= G or T, S= G or C, and W= A or T; TPO standard ST.25).

SEQ ID NOS:2-8, and their complements, are preferred sequences comprising allele-specific oligonucleotide (ASO) probes for detection of the alleles present at the polymorphic sites of the predictive haplotypes presented herein. SEQ ED NOS:9-22 are preferred sequences comprising ASO primers for detection of the alleles present at the polymorphic sites of the predictive haplotypes presented herein..

SEQ ID NOS:23-36 are preferred sequences comprising primer extension oligonucleotides for detection of the alleles present at the polymorphic sites of the predictive haplotypes presented herein. JUU I S:-5 /- s are me universal 'tail' sequences attached to trie 5 ^' end ot each umque sequence for the forward and reverse PCR primers, respectively, used to amplify genomic target regions comprising one or more of the polymorphic sites of interest herein.

DEFINITIONS

In the context of this disclosure, the terms below shall be defined as follows unless otherwise indicated:

Allele - A particular form of a genetic locus, distinguished from other forms by its particular nucleotide sequence, or one of the alternative polymorphisms found at a polymorphic site. Gene - A segment of DNA that contains the coding sequence for a protein, wherein the segment may include promoters, exons, introns, and other untranslated regions that control expression. Genotype - An unphased 5 ' to 3 ' sequence of nucleotide pair(s) found at one or more polymorphic sites in a locus on a pair of homologous chromosomes in an individual. As used herein, genotype includes a full-genotype and/or a sub-genotype as described below. Genotyping - A process for determining a genotype of an individual.

Haplotype - A 5 ' to 3 ' sequence of nucleotides found at one or more polymorphic sites in a locus on a single chromosome from a single individual.

Haplotype pair - The two haplotypes found for a locus in a single individual. Haplotyping - A process for determining one or more haplotypes in an individual and includes use of family pedigrees, molecular techniques and/or statistical inference.

Haplotype data - Information concerning one or more of the following for a specific gene: a listing of the haplotype pairs in an individual or in each individual in a population; a listing of the different haplotypes in a population; frequency of each haplotype in that or other populations, and any known associations between one or more haplotypes and a trait. Isolated — As applied to a biological molecule such as R A, DNA, oligonucleotide, or protein, isolated means the molecule is substantially free of other biological molecules such as nucleic acids, proteins, lipids, carbohydrates, or other material such as cellular debris and growth media. Generally, the term "isolated" is not intended to refer to a complete absence of such material or to absence of water, buffers, or salts, unless they are present in amounts that substantially interfere with the methods of the present invention.

Locus - A location on a chromosome or DNA molecule corresponding to a gene or a physical or phenotypic feature, where physical features include polymorphic sites.

Nucleotide pair - The nucleotides found at a polymorphic site on the two copies of a chromosome from an individual. Phased - As applied to a sequence of nucleotide pairs for two or more polymorphic sites in a locus, phased means the combination of nucleotides present at those polymorphic sites on a single copy of the locus is known.

Polymorphic site (PS) - A position on a chromosome or DNA molecule at which at least two alternative sequences are found in a population.

Polymorphism — The sequence variation observed in an individual at a polymorphic site. Polymorphisms include nucleotide substitutions, insertions, deletions and microsatellites and may, but need not, result in detectable differences in gene expression or protein function. Polynucleotide - A nucleic acid molecule comprised of single-stranded RNA or DNA or comprised of complementary, double-stranded DNA.

Population Group - A group of individuals sharing a common ethnogeographic origin.

Reference Population - A group of subjects or individuals who are predicted to be representative of the genetic variation found in the general population. Typically, the reference population represents the genetic variation in the population at a certainty level of at least 85%, preferably at least 90%, more preferably at least 95% and even more preferably at least 99%.

Single Nucleotide Polymorphism (SNP) - Typically, the specific pair of nucleotides observed at a single polymorphic site. In rare cases, three or four nucleotides may be found.

Subject - A human individual whose genotypes or haplotypes or response to treatment or disease state are to be determined.

Treatment - A stimulus administered internally or externally to a subject.

Unphased - As applied to a sequence of nucleotide pairs for two or more polymorphic sites in a locus, unphased means the combination of nucleotides present at those polymorphic sites on a single copy of the locus is not known.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

Each statin response marker of the invention is a combination of a particular genetic marker, or haplotype, and the copy number for that genetic marker. Preferably the genetic marker component of the statin response marker is one of the CETP haplotypes shown in Table 1. The CETP polymorphic sites in these CETP haplotypes are referred to herein as PS20, PS22, PS28, PS32, PS35, PS46 and PS47 and are located in the CETP gene at positions corresponding to those identified in SEQ ID NO: 1. In describing the polymorphic sites in the statin response markers of the invention, reference is made to the sense strand of a gene for convenience. However, as recognized by the skilled artisan, nucleic acid molecules containing a particular gene may be complementary double stranded molecules and thus reference to a particular site or haplotype on the sense strand refers as well to the corresponding site or haplotype on the complementary antisense strand. Further, reference may be made to detecting a genetic marker or haplotype for one strand and it will be understood by the skilled artisan that this includes detection of the complementary haplotype on the other strand. As described in more detail in Example 1, the statin response markers of the invention are based on the discovery by the inventors of correlations between certain haplotypes in the CETP gene and statin- specific HDLC response to statin treatment in a cohort of individuals participating in a a randomized, 16- week, open-label investigation of drug response in relationship to gene variants in adult subjects with primary hypercholesterolemia. In particular, the inventors herein discovered that a CETP haplotype compπsmg guanine at JPS22 and guanine at PS47 (CETP haplotype (a) in Table 1) affected the percent change in HDLC levels observed in patients participating in the study following treatment with one of three statins: Zocor® (simvastatin), Pravachol®(pravastatin sodium) or Lipitor® (atorvastatin calcium). The group of patients lacking this haplotype experienced a better HDLC response to Zocor® than the patient group having at least one copy of the haplotype. Conversely, the group of patients that had at least one copy of this haplotype experienced a better HDLC response to Lipitor® than the patient group lacking this haplotype. Mean HDLC response to Pravachol® (pravastatin sodium) in the patient cohort was not affected by the presence or absence of this haplotype at a statiscally significant level, however the general trend of the adjusted least squares means showed that patients having at least one copy of this haplotype experienced a better HDLC response to Pravachol® treatment than the patient group lacking a copy of this haplotype. Analogous results were obtained for the CETP haplotype comprising guanine at PS20 and guanine at PS47 (CETP haplotype (b) in Table 1).

Additionally, for the patient cohort lacking this haplotype, the ordering of the three drugs with respect to the best HDLC response after treatment was Zocor®>Pravachol®>Lipitor® at both the high and low doses examined in the study (see Table 7). For the patient cohort in the study having at least one copy of this haplotype, the ordering of the three drugs with respect to the best HDLC response after treatment was Pravachol®> Lipitor®> Zocor®, where Pravachol® and Lipitor® are roughly equivalent at the low dose of each statin, but Pravachol® provides a much better HDLC result than Lipitor® at the highest dose of each statin used in the study. Moreover, as shown by the marker-by statin interaction p value shown in Table 7, the different effect of copy number of this haplotype on HDLC response after treatment with each of the three drugs is statistically significant. Therefore this haplotype, in combination with the haplotype copy number, can be used to differentiate the HDLC response that might be observed in an individual or a trial population after treatment with a given statin. Consequently, zero copy of CETP haplotype (a) is referred to herein as a statin response marker I, while at least one copy of CETP haplotype (a) is referred to herein as a statin response marker II.

In addition to CETP haplotype (a), consisting of guanine at each of PS22 and PS47, several other polymorphic sites in the CETP gene have alleles in high linkage disequilibrium (LD) with the guanine at PS22 or the guanine at PS47. Two particular nucleotide alleles at different polymorphic sites are said to be in LD if the presence of one of the alleles at one of the sites tends to predict the presence of the other allele at the other site on the same chromosome (Stevens, JC, Mol. Diag. 4: 309-17, 1999). One of the most frequently used measures of linkage disequilibrium is Δ², which is calculated using the formula described in Devlin, B. and Risch, N. (l995,Genomics, 29(2):311-22). Basically, Δ² measures how well an allele X at a first polymorphic site predicts the occurrence of an allele Y at a second polymorphic site on the same chromosome. The measure only reaches 1.0 when the prediction is perfect (e.g., X if and only if Y).

For example, as shown in Table 2 below, essentially perfect LD exists between guanine at PS22 and guanine at PS20 (Δ² = 1.00 for the total experimental population examined herein). Thus, the skilled artisan would have expected that the presence or absence of a haplotype of guanine at each of PS20 and PS47 (CETP haplotype (b)) would be predictive of the presence or absence of CETP haplotype (a) and therefore predictive of an individual's HDLC response after statin treatment. Indeed, calculations on CETP haplotype (b) discussed in Example 3 confirmed this expectation. Consequently, zero copy of CETP haplotype (b) also comprises a statin response marker I, while at least one copy of CETP haplotype (b) also comprises a statin response marker II.

The inventors herein identified other alleles at polymorphic sites that are also in high LD with the allele at one of the two polymorphic sites in CETP haplotype (a) and believe that the presence or absence of haplotypes containing one of these alleles in high LD with the guanine at PS22 substituted for guanine at PS22 or containing guanine at PS46 substituted for guanine at PS47 can also function to predict the presence or absence of CETP haplotype (a). Such haplotypes containing a substitute polymorphism replacing one or more of the polymorphisms in the original haplotype are referred to herein as "substitute haplotypes". The LD relationships identified by the inventors herein are shown in Table 2 below, which lists the values for Δ² in the total experimental population and for each of the 4 ethnic population groups within that population.

Table 2. Linkage disequilibrium with polymorphisms in CETP haplotype (a).

Allele in Allele in LD A^" for various populations in the study³ response marker 1 Total CA AF HL AS

G at PS22 G at PS20 1.00 1.00 0.95 1.00 1.00

G at PS22 G at PS28 0.77 0.90 0.12 0.50 0.47

G at PS22 C at PS32 0.79 0.91 0.12 0.52 0.62

G at PS22 C at PS35 0.79 0.91 0.12 0.52 0.62

G at PS47 G at PS46 0.88 0.93 0.54 0.96 0.93 ^a CA, AF, HL, and AS stand for Caucasian, African-American, Hispanic-Latino, and Asian, respectively.

The substitute haplotypes (b)-(j) for haplotype (a) that comprise a substitute polymorphism for one or both of the polymorphisms in haplotype (a) are shown in Table 1. Thus, a statin response marker I is defined herein as comprising zero copy of any of haplotypes (a)-(j), while a statin response marker II is defined herein as comprising at least one copy of any of haplotypes (a)-(j). In addition, the skilled artisan would expect that there might be additional polymorphisms in the

CETP gene or elsewhere on chromosome 16 that are in high LD with one or more of the polymorphisms in the haplotypes comprising a statin response marker I or a statin response marker II. Thus, the skilled artisan would expect that all of the embodiments of the invention described herein may frequently be practiced by substituting any (or all) of the specifically identified CETP polymorphisms in a statin response marker with another polymorphism that is in high LD with the specifically identified polymorphism. This "substitute polymorphism" may be one that is currently known or subsequently discovered and may be present at a polymorphic site in the CETP gene or elsewhere on chromosome 16.

Further, the inventors contemplate that there will be other haplotypes in the CETP gene or elsewhere on chromosome 16 that are in high LD with one or more of the CETP haplotypes in Table 1 that would therefore also be predictive of the statin-specific HDLC response. Preferably, the linked -uαjjiui jjo 10 iccc L iii lac ^m gene or m a genomic region ot about 1UU J iobases spanning the CETP gene. The linkage disequilibrium between the CETP haplotypes from Table 1 and linked haplotypes can also be measured using Δ².

In preferred embodiments, the linkage disequilibrium between a polymorphism in any of the CETP haplotypes in Table 1 and a substitute polymorphism to replace it, or between any of the CETP haplotypes in Table 1 and a linked haplotype, has a Δ² value, as measured in a suitable reference population, of at least 0.75, more preferably at least 0.80, even more preferably at least 0.85 or at least 0.90, yet more preferably at least 0.95, and most preferably 1.0. A suitable reference population for this Δ² measurement is preferably selected from a population with the distribution of the ethnic background of its members reflecting the population of patients to be treated with statins, which may be the general population, a population using statins, a population with cardiovascular disease (CVD) or CVD risk factors, and the like.

LD patterns in genomic regions are readily determined empirically in appropriately chosen samples using various techniques known in the art for determining whether any two alleles (either two polymorphisms at different polymorphic sites or two haplotypes) are in linkage disequilibrium (Weir B.S. 1996 Genetic Data Analysis II, Sinauer Associates, Inc. Publishers, Sunderland, MA). The skilled artisan may readily select which method of determining LD will be best suited for a particular sample size and genomic region. Similarly, the ability of substitute haplotypes, that contain one or more substitute polymorphisms or that are in high LD with one or more of haplotypes (a) to (j), to predict the HDLC response to one of the statins studied herein may also be readily tested by the skilled artisan.

Thus, reference herein to a statin response marker I is deemed to include zero copy of haplotypes that (a) either (1) have a polymorphism sequence that is similar to a haplotype shown in Table 1, but in which at least one of the specifically identified CETP polymorphisms in that haplotype has been substituted with a polymorphism in high LD with the specifically identified polymorphism (a "substitute haplotype"; or (2) are in high linkage disequilibrium with a haplotype shown in Table 1; and (b) behave similarly to the statin response marker I defined as zero copy of haplotype (a) or haplotype (b) in terms of predicting an individual's HDLC response to different statins. Such similar statin response markers I are referred to herein as "alternative statin response markers I". Similarly, reference herein to a statin response marker II is deemed to include at least one copy of haplotypes that (a) either (1) have a polymorphism sequence that is similar to a haplotype shown in Table 1, but in which at least one of the specifically identified CETP polymorphisms in that haplotype has been substituted with a polymorphism in high LD with the specifically identified polymorphism; or (2) are in high linkage disequilibrium with a haplotype shown in Table 1; and (b) behave similarly to the statin response marker II defined as at least one copy of haplotype (a) or haplotype (b) in terms of predicting an individual's HDLC response to different statins. Such similar statin response markers Et are referred to herein as "alternative statin response markers II".

