WO2013065072A1 - Variantes de risque du cancer de la prostate - Google Patents

Variantes de risque du cancer de la prostate Download PDF

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Publication number
WO2013065072A1
WO2013065072A1 PCT/IS2012/000006 IS2012000006W WO2013065072A1 WO 2013065072 A1 WO2013065072 A1 WO 2013065072A1 IS 2012000006 W IS2012000006 W IS 2012000006W WO 2013065072 A1 WO2013065072 A1 WO 2013065072A1
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Prior art keywords
allele
chr8
marker
prostate cancer
susceptibility
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PCT/IS2012/000006
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English (en)
Inventor
Julius Gudmundsson
Patrick Sulem
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Decode Genetics Ehf
Illumina Inc.
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Publication of WO2013065072A1 publication Critical patent/WO2013065072A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

Definitions

  • Prostate cancer is the most frequently diagnosed non-cutaneous malignancy among men in industrialized countries, and in the United States, 1 in 8 men will develop prostate cancer during his life (Simard, J. et al., Endocrinology 143(6): 2029-40 (2002)).
  • prostate cancer An average 40% reduction in life expectancy affects males with prostate cancer. If detected early, prior to metastasis and local spread beyond the capsule, prostate cancer can be cured (e.g., using surgery). However, if diagnosed after spread and metastasis from the prostate, prostate cancer is typically a fatal disease with low cure rates. While prostate-specific antigen (PSA)-based screening has aided early diagnosis of prostate cancer, it is neither highly sensitive nor specific (Punglia et.al., N Engl 3 Med. 349(4) :335-42 (2003)). This means that a high percentage of false negative and false positive diagnoses are associated with the test. The consequences are both too many instances of missed cancers and unnecessary follow-up biopsies for those without cancer.
  • PSA prostate-specific antigen
  • PSA testing also has difficulty with specificity and predicting prognosis.
  • PSA levels can be abnormal in those without prostate cancer.
  • benign prostatic hyperplasia BPH
  • a variety of non-cancer conditions may elevate serum PSA levels, including urinary retention, prostatitis, vigorous prostate massage and ejaculation.
  • DRE Digital rectal examination
  • markers are associated with risk of prostate cancer. Such markers are useful in a number of prognostic and diagnostic applications, as described further herein.
  • the markers can also be used in certain aspects that relate to development of markers for diagnostic use, systems and apparati for diagnostic use, as well as in methods that include selection of individuals based on their genetic status with respect to such variants. These and other aspects of the invention are described in more detail herein.
  • the invention relates to a method of determining a susceptibility to prostate cancer, the method comprising analyzing nucleic acid sequence data about a human individual identifying at least one allele of at least one polymorphic marker, wherein different alleles of the at least one polymorphic marker are associated with different susceptibilities to prostate cancer in humans, and determining a susceptibility to prostate cancer from the nucleic acid sequence data, wherein the at least one polymorphic marker is selected from the group consisting of chr8: 128173299 (rsl83373024) (SEQ ID NO: l) and rsl46851087 (SEQ ID NO:8), and correlated markers in linkage disequilibrium therewith.
  • the at least one polymorphic marker is selected from the group consisting of rsl83373024, and markers in linkage disequilibrium therewith.
  • the markers in linkage disequilibrium with rsl83373024 are selected from the markers listed in Table 1(A).
  • the nucleic acid sequence data is sequence data from-a nucleic acid sample from the human individual.
  • polymorphic markers can comprise variations comprising one or more nucleotides at the nucleotide level. Sequence data indicative of a particular polymorphisms, in particular with respect to specific alleles of a polymorphism, is thus indicative of the nucleotides that are present at the specific polymorphic site(s) that characterize the polymorphism. For polymorphisms that comprise a single nucleotide, (so called single nucleotide polymorphisms (SNPs)), the sequence data thus includes at least sequence for the single nucleotide characteristic of the polymorphism.
  • SNPs single nucleotide polymorphisms
  • the invention in another aspect relates to a method for determining a susceptibility to prostate cancer in a human individual, comprising determining the presence or absence of at least one allele of at least one polymorphic marker in a nucleic acid sample obtained from the individual, or in a genotype dataset from the individual, wherein the at least one polymorphic marker is chr8: 128173299 and rsl46851087, and markers in linkage disequilibrium therewith, and wherein determination of the presence of the at least one allele is indicative of a susceptibility to prostate cancer.
  • the invention also provides an assay for determining a susceptibility to prostate cancer in a human subject, the assay comprising steps of (i) obtaining a nucleic acid sample from the human subject (ii) assaying the nucleic acid sample to determine the presence or absence of at least one allele of at least one polymorphic marker conferring increased susceptibility to prostate cancer in humans, and (iii) determining a susceptibility to prostate cancer for the human subject from the presence or absence of the at least one allele, wherein the at least one allele is selected from the group consisting of chr8: 128173299 allele G and rsl46851087 allele A, and marker alleles correlated therewith by values of of r 2 > 0.5, wherein determination of the presence of the at least one allele is indicative of an increased susceptibility to prostate cancer for the subject.
  • kits for assessing susceptibility to prostate cancer in a human individual comprising (i) reagents for selectively detecting at least one allele of at least one polymorphic marker in the genome of the individual, wherein the polymorphic marker is selected from the group consisting chr8: 128173299 (SEQ ID NO: l) and rsl46851087 (SEQ ID NO:8), and correlated markers in linkage disequilibrium therewith, and (ii) a collection of data comprising correlation data between the polymorphic markers assessed by the kit and susceptibility to prostate cancer.
  • the invention also provides computer-implemented aspects for carrying out the methods described herein.
  • the invention provides a system for identifying susceptibility to prostate cancer in a human subject, the system comprising (1) at least one processor; (2) at least one computer-readable medium; (3) a susceptibility database operatively coupled to a computer-readable medium of the system and containing population information correlating the presence or absence of at least one marker allele and susceptibility to prostate cancer in a population of humans; (4) a measurement tool that receives an input about the human subject and generates information from the input about the presence or absence of the at least one allele in the human subject; and (5) an analysis tool that (i) is operatively coupled to the susceptibility database and the measurement tool, (ii) is stored on a computer-readable medium of the system, (iii) is adapted to be executed on a processor of the system, to compare the information about the human subject with the population information in the susceptibility database and generate a conclusion with respect to susceptibility to prostate cancer for the human subject
  • Another computer-implemented aspect provided herein relates to a system for assessing or selecting a treatment protocol for a subject diagnosed with prostate cancer, comprising ( 1) at least one processor; (2) at least one computer-readable medium; (3) a medical treatment database operatively connected to a computer-readable medium of the system and containing information correlating the presence or absence of at least one marker allele selected from the group consisting of chr8: 128173299 allele G and rsl46851087 allele A, and marker alleles correlated therewith by values of r 2 > 0.5, and efficacy of treatment regimens for prostate cancer; (4) a measurement tool to receive an input about the human subject and generate information from the input about the presence or absence of the at least one marker allele in a human subject diagnosed with prostate cancer; and (5) a medical protocol tool operatively coupled to the medical treatment database and the measurement tool, stored on a computer- readable medium of the system, and adapted to be executed on a processor of the system, to compare the information with respect to presence or
  • FIG 1 provides a diagram illustrating a system comprising computer implemented methods utilizing risk variants as described herein.
  • FIG 2 shows an exemplary system for determining risk of prostate cancer as described further herein.
  • FIG 3 shows a system for selecting a treatment protocol for a subject diagnosed with prostate cancer.
  • FIG 4 shows schematic view of a region on chromosome 8q24 containing several prostate cancer risk variants
  • Triangles denote imputed association results (-log(P-value) of the unconditional analysis.
  • Crosses denote association results after step-1 of the conditional analysis (the logistic regression), boxed denote ass results after step-2, and filled circles denote association result after step-3 (See Example diamonds below the X-axis denote the previously reported prostate cancer risk SNPs on 8q24 in populations of European descent (1, rsl2543663; 2, rsl0086908; 3, rsl016343; 4, rsl3252298; 5, rsl6901979; 6, rsl6902094; 7, rs445114 / rs620861; 8, rs6983267; 9, rsl447295) and the novel SNPs (A, rsl83373024 and B, rs 188140481).
  • Shown are the CEU HapMap population recombination rate and the pairwise correlation coefficient (r 2 ) for SNPs in the 1 megabase (Mb) region depicted.
  • nucleic acid sequences are written left to right in a 5' to 3' orientation.
  • Numeric ranges recited within the specification are inclusive of the numbers defining the range and include each integer or any non-integer fraction within the defined range.
  • all technical and scientific terms used herein have the same meaning as commonly understood by the ordinary person skilled in the art to which the invention pertains.
  • the marker can comprise any allele of any variant type found in the genome, including SNPs, mini- or microsatellites, translocations and copy number variations (insertions, deletions, duplications).
  • Polymorphic markers can be of any measurable frequency in the population. For mapping of disease genes, polymorphic markers with population frequency higher than 5-10% are in general most useful. However, polymorphic markers may also have lower population frequencies, such as 1-5% frequency, or 0.1-1% frequence, or even lower frequency. The term shall, in the present context, be taken to include polymorphic markers with any population frequency.
  • an “allele” refers to the nucleotide sequence of a given locus (position) on a chromosome.
  • a polymorphic marker allele thus refers to the composition (i.e., sequence) of the marker on a chromosome.
  • CEPH sample (Centre d'Etudes du Polymorphisme Humain, genomics repository, CEPH sample 1347-02) is used as a reference, the shorter allele of each microsatellite in this sample is set as 0 and all other alleles in other samples are numbered in relation to this reference.
  • allele 1 is 1 bp longer than the shorter allele in the sample
  • allele 2 is 2 bp longer than the shorter allele in the CEPH sample
  • allele 3 is 3 bf than the lower allele in the CEPH sample
  • allele -1 is 1 bp shorter than the shorter allele in the CEPH sample
  • allele -2 is 2 bp shorter than the shorter allele in the CEPH sample, etc.
  • Sequence conucleotide ambiguity as described herein is as proposed by IUPAC-IUB. These codes are compatible with the codes used by the EMBL, GenBank, and PIR databases.
  • a nucleotide position at which more than one sequence is possible in a population is referred to herein as a "polymorphic site”.
  • SNP Single Nucleotide Polymorphism
  • SNP Single Nucleotide Polymorphism
  • rsl83373024 ID identification tag as assigned to each unique SNP by the National Center for Biotechnological Information (NCBI).
  • NCBI National Center for Biotechnological Information
  • Another terminology for identification of a SNP or polymorphic marker reported herein indicates the chromosome on which the SNP is located, followed by its position on the chromosome.
  • chr8: 128173299 refers to a DNA sequence variation on chromosome 8 at base pair position 128173299 according to build 36 of the human genome assembly.
  • the SNPs disclosed herein are further identified by their genomic flanking sequences, as illustrated in the accompanying sequence listing.
  • a “variant”, as described herein, refers to a segment of DIMA that differs from the referer
  • a “marker” or a “polymorphic marker”, as defined herein, is a variant. Alleles that differ the reference are referred to as “variant” alleles. A “variant” is sometimes also referred to as a "mutant”.
  • a "microsatellite” is a polymorphic marker that has multiple small repeats of bases that are 2-8 nucleotides in length (such as CA repeats) at a particular site, in which the number of repeat lengths varies in the general population.
  • An “indel” is a common form of polymorphism comprising a small insertion or deletion that is typically only a few nucleotides long.
  • haplotype refers to a segment of genomic DIMA that is characterized by a specific combination of alleles arranged along the segment.
  • a haplotype comprises one member of the pair of alleles for each polymorphic marker or locus along the segment.
  • the haplotype can comprise two or more alleles, three or more alleles, four or more alleles, or five or more alleles.
  • susceptibility refers to the proneness of an individual towards the development of a certain state (e.g. , a certain trait, phenotype or disease), or towards being less able to resist a particular state than the average individual.
  • the term encompasses both increased susceptibility and decreased susceptibility.
  • particular alleles at polymorphic markers and/or haplotypes of the invention as described herein may be characteristic of increased susceptibility (i.e., increased risk) of prostate cancer, as characterized by a relative risk (RR) or odds ratio (OR) of greater than one for the particular allele or haplotype.
  • the markers and/or haplotypes of the invention are characteristic of decreased susceptibility (i.e., decreased risk) of prostate cancer, as characterized by a relative risk of less than one.
  • look-up table is a table that correlates one form of data to another form, or one or more forms of data to a predicted outcome to which the data is relevant, such as phenotype or trait.
  • a look-up table can comprise a correlation between allelic data for at least one polymorphic marker and a particular trait or phenotype, such as a particular disease diagnosis, that an individual who comprises the particular allelic data is likely to display, or is more likely to display than individuals who do not comprise the particular allelic data.
  • Look-up tables can be multidimensional, i.e. they can contain information about multiple alleles for single markers simultaneously, or they can contain information about multiple markers, and they may also comprise other factors, such as particulars about diseases diagnoses, racial information, biomarkers, biochemical measurements, therapeutic methods or drugs, etc.
  • a "computer-readable medium” is an information storage medium that can be accessed by a computer using a commercially available or custom-made interface.
  • Exemplary computer- readable media include memory (e.g., RAM, ROM, flash memory, etc.), optical storage media (e.g. , CD-ROM), magnetic storage media (e.g., computer hard drives, floppy disks, etc. ) cards, or other commercially available media.
  • Information may be transferred between i of interest and a medium, between computers, or between computers and the computer- readable medium for storage or access of stored information. Such transmission can be electrical, or by other available methods, such as IR links, wireless connections, etc.
  • nucleic acid sample refers to a sample obtained from an individual that contains nucleic acid (DNA or RNA).
  • the nucleic acid sample comprises genomic DNA.
  • a nucleic acid sample can be obtained from any source that contains genomic DNA, including a blood sample, sample of amniotic fluid, sample of cerebrospinal fluid, or tissue sample from skin, muscle, buccal or conjunctival mucosa, placenta, gastrointestinal tract or other organs.
  • prostate cancer therapeutic agent refers to an agent that can be used to ameliorate or prevent symptoms associated with prostate cancer.
  • antisense agent or “antisense oligonucleotide” refers, as described herein, to molecules, or compositions comprising molecules, which include a sequence of purine an pyrimidine heterocyclic bases, supported by a backbone, which are effective to hydrogen bond to a corresponding contiguous bases in a target nucleic acid sequence.
  • the backbone is composed of subunit backbone moieties supporting the purine and pyrimidine heterocyclic bases at positions which allow such hydrogen bonding. These backbone moieties are cyclic moieties of 5 to 7 atoms in size, linked together by phosphorous-containing linkage units of one to three atoms in length.
  • the antisense agent comprises an oligonucleotide molecule.
  • polymorphic variants on chromosome 8q24 are associated with risk of developing prostate cancer.
  • Certain alleles of certain polymorphic marker have been found to be present at increased frequency in individuals with diagnosis of prostate cancer compared with controls. These polymorphic markers are thus associated with risk of prostate cancer.
  • the particular polymorphic marker described herein are contemplated to be useful as a marker for determining susceptibility to prostate cancer. These markers are believed to be useful in a range of diagnostic applications, as described further herein.
  • chr8: 128173299 (rsl83373024) (SEQ ID NO: l) represents one such variant.
  • the risk allele G was found to be in excess in individuals with prostate cancer, leading to increased risk of prostate cancer with an OR value of 2.69 and a corresponding P-value of association of 4.0xl0 '23 .
  • This marker and correlated markers in linkage disequilibrium therewith are therefore useful markers for determining susceptibility to prostate cancer.
  • Exemplary surrogate markers of rsl83373024 are shown in Table 1A herein. Such surrogate markers, which are correlated with rsl83373024, are all also useful for assessing risk of prostate c because they are all detecting the same underlying association.
  • a second signal was founded on chromosome 8q24, manifested by marker rsl46851087 (SEQ ID NO:8). This marker has been found to lead to increased risk of prostate cancer with an OR of 2.25 and a corresponding P-value of association of 5.21xl0 "14 .
