1. Saint Louis University School of Nursing 14 th Annual Advanced Practice Nursing Conference and Workshops February 17 – 18, 2011 Mary Ann Lavin, ScD, APRN, ANP-BC, FAAN Introduction to Where Genetics Meets Pharmacology

2. Basic Text and Online References Wecker, L. (2010). Brody’s Human Pharmacology: Molecular to Clinical. St. Louis: Elsevier/Mosby:49-56 Access online references within context of slide presentation 2

3. Objectives At the conclusion of this presentation, participants will be able to: Better integrate six basic concepts of gene therapy into their repertoire of pharmacologic language and practice Discuss genetic testing, especially, warfarin resistance, clopidrogel resistance. Gains comfort with CLIA standards for molecular laboratory testing Access online continuing education and self-learning applications 3

4. Basic concepts of gene therapy Concept #1: Gene therapy requirements Concept #2: Gene addition therapies Concept #3: Gene repair (editing) therapies Concept #4: DNA and RNA targeting and delivery methods Concept #5. Measuring Gene Expression, i.e., pharmacogentic outcomes (therapeutic effectiveness) Concept #6: Vectors for gene transfer 4

5. Concept #1: Gene therapy requirements Disease candidates An identified mutation Known molecular/cellular pathophysiology Available gene expression tools 5

6. Examples of diseases being studied 6 Bookshelf from NLM (NCBI at NLM at NIH): http://www.ncbi.nlm.nih.gov/books/NBK22183/ http://www.ncbi.nlm.nih.gov/books/NBK22183/ Select neonatal Select Marfan’s syndrome Find genetic site of mutation: from NLM website Find tests for mutation: from Cleveland Clinic website http://my.clevelandclinic.org/heart/disorder s/aorta_marfan/marfan.aspx http://my.clevelandclinic.org/heart/disorder s/aorta_marfan/marfan.aspx Current treatment? Not gene therapy, but BBs, ACEIs, ARBs

7. 7 Select Fragile X syndrome Find background info: http://www.ghr.nlm.nih.gov/conditio n/fragile-x-syndrome http://www.ghr.nlm.nih.gov/conditio n/fragile-x-syndrome Scroll down and click on “gene tests” But, is gene therapy available? Access: http://clinicaltrials.gov/search/condit ion=%22fragile+x+syndrome%22 http://clinicaltrials.gov/search/condit ion=%22fragile+x+syndrome%22

8. Basic concept #2: Gene addition therapies Exogenous DNA is ADDED to complement mutant DNA, Example: Adding genes that produce ApoE to treat hyperlipidemia and atherosclerosis. 8

9. Abstract Curr Opin Mol Ther. 2006 Aug;8(4):275-87. ApoE gene therapy to treat hyperlipidemia and atherosclerosis. Harris JD, Evans V, Owen JS. Harris JDEvans VOwen JS Royal Free & University College Medical School, The UCL Institute of Hepatology, Hampstead Campus, Upper 3rd Floor, Rowland Hill Street, London NW3 2PF, UK. j.harris@medsch.ucl.ac.uk 9

10. Abstract (con’t) Atherosclerosis is the leading cause of death in industrialized countries and is becoming an increasingly worldwide risk to health. Apolipoprotein E (ApoE) is a blood circulating protein with pleiotropic (many effects) atheroprotective properties that has emerged as a strong candidate for treating hypercholesterolemia and cardiovascular disease. In this review, authors discuss the major developments in both viral and non-viral vectors aimed at achieving efficient delivery and sustained expression of an ApoE transgene. 10

11. Gene addition 11 Add genetic material that produces ApoE

12. Note: Before we go on…. 12 To deliver DNA to target, vectors or vehicles are use (think of cars for transportation purposes) Viral vectors Non-viral vectors

13. Abstract (con’t): Gene addition, using viral vectors to deliver DNA The technological advances in engineering viruses allow pharmacogeneticists to Generate different serotypes (surface antigens) of recombinant adeno-associated virus Minimize immune responses Package the requisite DNA (ApoE genetic material) 13

14. Abstract (con’t): Gene addition, using non-viral vectors to deliver DNA Non-viral ApoE delivery systems, including plasmids and cell-based therapy for gene addition purposes 14

15. Abstract (con’t): Targeted gene editing, using oligonucelotides as vectors 15 Finally, a radical alternative to gene addition that has the potential for permanent cure in many genetic diseases--'targeted gene editing'--is reviewed. This technology uses synthetic oligonucleotides to correct underlying point mutations in situ and has been evaluated for repairing dysfunctional ApoE genes.