As described above and in the Examples below, the statin response markers of the invention are associated with statin-specific effects on mean percent changes HDLC in response to treatment. Thus, the invention provides a method and kit for determining whether an individual has a statin response marker I or a statin response marker II.

In one embodiment, the invention provides a method for determining whether an individual has a statin response marker I or II. The method comprises determining whether the individual has zero or at , least one copy of a haplotype selected from the group consisting of haplotypes (a) to (j) in Table 1, a linked haplotype for one of haplotypes (a) to (j); and a substitute haplotype for any one of haplotypes (a) to (j). In another embodiment, the invention provides a method for assigning an individual to a first or second statin response marker group. The method comprises determining whether the individual has zero or at least one copy of a haplotype selected from the group consisting of haplotypes (a) to (j) in Table 1, a linked haplotype for any one of haplotypes (a) to (j); and a substitute haplotype for any one of haplotype (a) to (j), and assigning the individual to the first statin response marker group if the individual has zero copies of the selected haplotypes and to the second statin response marker group if the individual has at least one copy of the selected haplotype. In preferred embodiments of the above methods, the selected haplotype is haplotype (a), haplotype (b), a linked haplotype for haplotype (a) or haplotype (b), or a substitute haplotype for (a) or (b) in which at least one of guanine at PS20, guanine at PS22 or guanine at PS47 is replaced with a substitute polymorphism in linkage disequilibrium with the replaced polymorphism. In some embodiments, the individual is Caucasian and may be diagnosed with a coronary artery disease or a cardiovascular disease, such as Type Ila or Type lib hypercholesterolemia, may have risk factors associated with cardiovascular disease, or may be a candidate for treatment with a statin for an alternative reason.

The presence in an individual of a statin response marker I or II may be determined by a variety of indirect or direct methods well known in the art for determining haplotypes or haplotype pairs for a set of polymorphic sites in one or both copies of the individual's genome, including those discussed below. The genotype for a polymorphic site in an individual may be determined by methods also described below.

One indirect method for determining whether zero or at least one copy of a CETP haplotype is present in an individual is by prediction based on the individual's genotype determined at one or more of the polymorphic sites comprising the haplotype and using the determined genotype at each site to determine the CETP haplotypes present in the individual. The presence of zero, one or two copies of a CETP haplotype of interest can be determined by visual inspection of the alleles at the PS that comprise the haplotype. The CETP haplotype pair is assigned by comparing the individual's genotype with the genotypes at the same set of PS corresponding to the haplotype pairs known to exist in the general population or in a specific population group or to the haplotype pairs that are theoretically possible based on the alternative alleles possible at each PS, and determining which haplotype pair is most likely to exist in the individual.

In a related indirect haplotyping method, the presence in an individual of zero copy or at least one copy of a CETP haplotype is predicted from the individual's genotype for a set of PS comprising the selected haplotype using information on haplotype pairs known to exist in a reference population. In one embodiment, this haplotype pair prediction method comprises identifying a genotype for the individual at the set of polymorphic sites comprising the selected haplotype, accessing data containing haplotype pairs identified in a reference population for a set of polymorphic sites comprising the polymorphic sites of the selected haplotype, and assigning to the individual a haplotype pair that is consistent with the individual's genotype. Whether the individual has a statin response marker I or a statin response marker II can be subsequently determined based on the assigned haplotype pair. The haplotype pair can be assigned by comparing the individual's genotype with the genotypes corresponding to the haplotype pairs known to exist in the general population or in a specific population group, and determining which haplotype pair is consistent with the genotype of the individual. In some embodiments, the comparing step may be performed by visual inspection. When the genotype of the individual is consistent with more than one haplotype pair, frequency data may be used to determine which of these haplotype pairs is most likely to be present in the individual. If a particular haplotype pair consistent with the genotype of the individual is more frequent in the reference population than others consistent with the genotype, then that haplotype pair with the highest frequency is the most likely to be present in the individual. This determination may also be performed in some embodiments by visual inspection. In other embodiments, the comparison may be made by a computer-implemented algorithm with the genotype of the individual and the reference haplotype data stored in computer-readable formats. For example, as described in WO 01/80156, one computer-implemented algorithm to perform this comparison entails enumerating all possible haplotype pairs which are consistent with the genotype, accessing data containing CETP haplotype pairs frequency data determined in a reference population to determine a probability that the individual has a possible haplotype pair, and analyzing the determined probabilities to assign a haplotype pair to the individual. Typically, the reference population is composed of randomly-selected individuals representing the major ethnogeographic groups of the world. A preferred reference population for use in the methods of the present invention consists of Caucasian individuals, the number of which is chosen based on how rare a haplotype is that one wants to be guaranteed to see. For example, if one wants to have a q% chance of not missing a haplotype that exists in the population at a p% frequency of occurring in the reference population, the number of individuals (n) who must be sampled is given by 2n=log(l-q)/log(l-p) where p and q are expressed as fractions. A preferred reference population allows the detection of any haplotype whose frequency is at least 10% with about 99% certainty. A particularly preferred reference population includes a 3 -generation Caucasian family to serve as a control for checking quality of haplotyping procedures.

If the reference population comprises more than one ethnogeographic group, the frequency data for each group is examined to determine whether it is consistent with Hardy- Weinberg equilibrium. Hardy- Weinberg equilibrium (D.L. Hartl et al., Principles of Population Genomics, Sinauer Associates (Sunderland, MA), 3^rd Ed., 1997) postulates that the frequency of finding the haplotype pair H_x I H₂ is equal to p_H__w(H l H₂) = 2p(H_l)p(H₂) if H, ≠ H₂ and p_H__w(H H₂) = p(H_λ)p(H₂) if H_l = H₂ . A statistically significant difference between the observed and expected haplotype frequencies could be due to one or more factors including significant inbreeding in the population group, strong selective pressure on the gene, sampling bias, and/or errors in the genotyping process. If large deviations from Hardy- Weinberg equilibrium are observed in an ethnogeographic group, the number of individuals in that group can be increased to see if the deviation is due to a sampling bias. If a larger sample size does not reduce the difference between observed and expected haplotype pair frequencies, then one may wish to consider haplotyping the individual using a direct haplotyping method such as, for example, CLASPER System technology (U.S. Patent No. 5,866,404), single molecule dilution, or allele- specific long-range PCR (Michalotos-Beloin et al., Nucleic Acids Res. 24:4841-4843, 1996).

In one embodiment of this method for predicting a haplotype pair for an individual, the assigning step involves performing the following analysis. First, each of the possible haplotype pairs is compared to the haplotype pairs in the reference population. Generally, only one of the haplotype pairs in the reference population matches a possible haplotype pair and that pair is assigned to the individual. Occasionally, only one haplotype represented in the reference haplotype pairs is consistent with a possible haplotype pair for an individual, and in such cases the individual is assigned a haplotype pair containing this known haplotype and a new haplotype derived by subtracting the known haplotype from the possible haplotype pair. Alternatively, the haplotype pair in an individual may be predicted from the individual's genotype for that gene using reported methods (e.g., Clark et al. 1990, Mol Bio Evol 7: 111-22 or WO 01/80156) or through a commercial haplotyping service such as offered by Genaissance Pharmaceuticals, Inc. (New Haven, CT). In rare cases, either no haplotypes in the reference population are consistent with the possible haplotype pairs, or alternatively, multiple reference haplotype pairs are consistent with the possible haplotype pairs. In such cases, the individual is preferably haplotyped using a direct molecular haplotyping method such as, for example, CLASPER System technology (U.S. Patent No. 5,866,404), SMD, or allele-specific long-range PCR (Michalotos-Beloin et al., supra).

Determination of the number of haplotypes present in the individual from the genotypes is illustrated here for CETP haplotype (a). The table below shows the 9 (3ⁿ, where each of n=2 bi-allelic polymorphic sites may have one of 3 different genotypes present) genotypes that may be detected at PS22 and PS47, using both chromosomal copies from an individual. Eight of the nine possible genotypes for the two sites allow unambiguous determination of the number of copies of the CETP haplotype (a) present in the individual and therefore would allow unambiguous determination of whether the individual has a statin response marker I or II. However, an individual with the A/G A/G genotype could possess either of the following haplotype pairs: GG/AA or AG/GA, and thus could have either 1 copy of the CETP haplotype (a) (GG/AA haplotype pair) corresponding to a statin response marker II, or 0 copy (AG/GA haplotype pair) of the CETP haplotype (a) corresponding to a statin response marker I. For this instance where there is ambiguity in the haplotype pair underlying the determined genotype A/G A/G, frequency information may be used to determine the most probable haplotype pair and therefore the most likely number of copies of CETP haplotype (a) in the individual. If a particular CETP haplotype pair consistent with the genotype of the individual is more frequent in the reference population than others consistent with the genotype, then that haplotype pair with the highest frequency is the most likely to be present in the individual. The copy number of the haplotype of interest in this haplotype pair can then be determined by visual inspection of the alleles at the PS that comprise the response marker for each haplotype in the pair.

Alternatively, for the ambiguous double heterozygote, genotyping of one or more additional sites in CETP may be performed to eliminate the ambiguity in deconvoluting the haplotype pairs underlying the genotype at PS22 and PS47. The skilled artisan would recognize that these one or more additional sites would need to have sufficient linkage with the alleles in at least one of the possible haplotypes in the pair to peπnit unambiguous assignment of the haplotype pair. Although this illustration has been directed to the particular instance of determining the number of CETP haplotype (a) present in an individual, the process would be analogous for the other CETP haplotypes shown in Table 1 or for the haplotypes comprising any alternative statin response markers I or II.

Table 3. Possible copy numbers of CETP Haplotype (a) based on the genotypes at PS22 and PS47

The individual's genotype for the desired set of PS may be determined using a variety of methods well-known in the art. Such methods typically include isolating from the individual a genomic DNA sample comprising both copies of the gene or locus of interest, amplifying from the sample one or more target regions containing the polymorphic sites to be genotyped, and detecting the nucleotide pair present at each PS of interest in the amplified target region(s). It is not necessary to use the same procedure to determine the genotype for each PS of interest.

In addition, the identity of the allele(s) present at any of the novel polymorphic sites described herein may be indirectly determined by haplotyping or genotyping another polymorphic site that is in linkage disequilibrium with the polymorphic site that is of interest. Polymorphic sites in linkage disequilibrium with the presently disclosed polymorphic sites may be located in regions of the gene or in other genomic regions not examined herein. Detection of the allele(s) present at a polymorphic site in linkage disequilibrium with the novel polymorphic sites described herein may be performed by, but is not limited to, any of the above-mentioned methods for detecting the identity of the allele at a polymorphic site.

Alternatively, the presence in an individual of a haplotype or haplotype pair for a set of PS comprising a statin response marker may be determined by directly haplotyping at least one of the copies of the individual's genomic region of interest, or suitable fragment thereof, using methods known in the art. Such direct haplotyping methods typically involve treating a genomic nucleic acid sample isolated from the individual in a manner that produces a hemizygous DNA sample that only has one of the two "copies" of the individual's genomic region which, as readily understood by the skilled artisan, may be the same allele or different alleles, amplifying from the sample one or more target regions containing the polymorphic sites to be genotyped, and detecting the nucleotide present at each PS of interest in the amplified target region(s). The nucleic acid sample may be obtained using a variety of methods known in the art for preparing hemizygous DNA samples, which include: targeted in vivo cloning (TIVC) in yeast as described in WO 98/01573, U.S. Patent No. 5,866,404, and U.S. Patent No. 5,972,614; generating hemizygous DNA targets using an allele specific oligonucleotide in combination with primer extension and exonuclease degradation as described in U.S. Patent No. 5,972,614; single molecule dilution (SMD) as described in Ruano et al., Proc. Natl. Acad. Sci. 87:6296-6300, 1990; and allele specific PCR (Ruano et al., 1989, supra; Ruano et al., 1991, supra; Michalatos-Beloin et al., supra).

As will be readily appreciated by those skilled in the art, any individual clone will typically only provide haplotype information on one of the two genomic copies present in an individual. If haplotype information is desired for the individual's other copy, additional clones will usually need to be exammed. Typically, at least five clones should be examined to have more than a 90% probability of haplotyping both copies of the genomic locus in an individual. In some cases, however, once the haplotype for one genomic allele is directly determined, the haplotype for the other allele may be inferred if the individual has a known genotype for the polymorphic sites of interest or if the haplotype frequency or haplotype pair frequency for the individual's population group is known.

While in direct haplotyping of both copies of the gene, the analysis is preferably performed with each copy of the gene being placed in separate containers, it is also envisioned that if the two copies are labeled with different tags, or are otherwise separately distinguishable or identifiable, it could be possible in some cases to perform the haplotyping in the same container. For example, if first and second copies of the gene are labeled with different first and second fluorescent dyes, respectively, and an allele-specific oligonucleotide labeled with yet a third different fluorescent dye is used to assay the polymorphic site(s), then detecting a combination of the first and third dyes would identify the polymorphism in the first gene copy while detecting a combination of the second and third dyes would identify the polymorphism in the second gene copy. The nucleic acid sample used in the above indirect and direct haplotyping methods is typically isolated from a biological sample taken from the individual, such as a blood sample or tissue sample. Suitable tissue samples include whole blood, saliva, tears, urine, skin and hair.

The target region(s) containing the PS of interest may be amplified using any oligonucleotide- directed amplification method, including but not limited to polymerase chain reaction (PCR) (U.S. Patent No. 4,965,188), ligase chain reaction (LCR) (Barany et al., Proc. Natl Acad. Sci. USA 88:189-193, 1991; WO90/01069), and oligonucleotide ligation assay (OLA) (Landegren et al., Science 241: 1077-1080, 1988). Other known nucleic acid amplification procedures may be used to amplify the target region(s) including transcription-based amplification systems (U.S. Patent No. 5,130,238; EP 329,822; U.S. Patent No. 5,169,766, WO89/06700) and isothermal methods (Walker et al., Proc. Natl. Acad. Sci. USA 89:392- 396, 1992).