  • the present invention provides a method of determining a
  • the method comprising analyzing nucleic acid sequence data about a human individual identifying at least one allele of at least one polymorphic marker, wherein different alleles of the at least one polymorphic marker are associated with different susceptibilities to prostate cancer in humans, and determining a susceptibility to prostate cancer from the nucleic acid sequence data, wherein the at least one polymorphic marker is selected from the group consisting of chr8: 128173299 (rsl83373024) (SEQ ID NO: l) and rsl46851087 (SEQ ID NO:8), and correlated markers in linkage disequilibrium therewith.
  • markers in linkage disequilibrium with chr8: 128173299 are correlated therewith by values of r 2 > 0.5.
  • Exemplary correlated markers with chr8: 128173299 are in one embodiment selected from the group consisting of chr8: 127193484, chr8: 128010975, chr8: 128041317, chr8: 128260854, chr8: 128277551, chr8: 128354590 and chr8: 128449566.
  • surrogate markers in linkage disequilibrium with chr8: 128173299 are selected from the group consisting of chr8: 128260854 (rsl88140481), chr8: 128277551 (rsl38042437), chr8: 128354590 and chr8: 128010975 (rsl82352457).
  • exemplary correlated markers in linkage disequilibrium with rsl46851087 are selected from the group consisting of rs78072686, rsll6929665, and rsl38287183.
  • the identification of allele G of chr8: 128173299 is indicative of increased susceptibility of prostate cancer for the human individual. In certain embodiments, identification of allele A of rsl46851087 is indicative of increased susceptibility of prostate cancer for the human individual.
  • identification of surrogate markers alleles of allele G of marker chr8: 128173299 and allele A of marker rsl46851087 are predictive of increased risk of prostate cancer.
  • such surrogate marker alleles are selected from the predicted risk alleles set forward in Table 1, A and B.
  • surrogate markers alleles of allele G of marker chr8: 128173299 which are predictive of increased risk of prostate cancer are selected from the group consisting of allele A of marker chr8: 127193484, allele A of marker chr8: 128010975, allele T of marker chr8: 128041317, allele A of marker chr8: 128260854, allele G of marker chr8: 128277551, allele A of marker chr8: 128449566, and allele G of marker chr8: 128354590.
  • surrogate marker alleles of allele G of chr8: 128173299 which are predictive of increased risk of prostate cancer are selected from the group consisting of allele A of marker chr8: 128260854, allele G of chr8: 128277551, allele G of marker chr8: 128354590, and allele A of marker chr8: 128010975.
  • a suitable surrogate marker of chr8: 128173299 is rsl88140481.
  • determination of the presen allele A in rsl88140481 is indicative of increased risk of prostate cancer.
  • a method of determining a susceptibility to prostate cancer comprising analyzing nucleic acid sequence data about a human individual identifying at least one allele of at least one polymorphic marker, wherein different alleles of the at least one polymorphic marker are associated with different
  • the at least one polymorphic marker is selected from the group consisting of chr8: 128173299, chr8: 128260854, chr8: 128277551,
  • chr8: 128354590 and chr8: 128010975 and wherein determination of the presence of an allele selected from the group consisting of allele G of chr8: 128173299, allele A of marker
  • chr8: 128260854 allele G of chr8: 128277551, allele G of marker chr8: 128354590, and allele A of marker chr8: 128010975 is predictive of increased susceptibility to prostate cancer for the individual.
  • the polymorphic marker is chr8: 128260854. In one such embodiment, determination of the presence of allele A is predictive of increased susceptibility to prostate cancer for the individual.
  • determination of the presence of an allele selected from the group consisting of allele G of chr8: 128173299, allele A of marker chr8: 128260854, allele G of chr8: 128277551, allele G of marker chr8: 128354590, and allele A of marker chr8: 128010975 is predictive of increased susceptibility to prostate cancer with an early onset for the individual.
  • the polymorphic marker is chr8: 128260854.
  • Another aspect of the invention provides a method of determining a susceptibility to prostate cancer, the method comprising obtaining nucleic acid sequence data about a human individual identifying at least one allele of at least one polymorphic marker, wherein different alleles of that polymorphic marker are associated with different susceptibilities to prostate cancer, in humans, and determining a susceptibility to prostate cancer from the nucleic acid sequence data, wherein the at least one polymorphic marker is the chr8: 128173299, and markers in linkage
  • nucleic acid sequence data identifying particular alleles of polymorphic markers is sometimes also referred to as genotype data.
  • nucleic acid sequence data is obtained from a biological sample from the individual.
  • the nucleic acid sequence data can thus be sequence data obtained by analysis of a biological sample from an individual.
  • the biological sample in one embodiment is a nucleic acid sample, i.e. the sample contains nucleic acid from the individual.
  • Nucleic acid sequence data can be obtained for example by analyzing sequence of the at least one polymorphic marker in a nucleic acid sample from the individual.
  • nucleic acid sequence data can be obtained in a genotype dataset from the human individual and by analyzing sequence of the at least one polymorphic marker in the dataset. Such analysis in certain embodiments comprises determining the presence or absence of a particular alle specific polymorphic markers.
  • the method of the invention comprises steps of (i) obtaining a nucleic acid sample from an individual; (ii) determining the nucleic acid sequence of at least one polymorphic marker in the nucleic acid sample; and (iii) determining a susceptibility to prostate cancer from the nucleic acid sequence of the at least one polymorphic marker.
  • the correlated markers in linkage disequilibrium with chr8: 128173299 are selected from the group consisting of chr8: 127193484, chr8: 128010975, chr8 : 128041317, chr8: 128260854, chr8: 128277551, chr8 : 128449566 and chr8 : 128354590, which are the markers listed in Table 1(A).
  • Surrogate markers can be selected based on certain values of the linkage disequilibrium measures D' and r 2 , as described further herein. Markers that are in linkage disequilibrium with the marker chr8: 128173299 and rsl46851087 are exemplified by the markers listed in Table 1 A and B herein, but the skilled person will appreciate that other correlated markers in linkage disequilibrium with these markers may also be used in the diagnostic and prognostic applications described herein. Further, the skilled person will appreciate that since linkage disequilibrium is a continuous measure, certain values of the LD measure r 2 may be suitably chosen to define markers that are useful as surrogate markers in LD with the markers described herein.
  • suitable markers in linkage disequilibrium are correlated with the anchor marker by values of r 2 greater than 0.2. Ir such embodiment, suitable markers in linkage disequilibrium are correlated with the anchor marker by values of r 2 greater than 0.5. In yet another such embodiment, suitable markers in linkage disequilibrium are correlated with the anchor marker by values of r 2 greater than 0.8. In one embodiment, suitable markers in linkage disequilibrium are correlated with the anchor marker by values of r 2 of 1.0. Such markers are perfect surrogates of the anchor marker, and will give identical association results, i.e. they provide identical genetic information.
  • markers that are useful in diagnostic for determining a susceptibility to prostate cancer it may be useful to compare the frequency of markers alleles in individuals with prostate cancer to their corresponding frequency in control individuals.
  • an increase in frequency of the at least one allele in the at least one polymorphism in individuals diagnosed with prostate cancer, as compared with the frequency of the at least one allele in the control group is indicative of the at least one allele being useful for assessing increased susceptibility to prostate cancer.
  • a decrease in frequency of the at least one allele in the at least one polymorphism in individuals diagnosed with prostate cancer, as compared with the frequency of the at least one allele in the control sample is indicative of the at least one allele being useful for assessing decreased susceptibility to, or protection against, prostate cancer.
  • sequence data can be obtained by analyzing a sample from an individual, or by analyzing information about specific markers in a genotype database.
  • sequence data can be obtained through nucleic acid sequence information or amino acid sequence information from a preexisting record about a human individual.
  • a preexisting record can be any documentation, database or other form of data storage containing such information.
  • Determination of a susceptibility or risk of a particular individual in general comprises
  • determining susceptibility comprises comparing the sequence data to a database containing correlation data between the at least one polymorphic marker and susceptibility to prostate cancer.
  • the database comprises at least one measure of susceptibility to prostate cancer for the at least one polymorphic marker.
  • the database comprises a look-up table comprising at least one measure of susceptibility to prostate cancer for the at least one polymorphic marker. Determination of susceptibility is based on sequence information about particular markers identifying particular alleles at those markers. A calculation of susceptibility (risk) of prostate cancer is performed based on the information, using risk measures that have been determined for the particular alleles or combination of alleles. The measure of
  • susceptibility may in the form of relative risk ( ), absolute risk (AR), percentage (%) or other convenient measure for describing genetic susceptibility of individuals.
  • Another aspect of the invention relates to a method for determining a susceptibility to pr cancer in a human individual, comprising determining the presence or absence of at leas allele of at least one polymorphic marker in a nucleic acid sample obtained from the individual, or in a genotype dataset from the individual, wherein the at least one polymorphic marker is chr8: 128173299, and correlated markers in linkage disequilibrium therewith, and wherein determination of the presence of the at least one allele is indicative of a susceptibility to prostate cancer.
  • Determination of the presence of an allele that correlates with prostate cancer is indicative of an increased susceptibility (increased risk) to prostate cancer.
  • Individuals who are homozygous for such alleles are particularly susceptible to prostate cancer.
  • individuals who do not carry such at-risk alleles are at a decreased susceptibility of developing prostate cancer.
  • SNPs such individuals will be homozygous for the alternate (protective) allele of the polymorphism.
  • the at least one marker is selected from the group consisting of
  • the at least one marker is selected from the group consisting of chr8: 128173299, chr8: 128260854, chr8: 128277551, chr8: 128354590 and chr8: 128010975, wherein determination of the presence of an allele selected from the group consisting of allele G of chr8: 128173299, allele A of marker chr8: 128260854, allele G of chr8: 128277551, allele G of marker chr8: 128354590, and allele A of marker chr8: 128010975 is predictive of increased susceptibility to prostate cancer for the individual.
  • Determination of susceptibility is in some embodiments reported using non-carriers of the at-risk alleles of polymorphic markers as a reference. In certain embodiments, susceptibility is reported based on a comparison with the general population, e.g. compared with a random selection of individuals from the population. Such embodiments thus reflect the susceptibility (risk) of an individual compared with a randomly selected individual from the population.
  • the individual for whom susceptibility is determined is of a particular ethnicity or of a particular ethnical background.
  • the ethnicity is in certain embodiments by genetic analysis of polymorphic markers that identify the particular ethnicity.
  • the ethnicity is self-reported, i.e. the ethnicity is reported by the individual himself/herself.
  • the ethnicity is in certain embodiments selected from the group consisting of Caucasian ancestry, white ancestry, black ancestry, american indian ancestry, Alaska native ancestry, african american ancestry, african ancestry, hispanic ancestry, native Hawaian ancestry, Pacific Islander ancestry, asian ancestry and asian-american ancestry.
  • Other descriptions of ancestry that are commonly used in the art are however also contemplated, and within scope of the invention.
  • polymorphic markers are detected by sequencing technologies.
  • sequence information about an individual identifies particular nucleotides in the context of a sequence.
  • sequence information about a single unique sequence site is sufficient to identify alleles at that particular SNP.
  • sequence information about the genomic region of the individual that contain polymorphic site identifies the alleles of the individual for the particular site.
  • the sequence information can be obtained from a sample from the individual.
  • the sample is a nucleic acid sample.
  • the sample is a protein sample.
  • Sequence data can be nucleic acid sequence data, which may be obtained by means known in the art. Sequence data is suitably obtained from a biological sample of genomic DNA, RNA, or cDNA (a "test sample") from an individual ("test subject). For example, nucleic acid sequence data may be obtained through direct analysis of the sequence of the polymorphic position (allele) of a polymorphic marker.
  • Suitable methods include, for instance, whole genome sequencing methods, whole genome analysis using SNP chips (e.g., Infinium HD BeadChip), cloning for polymorphisms, non-radioactive PCR-single strand conformation polymorphism analysis, denaturing high pressure liquid chromatography (DHPLC), DNA hybridization, computational analysis, single-stranded conformational polymorphism
  • SSCP restriction fragment length polymorphism
  • RFLP restriction fragment length polymorphism
  • CDGE clamped denaturing gel electrophoresis
  • DGGE denaturing gradient gel electrophoresis
  • CMC chemical mismatch cleavage
  • RNase protection assays use of polypeptides that recognize nucleotide mismatches, such as E. coli mutS protein, allele-specific PCR, and direct manual and automated sequencing.
  • sequencing to be performed in relatively condensed format. These technologies share sequencing-by-synthesis principle for generating sequence information, with different
  • Exemplary technologies include 454 pyrosequencing technology (IMyren, P. et al. Anal Biochem 208: 171-75 (1993); http://www.454.com), Illumina Solexa sequencing technology (Bentley, D.R. Curr Opin Genet Dev 16:545-52 (2006); http://www.illumina.com), and the SOLiD technology developed by Applied Biosystems (ABI) (http://www.appliedbiosystems.com; see also Strausberg, R.L., et al. Drug Disc Today 13: 569-77 (2008)).
  • Other sequencing technologies include those developed by Pacific Biosciences (http://www.pacificbiosciences.com), Complete Genomics
  • sequence data useful for performii present invention may be obtained by any such sequencing method, or other sequencing methods that are developed or made available.
  • any sequence method that provides the allelic identity at particular polymorphic sites e.g., the absence or presence of particular alleles at particular polymorphic sites is useful in the methods described and claimed herein .
  • hybridization methods may be used (see Current Protocols in Molecular Biology, Ausubel et al., eds., John Wiley & Sons, including all supplements).
  • a biological sample of genomic DNA, RNA, or cDNA (a "test sample") may be obtained from a test subject. The subject can be an adult, child, or fetus. The DNA, RNA, or cDNA sample is then examined. The presence of a specific marker allele can be indicated by sequence-specific hybridization of a nucleic acid probe specific for the particular allele. The presence of more than one specific marker allele or a specific haplotype can be indicated by using several sequence-specific nucleic acid probes, each being specific for a particular allele. A sequence-specific probe can be directed to hybridize to genomic DNA, RNA, or cDNA.
  • a "nucleic acid probe" as used herein, can be a
  • DNA probe or an RNA probe that hybridizes to a complementary sequence One of skill in the art would know how to design such a probe so that sequence specific hybridization will occur only if a particular allele is present in a genomic sequence from a test sample.
  • determination of a susceptibility to prostate cancer comprises forming a hybridization sample by contacting a test sample, such as a genomic DNA sample, with at least one nucleic acid probe.
  • a probe for detecting mRNA or genomic DNA is a labeled nucleic acid probe that is capable of hybridizing to mRNA or genomic DNA sequences described herein.
  • the nucleic acid probe can be, for example, a full-length nucleic acid molecule, or a portion thereof, such as an oligonucleotide of at least 10, 15, 30, 50, 100, 250 or 500 nucleotides in length that is sufficient to specifically hybridize under stringent conditions to appropriate mRNA or genomic DNA.
  • the nucleic acid probe can comprise all or a portion of the nucleotide sequence of the Asporin gene, or the probe can be the complementary sequence of such a sequence.
  • Hybridization can be performed by methods well known to the person skilled in the art (see, e.g., Current Protocols in Molecular Biology, Ausubel et al . , eds. , John Wiley & Sons, including all supplements).
  • hybridization refers to specific hybridization, i.e., hybridization with no mismatches (exact hybridization).
  • the hybridization conditions for specific hybridization are high stringency.
  • Specific hybridization if present, is detected using standard methods. If specific hybridization occurs between the nucleic acid probe and the nucleic acid in the test sample, then the sample contains the allele that is complementary to the nucleotide that is present in the nucleic acid probe.
  • a peptide nucleic acid (PNA) probe can be used in addition to, or instead of, a nucleic acid probe in the hybridization methods described herein.
  • a PNA is a DNA mimic having a peptide-like, inorganic backbone, such as N-(2-aminoethyl)glycine units, with an organic base (A, G, C, T or U) attached to the glycine nitrogen via a methylene carbonyl linker (see, for example, Nielsen et al., Bioconjug. Chem. 5:3-7 (1994)).