16. Basic concept #3: Gene REPAIR therapies and background Technically feasible and more definitive therapy, but Technique usually rely on homologous recombination Naturally occurring homologous recombination occurring during miosis in the cell cycle. Homologous recombination repairs cells naturally. Deficiency in homologous recombination leads to failure to repair gene and may lead to some forms of cancer. 16

17. Gene repair therapies Methods Directly correct mutant gene by homologous recombination methods (currently, only very inefficient methods are available) Protein engineering uses homologous (or heterologous) recombinations, which, for example, are capable of producing enzymes, e.g., chimeric P450 isoenzymes from parent P450 isoenzymes. 17

18. Gene repair therapies (con’t) 18 Term “chimera”: http://www.theoi.com/Ther/Khimaira.html http://www.theoi.com/Ther/Khimaira.html Chimera in genetics: Example: When a an organism, eg., a human, has two sources of DNA as when fraternal twins share a placenta and the blood from the two mix. The blood of the infants will then be chimeric. It will reflect blood cells with two different DNA patterns

19. Gene repair therapies: recombinant technologies used to engineer modified P450 Appl Environ Microbiol. 2007 Nov;73(22):7246-51. Epub 2007 Sep 28. Engineering of artificial plant cytochrome P450 enzymes for synthesis of isoflavones by Escherichia coli. Leonard E, Koffas MA. Leonard EKoffas MA Department of Chemical and Biological Engineering, The State University of New York, Buffalo, NY 14260, USA. 19

20. Diversion from abstract to review background on isoflavones from NCI Remember prodrugs and how prodrugs produce active metabolites. The P450 system metabolizes isoflavones per CYP1A2, primarily. NCI study using human liver isoenzymes (Hu, Krausz, Chen et al., 2003) Genistein ( a soybean isoflavone) → CYP1A2 → 3'-OH- genistein (metabolite of genistein) Methylated isoflavones biochanin A, prunetin, and formononetin, which are less active forms of genstein → CYP1A2 → active genistein and active daidzein (another soy isoflavone. So genistein and methylated isoflavones are prodrugs 20

21. Back to the work of Leonard & Koffas (2007), using bacterial and plant P450 isoenzymes 21 Bacterial P450 enzymes have industrial applications, insofar as bacteria produce rapidly, so their P450 enzyme metabolites can be produced at a rate proportional to bacterial reproduction rates. Plant P450 enzymes synthesize or yield many active pharm agents and other chemicals, in nature. But, plant P450 enzymes are difficult to isolate and their yield is low and when manipulated (repaired) lose enzymatic activity. So, their industrial use is low. But, wonder if we can repair/manipulate plant P450 enzymes to produce an agent, e.g., estrogen, within a rapidly growing bacteria, e.g., E. coli. But, plant P450 enzymes do not function within E. coli, so a “heterologous” recombinant approach was used.

22. Summary of work of Leonard & Koffas (2007) – con’t So, the genetic laboratory methodology was refined, resulting in a chimeric (two separate sources, bacterial and plant) P450 that metabolized soy isoflavones in E. Coli, yielding estrogen Better than the best of the previously synthesized product developed by a wild-type P450 enzyme (found in nature) expressed in yeast 22

23. Summary of work of Leonard & Koffas (2007) – con’t 23 Compared well with the production achieved by P450 enzymes in soy plants themselves, plants specially engineered or manipulated to increase isofavone yield/seed. But, lots of bacteria can produce more isoflavones more rapidly rather than waiting for slow growing plant. They manipulated genes of the P450 isoenzyme in E. Coli to produces more estrogen more rapidly than any other source.

24. Basic concept #4: DNA and RNA targeting and delivery methods Delivery Ex vivo In vivo Targeting Receptor-mediated targeting Antibody-mediated targeting 24

25. Ex vivo delivery Cells capable of removal i.e., capable of surviving ex vivo oTreated Reimplanted SQ or IM OR, alternatively Use patient’s own skin, muscle or bone marrow “transfected” with selected DNA, e.g., Ex vivo insulin production Ex vivo cytokine (IL-4) production Then cell implant is delivered SQ 25

26. In vivo delivery of Selected DNA or RNA delivered per Catheterization of organ Surgical methods, Fiberoptic guided method of gene therapy to brain, liver, bone marrow, etc. 26

27. Targeting Receptor targeting Antibody targeting 27

28. Receptor targeting for gene transfer/gene expression purposes Receptors are proteins located on the surface of the cell DNA therapeutic substance is attached to a ligand (an ion or molecule that binds to receptor) Receptor-mediated endocytosis occurs. 28

29. Antibody-mediated targeting for gene transfer/gene expression purposes Antigens are proteins located on the surface of the cell DNA therapeutic substance is attached to an antibody or antibody fragment that binds to an antigen (see abstract) 29

30. Basic Concept #5. Measuring Gene Expression, i.e., pharmacogentic outcomes (therapeutic effectiveness) Quantify magnitude and duration of gene expression to determine the following outcomes Amount of DNA reaching target cell Amount of RNA and protein production Functionality of the protein produced (does it work?) 30