In both the direct and indirect haplotyping methods, the identity of a nucleotide (or nucleotide pair) at a polymorphic site(s) in the amplified target region may be determined by sequencing the amplified region(s) using conventional methods. If both copies of the gene are represented in the amplified target, it will be readily appreciated by the skilled artisan that only one nucleotide will be detected at a polymorphic site in individuals who are homozygous at that site, while two different nucleotides will be detected if the individual is heterozygous for that site. The polymorphism may be identified directly, known as positive-type identification, or by inference, referred to as negative-type identification. For example, where a polymorphism is known to be guanine and cytosine in a reference population, a site may be positively determined to be either guanine or cytosine for an individual homozygous at that site, or both guanine and cytosine, if the individual is heterozygous at that site. Alternatively, the site may be negatively determined to be not guanine (and thus cytosine/cytosine) or not cytosine (and thus guanine/guanine). A polymorphic site in the target region may also be assayed before or after amplification using one of several hybridization-based methods known in the art. Typically, allele-specific oligonucleotides are utilized in performing such methods. The allele-specific oligonucleotides may be used as differently labeled probe pairs, with one member of the pair showing a perfect match to one variant of a target sequence and the other member showing a perfect match to a different variant. In some embodiments, more than one polymorphic site may be detected at once using a set of allele-specific oligonucleotides or oligonucleotide pairs. Preferably, the members of the set have melting temperatures within 5°C, and more preferably within 2°C, of each other when hybridizing to each of the polymorphic sites being detected.

Hybridization of an allele-specific oligonucleotide to a target polynucleotide may be performed with both entities in solution, or such hybridization may be performed when either the oligonucleotide or the target polynucleotide is covalently or noncovalently affixed to a solid support. Attachment may be mediated, for example, by antibody-antigen interactions, poly-L-Lys, streptavidin or avidin-biotin, salt bridges, hydrophobic interactions, chemical linkages, UV cross-linking baking, etc. Allele-specific oligonucleotides may be synthesized directly on the solid support or attached to the solid support subsequent to synthesis. Solid-supports suitable for use in detection methods of the invention include substrates made of silicon, glass, plastic, paper and the like, which may be formed, for example, into wells (as in 96-well plates), slides, sheets, membranes, fibers, chips, dishes, and beads. The solid support may be treated, coated or derivatized to facilitate the immobilization of the allele-specific oligonucleotide or target nucleic acid. Detecting the nucleotide or nucleotide pair at a PS of interest may also be determined using a mismatch detection technique, including but not limited to the RNase protection method using riboprobes (Winter et al., Proc. Natl. Acad. Sci. USA 82:7575, 1985; Meyers et al., Science 230:1242, 1985) and proteins which recognize nucleotide mismatches, such as the E. coli mutS protein (Modrich, P. Ann. Rev. Genet. 25:229-253, 1991). Alternatively, variant alleles can be identified by single strand conformation polymorphism (SSCP) analysis (Orita et al., Genomics 5:874-879, 1989; Humphries et al., in Molecular Diagnosis of Genetic Diseases, R. Elles, ed., pp. 321-340, 1996) or denaturing gradient gel electrophoresis (DGGE) (Wartell et al., Nucl. Acids Res. 18:2699-2706, 1990; Sheffield et al., Proc. Natl. Acad. Sci. USA 86:232-236, 1989).

A polymerase-mediated primer extension method may also be used to identify the polymorphism(s). Several such methods have been described in the patent and scientific literature and include the "Genetic Bit Analysis" method (W092/15712) and the ligase/polymerase mediated genetic bit analysis (U.S. Patent 5,679,524. Related methods are disclosed in W091/02087, WO90/09455, W095/17676, U.S. Patent Nos. 5,302,509, and 5,945,283. Extended primers containing the complement of the polymorphism may be detected by mass spectrometry as described in U.S. Patent No. 5,605,798. Another primer extension method is allele-specific PCR (Ruano et al., Nucl. Acids Res. 17:8392, 1989; Ruano et al., Nucl. Acids Res. 19, 6877-6882, 1991; WO 93/22456; Turki et al., J. Clin. Invest. 95:1635- 1641, 1995). In addition, multiple polymorphic sites may be investigated by simultaneously amplifying multiple regions of the nucleic acid using sets of allele-specific primers as described in Wallace et al. (WO89/10414).

The genotype or haplotype for the CETP gene of an individual may also be determined by hybridization of a nucleic acid sample containing one or both copies of the gene, mRNA, cDNA or fragment(s) thereof, to nucleic acid arrays and subarrays such as described in WO 95/11995. The arrays would contain a battery of allele-specific oligonucleotides representing each of the polymorphic sites to be included in the genotype or haplotype.

The invention also provides a kit for determining whether an- individual has a statin response marker I or a statin response marker II. The kit comprises a set of oligonucleotides designed for determining the allele(s) present at a set of polymorphic sites (PS) comprising the set of polymorphic sites for one of the CETP haplotypes (a) to (j) in Table 1, the set of PS for a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j) in Table 1 or the set of PS for a substitute haplotype for any of haplotypes (a) to (j) in which one or both of the polymorphisms in the original haplotype is replaced with a substitute polymorphism in linkage disequilibrium with the replaced polymorphism. In one embodiment, the kit comprises oligonucleotides for detecting at least one allele for a PS selected from the group consisting of PS20, PS22, PS28, PS32, and PS35 and at least one allele for one additional PS selected from the group consisting of PS46 and PS47. In one preferred embodiment, oligonucleotides to detect both alleles at the selected PS are included in the kit. In another preferred embodiment, oligonucleotides to detect both alleles at each of PS22 and PS47 or each of PS20 and PS47 are included in the kit. More preferably, oligonucleotides to detect both alleles at each of PS22 and PS47 are included. In another preferred embodiment, oligonucleotides to genotype PS20, PS22, PS28, PS32, PS35, PS46, and PS47 are included in the kit. In other embodiments, the oligonucleotides for genotyping one or more of the CETP polymorphic sites PS20, PS22, PS28, PS32, PS35, PS46, and PS47 are substituted with oligonucleotides designed to detect a polymorphism at a different PS in the CETP gene or elsewhere on cnromosome lb at is m linkage. disequilibrium with the replaced polymorphism. Alternatively, a kit of the invention may comprise oligonucleotides designed for detecting the nucleotide(s) or nucleotide pair(s) at the substitute polymorphic site(s) as well as at the CETP polymorphic site that comprise a haplotype for an alternative statin response marker I or II. Each genotyping oligonucleotide provided in the kit may be placed in the same or separate receptacles and may be provided together in a package.

As used herein, a genotyping oligonucleotide is a probe or primer capable of hybridizing to a target region that contains, or that is located close to, a polymorphic site of interest such as one of the polymorphic sites comprising a statin response marker described herein. The term "oligonucleotide" refers to a polynucleotide molecule having less than about 100 nucleotides. A preferred oligonucleotide of the invention is 10 to 35 nucleotides long. More preferably, the oligonucleotide is between 15 and 30, and most preferably, between 20 and 25 nucleotides in length. The exact length of the oligonucleotide will depend on many factors that are routinely considered and practiced by the skilled artisan.

The oligonucleotides used to practice the invention may be comprised of any phosphorylation state of ribonucleotides, deoxyribonucleotides, and acyclic nucleotide derivatives, and other functionally equivalent derivatives. Alternatively, oligonucleotides may have a phosphate-free backbone, which may be comprised of linkages such as carboxymethyl, acetamidate, carbamate, polyamide (peptide nucleic acid (PNA)) and the like (Varma, R. in Molecular Biology and Biotechnology, A Comprehensive Desk Reference, Ed. R. Meyers, VCH Publishers, Inc. (1995), pages 617-620). Oligonucleotides of the invention may be prepared by chemical synthesis using any suitable methodology known in the art, or may be derived from a biological sample, for example, by restriction digestion. The oligonucleotides may be labeled, according to any technique known in the art, including use of radiolabels, fluorescent labels, enzymatic labels, proteins, haptens, antibodies, sequence tags and the like.

Oligonucleotides of the invention must be capable of specifically hybridizing to a target region of a polynucleotide containing a desired locus. As used herein, specific hybridization means the oligonucleotide forms an anti-parallel double-stranded structure with the target region under certain hybridizing conditions, while failing to form such a structure when incubated with another region in the polynucleotide or with a polynucleotide lacking the desired locus under the same hybridizing conditions. Preferably, the oligonucleotide specifically hybridizes to the target region under conventional high stringency conditions. The skilled artisan can readily design and test oligonucleotide probes and primers suitable for detecting polymorphisms in the CETP gene or adjacent regions of chromosome 16 in linkage disequilibrium with one of the haplotypes (a) to (j), using the polymorphism information provided herein in conjunction with the known sequence information for the CETP gene, and adjacent regions of chromosome 16, and routine techniques.

A nucleic acid molecule such as an oligonucleotide or polynucleotide is said to be a "perfect" or "complete" complement of another nucleic acid molecule if every nucleotide of one of the molecules is complementary to the nucleotide at the corresponding position of the other molecule. A nucleic acid molecule is "substantially complementary" to another molecule if it hybridizes to that molecule with sufficient stability to remain in a duplex form under conventional low-stringency conditions. nyuπuizauon conditions are described, tor example, by Sambrook J. et al., in Molecular Cloning, A Laboratory Manual, 2^nd Edition, Cold Spring Harbor Press, Cold Spring Harbor, NY (1989) and by Haymes, B.D. et al. in Nucleic Acid Hybridization, A Practical Approach, IRL Press, Washington, D.C. (1985). While perfectly complementary oligonucleotides are preferred for detecting polymorphisms, departures from complete complementarity are contemplated where such departures do not prevent the molecule from specifically hybridizing to the target region. For example, an oligonucleotide primer may have a non-complementary fragment at its 5' end, with the remainder of the primer being complementary to the target region. Alternatively, non-complementary nucleotides may be interspersed into the probe or primer as long as the resulting probe or primer is still capable of specifically hybridizing to the target region.

Preferred oligonucleotides of the invention, useful in determining if an individual has a statin response marker I or II, are allele-specific oligonucleotides. As used herein, the term allele-specific oligonucleotide (ASO) means an oligonucleotide that is able, under sufficiently stringent conditions, to hybridize specifically to one allele of a gene, or other locus, at a target region containing a polymorphic site while not hybridizing to the corresponding region in another allele(s). As understood by the skilled artisan, allele-specificity will depend upon a variety of readily optimized stringency conditions, including salt and formamide concentrations, as well as temperatures for both the hybridization and washing steps. Examples of hybridization and washing conditions typically used for ASO probes are found in Kogan et al., "Genetic Prediction of Hemophilia A" in PCR Protocols, A Guide to Methods and Applications, Academic Press, 1990 and Ruano et al., 87 Proc. Natl. Acad. Sci. USA 6296-6300, 1990. Typically, an ASO will be perfectly complementary to one allele while containing a single mismatch for another allele.

Allele-specific oligonucleotides of the invention include ASO probes and ASO primers. ASO probes which usually provide good discrimination between different alleles are those in which a central position of the oligonucleotide probe aligns with the polymorphic site in the target region (e.g., approximately the 7^th or 8^th position in a 15mer, the 8^th or 9^th position in a 16mer, and the 10^th or 11^th position in a 20mer). An ASO primer of the invention has a 3 ' terminal nucleotide, or preferably a 3 ' penultimate nucleotide, that is complementary to only one nucleotide of a particular SNP, thereby acting as a primer for polymerase-mediated extension only if the allele containing that nucleotide is present. ASO probes and primers hybridizing to either the coding or noncodrng strand are contemplated by the invention. ASO probes and primers listed below use the appropriate nucleotide symbol (R= G or A, Y= T or C, M= A or C, K= G or T, S= G or C, and W= A or T; WIPO standard ST.25) at the position of the polymorphic site to represent that the ASO contains either of the two alternative allelic variants observed at that polymorphic site.

Preferred ASO probes for detecting the alleles at PS20, PS22, PS28, PS32, PS35, PS46, and PS47 in the haplotypes comprising the preferred embodiments of the statin response markers I and LT comprise a nucleotide sequence selected from the group consisting of:

CTTTGGTRAGAAGGT (SEQ ID NO: 2) and its complement, GGGGTTCRAGTTAGG (SEQ ID NO: 3) and its complement, ACTAGGCRCTCCATG (SEQ ID NO: 4) and its complement, GTGAGTGYGTTTCTG (SEQ ID NO 5) and its complement,

CTGCAGCMTCACAAG (SEQ ID NO 6) and its complement,

GGGCTGCKAGGGGAT (SEQ ID NO 7) and its complement, and

GCTAGGGRATCCAGA (SEQ ID NO 8) and its complement.

Preferred ASO primers for detecting the alleles at PS20, PS22, PS28, PS32, PS35, PS46, and PS47 in the haplotypes comprising the preferred embodiments of the statin response markers I and II comprise a nucleotide sequence selected from the group consisting of:

CAAGTTCTTTGGTRA (SEQ ID NO 9); GCTAGGACCTTCTYA (SEQ ID NO:10);

A ATTCCAACCTTGGGGGGGGTTTTCCRRAA ( (SSEEQQ I IDD N NOO: 1 111)); CTGAACCCTAACTYG (SEQ ID NO:12_.

GTACACACTAGGCRC (SEQ ID NO 13); TGCATCCATGGAGYG (SEQ ID NO:14);

GGGCTGGTGAGTGYG (SEQ ID NO 15); TGCAGACAGAAACRC (SEQ ID NO: 16

TTTGGGCTGCAGCMT (SEQ ID NO 17) ; ACACAGCTTGTGAKG (SEQ ID NO: 18

GGGGCTGGGCTGCKA (SEQ ID NO 19) ; ATCTGGATCCCCTMG (SEQ ID NO: 20 T TGGGGGGCCTTGGCCTTAAGGGGGGRRAA ( (SSEEQQ I IDD N NOO: 2 211)) and ATGCCATCTGGATYC (SEQ ID NO: 22

Other oligonucleotides useful in practicing the invention hybridize to a target region located one to several nucleotides downstream of a polymorphic site in a statin response marker. Such oligonucleotides are useful in polymerase-mediated primer extension methods for detecting one of the novel polymorphisms described herein and therefore such oligonucleotides are referred to herein as "primer-extension oligonucleotides". In a preferred embodiment, the 3 '-terminus of a primer-extension oligonucleotide is a deoxynucleotide complementary to the nucleotide located immediately adjacent to the polymorphic site. Particularly preferred primer extension oligonucleotides for detecting CETP gene polymorphisms at the different polymorphic sites in the set comprising a preferred statin response marker haplotype terminate in a nucleotide sequence selected from the group consisting of:

GTTCTTTGGT (SEQ ID NO 23) AGGACCTTCT (SEQ ID NO: 24

ACTGGGGTTC (SEQ ID NO 25) AACCCTAACT (SEQ ID NO: 26

CACACTAGGC (SEQ ID NO 27) ATCCATGGAG (SEQ ID NO: 28

CTGGTGAGTG (SEQ ID NO 29) AGACAGAAAC (SEQ ID NO: 30

GGGCTGCAGC (SEQ ID NO 31) CAGCTTGTGA (SEQ ID NO: 32

GCTGGGCTGC (SEQ ID NO 33) TGGATCCCCT (SEQ ID NO: 34

GCTGCTAGGG (SEQ ID NO 35) and CCATCTGGAT (SEQ ID NO: 36

Termination mixes are chosen to terminate extension of the oligonucleotide at the polymorphic site of interest, or one base thereafter, depending on the alternative nucleotides present at the polymorphic site.