  • the PNA probe can t designed to specifically hybridize to a molecule in a sample suspected of containing one of the at-risk alleles (mutations) shown herein to be associated with risk of prostate cancer..
  • Allele-specific oligonucleotides can also be used to detect the presence of a particular allele in a nucleic acid.
  • An "allele-specific oligonucleotide” (also referred to herein as an “allele-specific oligonucleotide probe”) is an oligonucleotide of any suitable size, for example an oligonucleotide of approximately 10-50 base pairs or approximately 15-30 base pairs, that specifically hybridizes to a nucleic acid which contains a specific allele at a polymorphic site (e.g., a polymorphic marker).
  • An allele-specific oligonucleotide probe that is specific for one or more particular alleles at polymorphic markers can be prepared using standard methods (see, e.g., Current Protocols in Molecular Biology, supra). PCR can be used to amplify the desired region. Specific hybridization of an allele-specific oligonucleotide probe to DNA from a subject is indicative of the presence of a specific allele at a polymorphic site (see, e.g., Gibbs et al., Nucleic Acids Res. 17:2437-2448 (1989) and WO 93/22456).
  • LNAs locked nucleic acids
  • oxy-LNA O-methylene
  • thio-LNA S-methylene
  • amino-LNA amino methylene
  • Tm melting temperatures
  • 74°C melting temperatures
  • Tm melting temperatures
  • primers and probes depending on where the LNA monomers are included (e.g., the 3' end, the 5' end, or in the middle), the Tm could be increased considerably. It is therefore contemplated that in certain embodiments, LNAs are used to detect particular at-risk alleles, as described herein.
  • arrays of oligonucleotide probes that are complementary to target nucleic acid sequence segments from a subject can be used to identify particular alleles in a nucleic acid.
  • an oligonucleotide array can be used.
  • Oligonucleotide arrays typically comprise a plurality of different oligonucleotide probes that are coupled to a surface of a substrate in different known locations. These arrays can generally be produced using mechanical synthesis methods or light directed synthesis methods that incorporate a combination of photolithographic methods and solid phase oligonucleotide synthesis methods, or by other methods known to the person skilled in the art (see, e.g., Bier et al., Adv Biochem Eng
  • markers alleles can be detected by fluorescence-based techniques (e.g., Chen et al., Genome Res. 9(5): 492-98 (1999); Kutyavin et al., Nucleic Acid Res. 34:el28 (2006)), utilizing PCR, LCR, Nested PCR and other techniques for nucleic acid amplification.
  • fluorescence-based techniques e.g., Chen et al., Genome Res. 9(5): 492-98 (1999); Kutyavin et al., Nucleic Acid Res. 34:el28 (2006)
  • PCR e.g., LCR, Nested PCR and other techniques for nucleic acid amplification.
  • SNP genotyping include, but are not limited to, TaqMan genotyping assays and SNPlex platforms (Applied Biosystems), gel electrophoresis (Applied Biosystems), mass spectrometry (e.g., MassARRAY system from Sequenom), minisequencing methods, real-time PCR, Bio-Plex system (BioRad), CEQ and SNPstream systems (Beckman), array hybridization technology(e.g., Affymetrix GeneChip; Perlegen ), BeadArray Technologies (e.g., Illumina GoldenGate and Infinium assays), array tag technology (e.g., Parallele), and endonuclease-based fluorescence hybridization technology (Invader; Third Wave).
  • Applied Biosystems Applied Biosystems
  • Gel electrophoresis Applied Biosystems
  • mass spectrometry e.g., MassARRAY system from Sequenom
  • minisequencing methods minisequencing methods, real-time PCR, Bio-
  • Suitable biological sample in the methods described herein can be any sample containing nucleic acid (e.g., genomic DNA) and/or protein from the human individual.
  • the biological sample can be a blood sample, a serum sample, a leukapheresis sample, an amniotic fluid sample, a cerbrospinal fluid sample, a hair sample, a tissue sample from skin, muscle, buccal, or conjuctival mucosa, placenta, gastrointestinal tract, or other organs, a semen sample, a urine sample, a saliva sample, a nail sample, a tooth sample, and the like.
  • the sample is a blood sample, a saliva sample or a buccal swab.
  • genotypes of un-genotyped relatives For every un-genotyped case, it is possible to calculate the probability of the genotypes of its relatives given its four possible phased genotypes. In practice it may be preferable to include only the genotypes of the case's parents, children, siblings, half-siblings (and the half-sibling's parents), grand-parents, grand-children (and the grand-children's parents) and spouses. It will be assumed that the individuals in the small sub-pedigrees created around each case are not related through any path not included in the pedigree. It is also assumed that alleles that are not transmitted to the case have the same frequency - the population allele frequency. The probability of the genotypes of the case's relatives can then be computed by:
  • Pr(genotypes of relatives; ⁇ ) ⁇ Pr( z; ⁇ ) Pr(genotypes of relatives I h) ,
  • ⁇ denotes the A allele's frequency in the cases. Assuming the genotypes of each set of relatives are independent, this allows us to write down a likelihood function for ⁇ :
  • nucleic acid sequence data may be obtained through indirect analysis of the nucleic acid sequence of the allele of the polymorphic marker, i.e. by detecting a protein variation.
  • Methods of detecting variant proteins are known in the art. For example, direct amino acid sequencing of the variant protein followed by comparison to a reference amino acid sequence can be used. Alternatively, SDS-PAGE followed by gel staining can be used to detect variant proteins of different molecular weights. Also, Immunoassays, e.g., antibody assays, e.g., immunofluorescent immunoassays, immunoprecipitations, radioimmunoasays, ELISA, and Western blotting, in which an antibody specific for an epitope comprising the variant sequence among the variant protein and non-variant or wild-type protein can be used. In certain embodiments, the amino acid sequence data about Asporin protein is obtained or dedua a preexisting record.
  • the methods can comprise obtaining sequence data about any number of polymorphic markers and/or about any number of genes.
  • the method can comprise obtaining sequence data for about at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, 100, 500, 1000, 10,000 or more polymorphic markers.
  • the sequence data is obtained from a microarray comprising probes for detecting a plurality of markers.
  • the sequence data is obtained through nucleic acid sequencing, for example by high-throughput nucleic acid sequencing.
  • the sequence data may also be obtained by imputation of nucleic acid sequence using known methods, such as those described herein.
  • the polymorphic markers can be the ones of the group specified herein or they can be different polymorphic markers that are not specified herein.
  • the method comprises obtaining sequence data about at least two polymorphic markers.
  • each of the markers may be associated with a different gene.
  • the method comprises obtaining nucleic acid data about a human individual identifying at least one allele of a polymorphic marker, then the method comprises identifying at least one allele of at least one polymorphic marker.
  • the method can comprise obtaining sequence data about a human individual identifying alleles of multiple, independent markers, which are not in linkage disequilibrium.
  • Linkage Disequilibrium refers to a non-random assortment of two genetic elements. For example, if a particular genetic element (e.g., an allele of a polymorphic marker, or a haplotype) occurs in a population at a frequency of 0.50 (50%) and another element occurs at a frequency of 0.50 (50%), then the predicted occurrance of a person's having both elements is 0.25 (25%), assuming a random distribution of the elements.
  • a particular genetic element e.g., an allele of a polymorphic marker, or a haplotype
  • Allele or haplotype frequencies can be determined in a population by genotyping individuals in a population and determining the frequency of the occurence of each allele or haplotype in the population. For populations of diploids, e.g., human populations, individuals will typically have two alleles or allelic combinations for each genetic element ⁇ e.g., a marker, haplotype or gene).
  • is defined in such a way that it is equal to 1 if just two or three of the possible haplotypes are present, and it is ⁇ 1 if all four possible haplotypes are present. Therefore, a value of
  • the measure r 2 represents the statistical correlation between two sites, and takes the value of 1 if only two haplotypes are present.
  • the r 2 measure is arguably the most relevant measure for association mapping, because there is a simple inverse relationship between r 2 and the sample size required to detect association between susceptibility loci and SNPs. These measures are defined for pairs of sites, but for some applications a determination of how strong LD is across an entire region that contains many polymorphic sites might be desirable (e.g., testing whether the strength of LD differs significantly among loci or across populations, or whether there is more or less LD in a region than predicted under a particular model). Measuring LD across a region is not straightforward, but one approach is to use the measure r, which was developed in population genetics.
  • a significant r 2 value can be at least 0.1 such as at least 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.91, 0.92, 0.93, 0.94, 0.95, 0.96, 0.97, 0.98, or at lesat 0.99.
  • the significant r 2 value can be at least 0.2.
  • linkage disequilibrium refers to linkage disequilibrium characterized by values of
  • linkage disequilibrium represents a correlation between alleles of distinct markers. It is measured by correlation coefficient or
  • linkage disequilibrium is defined in terms of values for both the r 2 and
  • a significant linkage diseqc is defined as r 2 > 0.1 and
  • a significant linkage diseqc is defined as r 2 > 0.1 and
  • Linkage disequilibrium is defined as r 2 > 0.2 and
  • Linkage disequilibrium can be determined in a single human population, as defined herein, or it can be determined in a collection of samples comprising individuals from more than one human population.
  • LD is determined in a sample from one or more of the HapMap populations (Caucasian, african, Japanese, Chinese), as defined (http://www.hapmap.org). In one such embodiment, LD is " determined in the CEU population of the HapMap samples. In another embodiment, LD is determined in the YRI population. In yet another embodiment, LD is determined in samples from the Icelandic population.
  • Haplotype blocks can be used to map associations between phenotype and haplotype status, using single markers or haplotypes comprising a plurality of markers.
  • the main haplotypes can be identified in each haplotype block, and then a set of "tagging" SNPs or markers (the smallest set of SNPs or markers needed to distinguish among the haplotypes) can then be identified. These tagging SNPs or markers can then be used in assessment of samples from groups of individuals, in order to identify association between phenotype and haplotype. If , desired, neighboring haplotype blocks can be assessed concurrently, as there may also exist linkage disequilibrium among the haplotype blocks.
  • markers used to detect association thus in a sense represent "tags" for a genomic region (i.e., a haplotype block or LD block) that is associating with a given disease or trait, and as such are useful for use in the methods and kits of the present invention.
  • One or more causative (functional) variants or mutations may reside the region found to be associating to the disease or trait.
  • the functional variant may be
  • SNP SNP
  • a tandem repeat polymorphism such as a minisatellite or a microsatellite
  • a transposable element such as a transposable element
  • a copy number variation such as an inversion, deletion or insertion.
  • RR relative risk
  • OR odds ratio
  • the present invention thus refers to the markers used for detecting association to the disease, as described herein, as well as markers in linkage disequilibrium with the markers.
  • markers that are in LD with the markers and/or haplotypes of the invention, as described herein may be used as surrogate markers.
  • the surrogate markers have in one embodiment relative risk (RR) and/or odds ratio (OR) values smaller than for the markers or haplotypes initially found to be associating with the disease, as described herein.
  • the surrogate markers have RR or OR values greater than those initially determined for the markers initially found to be associating with the disease, as described herein.
  • An example of such an embodiment would be a rare, or relatively rare (such as ⁇ 10% allelic population frequency) variant in LD with a more common variant (> 10% population frequency) initially found to be associating with the disease, such as the variants described herein. Identifying and using such markers for detecting the association discovered by the inventors as described herein can be performed by routine methods well known to the person skilled in the art, and are therefore within the scope of the present invention.
  • Suitable surrogate markers may be selected using public information, such as from the
  • the markers may also be suitably selected from results of whole-genome sequencing.
  • Markers with values of r 2 equal to 1 are perfect surrogates for the at-risk variants, i.e. genotypes for one marker perfectly predicts genotypes for the other. In other words, the surrogate will, by necessity, give exactly the same association data to any particular disease as the anchor marker. Markers with smaller values of r 2 than 1 can also be surrogates for the at- risk anchor variant.
  • the Fisher exact test can be used to calculate two- sided p-values for each individual allele. Correcting for relatedness among patients can be done by extending a variance adjustment procedure previously described (Risch, N. & Teng, J.
  • the method of genomic controls (Devlin, B. & Roeder, K. Biometrics 55:997 (1999)) can also be used to adjust for the relatedness of the individuals and possible
  • relative risk RR
  • PAR population att risk
  • haplotypes are independent, i.e., in Hardy-Weinberg equilibrium, within the affected population as well as within the control population.
  • an individual who is at an increased susceptibility (i.e., increased risk) for prostate cancer is an individual in whom at least one specific allele at one or more polymorphic marker or haplotype conferring increased susceptibility (increased risk) for prostate cancer is identified (i.e., at-risk marker alleles or haplotypes).
  • the at-risk marker or haplotype is one that confers an increased risk (increased susceptibility) of prostate cancer.
  • significance associated with a marker or haplotype is measured by a relative risk (RR).
  • significance associated with a marker or haplotype is measured by an odds ratio (OR).
  • the significance is measured by a percentage.
  • a significant increased risk is measured as a risk (relative risk and/or odds ratio) of at least 1.05, including but not limited to: at least 2.0, at least 2.1, at least 2.2, at least 2.3, at least 2.4, at least 2.5, at least 2.6, at least 2.7 and at least 2.8.
  • a risk (relative risk and/or odds ratio) of at least 2.5 is significant.
  • a risk of at least 2.6 is significant.
  • a risk of at least 2.7 is significant.
  • Other cutoffs are also contemplated, e.g., at least 2.15, 2.25, 2.35, and so on, and such cutoffs are also within scope of the present invention.
  • a significant increase in risk is at least about 110%, including but not limited to about 120%, 130%, 140%, 150%, 160%, 170%, and at least 180%. In one particular embodiment, a significant increase in risk is at least 150%. In another particular embodiment, a significant increase in risk is at least 160%. In another particular embodiment, a significant increase in risk is at least 170%. Other cutoffs or ranges as deemed suitable by the person skilled in the art to characterize the invention are however also contemplated, and those are also within scope of the present invention.
  • a significant increase in risk is characterized by a p- value, such as a p-value of less than 0.05, less than 0.01, less than 0.001, less than 0.0001, less than 0.00001, less than 0.000001, less than 0.0000001, less than 0.00000001, or less than 0.000000001.
  • An at-risk polymorphic marker or haplotype as described herein is one where at least on of at least one marker or haplotype is more frequently present in an individual at risk foi prostate cancer (affected), or diagnosed with prostate cancer, compared to the frequency of its presence in a comparison group (control), such that the presence of the marker or haplotype is indicative of susceptibility to prostate cancer.
  • the control group may in one embodiment be a population sample, i.e.
  • control group is represented by a group of individuals who are disease-free.
  • disease-free controls may in one embodiment be characterized by the absence of one or more specific disease-associated symptoms.
  • the disease-free controls are those that have not been diagnosed with prostate cancer.
  • the disease-free control group is characterized by the absence of one or more disease-specific risk factors.
  • risk factors are in one embodiment at least one environmental risk factor. Representative environmental factors are risk factors related to lifestyle, including but not limited to food and drink habits,
  • the risk factors comprise at least one additional genetic risk factor for prostate cancer.
  • the two by two Table is constructed out of the number of chromosomes that include both of the markers or haplotypes, one of the markers or
  • haplotypes but not the other and neither of the markers or haplotypes.
  • Other statistical tests of association known to the skilled person are also contemplated and are also within scope of the invention.
  • an individual who is at a decreased susceptibility (i.e., at a decreased risk) for a disease is an individual in whom at least one specific allele at one or more polymorphic marker or haplotype conferring decreased susceptibility for the disease or trait is identified.
  • the marker alleles and/or haplotypes conferring decreased risk are also said to be protective.
  • the protective marker or haplotype is one that confers a significant decreased risk (or susceptibility) of the disease or trait.
  • significant decreased risk is measured as a relative risk (or odds ratio) of less than 0.95, including but not limited to less than 0.9, less than 0.8, less than 0.7, less than 0.6, less than 0.5, less than 0.4, less than 0.3, less than 0.2 and less than 0.1. In one particular embodiment, significant decreased risk is less than 0.90. In another embodiment, significant decreased risk is less than 0.6. In yet another embodiment, significant decreased risk is less than 0.5. In another embodiment, the decrease in risk (or susceptibility) is at least 50%, including but not limited to at least 50%, at least 55%, at least 60%, at least 65%, and at least 70% . Other cutoffs or ranges as deemed suitable by the person skilled in the art to characterize the invention are however also contemplated, and those are also within scope of the present invention.
  • the person skilled in the art will appreciate that for markers with two alleles present in the population being studied (such as SNPs), and wherein one allele is found in increased frequency in a group of individuals with prostate cancer, compared with controls, the other allele of the marker will be found in decreased frequency in the group of individuals with prostate cancer, compared with controls. In such a case, one allele of the marker (the one found in increased frequency individuals with prostate cancer) will be the at-risk allele, while the other allele will be a
  • a genetic variant associated with a disease or a trait can be used alone to predict the risk of the disease for a given genotype.
  • a biallelic marker such as a SIMP
  • Risk associated with variants at multiple loci can be used to estimate overall risk.
  • For multiple SNP variants, there are k possible genotypes k 3 n x 2 P ; where n is the number autosomal loci and p the number of gonosomal (sex chromosomal) loci.
  • Overall risk assessment calculations for a plurality of risk variants usually assume that the relative risks of different genetic variants multiply, i.e.
  • the overall risk (e.g., RR or OR) associated with a particular genotype combination is the product of the risk values for the genotype at each locus. If the risk presented is the relative risk for a person, or a specific genotype for a person, compared to a reference population with matched gender and ethnicity, then the combined risk - is the product of the locus specific risk values - and which also corresponds to an overall risk estimate compared with the population. If the risk for a person is based on a comparison to non-carriers of the at risk allele, then the combined risk corresponds to an estimate that compares the person with a given combination of genotypes at all loci to a group of individuals who do not carry risk variants at any of those loci.
  • the risk presented is the relative risk for a person, or a specific genotype for a person, compared to a reference population with matched gender and ethnicity
  • the combined risk - is the product of the locus specific risk values - and which also corresponds to an overall risk estimate
  • the group of non-carriers of any at risk variant has the lowest estimated risk and has a combined risk compared with itself ( .e., non-carriers) of 1.0, but has an overall risk, compare with the population, of less than 1.0. It should be noted that the group of non-carriers can potentially be very small, especially for large number of loci, and in that case, its relevance is correspondingly small.
  • multiplicative model is a parsimonious model that usually fits the data of complex traits reasonably well. Deviations from multiplicity have been rarely described in the context of common variants for common diseases, and if reported are usually only suggestive since very large sample sizes are usually required to be able to demonstrate statistical interactions between loci. Risk assessment and Diagnostics
  • an absolute risk of developing a disease or trait defined as the chance of a person developing the specific disease or trait over a specified time-period.
  • a woman's lifetime absolute risk of breast cancer is one in nine. That is to say, one woman in every nine will develop breast cancer at some point in their lives.
  • Risk is typically measured by looking at very large numbers of people, rather than at a particular individual. Risk is often presented in terms of Absolute Risk (AR) and Relative Risk (RR).
  • AR Absolute Risk
  • RR Relative Risk
  • Relative Risk is used to compare risks associating with two variants or the risks of two different groups of people. For example, it can be used to compare a group of people with a certain genotype with another group having a different genotype.
  • a relative risk of 2 means that one group has twice the chance of developing a disease as the other group.
  • the creation of a model to calculate the overall genetic risk involves two steps: i) conversion of odds-ratios for a single genetic variant into relative risk and ii) combination of risk from multiple variants in different genetic loci into a single relative risk value.
  • allelic odds-ratio equals the risk factor:
  • RR(aa) Pr(A
  • aa)/Pr(A) (Pr(A
  • RR(g l,g2) RR(g l)RR(g2)
  • g l,g2) Pr(A
  • g2)/Pr(A) and Pr(g l,g2) Pr(g l)Pr(g2)
  • the model applied is not expected to be exactly true since it is not based on an underlying bio-physical model.
  • the multiplicative model has so far been found to fit the data adequately, i.e. no significant deviations are detected for many common diseases for which many risk variants have been discovered .
  • Other genetic markers in different genomic locations have been found to be associated v prostate cancer, in addition to the marker shown herein to be associated with risk of pro cancer. It can be useful to estimate genetic risk of prostate cancer for combinations of such markers, optionally including any one, or a combination of, the markers described herein.
  • Determining risk for multiple markers captures a greater percentage of the genetic risk of prostate cancer in the population. For example, by combining risk for 22 prostate cancer risk variants typed in the Icelandic population, carriers belonging to the top 1.3% of the risk distribution have a risk of developing the disease that is more than 2.5 times greater than the population average risk estimates. For these individuals this corresponds to a lifetime risk of over 25% of being diagnosed with prostate cancer, compared with a population average life time risk of about 10% in Iceland.
  • the lifetime risk of an individual is derived by multiplying the overall genetic risk relative to the population with the average life-time risk of the disease in the general population of the same ethnicity and gender and in the region of the individual's geographical origin. As there are usually several epidemiologic studies to choose from when defining the general population risk, we will pick studies that are well-powered for the disease definition that has been used for the genetic variants.
  • the methods and kits of the invention can be utilized from samples containing nucleic acid material (DNA or RNA) from any source and from any individual, or from genotype data derived from such samples.
  • the individual is a human individual.
  • the individual can be an adult, child, or fetus.
  • the nucleic acid source may be any sample comprising nucleic acid material, including biological samples, or a sample comprising nucleic acid material derived therefrom.
  • the present invention also provides for assessing markers and/or haplotypes in individuals who are members of a target population.
  • Such a target population is in one embodiment a population or group of individuals at risk of developing the disease, based on other genetic factors, biomarkers (e.g., PSA), biophysical parameters, or general health and/or lifestyle parameters [e.g., history of prostate cancer or related cancer, previous diagnosis of prostate cancer, family history of prostate cancer).
  • biomarkers e.g., PSA
  • biophysical parameters e.g., biophysical parameters
  • general health and/or lifestyle parameters e.g., history of prostate cancer or related cancer, previous diagnosis of prostate cancer, family history of prostate cancer.
  • the invention provides for embodiments that include individuals from specific age subgr ⁇ such as those over the age of 40, over age of 45, or over age of 50, 55, 60, 65, 70, 75,
  • Other embodiments of the invention pertain to other age groups, such as individuals aged less than 85, such as less than age 80, less than age 75, or less than age 70, 65, 60, 55, 50, 45, 40, 35, or age 30.
  • Other embodiments relate to individuals with age at onset of prostate cancer in any of the age ranges described in the above. It is also contemplated that a range of ages may be relevant in certain embodiments, such as age at onset at more than age 45 but less than age 60. Other age ranges are however also contemplated, including all age ranges bracketed by the age values listed in the above.
  • the invention furthermore relates to individuals of either gender, males or females.
  • the Icelandic population is a Caucasian population of Northern European ancestry.
  • a large number of studies reporting results of genetic linkage and association in the Icelandic population have been published in the last few years. Many of those studies show replication of variants, originally identified in the Icelandic population as being associating with a particular disease, in other populations (Styrkarsdottir, U., et al. N Engl J Med Apr 29 2008 (Epub ahead of print);
  • Such embodiments relate to human subjects that are from one or more human population including, but not limited to, Caucasian populations, European populations, White populations, Black population, American populations, Eurasian populations, Asian populations, Central/South Asian populations, East Asian populations, Middle Eastern populations, African populations, Hispanic populations, and Oceanian populations.
  • European populations include, but are not limited to, Swedish, Norwegian, Finnish, Russian, Danish, Icelandic, Irish, Kelt, English, Scottish, Dutch, Belgian, French, German, Spanish, Portuguese, Italian, Polish, Bulgarian, Slavic, Serbian, Laun, Czech, Greek and Vietnamese populations.
  • the invention relates to markers and/or haplotypes identified in specific populations, as described in the above.
  • measures of linkage disequilibrium (LD) may give different results when applied to different populations. This is due to different population history of different human populations as well as differential selective pressures that may have led to differences in LD in specific genomic regions.
  • certain markers e.g. SNP markers, have different population frequency in different populations, or are polymorphic in one population but not in another. The person skilled in the art will however apply the methods available ar thought herein to practice the present invention in any given human population.
  • This m include assessment of polymorphic markers in the LD region of the present invention, so as to identify those markers that give strongest association within the specific population.
  • the at-risk variants of the present invention may reside on different haplotype background and in different frequencies in various human populations.
  • the invention can be practiced in any given human population. Diagnostic Methods
  • Polymorphic markers associated with increased susceptibility of prostate cancer are useful in diagnostic methods. While methods of diagnosing prostate cancer are known in the art, the detection risk markers for the condition advantageously may be useful for detection of prostate cancer at its early stages and may also reduce the occurrence of misdiagnosis. In this regard, the invention further provides methods of diagnosing prostate cancer comprising obtaining sequence data identifying at least one risk allele as described herein, in conjunction with carrying out one or more clinical diagnostic steps for the identification of the condition. One such clinical diagnostic step is PSA testing. Other clinical diagnostic steps are described in more detail in the following.
  • the application of a genetic test for prostate cancer can provide an opportunity for the detection of the disease at an earlier stage which leads to higher cure rates, if found locally, and increases survival rates by minimizing regional and distant spread of the tumor.
  • a genetic test will most likely increase the sensitivity and specificity of the already generally applied Prostate Specific Antigen (PSA) test and Digital Rectal Examination (DRE). This can lead to lower rates of false positives (thus minimize unnecessary procedures such as needle biopsies) and false negatives (thus increasing detection of occult disease and minimizing morbidity and mortality due to PCA).
  • PSA Prostate Specific Antigen
  • DRE Digital Rectal Examination
  • Genetic testing can provide information about pre-diagnostic prognostic indicators and enable the identification of individuals at high or low risk for aggressive tumor types that can lead to modification in screening strategies. For example, an individual determined to be a carrier of a high risk allele for the development of aggressive prostate cancer will likely undergo more frequent PSA testing, examination and have a lower threshold for needle biopsy in the presence of an abnormal PSA value.
  • identifying individuals that are carriers of high or low risk alleles for aggressive tumor types will lead to modification in treatment strategies. For example, if prostate cancer is diagnosed in an individual that is a carrier of an allele that confers increased risk of developing an aggressive form of prostate cancer, then the clinician would likely advise a more aggr treatment strategy such as a prostatectomy instead of a less aggressive treatment strati
  • Prostate Specific Antigen is a protein that is secreted by the epithelial cells of the prostate gland, including cancer cells. An elevated level in the blood indicates an abnormal condition of the prostate, either benign or malignant. PSA is used to detect potential problems in the prostate gland and to follow the progress of prostate cancer therapy. PSA levels above 4 ng/ml are indicative of the presence of prostate cancer (although as known in the art and described herein, the test is neither very specific nor sensitive).
  • the method of the invention is performed in combination with (either prior to, concurrently or after) a PSA assay.
  • the presence of an at-risk marker or haplotype, in conjunction with the subject having a PSA level greater than 4 ng/ml is indicative of a more aggressive prostate cancer and/or a worse prognosis.
  • particular markers and haplotypes are associated with high Gleason (i.e., more aggressive) prostate cancer.
  • the presence of a marker or haplotype, in a patient who has a normal PSA level is indicative of a high Gleason (i.e.
  • a "worse prognosis” or “bad prognosis” occurs when it is more likely that the cancer will grow beyond the boundaries of the prostate gland, metastasize, escape therapy and/or kill the host.
  • the presence of a marker or haplotype is indicative of a predisposition to a somatic rearrangement (e.g., one or more of an amplification, a translocation, an insertion and/or deletion) in a tumor or its precursor.
  • a somatic rearrangement e.g., one or more of an amplification, a translocation, an insertion and/or deletion
  • the somatic rearrangement itself may subsequently lead to a more aggressive form of prostate cancer (e.g., a higher histologic grade, as reflected by a higher Gleason score or higher stage at diagnosis, an increased progression of prostate cancer (e.g., to a higher stage), a worse outcome (e.g., in terms of morbidity, complications or death)).
  • the Gleason grade is a widely used method for classifying prostate cancer tissue for the degree of loss of the normal glandular architecture (size, shape and differentiation of glands).
  • a grade from 1-5 is assigned successively to each of the two most predominant tissue patterns present in the examined tissue sample and are added together to produce the total or combined Gleason grade (scale of 2-10). High numbers indicate poor differentiation and therefore more aggressive cancer.
  • Aggressive prostate cancer is cancer that grows beyond the prostate, metastasizes and eventually kills the patient.
  • one surrogate measure of aggressiveness is a high combined Gleason grade. The higher the grade on a scale of 2-10 the more likely it is that a patient has aggressive disease.
  • a sample containing genomic DNA or protein from an individual is collected.
  • sample can for example be a buccal swab, a saliva sample, a blood sample, or other suitable samples containing genomic DNA or protein, as described further herein.
  • the sample is obtained by non-invasive means (e.g., for obtaining a buccal sample, saliva sample, hair sample or skin sample).
  • non-surgical means i.e. in the absence of a surgical intervention on the individual that puts the individual at substantial health risk.
  • Such embodiments may, in addition ti invasive means also include obtaining sample by extracting a blood sample (e. g., a venc sample).
  • genomic DNA or protein obtained from the individual is then analyzed using any common technique available to the skilled person, such as high-throughput technologies for genotyping and/or sequencing.
  • Results from such methods are stored in a convenient data storage unit, such as a data carrier, including computer databases, data storage disks, or by other convenient data storage means.
  • the computer database is an object database, a relational database or a post-relational database.
  • the genotype data is subsequently analyzed for the presence of certain variants known to be susceptibility variants for a particular human condition, such as the genetic variants described herein associated with risk of prostate cancer.
  • Genotype and/or sequencing data can be retrieved from the data storage unit using any convenient data query method.
  • Calculating risk conferred by a particular genotype for the individual can be based on comparing the genotype of the individual to previously determined risk (expressed as a relative risk (RR) or and odds ratio (OR), for example) for the genotype, for example for a heterozygous carrier of an at-risk variant.
  • the calculated risk for the individual can be the relative risk for a person, or for a specific genotype of a person, compared to the average population with matched gender and ethnicity.
  • the average population risk can be expressed as a weighted average of the risks of different genotypes, using results from a reference population, and the appropriate calculations to calculate the risk of a genotype group relative to the population can then be performed.
  • the risk for an individual is based on a comparison of particular genotypes, for example heterozygous carriers of an at-risk allele of a marker compared with non-carriers of the at-risk allele.
  • the calculated risk estimated can be made available to the customer via a website, preferably a secure website. Kits
  • Kits useful in the methods of the invention comprise components useful in any of the methods described herein, including for example, primers for nucleic acid amplification, hybridization probes, restriction enzymes (e.g. , for RFLP analysis), allele-specific oligonucleotides, antibodies that bind to an altered polypeptide encoded by a nucleic acid of the invention as described herein (e.g.
  • kits can for example include necessary buffers, nucleic acid primers for amplifying nucleic acids of the invention (e.g. , a nucleic acid segment
  • kits can provide reagents for assays to be used in combination with the methods of the present invention, e.g. , reagents for use with other diagnostic assays for prostate cancer.
  • the invention pertains to a kit for assaying a sample from a subject to detect a susceptibility to prostate cancer in a subject, wherein the kit comprises reagents necessary for selectively detecting at least one allele of at least one polymorphism of the present invention in the genome of the individual.
  • the reagents comprise at least one contiguous oligonucleotide that hybridizes to a fragment of the genome of the individual comprising at least one polymorphism of the present invention.
  • the reagents comprise at least one pair of oligonucleotides that hybridize to opposite strands of a genomic segment obtained from a subject, wherein each oligonucleotide primer pair is designed to selectively amplify a fragment of the genome of the individual that includes at least one polymorphism associated with prostate cancer risk.