In some embodiments, the genotyping oligonucleotides in a kit of the invention have different labels to allow probing of the identity of nucleotides or nucleotide pairs at two or more polymorphic sites simultaneously. It is also contemplated that a kit of the invention may contain two or more sets of allele- specific primer pairs to allow simultaneous targeting and amplification of two or more regions containing a polymorphic site in a statin response marker.

The oligonucleotides comprising a kit of the invention may also be immobilized on or synthesized on a solid surface such as a microchip, bead, or glass slide (see, e.g., WO 98/20020 and WO 98/20019). Such immobilized oligonucleotides may be used in a variety of polymorphism detection assays, including but not limited to probe hybridization and polymerase extension assays. Immobilized oligonucleotides useful in practicing the invention may comprise an ordered array of oligonucleotides designed to rapidly screen a nucleic acid sample for polymorphisms in multiple genes at the same time.

Kits of the invention may also contain other components such as hybridization buffer (e.g., where the oligonucleotides are to be used as allele-specific probes) or dideoxynucleotide triphosphates (ddNTPs; e.g., where the alleles at the polymorphic sites are to be detected by primer extension). In a preferred embodiment, the set of oligonucleotides consists of primer extension oligonucleotides. The kit may also contain a polymerase and a reaction buffer optimized for primer extension mediated by the polymerase. Preferred kits may also include detection reagents, such as biotin- or fluorescent-tagged oligonucleotides or ddNTPs and/or an enzyme-labeled antibody and one or more substrates that generate a detectable signal when acted on by the enzyme. It will be understood by the skilled artisan that the set of oligonucleotides and reagents for performing the genotyping or haplotyping assay will be provided in separate receptacles placed in the container if appropriate to preserve biological or chemical activity and enable proper use in the assay.

In a particularly preferred embodiment, each of the oligonucleotides and all other reagents in the kit have been quality tested for optimal performance in an assay for determining the alleles at a set of polymorphic sites comprising a statin response marker I or statin response marker II. In a further embodiment, the kit comprises a manual with instructions for performing genotyping assays on a nucleic acid sample from an individual and determining if the individual has a statin response marker I or a statin response marker π based on the results of the assay. The instructions may also contain information to help a physician determine whether or how to use particular statins, alone or in combination with other therapies affecting HDLC levels, to treat an individual with the determined statin response marker.

The methods and kits of the invention are useful for helping physicians make decisions about how to treat an individual. They can be used to predict the HDLC response of an individual to particular statins, in selecting a statin treatment for an individual to achieve an optimal HDLC response, and in choosing a statin treatment appropriate for an individual needing to maintain or increase their HDLC level.

Thus, the invention provides a method for predicting the HDL cholesterol response of an individual to treatment with a statin. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and making a response prediction based on the results of the determining step. The statin is preferably simvastatin, pravastatin sodium or atorvastatin calcium. The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymorphisms is replaced with a substitute polymorphism in linkage disequilibrium with the replaced polymorphism. The statin response marker I or II preferably comprises haplotype (a) or (b) in Table 1. More preferably the haplotype is haplotype (a) (guanine at PS22 and guanine at PS47). The determination of the statin response marker present in an individual can be made using one of the direct or indirect methods described herein. In some preferred embodiments, the determining step comprises ldenurymg tor one or ootn copies ot e genomic locus present m the individual the identity of the nucleotide or nucleotide pair at the set of polymorphic sites comprising the selected statin response marker. Alternatively, the determining step may comprise consulting a data repository that states the individual's copy number for the haplotypes comprising one of the satin response markers I or II. The data repository may be the individual's medical records or a medical data card. In preferred embodiments, the individual is Caucasian.

In some embodiments, if the individual is determined to have a statin response marker I, then the response prediction is that the individual will likely experience an increase in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a decrease in HDLC if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and a negligible change in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day. While if the individual is determined to have a statin response marker II, then the response prediction is that the individual will likely experience a decrease in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day or atorvastatin calcium at 80 mg/day, an increase in HDLC if treated with atorvastatin calcium at 10 mg/day and an increase in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day.

In other embodiments, if the individual is determined to have a statin response marker I, then the response prediction is that the individual will likely experience a better HDLC response if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a worse HDLC response if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and about the same or slightly worse HDLC response if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day compared to an individual determined to have a statin response marker II. Conversely, if the individual is determined to have a statin response marker II, then the response prediction is that the individual will likely experience a worse HDLC response if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a better HDLC response if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and about the same or slightly better HDLC response if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day compared to an individual determined to have a statin response marker I.

The invention also provides a method of selecting a statin to provide an optimal HDL cholesterol response in a human individual in need of statin therapy. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and selecting a statin based on the results of the determining step. The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymorphisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. The haplotype comprising the statin response marker I or II is preferably haplotype (a) or (b) in Table 1. More preferably the haplotype comprises haplotype (a) (guanine at PS22 and guanine at PS47). The determination of whether the individual has a statin response marker I or a statin response marker II can be made using one of the direct or indirect methods described herein. In some preferred embodiments, the determining step comprises identifying for one or both copies of the genomic locus present in the individual the identity of the nucleotide or nucleotide pair at the set of polymoφhic sites comprising the selected haplotype comprising the statin response marker I or II. Alternatively, the determining step may comprise consulting a data repository that states the individual's copy number for one or more haplotypes comprising a statin response marker I or II. The data repository may be the individual's medical records or a medical data card. In preferred embodiments, the individual is Caucasian.

If the individual has a statin response marker I, then the selected statin is simvastatin, a pharmaceutically acceptable salt of simvastatin acid, pravastatin, or a pharmaceutically acceptable salt of pravastatin acid. Preferably, the selected statin is simvastatin or pravastatin sodium. If the individual has a statin response marker II, then the selected statin is pravastatin, a pharmaceutically acceptable salt of pravastatin acid, atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid. Preferably, the selected statin is pravastatin sodium or atorvastatin calcium.

The invention further provides a method for treating an individual in need of maintaining or increasing his or her level of HDL cholesterol with a statin. The method comprises determining whether the individual has a statin response marker I or a statin response marker II and choosing a treatment for the individual based on the results of the determining step. The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. The haplotype comprising the statin response marker I or II is preferably haplotype (a) or (b) in Table 1. More preferably the haplotype comprises haplotype (a) (guanine at PS22 and guanine at PS47). In some embodiments, the determining step comprises identifying for one or both copies of the genomic locus present in the individual the identity of the nucleotide or nucleotide pair at the set of polymoφhic sites comprising the selected haplotype. Alternatively, the determining step may comprise consulting a data repository that states the individual's copy number for a haplotype comprising a statin response marker I or II. The data repository may be the individual's medical records or a medical data card. In preferred embodiments, the individual is Caucasian.

If the individual has a statin response marker II, then the chosen treatment is selected from the group consisting of (a) prescribing pravastatin sodium, (b) prescribing atorvastatin calcium at a dose no greater than 10 mg/day, (c) prescribing simvastatin in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker II, and (d) prescribing 80 mg/day of atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker LI. If the individual has a statin response marker I, then the chosen treatment is selected from the group consisting of (i) prescribing simvastatin, (ii) prescribing pravastatin sodium and (iii) prescribing atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker 1. Therapies capable of increasing mean HDLC levels are well-known to the physician. These therapies include drugs that have this indication on their approved label, e.g., niacin, fibrates, other drugs that mention HDLC increase in their labels (e.g., glitazones, metformin, etc.), and some nutriceuticals, including Vitamin E and supplements containing Vitamin E, or alcohol. Additional therapies that may be recommended are exercise, weight loss and smoking cessation. Preferred therapies capable of increasing or maintaining HDLC are prescription of niacin or a fibrate.Preferred therapies capable of increasing or maintaining HDLC are prescription of niacin or a fibrate.

In other aspects, the invention provides an article of manufacture. In one embodiment, an article of manufacture comprises a pharmaceutical formulation and at least one indicium identifying a population for which the pharmaceutical formulation is indicated. The pharmaceutical formulation comprises a statin as at least one active ingredient. Additionally, the pharmaceutical formulation may be regulated and the indicium may comprise the approved label for the pharmaceutical formulation. The identified population is partially or wholly defined by having a statin response marker I or a statin response marker II. The identified population preferably may be further defined as Caucasian. A population wholly defined by having a statin response marker I or II is one for which there are no other factors which should be considered in identifying the population for which the pharmaceutical formulation is indicated. In contrast, a population that is partially defined by having a statin response marker is one for which other factors may be pertinent to identification of the population for which the pharmaceutical formulation is indicated. Examples of other such factors are age, weight, gender, disease state, possession of other genetic markers or biomarkers, or the like.

The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. Preferably, the haplotype comprises (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b); or (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. More preferably the haplotype comprises guanine at PS20 or PS22 and guanine at PS47. The pharmaceutical formulation may be formulated, in any way known in the art, as a sustained release formulation, but most preferably as a transdermal patch. In some embodiments, the pharmaceutical formulation is a tablet or capsule and the article may further comprise an additional indicium comprising the color or shape of the table or capsule. In other embodiments, the article may further comprise an additional indicium comprising a symbol stamped on the tablet or capsule, or a symbol or logo printed on the approved label.

In some embodiments of this article, in a trial population, the group of individuals having the defining statin response marker exhibits a better mean HDL cholesterol response to the statin than the group of individuals lacking the defining statin response marker. The approved label may comprise a statement about the identified population having the defining statin response marker for which the pharmaceutical formulation is indicated or may further comprise a statement that the pharmaceutical formulation is contraindicated for individuals lacking the defining statin response marker. In some embodiments, the approved label may comprise a statement that the pharmaceutical formulation is indicated only for individuals who score positive for having the defining statin response marker on a specified test, preferably a specified genetic test. In some or all of these embodiments, the label may describe the mean percent change in HDLC expected for the identified population. Additionally, in some or all of these embodiments, the statin is present in the pharmaceutical formulation at an amount effective to reduce LDL cholesterol levels in a population having the defining statin response marker. In one preferred embodiment, the statin comprises simvastatin, a pharmaceutically acceptable salt of simvastatin acid, lovastatin or a pharmaceutically acceptable salt of lovastatin acid, and the defining statin response marker is a statin response marker I. More preferably, the statin is simvastatin or lovastatin, and most preferably the statin is simvastatin and the effective amount is ranging from 20 to 80 mg. In another preferred embodiment, the statin comprises atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker comprises a statin response marker II. More preferably, the statin is atorvastatin calcium and the effective amount is ranging from 10 to 80 mg. In yet another embodiment of this article of manufacture, the pharmaceutical formulation comprises at least a statin and an HDL cholesterol-modulating agent as active ingredients. In this embodiment, in a trial population, the group of individuals having the defining statin response marker exhibits a worse mean HDL cholesterol response to the statin than the group of individuals lacking the defining statin response marker. The HDLC modulating agent may be any known in the art, and preferably is niacin or a fibrate. The approved label may comprise a statement that the pharmaceutical formulation provides a better HDLC response in a population having the defining statin response marker than another pharmaceutical formulation comprising only the statin as the active ingredient. In other embodiments, the approved label may comprise a statement that the pharmaceutical formulation is indicated for individuals who score positive for having the defining statin response marker on a specified test, preferably a specified genetic test. In some or all of these embodiments, the label may describe the mean percent change in HDLC expected for the identified population. Additionally, in some or all of these embodiments, the statin is present in the pharmaceutical formulation at an amount effective to reduce LDL cholesterol levels in a population having the defining statin response marker. In one preferred embodiment, the statin comprises atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker comprises a statin response marker π. More preferably, the statin is atorvastatin calcium and the HDLC modulating agent is niacin. The effective amount of atorvastatin calcium in the pharmaceutical formulation preferably ranges from 10 to 80 mg. In another preferred embodiment, the statin comprises simvastatin, a pharmaceutically acceptable salt of simvastatin acid, lovastatin or a pharmaceutically acceptable salt of lovastatin acid, and the defining statin response marker comprises a statin response marker I. More preferably the statin is simvastatin or lovastatin, and most preferably the statin is simvastatin and the HDLC modulating agent is niacin. The effective amount of simvastatin in the pharmaceutical formulation preferably ranges from 20 to 80 mg. An additional embodiment of the article of manufacture provided by the invention comprises packaging material and a pharmaceutical formulation contained within said packaging material. The pharmaceutical formulation comprises a statin as at least one active ingredient. The packaging material may comprise a label that may state that the pharmaceutical formulation is indicated for a population partly or wholly defined by having a statin response marker I or II. The indicated population preferably may be further defined as Caucasian. The statin response marker I comprises zero copy of a haplotype and the statm response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. Preferably, the haplotype comprises (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b); or (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. More preferably the haplotype comprises guanine at PS20 or PS22 and guanine at PS47. The label may further state that a specified test can be used to identify members of the indicated population. Preferably the specified test is a genetic test. In some embodiments, the statin may be simvastatin, a pharmaceutically acceptable salt of simvastatin acid, lovastatin, or a pharmaceutically acceptable salt of lovastatin acid and the defining statin response marker is a statin response marker I. A population having a statin response marker I exhibits a better mean HDLC response to the statin than a population lacking the defining statin response marker. The statin in this article is preferably simvastatin or lovastatin, and most preferably is simvastatin present in the pharmaceutical formulation at an amount ranging from 20 to 80 mg.