  • the polymorphism is selected from the group consisting of the markers described herein to be associated with risk of prostate cancer, and polymorphic markers in linkage disequilibrium therewith.
  • the fragment is at least 20 base pairs in size.
  • oligonucleotides or nucleic acids e.g., oligonucleotide primers
  • the kit comprises one or more labeled nucleic acids capable of allele-specific detection of one or more specific polymorphic markers or haplotypes, and reagents for detection of the label.
  • Suitable labels include, e.g. , a radioisotope, a fluorescent label, an enzyme label, an enzyme co-factor label, a magnetic label, a spin label, an epitope label.
  • the polymorphic marker to be detected by the reagents of the kit comprises at least the marker chr8: 128173299 and/or marker rsl46851087, and correlated markers in linkage disequilibrium therewith.
  • the marker to be detected comprises at least one marker from the group of markers in strong linkage disequilibrium, as defined by values of r 2 greater than 0.5, to marker chr8: 128173299.
  • the marker is selected from the group consisting of chr8: 128173299, chr8: 128260854,
  • the marker to be detected comprises at least one marker from the group of markers in strong linkage disequilibrium, as defined by values of r 2 greater than 0.5, to marker rsl46851087.
  • the DNA template containing the SNP polymorphism is amplified by Polymerase Chain Reaction (PCR) prior to detection, and primers for such amplification are included in the reagent kit.
  • PCR Polymerase Chain Reaction
  • the amplified DNA serves as the template for the detection probe and the enhancer probe.
  • the DNA template is amplified by means of Whole Genome Amplification (WGA) methods, prior to assessment for the presence of specific polymorphic markers as described herein. Standard methods well known to the skilled person for performing WGA may be utilized, and are within scope of the invention.
  • reagents for performing WGA are included in the reagent kit.
  • determination of the presence of a particular marker allele is i of a susceptibility (increased susceptibility or decreased susceptibility) to prostate cance another embodiment, determination of the presence of a marker allele is indicative of response to a therapeutic agent for prostate cancer.
  • the presence of the marker allele is indicative of prostate cancer prognosis.
  • the presence of the marker allele is indicative of progress of prostate cancer treatment. Such treatment may include intervention by surgery, medication or by other means ⁇ e.g. , lifestyle changes) .
  • a pharmaceutical pack comprising a therapeutic agent and a set of instructions for administration of the therapeutic agent to humans diagnostically tested for one or more variants of the present invention, as disclosed herein.
  • the therapeutic agent can be a small molecule drug, an antibody, a peptide, an antisense or RNAi molecule, or other therapeutic molecules.
  • an individual identified as a carrier of at least one variant of the present invention is instructed to take a prescribed dose of the therapeutic agent.
  • an individual identified as a homozygous carrier of at least one variant of the present invention is instructed to take a prescribed dose of the therapeutic agent.
  • an individual identified as a non-carrier of at least one variant of the present invention is instructed to take a prescribed dose of the therapeutic agent.
  • the kit further comprises a set of instructions for using the reagents comprising the kit.
  • the kit further comprises a collection of data comprising correlation data between the polymorphic markers assessed by the kit and susceptibility to prostate cancer.
  • antisense agents are comprised of single stranded oligonucleotides (RNA or DNA) that are capable of binding to a complimentary nucleotide segment.
  • RNA or DNA single stranded oligonucleotides
  • the antisense oligonucleotides are complementary to the sense or coding strand of a gene. It is also possible to form a triple helix, where the antisense oligonucleotide binds to duplex DNA.
  • antisense oligonucleotide binds to target RNA sites, activate intracellular nucleases (e.g., RnaseH or Rnase L), that cleave the target RNA.
  • Blockers bind to target RNA, inhibit protein translation by steric hindrance of the ribosomes. Examples of blockers include nucleic acids, morpholino compounds, locked nucleic acids and methylphosphonates (Thompson, Drug Discovery Today, 7:912-917 (2002)).
  • Antisense oligonucleotides are useful directly as therapeutic agents, and are also useful for determining and validating gene function, for by gene knock-out or gene knock-down experiments. Antisense technology is further described in Lavery et al., Curr. Opin. Drug Discov. Devel. 6: 561-569 (2003), Stephens et a/., Curr. Opin. Mol. Ther. 5: 118-122 (2003), Kurreck, Eur. J. Biochem. 270: 1628-44 (2003), Dias et al., Mot. Cancer Ter. 1:347-55 (2002), Chen, Methods Mol. Med. 75:621-636 (2003), Wang et al., Curr. Cancer Drug Targets 1 : 177-96 (2001), and Bennett, Antisense Nucleic Acid Drug. Dev. 12:215- 24 (2002).
  • Antisense nucleotides can be from 5-500 nucleotides in length, including 5-200 nucleotides, 5- 100 nucleotides, 10-50 nucleotides, and 10-30 nucleotides. In certain preferred embodiments, the antisense nucleotides are from 14-50 nucleotides in length, including 14-40 nucleotides and 14-30 nucleotides. In certain such embodiments, the antisense nucleotide is capable of binding to a nucleotide segment with sequence as set forth in any one of SEQ ID NO: 1-12.
  • the variants described herein can also be used for the selection and design of antisense reagents that are specific for particular variants. Using information about the variants described herein, antisense oligonucleotides or other antisense molecules that specifically target mRNA molecules that contain one or more variants of the invention can be designed. In this manner, expression of mRNA molecules that contain one or more variant of the present invention (markers and/or haplotypes) can be inhibited or blocked.
  • the antisense molecules are designed to specifically bind a particular allelic form (i.e., one or several variants (alleles and/or haplotypes)) of the target nucleic acid, thereby inhibiting translation of a product originating from this specific allele or haplotype, but which do not bind other or alternate variants at the specific polymorphic sites of the target nucleic acid molecule.
  • allelic form i.e., one or several variants (alleles and/or haplotypes)
  • the molecules can be used for disease treatment.
  • the methodology can involve cleavage by means of ribozymes containing nucleotide sequences complementary to one or more regions in the mRNA that attenuate the ability of the mRNA to be translated.
  • Such mRNA regions include, for example, protein-coding regions, in particular protein-coding regions corresponding to catalytic activity, substrate and/or ligand binding sites, or other functional domains of a protein.
  • RNA interference also called gene silencing, is based on using double-stranded RNA molecules (dsRNA) to turn off specific genes.
  • dsRNA double-stranded RNA molecules
  • siRNA small interfering RNA
  • the siRNA molecules are typically about 20, 21, 22 or 23 nucleotides in length.
  • one aspect of the invention relates to isolated nucleic acid molecules, and the use of those molecules for RNA interference, i.e. as small interfering RNA molecules (siRNA).
  • the isolated nucleic acid molecules are 18-26 nucleotides in length, pr ⁇ 19-25 nucleotides in length, more preferably 20-24 nucleotides in length, and more pref 21, 22 or 23 nucleotides in length.
  • RNAi-mediated gene silencing originates in endogenously encoded primary microRNA (pri-miRNA) transcripts, which are processed in the cell to generate precursor miRNA (pre-miRNA). These miRNA molecules are exported from the nucleus to the cytoplasm, where they undergo processing to generate mature miRNA molecules (miRNA), which direct translational inhibition by recognizing target sites in the 3' untranslated regions of mRNAs, and subsequent mRNA degradation by processing P-bodies (reviewed in Kim & Rossi, Nature Rev. Genet. 8: 173-204 (2007)).
  • pri-miRNA primary microRNA
  • pre-miRNA precursor miRNA
  • RNAi Clinical applications of RNAi include the incorporation of synthetic siRNA duplexes, which preferably are approximately 20-23 nucleotides in size, and preferably have 3' overlaps of 2 nucleotides. Knockdown of gene expression is established by sequence-specific design for the target mRNA. Several commercial sites for optimal design and synthesis of such molecules are known to those skilled in the art.
  • siRNA molecules typically 25-30 nucleotides in length, preferably about 27 nucleotides
  • shRNAs small hairpin RNAs
  • the latter are naturally expressed, as described in Amarzguioui et al. (FEBS Lett. 579: 5974-81 (2005)).
  • Chemically synthetic siRNAs and shRNAs are substrates for in vivo processing, and in some cases provide more potent gene-silencing than shorter designs
  • siRNAs provide for transient silencing of gene expression, because their intracellular concentration is diluted by subsequent cell divisions.
  • expressed shRNAs mediate long-term, stable knockdown of target transcripts, for as long as transcription of the shRNA takes place (Marques et al., Nature Biotechnol. 23: 559-565 (2006); Brummelkamp et al., Science 296: 550-553 (2002)).
  • RNAi molecules including siRNA, miRNA and shRNA
  • the variants presented herein can be used to design RNAi reagents that recognize specific nucleic acid molecules comprising specific alleles and/or haplotypes ⁇ e.g., the alleles and/or haplotypes of the present invention), while not recognizing nucleic acid molecules comprising other alleles or haplotypes.
  • RNAi reagents can thus recognize and destroy the target nucleic acid molecules.
  • RNAi reagents can be useful as therapeutic agents (i.e., for turning off disease-associated genes or disease-associated gene variants), but may also be useful for characterizing and validating gene function (e.g., by gene knock-out or gene knock- down experiments).
  • RNAi may be performed by a range of methodologies known to those skilled in the art. Methods utilizing non-viral delivery include cholesterol, stable nucleic acid-lipid particle (SNALP), heavy-chain antibody fragment (Fab), aptamers and nanoparticles. Viral delivery methods include use of lentivirus, adenovirus and adeno-associated virus.
  • the siRNA molecules are in some embodiments chemically modified to increase their stability. This can include modifications at the 2' position of the ribose, including 2'-0-methylpurines and 2'- fluoropyrimidin.es, which provide resistance to Rnase activity. Other chemical modificati possible and known to those skilled in the art.
  • the variants of the present invention may determine the manner in which a therapeutic agent and/or therapeutic method acts on the body, or the way in which the body metabolizes the therapeutic agent.
  • the presence of a particular at-risk allele for prostate cancer is indicative of a different response, e.g. a different response rate, to a particular treatment modality for prostate cancer.
  • a patient diagnosed with prostate cancer, and carrying a certain allele at a polymorphic or haplotype of the present invention e.g., the at-risk alleles described herein
  • the presence of a marker of the present invention may be assessed (e.g., through testing DNA derived from a blood sample, as described herein). If the patient is positive ,for a marker allele, then the physician recommends one particular therapy, while if the patient is negative for the at least one allele of a marker, or a haplotype, then a different course of therapy may be recommended (which may include recommending that no immediate therapy, other than serial monitoring for progression of the disease, be performed). Thus, the patient's carrier status could be used to help determine whether a particular treatment modality should be administered.
  • the value lies within the possibilities of being able to diagnose the disease at an early stage, to select the most appropriate treatment, and provide information to the clir about prognosis/aggressiveness of the disease in order to be able to apply the most appi treatment.
  • the treatment modality may be any suitable treatment modality used for treating prostate cancer. In certain embodiments, the treatment modality is surgical intervention.
  • susceptibility markers for prostate cancer e.g. the markers as described herein, is combined with assessment or assessment results for a biomarker indicative of prostate cancer, such as Prostate Specific Antigen (PSA).
  • PSA Prostate Specific Antigen
  • the present invention also relates to methods of monitoring progress or effectiveness of a treatment for prostate cancer. This can be done based on the genotype status (i.e., the presence or absence of particular alleles) of an individual for the at-risk alleles for prostate cancer described herein, i.e., by assessing the absence or presence of at least one allele of at least one polymorphic marker as disclosed herein. Alternatively, or concomitantly, the genotype status of at least one risk variant for prostate cancer as presented herein is determined before and during treatment to monitor its effectiveness.
  • the markers of the present invention can be used to increase power and effectiveness of clinical trials.
  • individuals who are carriers of at least one at-risk variant of the present invention may be more likely to respond favorably to a particular treatment modality.
  • Such an application can improve the safety of clinical trials, but can also enhance the chance that a clinical trial will demonstrate statistically significant efficacy, which may be limited to a certain sub-group of the population.
  • one possible outcome of such a trial is that carriers of certain genetic variants, e.g., the markers and haplotypes of the present invention, are statistically significantly likely to show positive response to the therapeutic agent, i.e.
  • the markers and haplotypes of the present invention can be used for targeting the selection of pharmaceutical agents for specific individuals.
  • Personalized selection of treatment modalities, lifestyle changes or combination of lifestyle changes and administration of particular treatment can be realized by the utilization of the at-risk variants of the present invention.
  • the knowledge of an individual's status for particular risk alleles for prostate cancer as described herein can be useful for selection of appropriate treatment options.
  • nucleic acids and polypeptides described herein can be used in methods and kits of the present invention.
  • An "isolated" nucleic acid molecule is one that is separated from nucleic acids that normally flank the gene or nucleotide sequence (as in genomic sequences) and/or has been completely or partially purified from other transcribed sequences (e.g., as in an RNA library).
  • an isolated nucleic acid of the invention can be substantially isolated with respect to the complex cellular milieu in which it naturally occurs, or culture medium when produced by recombinant techniques, or chemical precursors or ot chemicals when chemically synthesized.
  • the isolated material will for of a composition (for example, a crude extract containing other substances), buffer system or reagent mix.
  • the material can be purified to essential homogeneity, for example as determined by polyacrylamide gel electrophoresis (PAGE) or column chromatography (e.g. , HPLC).
  • An isolated nucleic acid molecule of the invention can comprise at least about 50%, at least about 80% or at least about 90% (on a molar basis) of all macromolecular species . present.
  • genomic DNA the term "isolated" also can refer to nucleic acid molecules that are separated from the chromosome with which the genomic DNA is naturally associated.
  • the isolated nucleic acid molecule can contain less than about 250 kb, 200 kb, 150 kb, 100 kb, 75 kb, 50 kb, 25 kb, 10 kb, 5 kb, 4 kb, 3 kb, 2 kb, 1 kb, 0.5 kb or 0.1 kb of the nucleotides that flank the nucleic acid molecule in the genomic DNA of the cell from which the nucleic acid molecule is derived.
  • nucleic acid molecule can be fused to other coding or regulatory sequences and still be considered isolated.
  • recombinant DNA contained in a vector is included in the definition of "isolated” as used herein.
  • isolated nucleic acid molecules include recombinant DNA molecules in heterologous host cells or heterologous organisms, as well as partially or
  • An isolated nucleic acid molecule or nucleotide sequence can include a nucleic acid molecule or nucleotide sequence that is synthesized chemically or by recombinant means. Such isolated nucleotide sequences are useful, for example, in the manufacture of the encoded polypeptide, as probes for isolating homologous sequences (e.g. , from other mammalian species), for gene mapping (e.g. , by in situ hybridization with chromosomes), or for detecting expression of the gene in tissue (e.g. , human tissue), such as by Northern blot analysis or other hybridization techniques.
  • homologous sequences e.g. , from other mammalian species
  • gene mapping e.g. , by in situ hybridization with chromosomes
  • tissue e.g. , human tissue
  • the invention also pertains to nucleic acid molecules that hybridize under high stringency hybridization conditions, such as for selective hybridization, to a nucleotide sequence described herein (e.g. , nucleic acid molecules that specifically hybridize to a nucleotide sequence
  • nucleic acid molecules can be detected and/or isolated by allele- or sequence-specific
  • hybridization e.g. , under high stringency conditions.
  • Stringency conditions and methods for nucleic acid hybridizations are well known to the skilled person (see, e.g. , Current Protocols in Molecular Biology, Ausubel, F. et al, John Wiley & Sons, (1998), and Kraus, M. and Aaronson, S., Methods Enzymol., 200:546-556 (1991), the entire teachings of which are incorporated by reference herein.
  • the length of a sequence aligned for comparison purpo least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%, of the length of the reference sequence.
  • Another example of an algorithm is BLAT (Kent, W.J. Genome Res. 12: 656-64 (2002)).
  • the percent identity between two amino acid sequences can be accomplished using the GAP program in the GCG software package (Accelrys, Cambridge, UK).
  • the present invention also provides isolated nucleic acid molecules that contain a fragment or portion that hybridizes under highly stringent conditions to a nucleic acid that comprises, or consists of, the nucleotide sequence of at least one polymorphic allele contained in the markers and haplotypes described herein.
  • the nucleic acid fragments of the invention are at least about 15, at least about 18, 20, 23 or 25 nucleotides, and can be 30, 40, 50, 100, 200, 500, 1000, 10,000 or more nucleotides in length. In certain embodiments, the nucleic acid fragments are from about 15 to about 1000 nucleotides in length.
  • the nucleic acid fragments are from about 18 to about 100 nucleotides in length, from about 12 to about 50 nucleotides in length, from about 12 to about 40 nucleotides in length, or from about 12 to about 30 nucleotides in length.
  • the present invention further provides isolated nucleic acid molecules that contain a fragment or portion that hybridizes under highly stringent conditions to a nucleic acid that comprises, or consists of, the nucleotide sequence of any one of SEQ ID NO: 1 - 12, as described herein.
  • the nucleic acid fragments can be from 10-600 nucleotides in length, such as from 10 - 500 nucleotides, 12 - 200 nucleotides, 12 - 100 nucleotides, 12 - 50 nucleotides and 12 - 30 nucleotides in length.
  • probes or primers are oligonucleotides that hybridize in a base- specific manner to a complementary strand of a nucleic acid molecule.
  • probes and primers include polypeptide nucleic acids (PNA), as described in Nielsen, P. et al.. Science 254: 1497-1500 (1991).
  • PNA polypeptide nucleic acids
  • a probe or primer comprises a region of nucleotide sequence that hybridizes to at least about 15, typically about 20-25, and in certain embodiments about 40, 50 or 75, consecutive nucleotides of a nucleic acid molecule.
  • probe or primer comprises at least one allele of at least one polymorphic marker or at le haplotype described herein, or the complement thereof.
  • a probe or primer can comprise 100 or fewer nucleotides; for example, in certain embodiments from 6 to 50 nucleotides, or, for example, from 12 to 30 nucleotides.
  • the probe or primer is at least 70% identical, at least 80% identical, at least 85% identical, at least 90% identical, or at least 95% identical, to the contiguous nucleotide sequence or to the complement of the contiguous nucleotide sequence.
  • the probe or primer is capable of selectively hybridizing to the contiguous nucleotide sequence or to the complement of the contiguous nucleotide sequence.
  • the probe or primer further comprises a label, e.g. , a radioisotope, a fluorescent label, an enzyme label, an enzyme co-factor label, a magnetic label, a spin label, an epitope label.
  • the nucleic acid molecules of the invention can be identified and isolated using standard molecular biology techniques well known to the skilled person.
  • the amplified DNA can be labeled (e.g. , radiolabeled, fluorescently labeled) and used as a probe for screening a cDNA library derived from human cells.
  • the cDNA can be derived from mRNA and contained in a suitable vector.
  • Corresponding clones can be isolated, DNA obtained following in vivo excision, and the cloned insert can be sequenced in either or both orientations by art- recognized methods to identify the correct reading frame encoding a polypeptide of the appropriate molecular weight. Using these or similar methods, the polypeptide and the DNA encoding the polypeptide can be isolated, sequenced and further characterized.
  • the invention also provides antibodies which bind to an epitope comprising either a variant amino acid sequence (e.g., comprising an amino acid substitution) encoded by a variant allele or the reference amino acid sequence encoded by the corresponding non-variant or wild-type allele.
  • antibody refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e., molecules that contain antigen-binding sites that specifically bind an antigen.
  • a molecule that specifically binds to a polypeptide of the invention is a molecule that binds to that polypeptide or a fragment thereof, but does not substantially bind other molecules in a sample, e.g., a biological sample, which naturally contains the polypeptide.
  • immunologically active portions of immunoglobulin molecules include F(ab) and F(ab') 2 fragments which can be generated by treating the antibody with an enzyme such as pepsin.
  • the invention provides polyclonal and monoclonal antibodies that bind to a polypeptide of the invention.
  • the term "monoclonal antibody” or “monoclonal antibody composition”, as used herein, refers to a population of antibody molecules that contain only one species of an antigen binding site capable of immunoreacting with a particular epitope of a polypeptide of the invention. A monoclonal antibody composition thus typically displays a single binding affinity for a particular polypeptide of the invention with which it immunoreacts.
  • Polyclonal antibodies can be prepared as described above by immunizing a suitable subj a desired immunogen, e.g., polypeptide of the invention or a fragment thereof.
  • a desired immunogen e.g., polypeptide of the invention or a fragment thereof.
  • the ant titer in the immunized subject can be monitored over time by standard techniques, such as with an enzyme linked immunosorbent assay (ELISA) using immobilized polypeptide.
  • ELISA enzyme linked immunosorbent assay
  • the antibody molecules directed against the polypeptide can be isolated from the mammal (e.g., from the blood) and further purified by well-known techniques, such as protein A
  • antibody-producing cells can be obtained from the subject and used to prepare monoclonal antibodies by standard techniques, such as the hybridoma technique originally described by Kohler and Milstein, Nature 256:495-497 (1975), the human B cell hybridoma technique (Kozbor et al., Immunol. Today 4: 72 (1983)), the EBV-hybridoma technique (Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss,1985, Inc., pp. 77-96) or trioma techniques.
  • hybridomas The technology for producing hybridomas is well known (see generally Current Protocols in Immunology (1994) Coligan et al., (eds.) John Wiley & Sons, Inc., New York, NY). Briefly, an immortal cell line (typically a myeloma) is fused to lymphocytes
  • splenocytes typically splenocytes
  • the culture supernatants of the resulting hybridoma cells are screened to identify a hybridoma producing a monoclonal antibody that binds a polypeptide of the invention.
  • a monoclonal antibody to a polypeptide of the invention can be identified and isolated by screening a recombinant combinatorial immunoglobulin library (e.g., an antibody phage display library) with the polypeptide to thereby isolate immunoglobulin library members that bind the polypeptide.
  • Kits for generating and screening phage display libraries are commercially available (e.g. , the
  • chimeric and humanized monoclonal antibodies can be produced by recombinant DNA techniques known in the art.
  • antibodies of the invention can be used to isolate a polypeptide of the invention by standard techniques, such as affinity chromatography or immunoprecipitation.
  • a polypeptide-specific antibody can facilitate the purification of natural polypeptide from cells and of recombinantly produced polypeptide expressed, in host cells.
  • an antibody specific for a polypeptide of the invention can be used to detect the polypeptide (e.g., in a cellular lysate, cell supernatant, or tissue sample) in order to evaluate the abundance and pattern of expression of the polypeptide.
  • Antibodies can be used diagnostically to monitor protein levels in tissue as part of a clinical testing procedure, e.g., to, for example, determine the efficacy of a given treatment regimen.
  • the antibody can be coupled to a detectable substance to facilitate its detection. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials,
  • bioluminescent materials and radioactive materials.
  • suitable enzymes include horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase;
  • suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin;
  • suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin;
  • an example of a luminescent material includes luminol;
  • examples of bioluminescent materials include luciferase, luciferin, and aequorin, and examples of suitable radioactive material include 125 I, 131 I, 3s S or 3 H.
  • Antibodies may also be useful in pharmacogenomic analysis.
  • antibodies against variant proteins encoded by nucleic acids according to the invention such as variant proteins that are encoded by nucleic acids that contain at least one polymorphic marker of the invention, can be used to identify individuals that require modified treatment modalities.
  • Antibodies can furthermore be useful for assessing expression of variant proteins in disease states, such as in active stages of a disease, or in an individual with a predisposition to a disease related to the function of the protein, in particular prostate cancer.
  • Antibodies specific for a variant protein of the present invention that is encoded by a nucleic acid that comprises at least one polymorphic marker or haplotype as described herein can be used to screen for the presence of the variant protein, for example to screen for a predisposition to prostate cancer as indicated by the presence of the variant protein.
  • Antibodies can be used in other methods. Thus, antibodies are useful as diagnostic tools for evaluating proteins, such as variant proteins of the invention, in conjunction with analysis by electropHoretic mobility, isoelectric point, tryptic or other protease digest, or for use in other physical assays known to those skilled in the art. Antibodies may also be used in tissue typing. In one such embodiment, a specific variant protein has been correlated with expression in a specific tissue type, and antibodies specific for the variant protein can then be used to id the specific tissue type.
  • Subcellular localization of proteins can also be determined using antibodies, and can be applied to assess aberrant subcellular localization of the protein in cells in various tissues. Such use can be applied in genetic testing, but also in monitoring a particular treatment modality. In the case where treatment is aimed at correcting the expression level or presence of the variant protein or aberrant tissue distribution or developmental expression of the variant protein, antibodies specific for the variant protein or fragments thereof can be used to monitor therapeutic efficacy.
  • Antibodies are further useful for inhibiting variant protein function, for example by blocking the binding of a variant protein to a binding molecule or partner. Such uses can also be applied in a therapeutic context in which treatment involves inhibiting a variant protein's function.
  • An antibody can be for example be used to block or competitively inhibit binding, thereby modulating (i.e. , agonizing or antagonizing) the activity of the protein .
  • Antibodies can be prepared against specific protein fragments containing sites required for specific function or against an intact protein that is associated with a cell or cell membrane.
  • an antibody may be linked with an additional therapeutic payload, such as radionuclide, an enzyme, an immunogenic epitope, or a cytotoxic agent, including bacterial toxins (diphtheria or plant toxins, such as ricin) .
  • an additional therapeutic payload such as radionuclide, an enzyme, an immunogenic epitope, or a cytotoxic agent, including bacterial toxins (diphtheria or plant toxins, such as ricin) .
  • the in vivo half-life of an antibody or a fragment thereof may be increased by pegylation through conjugation to polyethylene glycol.
  • kits for using antibodies in the methods described herein includes, but is not limited to, kits for detecting the presence of a variant protein in a test sample.
  • kits for detecting the presence of a variant protein in a test sample comprises antibodies such as a labelled or labelable antibody and a compound or agent for detecting variant proteins in a biological sample, means for determining the amount or the presence and/or absence of variant protein in the sample, and means for comparing the amount of variant protein in the sample with a standard, as well as instructions for use of the kit.
  • the methods and information described herein may be implemented, in all or in part, as computer executable instructions on known computer readable media.
  • the methods described herein may be implemented in hardware.
  • the method may be implemented in software stored in, for example, one or more memories or other computer readable medium and implemented on one or more processors.
  • the processors may be associated with one or more controllers, calculation units and/or other units of a computer system, or implanted in firmware as desired.
  • the routines may be stored in any computer readable memory such as in RAM, ROM, flash memory, a magnetic disk, a laser disk, or other storage medium, as is also known.
  • this software may be delivered to a computing device via any known delivery method including, for example, over a communication channel such as a telephone line, the Internet, a wireless connection, etc., or via a transportable medium, such as a computer readable d drive, etc.
  • a communication channel such as a telephone line, the Internet, a wireless connection, etc.
  • a transportable medium such as a computer readable d drive, etc.
  • the various steps described above may be implemented as various blocks, operations, tools, modules and techniques which, in turn, may be implemented in hardware, firmware, software, or any combination of hardware, firmware, and/or software.
  • some or all of the blocks, operations, techniques, etc. may be implemented in, for example, a custom integrated circuit (IC), an application specific integrated circuit (ASIC), a field programmable logic array (FPGA), a programmable logic array (PLA), etc.
  • the software When implemented in software, the software may be stored in any known computer readable medium such as on a magnetic disk, an optical disk, or other storage medium, in a RAM or ROM or flash memory of a computer, processor, hard disk drive, optical disk drive, tape drive, etc. Likewise, the software may be delivered to a user or a computing system via any known delivery method including, for example, on a computer readable disk or other transportable computer storage mechanism.
  • a system of the invention includes one or more machines used for analysis of biological material (e.g., genetic material), as described herein. In some variations, this analysis of the biological material involves a chemical analysis and/or a nucleic acid amplification.
  • biological material e.g., genetic material
  • an exemplary system of the invention which may be used to implement one or more steps of methods of the invention, includes a computing device in the form of a computer 110.
  • a computing device in the form of a computer 110.
  • Components shown in dashed outline are not technically part of the computer 110, but are used to illustrate the exemplary embodiment of Fig. 1.
  • Components of computer 110 may include, but are not limited to, a processor 120, a system memory 130, a
  • memory/graphics interface 121 also known as a Northbridge chip
  • I/O interface 122 also known as a Southbridge chip
  • the system memory 130 and a graphics processor 190 may be coupled to the memory/graphics interface 121.
  • a monitor 191 or other graphic output device may be coupled to the graphics processor 190.
  • a series of system busses may couple various system components including a high speed system bus 123 between the processor 120, the memory/graphics interface 121 and the I/O interface 122, a front-side bus 124 between the memory/graphics interface 121 and the system memory 130, and an advanced graphics processing (AGP) bus 125 between the memory/graphics interface 121 and the graphics processor 190.
  • the system bus 123 may be any of several types of bus structures including, by way of example, and not limitation, such architectures im
  • ISA Industry Standard Architecture
  • MCA Micro Channel Architecture
  • EISA ISA
  • Intel Intel Hub Architecture
  • HypertransportTM HypertransportTM
  • the computer 110 typically includes a variety of computer-readable media.
  • Computer-readable media can be any available media that can be accessed by computer 110 and includes both volatile and nonvolatile media, removable and non-removable media.
  • Computer readable media may comprise computer storage media.
  • Computer storage media includes both volatile and nonvolatile, removable and non-removable media implemented in any method or technology for storage of information such as computer readable instructions, data structures, program modules or other data.
  • Computer storage media includes, but is not limited to, RAM, ROM, EEPROM, flash memory or other memory technology, CD-ROM, digital versatile disks (DVD) or other optical disk storage, magnetic cassettes, magnetic tape, magnetic disk storage or other magnetic storage devices, or any other physical medium which can be used to store the desired information and which can accessed by computer 110.
  • the system memory 130 includes computer storage media in the form of volatile and/or nonvolatile memory such as read only memory (ROM) 131 and random access memory (RAM) 132.
  • the system ROM 131 may contain permanent system data 143, such as identifying and manufacturing information.
  • a basic input/output system (BIOS) may also be stored in system ROM 131.
  • RAM 132 typically contains data and/or program modules that are immediately accessible to and/or presently being operated on by processor 120.
  • Fig. 1 illustrates operating system 134, application programs 135, other program modules 136, and program data 137.
  • the I/O interface 122 may couple the system bus 123 with a number of other busses 126, 127 and 128 that couple a variety of internal and external devices to the computer 110.
  • a serial peripheral interface (SPI) bus 126 may connect to a basic input/output system (BIOS) memory 133 containing the basic routines that help to transfer information between elements within computer 110, such as during start-up.
  • BIOS basic input/output system
  • a super input/output chip 160 may be used to connect to a number of 'legacy' peripherals, such as floppy disk 152, keyboard/mouse 162, and printer 196, as examples.
  • the super I/O chip 160 may be connected to the I/O interface 122 with a bus 127, such as a low pin count (LPC) bus, in some embodiments.
  • a bus 127 such as a low pin count (LPC) bus, in some embodiments.
  • LPC low pin count
  • Various embodiments of the super I/O chip 160 are widely available in the commercial marketplace.
  • bus 128 may be a Peripheral Component Interconnect (PCI) bus, or a variation thereof, may be used to connect higher speed peripherals to the I/O interface 122.
  • PCI Peripheral Component Interconnect
  • a PCI bus may also be known as a Mezzanine bus.
  • Variations of the PCI bus include the Peripheral Component Interconnect-Express (PCI-E) and the Peripheral Component Interconnect - Extended (PCI-X) busses, the former having a serial interface and the latter being a backward compatible parallel interface.
  • bus 128 may be an advanced tech attachment (ATA) bus, in the form of a serial ATA bus (SATA) or parallel ATA (PATA).