In other embodiments, the statin may be atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker II. A population having the defining statin response marker exhibits a better mean HDLC response to the statin than a population lacking the defining statin response marker. The statin in this article is preferably atorvastatin calcium, most preferably present in the pharmaceutical formulation at an amount ranging from 10 to 80 mg.

Additionally, in other aspects of the invention, a method of manufacturing a drug product comprising a statin as at least one active ingredient is provided. The method comprises combining in a package a pharmaceutical formulation comprising the statin and a label that states that the formulation is indicated for treating a population partially or wholly defined by having a statin response marker I or II. In a trial population, the group of individuals having the defining statin response marker was shown to have a better mean HDLC response to the formulation than did those individuals lacking the defining statin response marker. The label may further state that the pharmaceutical formulation is contraindicated for individuals lacking the defining statin response marker. The indicated and/or contraindicated populations may be identified on the pharmaceutical formulation, on the label or on the package by at least one indicium, such as a symbol or logo, color, or the like. The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) i which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. Preferably, the haplotype comprises (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b); or (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. More preferably the haplotype comprises guanine at PS20 or PS22 and guanine at PS47. The indicated population having the defining statin response marker preferably may be further defined as Caucasian.

In some embodiments, the statin is simvastatin, a pharmaceutically acceptable salt of simvastatin acid, lovastatin, or a pharmaceutically acceptable salt of lovastatin acid and the defining statin response marker is a statin response marker I. More preferably in these embodiments, the statin is simvastatin or a pharmaceutically acceptable salt of simvastatin acid, and most preferably, the statin is simvastatin. In other embodiments, the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker I. In these embodiments, the statin is preferably atorvastatin calcium.

Detecting the presence of a statin response marker I or II in an individual is also useful in a method of seeking regulatory approval for marketing a pharmaceutical formulation for treating a disease or condition in a population defined by the statin response marker. The method comprises: (a) conducting at least one clinical trial which comprises administering the pharmaceutical formulation to first and second treatment groups of patients having the disease or condition, wherein each patient in the first treatment group has a statin response marker I and each patient in the second treatment group has a statin response marker II; (b) demonstrating that one of the treatment groups exhibits a mean percent change in HDLC that is better than the mean percent change in HDLC exhibited by the other treatment group; and (c) filing with a regulatory agency an application for marketing approval of the pharmaceutical formulation with a label stating that the pharmaceutical formulation is indicated for treating the disease or condition in patients having the same statin response marker as in the treatment group exhibiting the better mean change in HDLC. In some embodiments, the pharmaceutical formulation comprises simvastatin or lovastatin, or a pharmaceutically acceptable salt of simvastatin acid or lovastatin acid and the statin response marker in the treatment group exhibiting the better mean change in HDLC is a statin response marker I. More preferably the pharmaceutical formulation comprises simvastatin or lovastatin, and most preferably the pharmaceutical formulation comprises simvastatin. In other preferred embodiments the pharmaceutical formulation comprises atorvastatin or a pharmaceutically acceptable salt of atorvastatin and the statin response marker in the treatment group exhibiting the better mean change in HDLC is a statin response marker II. Most preferably the pharmaceutical formulation comprises atorvastatin calcium. The statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype. The haplotype comprising the statin response markers comprises any one of haplotypes (a) to (j) in table 1, (c) a haplotype in linkage disequilibrium with any one of haplotypes (a) to (j); or (d) a substitute haplotype for any one of haplotypes (a) to (j) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. Preferably, the haplotype comprises (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b); or (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism. More preferably the haplotype comprises guanine at PS20 or PS22 and guanine at PS47. The clinical trial may be conducted by recruiting patients with the disease or condition, determining whether they have a statin response marker I or II and assigning the patients to the first and second treatment groups based on the results of the determining step. The disease or condition may include any for which statin therapy is indicated, e.g., hyperlipidemia, hypercholesterolemia, cardiovascular disease (CVD), presence of CVD risk factors, coronary artery disease , and the like. The patients in each treatment group are preferably administered the same dose of the pharmaceutical formulation, which includes a statin compound as at least one active ingredient. The pharmaceutical formulation may contain other active ingredients, for example another compound known or believed to have therapeutic activity in treating the disease or condition examined in the study or a compound that serves to reduce or block one or more side effects caused by the statin compound. The regulatory agency may be any person or group authorized by the government of a country anywhere in the world to control the marketing or distribution of drugs in that country. Preferably, the regulatory agency is authorized by the government of a major industrialized country, such as Australia, Canada, China, a member of the European Union, Japan, and the like. Most preferably the regulatory agency is authorized by the government of the United States and the type of application for approval that is filed will depend on the legal requirements set forth in the last enacted version of the Food, Drug and Cosmetic Act that are applicable for the pharmaceutical formulation and may also include other considerations such as the cost of making the regulatory filing and the marketing strategy for the composition. For example, if the pharmaceutical formulation has previously been approved for the same cognitive function, then the application might be a paper NDA, a supplemental NDA or an abbreviated NDA, but the application would be a full NDA if the pharmaceutical formulation has never been approved before; with these terms having the meanings applied to them by those skilled in the pharmaceutical arts or as defined in the Drug Price Competition and Patent Term Restoration Act of 1984. Further, in perfoπning any of the methods described herein which require information on the haplotype content of the individual (i.e., the haplotypes and haplotype copy number present in the individual for the polymoφhic sites in haplotypes comprising a statin response marker I or II) or which require knowing if a statin response marker I or U is present in the individual, the individual's CETP haplotype content or statin response marker may be determined by consulting a data repository such as the individual's patient records, a medical data card, a file (e.g. a flat ASCII file) accessible by a computer or other electronic or non-electronic media on which information about the individual's CETP haplotype content or statin response marker can be stored. As used herein, a medical data card is a portable storage device such as a magnetic data card, a smart card, which has an on-board processing unit and which is sold by vendors such as Siemens of Munich Germany, or a flash-memory card. The medical data card may be, but does not have to be, credit-card sized so that it easily fits into pocketbooks, wallets and other such objects carried by the individual. The medical data card may be swiped through a device designed to access information stored on the data card. In an alternative embodiment, portable data storage devices other than data cards can be used. For example, a touch-memory device, such as the "i- button" produced by Dallas Semiconductor of Dallas, Texas can store information about an individual's CETP haplotype content or statin response marker, and this device can be incoφorated into objects such as jewelry. The data storage device may be implemented so that it can wirelessly communicate with routing/intelligence devices through IEEE 802.11 wireless networking technology or through other methods well known to the skilled artisan. Further, as stated above, information about an individual's CETP haplotype content or statin response marker can also be stored in a file accessible by a computer; such files may be located on various media, including: a server, a client, a hard disk, a CD, a DVD, a personal digital assistant such as a Palm Pilot, a tape, a zip disk, the computer's internal ROM (read-only- memory) or the internet or worldwide web. Other media for the storage of files accessible by a computer will be obvious to one skilled in the art. Any or all analytical and mathematical operations involved in practicing the methods of the present invention may be implemented by a computer. For example, the computer may execute a program that assigns CETP haplotype pairs and/or a statin response marker I or II to individuals based on genotype data inputted by a laboratory technician or treating physician. In addition, the computer may output the predicted change in one or more lipoprotein levels in response to a statin following input of the individual's CETP haplotype content or statin response marker, which was either determined by the computer program or input by the technician or physician. Data on which statin response markers were detected in an individual may be stored as part of a relational database (e.g., an instance of an Oracle database or a set of ASCII flat files) containing other clinical and/or haplotype data for the individual. These data may be stored on the computer's hard drive or may, for example, be stored on a CD ROM or on one or more other storage devices accessible by the computer. For example, the data may be stored on one or more databases in communication with the computer via a network.

It is also contemplated that the above described methods and compositions of the invention may be utilized in combination with identifying genotype(s) and/or haplotype(s) for other genomic regions. Preferred embodiments of the invention are described in the following examples. Other embodiments within the scope of the claims herein will be apparent to one skilled in the art from consideration of the specification or practice of the invention as disclosed herein. It is intended that the specification, together with the examples, be considered exemplary only, with the scope and spirit of the invention being indicated by the claims that follow the examples.

EXAMPLES

The Examples herein are meant to exemplify the various aspects of carrying out the invention and are not intended to limit the scope of the invention in any way. The Examples do not include detailed descriptions for conventional methods employed, such as in the performance of genomic DNA isolation, PCR and sequencing procedures. Such methods are well-known to those skilled in the art and are described in numerous publications, for example, Sambrook, Fritsch, and Maniatis, "Molecular Cloning: A Laboratory Manual", 2^nd Edition, Cold Spring Harbor Laboratory Press, USA, (1989).

Example 1

This example illustrates the clinical and biochemical characterization of 679 patients in the patient cohort.

A multicenter, 17-week, (16 weeks controlled), open-label, clinical discovery trial was designed to assess the relationship between genetic HAP ^M Markers and treatment response associated with 4 different commercially available medications, all of which act as 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (cerivastatin sodium [Baycol™], atorvastatin calcium [Lipitor®], simvastatin [Zocor®], and pravastatin sodium [Pravachol®]) in adult subjects with primary hypercholesterolemia. Study medications were packaged by their respective manufacturers and dispensed in a non-blinded fashion by a commercial pharmacist. Prior to randomization, all subjects underwent a screening and baseline period (up to 10 days).

Then subjects were randomly assigned to the recommended starting dose, as stated in the package insert, of 1 of 4 study medications: cerivastatin sodium, atorvastatin calcium, simvastatin, or pravastatin sodium. Following the initial 8 weeks of treatment, subjects proceeded to the highest allowed dose, as stated in the package insert, for an 8-week treatment period. As both periods incoφorate a fixed-dose design, dosing adjustments other than the Week 8 increase were not permitted in this study. Thus, the total duration of therapy was maximally 16 weeks/subject from the point of randomization (8 weeks at the recommended starting dose; plus 8 weeks at the highest allowed dose as stated in the package insert).

Male or female outpatients aged 18 to 75 years with a diagnosis of type Ila or lib hypercholesterolemia who have been on the American Heart Association (AHA) Step I or Step II diet for at least 6 weeks prior to the onset of screening were eligible to participate. Subjects were either treatment-naϊve or previously treated for hypercholesterolemia with any approved medications. Previously treated subjects must have discontinued antiliyperlipidemic medication 4 weeks prior to screening (8 weeks prior to screening if clofibrate [Atromid-S®] was in use) to be eligible. Subject inclusion criteria were based upon medical history assessments and laboratory determinations of cholesterol levels as described by the National Cholesterol Education Program (NCEP)- recommended goal for LDL-cholesterol (> 160 mg/dL for subjects with 0 to 1 coronary heart disease [CHD] risk factor, > 130 mg/dL for those with 2 or more CHD risk factors, or > 100 mg/dL for those with documented CHD or peripheral vascular disease) and had triglyceride levels < 400 mg/dL prior to randomization. Eligible subjects had an LDL-cholesterol level < 240 mg/dL at screening and baseline. Subjects had to demonstrate dietary compliance with the AHA Step I or Step II diet as measured by a food diary at baseline to be eligible for randomization.

The entire patient cohort comprised 679 patients. Subjects were randomly assigned to 1 of 4 treatment groups: 0.4 mg/day cerivastatin sodium, 10 mg/day atorvastatin calcium, 20 mg/day simvastatin, or 10 mg/day pravastatin sodium at baseline. All medication was taken once daily in the evening.

At the Week 8 visit, all subjects proceeded to the highest allowed dose, as stated in the package insert, of their assigned medication. The doses for the treatment groups were as follows: 0.8 mg/day cerivastatin sodium, 80 mg/day atorvastatin calcium, 80 mg/day simvastatin, and 40 mg/day pravastatin sodium. All medication was taken once daily in the evening.

The primary phenotypic endpoint used in the association of treatment response to genetic variability was the percent change from baseline in LDL-cholesterol values after 8 weeks and after 16 weeks of treatment, separately. The final Week 8 value was defined as the mean of the last 2 measurements (Weeks 6 and 8) during the first 8 weeks (low dose) of therapy. The final Week 16 value was defined as the mean of the last 2 measurements (Weeks 14 and 16) during the final 8 weeks (high dose) of therapy. Baseline was defined as the mean of the measurements taken at screening and baseline.

Secondary endpoints studied were the relationship of HAP^™ Markers to the percentage change and absolute change in total cholesterol and HDL-cholesterol; the absolute change from baseline in LDL- cholesterol values after the first 8 weeks (low dose) and after the last 8 weeks (high dose) of treatment, separately; and the change in ratios of total or LDL-cholesterol to HDL-cholesterol.

The patient cohort was characterized with respect to statin taken in treatment as shown below.

Table 4. Demographics, baseline characteristics, and lipid changes for the low-dose compliant population.

Atorvastatin Simvastatin Pravastatin Pooled

Characteristic

(n=155) (n=168) (n=153) (n=476)

Male 65 (41.9%) 91 (54.2%) 64 (41.8%) 220 (46.2%)

Caucasian 135 (87.1%) 146 (86.9%) 126 (82.4%) 407 (85.5%)

Smoker 27 (17.4%) 34 (20.2%) 31 (20.3%) 92 (19.3%)

Drinker 95 (61.3%) 104 (61.9%) 84 (54.9%) 283 (59.5%)

Age (yr)* 56.8 ± 9.7 56.0 ± 10.4 57.0 ± 10.4 56.6 ± 10.2

Height (cm)* 168 ± 11 169 ± 10 169 ± 10 169 ± 10

Weight (kg)* 81.8 ± 16.6 84.1 ± 17.3 84.1 ± 19.0 83.3 ± 17.6

BMI (kg/m2)* 28.8 ± 4.5 29.3 ± 5.1 29.3 ± 6.0 29.1 ±3.2

LDL-C* BL (mg/dL) 172 ± 27 175 ± 25 173 ± 25 173 ± 26

8-week % Δ -39.3 ± 10.0 -35.8 ± 11.0 -21.3 ± 11.3 -32.3 ± 13.2 Atorvastatin Simvastatin Pravastatin Pooled

Characteristic

(n=155) (n=168) (n=153) (n=476)

16-week % Δf -52.2 ± 11.9 -45.1 ± 11.4 -28.8 ± 11.9 -42.0 ± 15.2

HDL-C* BL (mg/dL) 50.6 ± 13.7 47.3 ± 11.2 48.9 ± 12.4 48.9 ± 12.5 8-week % Δ -0.3 ± 10.5 2.1 ± 9.7 0.5 ± 8.6 0.9 ± 9.7 16-week % Δf -3.3 ± 10.4 1.6 ± 10.9 1.2 ± 9.1 -0.1 ± 10.4

TGJ BL (mg/dL) 164 (70, 361) 173 (60, 384) 166 (54, 370) 167 (54, 384) 8-week % Δ -18 (-57, 52) -12 (-62, 234) -6 (-53, 183) -12 (-62, 234) 16-week % Δf -32 (-74, 45) -26 (-60, 70) -10 (-60, 300) -22 (-74, 300)

*BL = baseline; Mean ± Standard Deviation shown.