  • ATA advanced tech attachment
  • the computer 110 may also include other removable/non-removable, volatile/nonvolatile computer storage media.
  • Fig. 1 illustrates a hard disk drive 140 that reads from or writes to non-removable, nonvolatile magnetic media.
  • the hard disk drive 140 may be a conventional hard disk drive.
  • Removable media such as a universal serial bus (USB) memory 153, firewire (IEEE 1394), or CD/DVD drive 156 may be connected to the PCI bus 128 directly or through an interface 150.
  • a storage media 154 may be coupled through interface 150.
  • Other removable/non-removable, volatile/nonvolatile computer storage media that can be used in the exemplary operating environment include, but are not limited to, magnetic tape cassettes, flash memory cards, digital versatile disks, digital video tape, solid state RAM, solid state ROM, and the like.
  • the drives and their associated computer storage media discussed above and illustrated in Fig. 1, provide storage of computer readable instructions, data structures, program modules and other data for the computer 110.
  • hard disk drive 140 is illustrated as storing operating system 144, application programs 145, other program modules 146, and program data 147. Note that these components can either be the same as or different from operating system 134, application programs 135, other program modules 136, and program data 137. Operating system 144, application programs 145, other program modules 146, and program data 147 are given different numbers here to illustrate that, at a minimum, they are different copies.
  • a user may enter commands and information into the computer 20 through input devices such as a mouse/keyboard 162 or other input device combination.
  • Other input devices may include a microphone, joystick, game pad, satellite dish, scanner, or the like. These and other input devices are often connected to the processor 120 through one of the I/O interface busses, such as the SPI 126, the LPC 127, or the PCI 128, but other busses may be used. In some embodiments, other devices may be coupled to parallel ports, infrared interfaces, game ports, and the like (not depicted), via the super I/O chip 160.
  • the computer 110 may operate in a networked environment using logical connections to one or more remote computers, such as a remote computer 180 via a network interface controller (NIC) 170.
  • the remote computer 180 may be a personal computer, a server, a router, a network PC, a peer device or other common network node, and typically includes many or all of the elements described above relative to the computer 110.
  • the logical connection between the NIC 170 and the remote computer 180 depicted in Fig. 1 may include a local area network (LAN), a wide area network (WAN), or both, but may also include other networks.
  • LAN local area network
  • WAN wide area network
  • Such networking environments are commonplace in offices, enterprise-wide computer networks, intranets, and the Internet.
  • the remote computer 180 may also represent a web server supporting interactive sessions with the computer 110, or in the specific case of location-based applications may be a location server or an application server.
  • the network interface may use a modem (not depicted) when a broadband connection is not available or is not used. It will be appreciated that the network connection shown is exemplary and other means of establishing a communications link t the computers may be used.
  • the invention is a system for identifying susceptibility to a cancer in a human subject.
  • the system includes tools for performing at least one step, preferably two or more steps, and in some aspects all steps of a method of the invention, where the tools are operably linked to each other.
  • Operable linkage describes a linkage through which components can function with each other to perform their purpose.
  • a system of the invention is a system for identifying susceptibility to prostate cancer in a human subject, and comprises:
  • a susceptibility database operatively coupled to a computer-readable medium of the system and containing population information correlating the presence or absence of one or more alleles of at least one polymorphic marker is selected from the group consisting of chr8: 128173299 (rsl83373024,SEQ ID NO: l) and rsl46851087 (SEQ ID NO:8), and correlated markers in linkage disequilibrium therewith susceptibility to the cancer in a population of humans;
  • (iii) is adapted to be executed on a processor of the system, to compare the information about the human subject with the population information in the susceptibility database and generate a conclusion with respect to susceptibility to the cancer for the human subject.
  • the correlated markers are. selected from the markers set forth in Table 1 herein.
  • the at least one polymorphic marker is selected from the group consisting of chr8: 128173299, chr8: 128260854, chr8: 128277551, chr8: 128354590 and chr8: 128010975.
  • Exemplary processors include all variety of microprocessors and other processing units used in computing devices.
  • Exemplary computer-readable media are described above.
  • the system generally can be created where a single processor and/or computer readable medium is dedicated to a single component of the system; or where two or more functions share a single processor and/or share a single computer readable medium, such that the system contains as few as one processor and/or one computer readable medium.
  • it is advantageous to use multiple processors or media for example, wl convenient to have components of the system at different locations.
  • some components of a system may be located at a testing laboratory dedicated to laboratory or data analysis, whereas other components, including components (optional) for supplying input information or obtaining an output communication, may be located at a medical treatment or counseling facility (e.g., doctor's office, health clinic, HMO, pharmacist, geneticist, hospital) and/or at the home or business of the human subject (patient) for whom the testing service is performed.
  • a medical treatment or counseling facility e.g., doctor's office, health clinic, HMO, pharmacist, geneticist, hospital
  • an exemplary system includes a susceptibility database 208 that is operatively coupled to a computer-readable medium of the system and that contains population information correlating the presence or absence of at least one allele of at least one polymorphic marker selected from the group consisting of chr8: 128173299 and rsl46851087, and correlated markers in linkage disequilibrium therewith and susceptibility to prostate cancer in a population of humans.
  • the correlated markers are selected from the markers set forth in Table 1 herein.
  • the at least one polymorphic marker is selected from the group consisting of chr8: 128173299, chr8: 128260854, chr8: 128277551,
  • the susceptibility database contains 208 data relating to the frequency that a particular allele of the at least one marker has been observed in a population of humans with prostate cancer and a population of humans free of the cancer. Such data provides an indication as to the relative risk or odds ratio of developing the cancer for a human subject that is identified as having the allele in question.
  • the susceptibility database includes similar data with respect to two or more alleles of at least one marker, thereby providing a useful reference if the human subject has any of the two or more alleles.
  • the susceptibility database includes additional quantitative personal, medical, or genetic information about the individuals in the database diagnosed with prostate cancer or free of prostate cancer.
  • Such information includes, but is not limited to, information about parameters such as age, sex, ethnicity, race, medical history, weight, diabetes status, blood pressure, family history of the cancer, smoking history, and alcohol use in humans and impact of the at least one parameter on susceptibility to the cancer.
  • the information also can include information about other genetic risk factors for the cancer besides these aforementioned markers.
  • the system further includes a measurement tool 206 programmed to receive an input 204 from or about the human subject and generate an output that contains information about the presence or absence of the at least one allele of interest.
  • the input 204 is not part of the system per se but is illustrated in the schematic Figure 2.
  • the input 204 will contain a specimen or contain data from which the presence or absence of the at least one allele can be directly read, or analytically determined.
  • the input contains annotated information about genotypes or allele counts for the markers predictive of prostate cancer risk in the genome of the human subject, in which further processing by the measurement tool 206 is required, except possibly transforma the relevant information about the presence/absence of the at least one allele into a format compatible for use by the analysis routine 210 of the system.
  • the input 204 from the human subject contains data that is unannotated or insufficiently annotated with respect to the at least one marker, requiring analysis by the measurement tool 206.
  • the input can be genetic sequence of a chromosomal region or chromosome on which the marker resides, or whole genome sequence information, or unannotated information from a gene chip analysis of a variable loci in the human subject's genome.
  • the measurement tool 206 comprises a tool, preferably stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to receive a data input about a subject and determine information about the presence or absence of the at least one mutant allele in a human subject from the data.
  • the measurement tool 206 contains instructions, preferably executable on a processor of the system, for analyzing the unannotated input data . and determining the presence or absence of the allele of interest in the human subject.
  • the input data is genomic sequence information
  • the measurement tool optionally comprises a sequence analysis tool stored on a computer readable medium of the system and executable by a processor of the system with instructions for determining the presence or absence of the at least one mutant allele from the genomic sequence information.
  • the input 204 from the human subject comprises a biological sample, such as a fluid (e.g., blood) or tissue sample that contains genetic material that can be analyzed to determine the presence or absence of the allele of interest.
  • a biological sample such as a fluid (e.g., blood) or tissue sample that contains genetic material that can be analyzed to determine the presence or absence of the allele of interest.
  • an exemplary measurement tool 206 includes laboratory equipment for processing and analyzing the sample to determine the presence or absence (or identity) of the allele(s) in the human subject.
  • the measurement tool includes: an oligonucleotide microarray (e.g., "gene chip") containing a plurality of oligonucleotide probes attached to a solid support; a detector for measuring interaction between nucleic acid obtained from or amplified from the biological sample and one or more oligonucleotides on the oligonucleotide microarray to generate detection data; and an analysis tool stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to determine the presence or absence of the at least one allele of interest based on the detection data.
  • an oligonucleotide microarray e.g., "gene chip”
  • a detector for measuring interaction between nucleic acid obtained from or amplified from the biological sample and one or more oligonucleotides on the oligonucleotide microarray to generate detection data
  • an analysis tool stored on a computer-readable medium of the system and adapted to be executed on
  • the measurement tool 206 includes: a nucleotide sequencer (e.g., an automated DNA sequencer) that is capable of determining nucleotide sequence information from nucleic acid obtained from or amplified from the biological sample; and an analysis tool stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to determine the presence or absence of the at least one mutant allele based on the nucleotide sequence information.
  • a nucleotide sequencer e.g., an automated DNA sequencer
  • an analysis tool stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to determine the presence or absence of the at least one mutant allele based on the nucleotide sequence information.
  • the measurement tool 206 further includes additional equipment and/or chemical reagents for processing the biological sample to purify and/or amplify nucleic acid of the human subject for further analysis using a sequencer, gene chip, or other analytical equipment.
  • the exemplary system further includes an analysis tool or routine 210 that: is operatively coupled to the susceptibility database 208 and operatively coupled to the measurement tool 206, is stored on a computer-readable medium of the system, is adapted to be executed on a processor of the system to compare the information about the human subject with the population information in the susceptibility database 208 and generate a conclusion with respect to susceptibility to the cancer for the human subject.
  • the analysis tool 210 looks at the alleles identified by the measurement tool 206 for the human subject, and compares this information to the susceptibility database 208, to determine a susceptibility to the cancer for the subject.
  • the susceptibility can be based on the single parameter (the identity of one or more alleles of one or more markers predictive of risk of prostate cancer), or can involve a calculation based on other genetic and non-genetic data, as described above, that is collected and included as part of the input 204 from the human subject, and that also is stored in the susceptibility database 208 with respect to a population of other humans.
  • each parameter of interest is weighted to provide a conclusion with respect to susceptibility to prostate cancer.
  • Such a conclusion is expressed in the conclusion in any statistically useful form, for example, as an odds ratio, a relative risk, or a lifetime risk for subject developing prostate cancer.
  • system as just described further includes a
  • the communication tool is operatively connected to the analysis routine 210 and comprises a routine stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to: generate a communication containing the conclusion; and to transmit the communication to the human subject 200 or the medical practitioner 202, and/or enable the subject or medical practitioner to access the communication.
  • the communication tool 212 provides an interface for communicating to the subject, or to a medical practitioner for the subject (e.g., doctor, nurse, genetic counselor), the conclusion generated by the analysis tool 210 with respect to susceptibility to the cancer for the subject.
  • the medical practitioner will share the communication with the human subject 200 and/or counsel the human subject about the medical significance of the communication.
  • the communication tool is operatively connected to the analysis routine 210 and comprises a routine stored on a computer-readable medium of the system and adapted to be executed on a processor of the system, to: generate a communication containing the conclusion; and to transmit the communication to the human subject 200 or the medical practitioner 202, and/or enable the
  • the communication is provided in a tangible form, such as a printed report or report stored on a computer readable medium such as a flash drive or optical disk.
  • the communication is provided electronically with an output that is visible on a video display or audio output (e.g., speaker).
  • the communication is transmitted to the subject or the medical practitioner, e.g., electronically or through the mail.
  • the system is designed to permit the subject or medical practitioner to access the communication, e.g., by telephone or computer.
  • the system may include software residing on a memory and executed by a processor of a computer used by the human subject or the medical practitioner, with which the subject or practitioner can access the communication, prefer securely, over the internet or other network connection.
  • the syste computer will be located remotely from other components of the system, e.g., at a location of the human subject's or medical practitioner's choosing.
  • system as described further includes components that add a treatment or prophylaxis utility to the system. For instance, value is added to a determination of
  • susceptibility to a cancer when a medical practitioner can prescribe or administer a standard of care that can reduce susceptibility to prostate cancer; and/or delay onset of the cancer; and/or increase the likelihood of detecting the cancer at an early stage, to facilitate early treatment when the cancer has not spread and is most curable.
  • Exemplary lifestyle change protocols include loss of weight, increase in exercise, cessation of unhealthy behaviors such as smoking, and change of diet.
  • Exemplary medicinal and surgical intervention protocols include
  • exemplary diagnostic protocols include non-invasive and invasive imaging; monitoring metabolic biomarkers; and biopsy screening.
  • the system further includes a medical protocol database 214 operatively connected to a computer-readable medium of the system and containing information correlating the presence or absence of the at least one marker allele of interest and medical protocols for human subjects at risk for the cancer.
  • medical protocols include any variety of medicines, lifestyle changes, diagnostic tests, increased frequencies of diagnostic tests, and the like that are designed to achieve one of the aforementioned goals.
  • the information correlating a marker allele with protocols could include, for example, information about the success with which the cancer is avoided or delayed, or success with which the cancer is detected early and treated, if a subject has a marker susceptibility allele and follows a protocol.
  • the system of this embodiment further includes a medical protocol tool or routine 216, operatively connected to the medical protocol database 214 and to the analysis tool or routine 210.
  • the medical protocol tool or routine 216 preferably is stored on a computer-readable medium of the system, and adapted to be executed on a processor of the system, to: (i) compare (or correlate) the conclusion that is obtained from the analysis routine 210 (with respect to susceptibility to cancer for the subject) and the medical protocol database 214, and (ii) generate a protocol report with respect to the probability that one or more medical protocols in the medical protocol database will achieve one or more of the goals of reducing susceptibility to prostate cancer; delaying onset of the cancer; and increasing the likelihood of detecting the cancer at an early stage to facilitate early treatment.
  • the probability can be based on empirical evidence collected from a population of humans and expressed either in absolute terms (e.g., compared to making no intervention), or expressed in relative terms, to highlight the
  • the communication tool 212 includes the protoco in addition to, or instead of, the conclusion with respect to susceptibility.
  • Information about allele status of risk marker of prostate cancer not only can provide useful information about identifying or quantifying susceptibility to prostate cancer; it can also provide useful information about possible causative factors for a human subject identified with prostate cancer, and useful information about therapies for the cancer patient. In some variations, systems of the invention are useful for these purposes.
  • the invention is a system for assessing or selecting a treatment protocol for a subject diagnosed with prostate cancer.
  • An exemplary system schematically depicted in Figure 3, comprises:
  • a medical treatment database 308 operatively connected to a computer-readable medium of the system and containing information correlating the presence or absence of at least one marker allele selected from the group consisting of chr8: 128173299 allele G, rsl46851087 allele A, and marker alleles correlated therewith by values of r 2 > 0.5, and efficacy of treatment regimens for prostate cancer;
  • a measurement tool 306 to receive an input (304, depicted in Fig. 3 but not part of the system per se) about the human subject and generate information from the input 304 about the presence or absence of the at least one marker allele in a human subject diagnosed with prostate cancer;
  • a medical protocol routine or tool 310 operatively coupled to the medical treatment database 308 and the measurement tool 306, stored on a computer-readable medium of the system, and adapted to be executed on a processor of the system, to compare the information with respect to presence or absence of the at least one marker allele for the subject and the medical treatment database, and generate a conclusion with respect to at least one of:
  • such a system further includes a communication tool 312 operatively connected to the medical protocol tool or routine 310 for communicating the conclusion to the subject 300, or to a medical practitioner for the subject 302 (both depicted in the schematic of Fig. 3, but not part of the system per se).
  • An exemplary communication tool comprises a routine stored on a - computer-readable medium of the system and adapted to be executed on a processor of the system, to generate a communication containing the conclusion; and transmit the
  • the correlated markers are selected from the markers set forth in T (A) and (B) herein.