116-week percent changes are based on the high-dose compliant population: pooled n=409.

{Median (Min, Max).

Statin

Ethnicity Lipitor® Zocor® Pravacol

Afr Am 4 (2%) 7 (4%) 12 (8%)

Am lnd - 1 (0.6%) -

Asian 5 (3%) 3 (2%) 1 (0.6%)

Cauc 133 (79%) 135 (75%) 123 (77%)

Hisp-Lat 10 (6%) 9 (5%) 9 (6%)

Other 1 (0.6%) 2 (1%) 2 (1%)

Not Assigned 15 (9%) 24 (13%) 12 (8%)

Missing - 1 (0.6%) -

Example 2

This example illustrates determination of the genotype of 854 individuals for the polymoφhic sites of interest herein by sequencing. The population of 854 individuals subjected to genotyping comprised individuals initially recruited into the statin study as well as a reference population. The reference population included 93 human individuals, organized into population subgroups by their self- identified ethnogeographic origin. Within this reference population were 82 self-identified unrelated individuals belonging to one of four major population groups: Caucasian (21 individuals), African descent (20 individuals), Asian (20 individuals), or Hispanic/Latino (18 individuals). In addition, the reference population contained three unrelated indigenous American Indians (one from each of North, Central and South America), one three-generation Caucasian family (from the CEPH Utah cohort) and one two-generation African- American family.

Amplification of Target Regions

The following target regions of the CETP gene were amplified using 'tailed' PCR primers, each of which includes a universal sequence forming a noncomplementary 'tail' attached to the 5 ' end of each unique sequence in the PCR primer pairs. The universal 'tail' sequence for the forward PCR primers comprises the sequence 5 '-TGTAAAACGACGGCCAGT-3 ' (SEQ ID NO:37) and the universal 'tail' sequence for the reverse PCR primers comprises the sequence 5 '-AGGAAACAGCTATGACCAT-3 ' (SEQ ID NO:38). The nucleotide positions of the first and last nucleotide of the forward and reverse primers for each region amplified are presented below and correspond to positions in SEQ ID NO: 1 (Figure 1). PCR Primer Pairs ent No. Forward Primer Reverse Primer PCR Product ent 6 5298-5317 complement of 5898-5878 601 nt ent 7 5693-5715 complement of 6269-6248 577 nt ent 12 14442-14463 complement of 14889-14869 448 nt ent 13 14710-14729 complement of 15309-15290 600 nt ent 16 16750-16770 complement of 17225-17205 476 nt

These primer pairs were used in PCR reactions containing genomic DNA isolated from immortalized cell lines for each member of the Index Repository. The PCR reactions were carried out under the following conditions:

Reaction volume = 10 μl

10 x Advantage 2 Polymerase reaction buffer (Clontech) = 1 μl

100 ng of human genomic DNA = 1 μl lO mM dNTP = 0.4 μl

Advantage 2 Polymerase enzyme mix (Clontech) = 0.2 μl

Forward Primer (10 μM) = 0.4 μl

Reverse Primer (10 μM) = 0.4 μl

Water = 6.6μl

Amplification profile: 97°C - 2 min. 1 cycle

97°C - 15 sec. -ι 70°C - 45 sec. I 10 cycles

72°C - 45 sec. J

97°C - 15 sec. -.

64°C - 45 sec. I 35 cycles 72°C - 45 sec. J

Sequencing of PCR Products

The PCR products were purified using a Whatman/Polyfiltronics 100 μl 384 well unifilter plate essentially according to the manufacturers protocol. The purified DNA was eluted in 50 μl of distilled water. Sequencing reactions were set up using Applied Biosystems Big Dye Terminator chemistry essentially according to the manufacturers protocol. The purified PCR products were sequenced in both directions using the appropriate universal 'tail' sequence as a primer. Reaction products were purified by isopropanol precipitation, and run on an Applied Biosystems 3700 DNA Analyzer.

Analysis of Sequences for Alleles at the Polymoφhic Sites

Sequence information was analyzed for the presence of the alleles present at the polymoφhic sites of interest using the Polyphred program (Nickerson et al., Nucleic Acids Res. 14:2745-2751, 1997). The presence of the alleles on each strand was generally determined for each individual. The polymoφhisms studied and their locations in the CETP reference genomic sequence (SEQ ID NO: 1) are listed in Table 5 below. Table 5. Polymoφhic Sites in the CETP Gene correlated with statin-specific HDLC response

Polymoφhic Nucleotide Reference Variant

Site Number Poly Id(a) Position Allele Allele

PS20 14979599 5858 G A

PS22 14979593 5935 G A

PS28 8953014 14541 G A

PS32 8951196 14953 C T

PS35 8951476 15131 C A

PS46 11972308 17098 T G

PS47 8948348 17103 G A

(a) Polyld is a unique identifier assigned to each PS by Genaissance Pharmaceuticals, Inc. Example 3

This example illustrates analysis of the genetic data for the individuals for association with individuals' response to statin.

Haplotypes for the members of the experimental population were assigned using a computer- implemented algorithm for assigning haplotypes to unrelated individuals in a population sample described in WO 01/80156. However, for the analyses assessing the associations between genetic markers and percent change in HDL, the cohort included only patients assigned to the simvastatin, pravastatin sodium, and atorvastatin calcium treatment regimens; for low-dose analyses, patients who were not compliant in the taking of their statins (i.e., who had at least two consecutive visits with an answer "No" to the question of whether they had been between 80% and 120% compliant since the previous visit) during the low-dose period of the study were excluded; for the high-dose analyses, the patients who were not compliant during the low-dose period of the study were excluded as well as those who were not compliant during the high-dose period by the same definition (i.e., had at least two consecutive visits with an answer "No" to the question of whether they had been between 80%> and 120% compliant since the previous visit); patients with incomplete covariate information were excluded. The percent change in HDL cholesterol was determined from the clinical data using the formulas:

Low-dose percent change in HDL = (Low-dose follow-up HDL - Baseline HDL)/(Baseline HDL)* 100.

High-dose percent change in HDL = (High-dose follow-up HDL - Baseline HDL)/(Baseline HDL)* 100,

The parameters in these formulas were defined as follow Baseline HDL - the average of the screening and baseline visits' HDL values, unless the screening and baseline LDL values were more than 15% apart from one another, in which case a second baseline sample of all lipids was collected, and baseline HDL was the average of the two baseline HDL values.

Low-dose follow-up HDL - the average of the 6-week and 8-week values if both were available; otherwise the last single value from among the 4-week, 6-week, 8-week and early termination (if applicable) values was used. High-dose follow-up HDL - the average of the 14-week and 16-week values if both were available; otherwise the last single value from among the 12-week, 14- week, 16-week and early termination (if applicable) values was used.

Haplotype (a) comprising a guanine at PS22 and a guanine at PS47 and Haplotype (b) comprising ■ a guanine at PS20 and a guanine at PS47 in Table 1 were initially screened for association to HDLC changes in response to treatment using a linear regression model (with number of copies treated as a continuous variable), with covariates age, gender, statin assignment (in the all statins combined model only), ethnicity, baseline level of HDL, alcohol consumption, smoking status and body mass index (BMI). Association was tested for each of the 3 individual statins, as well as for the combined statin class. Results for haplotypes (a) and (b) (not shown) were identical except for the low-dose pravastatin sodium statin subset. Due to the high linkage disequilibrium (Δ² = 1.00 for the total experimental population examined herein) between guanine at PS20 and guanine at PS22 that inventors identified in the study herein, copy number for CETP haplotype (a) and (b) divide the population in this study cohort into exactly the same groups except for one individual in the pravastatin sodium arm of the study who was present for the low dose regimen but dropped out of the high dose regimen. Thus only results for CETP haplotype (a) are presented below. Applicants observed that the pattern of least squares means in percent change in HDLC as a function of haplotype copy number followed a dominant model (i.e., that 1 or 2 copies of the haplotype had a similar effect on percent change in HDLC), so the statistical analysis was re-run for haplotype (a) using a dominant model (with one degree of freedom for the marker - 0 vs. 1 and 2 copies combined) with the same covariates. Table 6, below, presents the results obtained when association with the percent change in HDLC was tested at each dose for each of the 3 individual statins, as well as for the combined statin class. The unadjusted p-values characterizing the significance of the difference in percent HDLC for each statin subset and the number of people (count) in the dose compliant cohort having a particular number of copies of the marker are also shown in Table 6.

Table 6. Associations of CETP haplotype (a) with per cent change in HDLC.

^aThe least squares mean percent change in HDLC is presented. (LC,HC) denotes the lower and upper 95% confidence limits on the mean. Count denotes the number of individuals with that number of markers in the indicated statin subset. " When data for all three statins were analyzed together as a class, no significant ditterence was observed in change in percent HDLC with 0 vs 1 or 2 copies of the CETP haplotype at either low or high dose (See Table 6). In contrast, analyzing the data by statin yielded a number of significant results. The group of patients lacking this haplotype experienced a better HDLC response to Zocor® than the patient group having the marker. Conversely, the group of patients that had at least one copy of this haplotype experienced a better HDLC response to Lipitor® than the patient group lacking this haplotype. Mean HDLC response to Pravachol® (pravastatin sodium) in the patient cohort was not affected by the presence or absence of this statin response marker at a statiscally significant level, however the general trend showed that patients having at least one copy of this haplotype experienced a better HDLC response to Pravachol® treatment than the patient group lacking any copy of this haplotype.

Table 7 presents the results obtained for the three individual statins reanalyzed using a single model, at low or high dose, including the statin main effects, the marker main effects and a marker-by- statin-interaction effect, using the same covariates as above. The adjusted least square mean percent change in HDLC in Tables 6 and 7 for the individual statins differ due to the differences between the models used in the two analyses.

Table 7. Associations of CETP haplotype (a) with per cent change in HDLC.

"The least squares mean percent change in HDLC is presented. (LC,HQ denotes the lower and upper 95% confidence limits on the mean. ^bCount denotes the number of individuals with that number of markers in the indicated statin subset.

Using this second model, the marker-by-statin interaction p-value could be calculated and was found to be 0.0005 at high dose and 1.0 x 10^"5 at low dose, demonstrating that the differences in marker effect observed between the three statins were statistically significant at each dosage. Additionally, for the high dose, comparing the statins for patients with 0 copies of the marker yielded a p value of 9.2 x 10^"7 while for patients with 1 or 2 copies the p-value was 0.0.024. At the low dose, comparing the statins for patients with 0 copies of the marker yielded a p value of 1.3 x 10^"4, while for patients with 1 or 2 copies the p-value was 0.0086. These p-values indicate that for patients with a given number of copies of the marker, the difference in the means observed for percent change in HDLC for the three statins is statistically significant at each of the two dosages.

Based on the results shown in Table 7, the three drugs examined were characterized by different marker effects on percent change in HDLC. Zocor® (simvastatin) provides the best HDLC response, at high or low dose, to patients with 0 copies of CETP haplotype (a), improving HDLC by at least 2.76 % relative to baseline. However, for patients with at least 1 copy of CETP haplotype (a), Zocor® provides the worst HDLC response, resulting in about a 3% decrease in HDLC relative to baseline at each dosage. Lipitor® (atorvastatin calcium) provides the worst HDLC response, at high or low dose, to patients with 0 copies of CETP haplotype (a), lowering HDLC by nearly 5% relative to baseline at high dosage. However, for patients with at least one copy of the marker, the effect of Lipitor® on percent change in HDLC is to provide a relatively small percent decrease (-0.41%) at the high dose and an increase of about 2% at the low dose. In contrast to Lipitor® and Zocor®, percent change in HDLC in patients after treatment with Pravachol® was not affected by the number of copies of CETP haplotype (a) at a statistically significant level.

Additionally, the data in Table 7 for patients with a given number of copies of the marker at a given dosage allow determination of the relative ordering of the three statins with respect to producing the best effect on HDLC. For patients with at least one copy of CETP haplotype (a), at high or low dose, Pravachol® provides the best HDLC response, Lipitor® provides the second best response (and is as effective as Pravachol® at the low dose in raising HDLC), while Zocor® provides the worst response. For patients with zero copy of CETP haplotype (a) at each dosage tested, Zocor® provides the best HDLC response, Pravachol® the next best HDLC response and Lipitor® the worst response.

In view of the above, it will be seen that the several advantages of the invention are achieved and other advantageous results attained.

For any and all embodiments of the present invention discussed herein, in which a feature is described in terms of a Markush group or other grouping of alternatives, the inventors contemplate that such feature may also be described by, and that their invention specifically includes, any individual member or subgroup of members of such Markush group or other group.

As various changes could be made in the above methods and compositions without departing from the scope of the invention, it is intended that all matter contained in the above description and shown in the accompanying drawings shall be interpreted as illustrative and not in a limiting sense. All references cited in this specification, including patents and patent applications, are hereby incorporated in their entirety by reference. The discussion of references herein is intended merely to summarize the assertions made by their authors and no admission is made that any reference constitutes prior art. Applicants reserve the right to challenge the accuracy and pertinency of the cited references.

Claims

What is Claimed is:

1. A kit for determimng whether an individual has a statin response marker I or a statin response marker II, the kit comprising a set of oligonucleotides designed for identifying at least one of the alleles at each polymorphic site (PS) in a set of polymorphic sites, wherein the set of PS comprises: (a) PS20 and PS47; (b) PS22 and PS47; (c) PS28 and PS47; (d) PS32 and PS47; (e) PS35 and PS47; (f) PS20 and PS46; (g) PS22 and PS46; (h) PS28 and PS46; (i) PS32 and PS46; (j) PS35 and PS46; (k) a set of polymorphic sites in a linked haplotype for a CETP haplotype selected from the group consisting of (1) guanine at PS20 and guanine at PS47; (2) guanine at PS22 and guanine at PS47; (3) guanine at PS28 and guanine at PS47 (4) cytosine at PS32 and guanine at PS47; (5) cytosine at PS35 and guanine at PS47; (6) guanine at PS20 and guanine at PS46; (7) guanine at PS22 and guanine at PS46;

(8) guanine at PS28 and guanine at PS46; (9) cytosine at PS32 and guanine at PS46; and (10) cytosine at PS35 and guanine at PS46; and (1) a set of PS in a substitute haplotype for any of the CETP haplotypes (1) to (10); wherein the enumerated polymorphic sites in sets (a) to (j) correspond to the following nucleotide positions in SEQ ID NO: 1: PS20, 5858; PS22, 5935; PS28, 14541; PS32, 14953; PS35, 15131; PS46,

17098 and PS47, 17103.

2. The kit of claim 1 , wherein the set of oligonucleotides is designed for identifying both alleles at each member of the selected set of PS.

3. The kit of claim 1, wherein the set of PS is set (a), set (b), set (k) or set (1).

4. The kit of claim 3, wherein the set of PS is set (a) or set (b).

5. The kit of claim 4, wherein the set of PS is set (b).

6. The kit of claim 1 , wherein the individual is Caucasian.

7. The kit of claim 1 , which further comprises a manual with instructions for performing one or more reactions on a human nucleic acid sample to identify the allele(s) present in the individual at each polymorphic site in the set and determining if the individual has a statin response marker I or a statin response marker II based on the identified allele(s).

8. The kit of claim 1 , wherein the linkage disequilibrium between the linked haplotype and the CETP haplotype has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least

0.90, at least 0.95, and 1.0.

9. The kit of claim 8, wherein the set of PS is set (a), set (b), or set (k) and Δ² for the linkage disequilibrium between the linked haplotype and either CETP haplotype (1) or CETP haplotype (2) is at least 0.95.

10. The kit of claim 1, wherein the linkage disequilibrium between a substitute polymorphism in the linked haplotype and a replaced polymorphism in any of the CETP haplotypes (1) to (10) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

11. The kit of claim 10, wherein the set of PS is set (a), set (b) or set (k) and the replaced polymorphism is in set (a) or set (b) and Δ² is at least 0.95.

12. The kit of claim 1, wherein at least one of the oligonucleotides in the set is an allele-specific oligonucleotide comprising a nucleotide sequence selected from the group consisting of SEQ ID NOS:2-8, and their complements.

13. The kit of claim 12, wherein the set of PS is set (b) and the set of oligonucleotides comprises first, second third and fourth allele-specific oligonucleotide (ASO) probes, wherein the first ASO probe comprises SEQ ID NO:3 or its complement, wherein R in SEQ ID NO:3 is guanine, the second ASO probe comprises SEQ ID NO: 3 or its complement, wherein R in SEQ ID NO: 3 is adenine, the third ASO probe comprises SEQ ID NO: 8 or its complement, wherein R in SEQ ID NO: 8 is guanine and ^• the fourth ASO probe comprises SEQ ED NO:8 or its complement, wherein R in SEQ ID NO:8 is adenine.

14. The kit of claim 1, wherein at least one of the oligonucleotides in the set is a primer extension oligonucleotide comprising a nucleotide sequence selected from the group consisting of SEQ ID NOS:9-36.

15. The kit of claim 14, wherein the set of PS is set (b) and the set of oligonucleotides comprises first, second, third and fourth primer-extension oligonucleotides, wherein the first primer-extension oligonucleotide comprises SEQ ID NO:25, the second primer-extension oligonucleotide comprises SEQ ID NO:26, the third primer-extension oligonucleotide comprises SEQ ED NO:35 and the fourth primer-extension oligonucleotide comprises SEQ ID NO:36.

16. A method for determining whether an individual has a statin response marker I or a statin response marker II, the method comprising determining whether the individual has zero or at least one copy of a haplotype selected from the group consisting of: (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j), wherein the polymorphic sites in haplotypes (a) to (j) correspond to the following nucleotide positions in SEQ ED NO:l: PS20, 5858; PS22, 5935; PS28, 14541; PS32, 14953; PS35, 15131; PS46, 17098 and PS47, 17103; and wherein if the individual has zero copies of any of haplotypes (a) to (1), then the individual has statin response marker I and if the individual has at least one copy of any of haplotypes (a) to (1), then the individual has statin response marker II.

17. The method of claim 16, wherein the determining step comprises examining a nucleic acid sample from the individual comprising both genomic copies of each of the polymorphic sites in the selected haplotype; and identifying the phased sequence of alleles present at the polymorphic sites in each genomic copy.

18. The method of claim 17, wherein the identifying step comprises performing an assay selected from the group consisting of (a) a primer extension assay; (b) an allele-specific PCR assay; (c) a nucleic acid amplification assay; (d) a hybridization assay; (e) a mismatch-detection assay; (f) an enzymatic nucleic acid cleavage assay; and (g) a sequencing assay.

19. The method of claim 16, wherein the determining step comprises consulting a data repository that provides information on the haplotypes and haplotype copy number present in the individual for the polymorphic sites in the haplotypes (a) to (1).

20. The method of claim 19, wherein the data repository is the individual's medical records or a medical data card.

21. The method of claim 16, wherein the selected haplotype is haplotype (a), haplotype (b), a linked haplotype for haplotype (a) or haplotype (b), or a substitute haplotype for haplotype (a) or haplotype (b).

22. The method of claim 21, wherein the selected haplotype is haplotype (a) or haplotype (b).

23. The method of claim 16, wherein the linkage disequilibrium between haplotype (k) and any of haplotypes (a) to (j) has a Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

24. The method of claim 23, wherein the selected haplotype is haplotype (a) or haplotype (b) and Δ² for the linkage disequilibrium between haplotype (k) and either haplotype (a) or haplotype (b) is at least 0.95.

25. The method of claim 16, wherein the linkage disequilibrium between a substitute polymoφhism in haplotype (1) and a replaced polymoφhism in any of haplotypes (a) to (j) has a Δ² selected from the group consisting of at least 0.75, least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

26. The method of claim 25, wherein the replaced polymoφhism is in haplotype (a) or haplotype haplotype (b) and Δ² is at least 0.95.

27. The method of claim 16, wherein the individual is Caucasian.

28. The method of claim 16, wherein the individual is diagnosed as having Type Ha or Type lib hypercholesterolemia.

29. The method of claim 16, wherein the individual is a candidate for treatment with a statin.

30. A method for assigning an individual to a first or second statin response marker group, the method comprising: determining whether the individual has zero or at least one copy of a haplotype selected from the group consisting of: (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j), wherein the polymoφhic sites in haplotypes (a) to (j) correspond to the following nucleotide positions in SEQ ID NO:l: PS20,

5858; PS22, 5935; PS28, 14541; PS32, 14953; PS35, 15131; PS46, 17098 and PS47, 17103; and assigning the individual to the first statin response marker group if the individual has zero copies of any of haplotypes (a) to(l) and assigning the individual to the second statin response marker group if the individual has at least one copy of any of haplotypes (a)-(l).

31. The method of claim 30, wherein the determining step comprises examining a nucleic acid sample from the individual comprising both genomic copies of each of the polymoφhic sites in the selected haplotype; and identifying the phased sequence of alleles present at the polymoφhic sites in each genomic copy.

32. The method of claim 31 , wherein the identifying step comprises performing an assay selected from the group consisting of: (a) a primer extension assay; (b) an allele-specific PCR assay; (c) a nucleic acid amplification assay; (d) a hybridization assay; (e) a mismatch-detection assay; (f) an enzymatic nucleic acid cleavage assay; and (g) a sequencing assay.

33. The method of claim 30, wherein the determining step comprises consulting a data repository that provides information on the haplotypes and haplotype copy number present in the individual for the polymoφhic sites in the haplotypes a to 1.

34. The method of claim 33, wherein the data repository is the individual's medical records or a medical data card.

35. The method of claim 30, wherein the selected haplotype is haplotype (a), haplotype (b), a linked haplotype for haplotype (a) or haplotype (b), or a substitute haplotype for haplotype (a) or haplotype (b).

36. The method of claim 35, wherein the selected haplotype is haplotype (a) or haplotype (b).

37. The method of claim 30, wherein the individual is Caucasian.

38. The method of claim 30, wherein the individual is diagnosed as having Type Ila or Type lib hypercholesterolemia.

39. The method of claim 30, wherein the individual is a candidate for treatment with a statin.

40. The method of claim 30, wherein the linkage disequilibrium between haplotype (k) and any of haplotypes (a) to (j) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

41. The method of claim 40, wherein the selected haplotype is haplotype (a) or haplotype (b) and the Δ² for the linkage disequilibrium between haplotype (k) and either of haplotype (a) or haplotype (b) is at least 0.95.

42. The method of claim 30, wherein the linkage disequilibrium between a substitute polymoφhism in haplotype (1) and a replaced polymoφhism in any of haplotypes (a) to (j) has Δ²selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

43. The method of claim 42, wherein the replaced polymoφhism is in haplotype (a) or haplotype (b) and Δ² is at least 0.95.

44. A method of selecting a statin to provide an optimal HDL cholesterol (HDLC) response in an individual in need of statin therapy, the method comprising: determining whether the individual has a statin response marker I or a statin response marker El and selecting a statin based on the results of the determining step.

45. The method of claim 44, wherein if the individual has a statin response marker I, then the selected statin is simvastatin, a pharmaceutically acceptable salt of simvastatin acid, pravastatin, or a pharmaceutically acceptable salt of pravastatin acid; and wherein if the individual has a statin response marker II, then the selected statin is pravastatin, a pharmaceutically acceptable salt of pravastatin acid, atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid.

46. The method of claim 45, wherein if the individual has a statin response marker I, then the selected statin is simvastatin or pravastatin sodium.

47. The method of claim 46, wherein the selected statin is simvastatin.

48. The method of claim 45, wherein if the individual has a statin response marker II, then the selected statin is pravastatin sodium or atorvastatin calcium.

49. The method of claim 44, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

50. The method of claim 49, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a linked haplotype for haplotype (a) or haplotype (b); and (d) a substitute haplotype for haplotype (a) or haplotype (b).

51. The method of claim 50, wherein the selected haplotype is haplotype (a) or (b).

52. The method of claim 50, wherein the linkage disequilibrium between haplotype (c) and haplotype (a) or haplotype (b) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

53. The method of claim 52, wherein Δ² is at least 0.95.

54. The method of claim 50, wherein the linkage disequilibrium between a substitute polymoφhism in haplotype (d) and a replaced polymoφhism in haplotype (a) or (b) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

55. The method of claim 54, wherein Δ² is at least 0.95.

56. The method of claim 44, wherein the individual is Caucasian.

57. The method of claims 44, wherein the individual has been diagnosed as having Type Ila or lib hypercholesterolemia.

58. A method for treating an individual with a statin, wherein the individual needs to maintain or increase her level of High Density Lipoprotein Cholesterol (HDLC), the method comprising: determining whether the individual has a statin response marker I or a statin response marker II; and choosing a treatment for the individual based on the results of the determining step.

59. The method of claim 58, wherein if the individual has a statin response marker H, then the chosen treatment is selected from the group consisting of (a) prescribing pravastatin sodium, (b) prescribing atorvastatin calcium at a dose no greater than 10 mg/day, (c) prescribing simvastatin in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker II, and (d) prescribing 80 mg/day of atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker II, and if the individual has a statin response marker I, then the chosen treatment is selected from the group consisting of (i) prescribing simvastatin, (ii) prescribing pravastatin sodium and (iii) prescribing atorvastatin calcium in combination with another therapy capable of increasing mean HDLC levels in a population having a statin response marker I.

60. The method of claim 58, wherein the determining step comprises consulting a data repository that states whether the individual has a statin response marker I or a statin response marker II.

61. The method of claim 60, wherein said data repository is the individual's medical records or a medical data card.

62. A method for predicting an individual's HDL cholesterol (HDLC) response to treatment with a statin, the method comprising: determining whether the individual has a statin response marker I or a statin response marker II; and making a response prediction based on the results of the determining step.

63. The method of claim 62, wherein if the individual is determined to have a statin response marker I, then the response prediction is that the individual will likely experience an increase in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a decrease in HDLC if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and a negligible change in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day; and wherein if the individual is determined to have a statin response marker II, then the response prediction is that the individual will likely experience a decrease in HDLC if treated with simvastatin at a dose ranging from 20 to 80 mg/day or atorvastatin calcium at 80 mg/day, an increase in HDLC if treated with atorvastatin calcium at 10 mg/day and an increase in HDLC if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day

64. The method of claim 62, wherein if the individual is determined to have a statin response marker I, then the response prediction is that the individual will likely experience a better HDLC response if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a worse HDLC response if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and about the same or slightly worse HDLC response if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day compared to an individual determined to have a statin response marker II; and if the individual is determined to have a statin response marker II, then the response prediction is that the individual will likely experience a worse HDLC response if treated with simvastatin at a dose ranging from 20 to 80 mg/day, a better HDLC response if treated with atorvastatin calcium at a dose ranging from 10 to 80 mg/day, and about the same or slightly better HDLC response if treated with pravastatin sodium at a dose ranging from 10 to 40 mg/day compared to an individual determined to have a statin response marker I.

65. The method of claim 62, wherein the determining step comprises consulting a data repository that states whether the individual has a statin response marker I or a statin response marker II.

66. The method of claim 65, wherein the data repository is the individual's medical records or a medical data card.

67. The method of claim 62, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

68. The method of claim 67, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a linked haplotype for haplotype (a) or haplotype (b); and (d) a substitute haplotype for haplotype (a) or haplotype (b).

69. The method of claim 68, wherein the selected haplotype is haplotype (a) or (b).

70. The method of claim 68, wherein the linkage disequilibrium between haplotype (c) and haplotype (a) or haplotype (b) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

71. The method of claim 70, wherein Δ^" is at least 0.95.

72. The method of claim 68, wherein the linkage disequilibrium between a substitute polymoφhism in haplotype (d) and a replaced polymoφhism in haplotype (a) or (b) has Δ² selected from the group consisting of at least 0.75, at least 0.80, at least 0.85, at least 0.90, at least 0.95, and 1.0.