  • the at least one polymorphic marker is sel from the group consisting of chr8: 128173299, chr8: 128260854, chr8: 128277551, chr8: 128354590 and chr8: 128010975.
  • genotyping information into individuals for whom neither SNP chip nor sequence data were available, a process referred to as "genealogy-based in silico genotyping".
  • chr8: 128173299 signal This second variant, identified as rsl46851087, is located at position 127916112 on chromosome 8 in NCBI Build 36 of the human genome assembly. The A allele of this variant was found to confer a risk of prostate cancer with OR of 2.25 and a P-value of 5.21X10 "14 .
  • Table 2 summarizes results obtained for the two signals on chromosome 8q24.
  • Illumina SNP Chip Genotyping The Icelandic chip-typed samples were assayed with the Illumina Human Hap300, Hap CNV370, Hap 610, 1 or Omni-1 Quad bead chips at deCODE genetics. Only the 317,503 SNPs from the Human Hap300 chip were used in the long range phasing and the subsequent SNP imputations.
  • SNPs were excluded if they had (i) yield lower than 95%, (ii) minor allele frequency less than 1% in the population or (iii) significant deviation from Hardy- Weinberg equilibrium in the controls (P ⁇ 0.001), (iv) if they produced an excessive inheritance error rate (over 0.001), (v) if there was substantial difference in allele frequency between chip types (from just a single chip if the problem that resolved all differences, but from all chips otherwise). All samples with a call rate below 97% were excluded from the analysis. The final set of SNPs used for long range phasing was composed of 297,835 autosomal SNPs.
  • SNPs were imputed based on whole genome sequence data from about 1176 Icelanders, selected for various neoplastic, cardiovascular and psychiatric conditions. All of the individuals were sequenced at a depth of at least 10X.
  • Enriched libraries were further purified using agarose (2%) gel electrophoresis as described above. The quality and concentration of the libraries were assessed with the Agilent 2100 Bioanalyzer using the DNA 1000 LabChip (Agilent). Barcoded libraries were stored at -20 °C. All steps in the workflow were monitored using an in-house laboratory information management system with barcode tracking of all samples and reagents.
  • Sequencing-by-synthesis was performed on Illumina GAIIx instruments equipped with p, end modules. Paired-end libraries were sequenced using 2 x 101 cycles of incorporation and imaging with Illumina sequencing kits, v4 or v5 (TruSeq). Each library or sample was initially run on a single lane for validation followed by further sequencing of >4 lanes with targeted raw cluster densities of 500-700 k/mm 2 , depending on the version of the data imaging and analysis packages. Imaging and analysis of the data was performed using either the SCS2.6 /RTA1.6 or SCS2.8/RTA1.8 software packages from Illumina, respectively. Real-time analysis involved conversion of image data to base-calling in real-time.
  • SNP identification and genotype calling A two-step approach was applied. The first step was to detect SNPs by identifying sequence positions where at least one individual could be determined to be different from the reference sequence with confidence (quality threshold of 20) based on the SNP calling feature of the pileup tool in SAMtools. SNPs that always differed heterozygous or homozygous from the reference were removed. The second step was to use the pileup tool to genotype the SNPs at the positions that were flagged as polymorphic. Because sequencing depth varies and hence the certainty of genotype calls also varies, genotype likelihoods rather than deterministic calls were calculated.
  • Long range phasing Long range phasing of all chip-genotyped individuals was performed with methods described previously (Kong, A. et al. Nat Genet 40, 1068-75 (2008); Holm, H. et al. Nat Genet 43, 316-20 (2011)). In brief, phasing is achieved using an iterative algorithm which phases a single proband at a time given the available phasing information about everyone else that shares a long haplotype identically by state with the proband. Given the large fraction of the Icelandic population that has been chip-typed, accurate long range phasing is available genome- wide for all chip-typed Icelanders. For long range phased haplotype association analysis, we then partitioned the genome into non-overlapping fixed 0.3cM bins. Within each bin, we observed the haplotype diversity described by the combination of all chip-typed markers in the bin. Hi with frequencies over 0.001 were tested in a case : control analysis.
  • Genotype imputation We imputed the SNPs identified and genotyped through sequencing into all Icelanders who had been phased with long range phasing using the same model as used by IMPUTE (Kong, A. et al. Nat Genet 40, 1068-75 (2008)). The genotype data from sequencing can be ambiguous due to low sequencing coverage. In order to phase the sequencing genotypes, an iterative algorithm was applied for each SNP with alleles 0 and 1. We let H be the long range phased haplotypes of the sequenced individuals and applied the following algorithm :
  • the genotype likelihood L g is the probability of the observed sequencing data at the SNP for a given individual assuming g is the true genotype at the SNP. If L 0 ,
  • Z-! and L 2 are the likelihoods of the genotypes 0, 1 and 2 in the individual that carries h, then set 3b .
  • step 3 when the maximum difference between iterations is greater than a
  • the above algorithm can easily be extended to handle simple family structures such as parent- offspring pairs and triads by letting the P distribution run over all founder haplotypes in the family structure.
  • the algorithm also extends trivially to the X-chromosome. If source genotype data are only ambiguous in phase, such as chip genotype data, then the algorithm is still applied, but all but one of the Ls will be 0.
  • the reference set was intentionally enriched for carriers of the minor allele of a rare SNP in order to improve imputation accuracy. In this case, expected allele counts will be biased toward the minor allele of the SNP.
  • Genotype imputation information The informativeness of genotype imputation was estimated by the ratio of the variance of imputed expected allele counts and the variance of the actual allele counts:
  • Genealogy-based in silico genotyping In addition to imputing sequence variants from the whole genome sequencing effort into chip genotyped individuals, we also performed a second imputation step where genotypes were imputed into relatives of chip genotyped individuals, creating in silico genotypes.
  • the inputs into the second imputation step are the fully phased (in particular every allele has been assigned its parent of origin (Kong, A. et al. Nature 462, 868-74 (2009)) imputed and chip type genotypes of the available chip typed individual.
  • the algorithm used to perform the second imputation step consists of:
  • the proband For each acheotyped individual (the proband), find all chip genotyped individuals within two meioses of the individual. The six possible types of two meiotic distance relatives of the proband are (ignoring more complicated relationships due to pedigree loops):
  • Haplotypes that are the same, except at most at a single SNP, are treated as identical.
  • haplotypes in the pedigree are incompatible over a bin, then a uniform probability distribution was used for that bin.
  • the most common causes for such incompatibilities are recombinations within the pedigree, phasing errors and genotyping errors.
  • the single point information is substantially more informative than for unphased genotyped, in particular one haplotype of the parent of a genotyped child is always known.
  • the single point distributions are then con using the multipoint algorithm to obtain multipoint sharing probabilities at the ce each bin. Genetic distances were obtained from the most recent version of the deCODE genetic map (Kong, A. et al. Nature 467, 1099-103 (2010)).
  • Oc + (1 - 0)0 is an estimate of the allele count for the proband's paternal haplotype. Similarly, an expected allele count can be obtained for the proband's maternal haplotype.
  • the mean informativeness values cluster into groups corresponding to the most common pedigree configurations used in the imputation, such as imputing from parent into child or from child into parent.
  • Inflation Factor Adjustment In order to account for the relatedness and stratification within the case and control sample sets we applied the method of genomic control based on chip typed markers. Quoted P values were adjusted accordingly.
  • the SNP, rsl88140481, is located telomeric to the previously published region-2 on 8q24;
  • rsl6901979 has the closest resemblance in terms of conferred risk and allele frequency (risk allele control frequency in Iceland is ⁇ 3% and ⁇ 1% for rsl6901979[A] and rsl88140481[A], respectively). This indicates that the two risk variants are never together on the same haplotype. Consequently, the association results for rsl6901979 and rsl88140481 remain significant after being adjusted for each other.
  • rsl88i40481 is only very weakly correlated (r 2 ⁇ 0.06) with the previously published leukemia 15 , prostate 7"10,14 , colon 16"18 , breast 19 , or bladder cancer 20 risk variants on 8q24 (Table 3), and the association results for rsl88140481 remain significant after being adjusted for all nine previously published prostate cancer risk SNPs at 8q24 (Table 8).
  • CLL chronic lymphocytic leukemia
  • c rsl88140481 128260854 1.0 1.0 A T 0.0111 rsl38042437 128277551 1.0 1.0 G A 0.0111 6 chr8: 128354590* 128354590 0.94 0.97 G GATAA 0.0114 1 rsl83373024 128173299 0.80 1.0 G A 0.0140 ⁇ rsl82352457 128010975 0.74 0.97 A G 0.00892
  • na denotes that the variants is being included in the logistic regression, hence, it is not applicable to show results for that variant and a particular step of the regression analysis.
  • prostate cancer diagnois Study populations Icelandic study population.
  • the ICR contains 5,141 Icelandic prostate cancer patients diagnosed from January 1, 1955, to December 31, 2010.
  • the Icelandic prostate cancer sample collection included 2,315 patients (diagnosed from December 1974 to December 2008) who were recruited from November 2000 until June 2010.
  • a total of 4,537 patients were included in the study of which 2,315 had genotypes from a genome wide SNP genotyping effort, using the Infinium II assay method and the Sentrix HumanHap300 BeadChip (Illumina, San Diego, CA, USA), and 2,222 had imputed genotypes based on genotypic information from first or second degree relatives that have been chip-genotyped.
  • the 4,537 prostate cancer patients 53 were among the 1,795 individuals who had been whole genome sequenced.
  • the mean age at diagnosis is 71 years for prostate cancer patients in the ICR.
  • aggressive prostate cancer is defined as: Gleason >7 and/or T3 or higher and/or node positive and/or metastatic disease, while the less aggressive disease is defined as Gleason ⁇ 7 and T2 or lower.
  • the 54,444 male controls (27,780 had variants imputed based on chip-genotypes and 26,664 had variants imputed with a family based methods) are comprised of individuals recruited through different genetic research projects at deCODE.
  • the controls have been diagnosed with common diseases of the cardio-vascular system (e.g. stroke or. myocardial infraction), psychiatric and neurological diseases (e.g. schizophrenia, bipolar disorder), endocrine and autoimmune system (e.g. type 2 diabetes, asthma), malignant diseases other than prostate cancer as well as individuals randomly selected from the Icelandic genealogical database.
  • the controls had a mean age of 84 years and the range was from 8 to 105 years.
  • the controls were absent from the nation-wide list of prostate cancer patients according to the ICR.
  • the total number of Dutch prostate cancer cases used in this study was 1,545.
  • the Dutch study population consisted of two recruitment-sets of prostate cancer cases; Group-A was comprised of 360 hospital-based cases recruited from January 1999 to June 2006 at the Urology Outpatient Clinic of the Radboud University Nijmegen Medical Centre (RUIMMC); Group-B consisted of 707 cases recruited from June 2006 to December 2006 through a population-based cancer registry held by the Comprehensive Cancer Centre IKO. Both groups were of self-reported European descent.
  • the average age at diagnosis for patients in Group-A was 63 years (median 63 years; range 43 to 83 years).
  • the average age at diagnosis for patients in Group-B was 65 years (median 66 years; range 43 to 75 years).
  • the 1,960 control individuals were cancer free and were matched for age with the cases. They were recruited within a project entitled "The Nijmegen Biomedical Study", in the Netherlands. This is a population-based survey conducted by the Department of Epidemiology and Biostatistics and the Department of Clinical Chemistry of RUNMC, in which 9,371 individuals participated from a total of 22,500 age and sex stratified, randomly selected inhabitants of Nijmegen. Control individuals from the Nijmegen Biomedical Study were invited to participate in a study on gene-environment interactions in multifactorial diseases, such as cancer. All the Dutch participants in the present study are of self-reported European descent and were fully informed about the goals and the procedures of the study. The study protocol was approved by the Institutional Review Board of Radboud University and all study subjects gave written informed consent.
  • Clinical information including age at onset, grade and stage was obtained from medical records.
  • the average age at diagnosis for the patients was 69 years (median 70 years) and the range was from 44 to 83 years.
  • the 1,635 Spanish control individuals were approached at the
  • the Romanian study population used in this study consisted of 738 prostate cancer cases.
  • the cases were recruited from the Urology Clinic “Theodor Burghele” of The University of Medicine and Pharmacy “Carol Davila” Bucharest, Romania, from May 2008 to November 2009. All patients were of self-reported European descent.
  • Clinical information including age at onset, grade and stage were obtained from medical records at the hospital.
  • the average age at diagnosis for the cases was 70 years (median 71 years) and the range was from 46 to 89 years.
  • the 932 Romanian controls were recruited at the General Surgery Clinic “St. Mary” and at the Urology Clinic "Theodor Burghele” of The University of Medicine and Pharmacy "Carol Davila”
  • Hong Kong The Hong Kong study population used in this study consisted of 498 prostate cancer cases. The cases were recruited from the Division of Urology, Department of Surgery, Prince of Wales Hospital in Hong Kong, China, from October 2007 to June 2009. All patients were of self-reported Chinese descent. Clinical information including age at onset, grade and stage was obtained from medical records. The average age at diagnosis for the patients was 70.3 years (median 71 years) and the range was from 46 to 92 years. Study protocol was approved by the joint ethics committee of The Chinese University of Hong Kong and New Territories East Cluster Clinical Research. All subjects gave written informed consent. Lichtenstein, P. et al. Environmental and heritable factors in the causation of can ⁇ . analyses of cohorts. of twins from Sweden, Denmark, and Finland. N Engl J Med 343, 78- 85. (2000).

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Abstract

Il a été découvert que certaines variantes génétiques peuvent prédire un risque du cancer de la prostate chez des êtres humains. La présente invention concerne de telles variantes et leur utilisation dans des procédés de détermination de la probabilité du cancer de la prostate chez des êtres humains. L'invention concerne également des trousses et des systèmes informatiques destinés à être utilisés dans des tels procédés.
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US9857328B2 (en) 2014-12-18 2018-01-02 Agilome, Inc. Chemically-sensitive field effect transistors, systems and methods for manufacturing and using the same
US9859394B2 (en) 2014-12-18 2018-01-02 Agilome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10006910B2 (en) 2014-12-18 2018-06-26 Agilome, Inc. Chemically-sensitive field effect transistors, systems, and methods for manufacturing and using the same
US10020300B2 (en) 2014-12-18 2018-07-10 Agilome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10429342B2 (en) 2014-12-18 2019-10-01 Edico Genome Corporation Chemically-sensitive field effect transistor
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WO2008050356A1 (fr) * 2006-10-27 2008-05-02 Decode Genetics Variants de prédisposition au cancer sur le chromosome 8q24.21
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WO2008050356A1 (fr) * 2006-10-27 2008-05-02 Decode Genetics Variants de prédisposition au cancer sur le chromosome 8q24.21
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Cited By (10)

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US9618474B2 (en) 2014-12-18 2017-04-11 Edico Genome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US9857328B2 (en) 2014-12-18 2018-01-02 Agilome, Inc. Chemically-sensitive field effect transistors, systems and methods for manufacturing and using the same
US9859394B2 (en) 2014-12-18 2018-01-02 Agilome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10006910B2 (en) 2014-12-18 2018-06-26 Agilome, Inc. Chemically-sensitive field effect transistors, systems, and methods for manufacturing and using the same
US10020300B2 (en) 2014-12-18 2018-07-10 Agilome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10429381B2 (en) 2014-12-18 2019-10-01 Agilome, Inc. Chemically-sensitive field effect transistors, systems, and methods for manufacturing and using the same
US10429342B2 (en) 2014-12-18 2019-10-01 Edico Genome Corporation Chemically-sensitive field effect transistor
US10494670B2 (en) 2014-12-18 2019-12-03 Agilome, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10607989B2 (en) 2014-12-18 2020-03-31 Nanomedical Diagnostics, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids
US10811539B2 (en) 2016-05-16 2020-10-20 Nanomedical Diagnostics, Inc. Graphene FET devices, systems, and methods of using the same for sequencing nucleic acids

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