73. The method of claim 72, wherein Δ² is at least 0.95.

73. A method of seeking regulatory approval for marketing a pharmaceutical formulation comprising a statin as at least one active ingredient for treating a disease or condition in a population partially or wholly defined by having a statin response marker, the method comprising conducting at least one clinical trial which comprises administering the pharmaceutical formulation to first and second treatment groups of patients having the disease or condition, wherein each patient in the first treatment group has a statin response marker I and each patient in the second treatment group has a statin response marker II; demonstrating that one of the treatment groups exhibits a mean percent change in HDLC that is better than the mean percent change in HDLC exhibited by the other treatment group; and filing with a regulatory agency an application for marketing approval of the pharmaceutical formulation with a label stating that the pharmaceutical formulation is indicated for treating the disease or condition in patients having the same statin response marker as in the treatment group exhibiting the better mean change in HDLC.

74. The method of claim 73, wherein the statin is selected from the group consisting of simvastatin or a pharmaceutically acceptable salt of simvastatin acid and the statin response marker in the treatment group exhibiting the better mean change in HDLC is a statin response marker I.

75. The method of claim 74, wherein the statin is simvastatin.

76. The method of claim 73, wherein the statin is atorvastatin, a pharmaceutically acceptable salt of atorvastatin acid, pravastatin, or a pharmaceutically acceptable salt of pravastatin acid and the statin response marker in the treatment group exhibiting the better mean change in HDLC is a statin response marker II.

77. The method of claim 76, wherein the statin is atorvastatin calcium.

78. The method of claim 76, wherein the statin is pravastatin sodium.

79. The method of claim 73, wherein the disease or condition is a cardiovascular disorder and the statin is present in the pharmaceutical formulation in an amount effective for lowering LDL cholesterol levels in each of the first and second treatment groups.

80. The method of claim 73, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

81. The method of claim 80, wherein the selected haplotype is (a) or (b).

82. The method of claim 80, wherein the linkage disequilibrium between haplotype (k) and any of haplotype (a) or (b) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

83. The method of claim 82, wherein Δ² is at least 0.95.

84. The method of claim 80, wherein the linkage disequilibrium between a substitute polymoφhism in hhaapplloottyyppee ((11)) aanndd aa rreeppllaacceedd ppoollyymmooφφhhiissmm iinn hhaapplloottyyppee ((aa)) ooir (b) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

85. The method of claim 84, wherein Δ² is at least 0.95.

86. The method of claim 73, wherein the individual is Caucasian.

87. The method of claim 73, wherein the disease or condition is hypercholesterolemia.

88. The method of claim 73, wherein the regulatory agency is the United States Food and Drug Administration or the European Agency for the Evaluation of Medicinal Products.

89. An article of manufacture, comprising a pharmaceutical formulation and at least one indicium identifying a population for whom the pharmaceutical formulation is indicated, wherein the pharmaceutical formulation comprises a statin as at least one active ingredient and the identified population is partially or wholly defined by having a statin response marker I or a statin response marker II, wherein a trial population having the defining statin response marker exhibits a better mean HDL cholesterol (HDLC) response to the statin than a trial population lacking the defining statin response marker.

90. The article of claim 89, wherein marketing of the pharmaceutical formulation is regulated and the indicium comprises the approved label for the pharmaceutical formulation, wherein the approved label states that the pharmaceutical formulation is indicated for individuals having the defining statin response marker.

91. The article of claim 90, wherein the approved label for the phannaceutical formulation further states that the pharmaceutical formulation is contraindicated for individuals lacking the defining statin response marker.

92. The article of claim 89, wherein marketing of the pharmaceutical formulation is regulated and the indicium comprises the approved label for the pharmaceutical formulation, wherein the approved label states that the pharmaceutical formulation is indicated only for individuals who score positive for having the defining statin response marker on a specified genetic test.

93. The article of claim 92, wherein the approved label describes the mean percent change in HDLC expected for the identified population.

94. The article of claim 89, wherein the statin is present in the pharmaceutical formulation at an amount effective to reduce mean LDL cholesterol levels in a population having the defining statin response marker.

95. The article of claim 89, wherein the statin is simvastatin or a pharmaceutically acceptable salt of simvastatin acid and the defining statin response marker is statin response marker I.

96. The article of claim 95, wherein the statin is simvastatin and the amount effective to reduce mean LDL cholesterol levels ragnes from 20 to 80 mg.

97. The article of claim 89, wherein the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is statin response marker I.

98. The article of claim 97, wherein the statin is atorvastatin calcium and the amount effective to reduce mean LDL cholesterol level ranges from 10 to 80 mg.

99. The article of claim 89, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker IE comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

100. The article of claim 99, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b); and (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism.

101. The article of claim 100, wherem the linkage disequilibrium between haplotypes (a) or (b) and haplotype (C) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

102. The article of claim 68101 wherein Δ² is at least 0.95.

103. The article of claim 100, wherein the linkage disequilibrium between a replaced polymoφhism in haplotype (a) or (b) and a substitute polymoφhism in haplotype (d) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

104. The article of claim 103, wherein Δ² is at least 0.95.

105. The article of claim 89, wherein the pharmaceutical formulation is a sustained release formulation.

106. The article of claim 89, wherein the article further comprises an additional indicium identifying the indicated population.

107. The article of claim 106, wherein the pharmaceutical formulation is a tablet or capsule and the additional indicium comprises the color or shape of the tablet or capsule.

108. The article of claim 106, wherein the pharmaceutical formulation is a tablet or capsule and the additional indicium comprises a symbol stamped on the tablet or capsule.

109. The article of claim 89, wherein the identified population is further defined as being Caucasian.

110. An article of manufacture, comprising packaging material and a pharmaceutical formulation contained within said packaging material, wherein the pharmaceutical formulation comprises a statin as at least one active ingredient and said packaging material comprises a label which states that the pharmaceutical formulation is indicated for a population partly or wholly defined by having a statin response marker I or a statin response marker IE, wherein a first trial population consisting of individuals having the defining statin response marker exhibits a better mean HDLC response to the statin than a second trial population consisting of individuals lacking the defining statin response marker.

111. The article of claim 110, wherein the statin response marker I comprises zero copy of a haplotype and the statm response marker II comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to ( ); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

112. The article of claim 111, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b);.and (d) a substitute haplotype for haplotype (a) or (b) in which one or both of the polymoφhisms is replaced with a substitute polymoφhism in linkage disequilibrium with the replaced polymoφhism.

113. The article of claim 112, wherein the selected haplotype is (a) or (b).

114. The article of claim 112, wherein the linkage disequilibrium between haplotypes (a) or (b) and the haplotype (c) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

115. The article of claim 114, wherein Δ² is at least 0.95.

116. The article of claim 112, wherein the linkage disequilibrium between a replaced polymoφhism in haplotype (a) or (b) and a substitute polymoφhism in haplotype (d) has Δ^z selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

117. The article of claim 116, wherein Δ² is at least 0.95.

118. The method of claim 110, wherein the statin is simvastatin or a pharmaceutically acceptable salt of simvastatin and the defining statin response marker is a statin response marker I.

119. The article of claim 118, wherein the statin is simvastatin.

120. The article of claim 119, wherein the simvastatin is present in the pharmaceutical formulation at an amount ranging from 20 to 80 mg.

121. The article of claim 110, wherein the label further states that a specified genetic test can be used to identify individuals having the defining statin response marker.

122. The article of claim 110, wherein the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker II.

123. The article of claim 122, wherein the statin is atorvastatin calcium which is present in an amount ranging from 10 to 80 mg.

124. An article of manufacture, comprising a pharmaceutical formulation and at least one indicium identifying a population for whom the pharmaceutical formulation is indicated, wherein the pharmaceutical formulation comprises as separate active ingredients a statin and an

HDL cholesterol (HDLC) modulating agent, wherein the identified population is partially or wholly defined by having a statin response marker I or a statin response marker II, wherein a trial population having the defining statin response marker exhibits a better mean HDLC response to the pharmaceutical formulation than to the statin alone.

125. The article of claim 124, wherein marketing of the pharmaceutical formulation is regulated and the indicium comprises the approved label for the pharmaceutical formulation, wherein the approved label states that the pharmaceutical formulation provides a better HDLC response in a population having the defining statin response marker than a pharmaceutical formulation comprising only the statin as the active ingredient.

126. The article of claim 125, wherein the approved label describes the mean percent change in HDLC expected for the identified population.

127. The article of claim 124, wherein marketing of the pharmaceutical formulation is regulated and the indicium comprises the approved label for the pharmaceutical formulation, wherein the approved label states that the pharmaceutical formulation is indicated for individuals who score positive for having the defining statin response marker on a specified genetic test.

128. The article of claim 124, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

129. The article of claim 128, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) a haplotype in linkage disequilibrium with either haplotypes (a) or (b);.and (d) a substitute haplotype for haplotype (a) or (b).

130. The article of claim 129, wherein the selected haplotype is (a) or (b).

131. The article of claim 129, wherein the linkage disequilibrium between haplotypes (a) or (b) and haplotype (c) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and

1.0.

132. The article of claim 131, wherein Δ² is at least 0.95.

133. The article of claim 129, wherein the linkage disequilibrium between a replaced polymoφhism in haplotype (a) or (b) and a substitute polymoφhism in haplotype (d) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

134. The article of claim 133, wherein Δ² is at least 0.95.

135. The article of claim 124, wherein the HDLC modulating agent is niacin or a fibrate.

136. The article of claim 124, wherein the statin is present in the pharmaceutical formulation at an amount effective to reduce LDL cholesterol levels in a population having the defining statin response marker.

137. The article of claim 124, wherein the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker I.

138. The article of claim 137, wherein the statin is atorvastatin calcium and the HDLC modulating agent is niacin.

139. The article of claim 124, wherein the statin is shnvastatin or a phaπnaceutically acceptable salt of simvastatin acid and the defining statin response marker is a statm response marker II.

140. The article of claim 139, wherein the statin is simvastatin and the HDLC modulating agent is niacin.

141. A method for-manufacturing a drug product comprising combining in a package a pharmaceutical formulation comprising a statin as at least one active ingredient and a label which states that the pharmaceutical formulation is indicated for treating a population partially or wholly defined by having a statin response marker I or a statin response marker II, wherein a trial population having the defining statin response marker exhibits a better mean HDL cholesterol (HDLC) response to the statin than a trial population lacking the defining statin response marker.

142. The method of claim 141, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker IE comprises at least one copy of the haplotype, wherein the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to G).

143. The method of claim 142, wherein the linkage disequilibrium between any of haplotypes (a) to (j) and haplotype (k) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

144. The method of claim 143, wherem Δ² is at least 0.95.

145. The method of claim 142, wherein the linkage disequilibrium between a replaced polymoφhism in any of haplotypes (a) to (j) and a substitute polymoφhism in haplotype (1) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1.0.

146. The method of claim 145, wherein Δ² is at least 0.95.

147. The method of claim 141, wherein the statin is simvastatin or a pharmaceutically acceptable salt of simvastatin acid and the defining statin response marker is a statin response marker 1.

148. The method of claim 147, wherein the statin is simvastatin.

149. The method of claim 141, wherein the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is statin response marker II.

150. The method of claim 149, wherein the statin is atorvastatin calcium.

151. The method of claim 141, wherein the label further states that the indicated population is further defined as being Caucasian.

152. The method of claim 141, wherein the label further states that the pharmaceutical formulation is contraindicated for individuals lacking the defining statin response marker.

153. A method for marketing a drug product comprising a statin as at least one active ingredient for treating a disease or condition in a population partially or wholly defined by having a statin response marker I or a statin response marker II, the method comprising promoting to a target audience the use of the drug product for treating individuals who belong to the defined population, wherein a trial population having the defining statin response marker exhibits a better mean HDL cholesterol (HDLC) response to the statin than a trial population lacking the defining statin response marker.

154. The method of claim 153, wherein the statin response marker I comprises zero copy of a haplotype and the statin response marker II comprises at least one copy of the haplotype, wherem the haplotype is selected from the group consisting of (a) guanine at PS20 and guanine at PS47; (b) guanine at PS22 and guanine at PS47; (c) guanine at PS28 and guanine at PS47 (d) cytosine at PS32 and guanine at PS47; (e) cytosine at PS35 and guanine at PS47; (f) guanine at PS20 and guanine at PS46; (g) guanine at PS22 and guanine at PS46; (h) guanine at PS28 and guanine at PS46; (i) cytosine at PS32 and guanine at PS46; (j) cytosine at PS35 and guanine at PS46; (k) a linked haplotype for any one of haplotypes (a) to (j); and (1) a substitute haplotype for any one of haplotypes (a) to (j).

155. The method of claim 154, wherein the linkage disequilibrium between any of haplotypes (a) to (j) and haplotype (k) has Δ² selected from the group consisting of at least 0.75,. at least 0.90, at least 0.95, and 1.0.

156. The method of claim 158, wherein Δ² is at least 0.95.

157. The method of claim 154, wherein the linkage disequilibrium between a replaced polymoφhism iinn aannyy ooff hhaapplloottyyppeess ((aa)) ttoo ((jj)) aanndd aa ssuubbssttiittuuttee ppoollyymmooφφhhiissmm iinn hhaappllotype (1) has Δ² selected from the group consisting of at least 0.75, at least 0.90, at least 0.95, and 1,0.

158. The method of 157, wherein Δ² is at least 0.95.

159. The method of claim 154, wherein the haplotype is (a) or (b).

160. The method of claim 153, wherein the statin is selected from the group consisting of simvastatin and a pharmaceutically acceptable salt of simvastatin acid and the defining statin response marker is a statin response marker I.

161. The method of claim 153, wherein the statin is atorvastatin or a pharmaceutically acceptable salt of atorvastatin acid and the defining statin response marker is a statin response marker II.

162. The method of claim 153, wherein the disorder or condition is dyslipidemia, hyperlipidemia, or hypercholesterolemia.

163. The method of claim 153, wherein the target audience is selected from physicians, pharmacists, patients with the disorder, and a government regulatory agency.

164. The method of claim 153, wherein the target audience comprises physicians and the promoting step comprises providing dosage and performance information from one or more clinical trials of individuals having the defining statin response marker.

165. The method of claim 164, wherein the promoting step further comprises providing physicians with a supply of free diagnostic kits for identifying patients who have the defining statin response marker.

166. The method of claim 153, wherein the individuals are Caucasian.