31. Basic Concept #5. Measuring Gene Expression, i.e., pharmacogentic outcomes (therapeutic effectiveness) 31 Factors that determine magnitude of gene expression Protein half life Therapeutic goal (Did the genes expressed attain desired outcomes?) Gene transfer efficacy Strength or potency of the gene promoter

32. Basic Concept #5. Measuring Gene Expression, i.e., pharmacogentic outcomes (therapeutic effectiveness) 32 Factors that determine duration of gene expression Function of the type of vector or vehicle used to carry the DNA or RNA

33. A primitive example of the magnitude and duration of gene expression in a mouse model: Imagine… 33 Purpose of therapy gene: To inhibit mitosis in cancer cells. How is therapeutic effectiveness of therapy gene determined? By monitoring gene expression, i.e., to measure the therapeutic effectiveness of the therapy gene in decreasing mitosis.

34. What are the steps in measuring the effectiveness (magnitude and duration of gene expression) of a therapeutic gene? One primitive example in mouse model (con’t) 34 Step 1. Use a microPET (a microPET scan) reporter gene – a radioactive gene that links up with the gene sequence that is causing increased mitosis within cancer cells and count the magnitude of the positrons being expressed, which is a function of the gene expression before therapy, e.g., number of cell divisions/unit time.

35. What are the steps in measuring the effectiveness (magnitude and duration of gene expression) of a therapeutic gene? One primitive example in mouse model (con’t) 35 Step 2. Introduce therapy gene, i.e., the genetic material delivered per a vector that will turn off the cells responsible for increased mitosis

36. What are the steps in measuring the effectiveness (magnitude and duration of gene expression) of a therapeutic gene? One primitive example in mouse model (con’t) 36 Step 3. Now again use the PET reporter gene to monitor the expression of the gene responsible for mitosis after the introduction of the therapy gene. Did the number of cell divisions/unit time decrease? What is the rate of decrease? How long (duration) did the decrease last?

37. Saved PubMed searches on gene addition and gene repair Go to www.pubmed.govwww.pubmed.gov Click on MyNCBI hyperlink in upper right hand corner ID: SLUSONgenepharm Password: search Click on whichever search you like. 37

38. Concept #6: Vectors for gene transfer Vectors Viral Non-viral 38

39. Viral Vectors or Vehicles used to transfer DNA or RNA Retrovirus packaging DNA or RNA for transfer Easy to produce Efficient DNA transfer Stable gene expression over time Adverse effects: low or limited immune response Use: Integrate therapeutic gene into host genome while cell is dividing Type of therapy: Ex vivo Problem: for a retrovirus to be introduced within a cell, it requires the enzyme integrase. Integrase does not discriminate well among the genes as to the insertion site. If it inserts at an oncogene site, it may stimulate the release of genes that yield disinhibited growth, leading to diseases, e.g., leukemia. 39

40. Viral Vectors or Vehicles used to transfer DNA or RNA Adenoviruses High volume production, therefore, good for in vivo transfer Efficient delivering to non-dividing cells Stable gene expression over time Adverse effects: Concomittant viral infection can trigger immune response, precluding further therapy. Uses: Used for gene transfer with viral infection mediated by a cell receptor and entry into the nucleus but not integrated into the genome itself. Therefore, repeat dosing at monthly intervals is needed. Therapy: In vivo 40

41. Viral Vectors or Vehicles used to transfer DNA or RNA Adeno-associated, herpes-associated, and lenti-associated viruses Adeno-associated viruses integrate DNA into genome of the target cells that are not dividing. Herpes simplex-associated viruses enhance delivery of genes to neurons. Lentiviruses are slowly replicating retroviruses that can infect, unlike other retroviruses, only non-replicating or non-dividing cells. They have the capability of delivering large amounts of genetic material and thus are one of the most efficient vectors. 41

42. Plasmid-based non-viral vectors or vehicles used to transfer DNA or RNA Naked DNA plasmids Liposome DNA 42

43. Plasmid-based non-viral vectors or vehicles used to transfer DNA or RNA: Naked DNA plasmids Plasmid – A DNA container or a circular-like piece of DNA, used as a vehicle to transfer therapeutic DNA or RNA Administration: Gene gun, because on their own, plasmids enter cells very inefficiently or syringe for IM injection. Example: Vaccine gene gun to deliver DNA vaccines to promote or trigger immune response. Still experimental. http://www.scientificamerican.com/media/inline/00 0052A5-43F6-1CBC-B4A8809EC588EEDF_arch2.gif http://www.scientificamerican.com/media/inline/00 0052A5-43F6-1CBC-B4A8809EC588EEDF_arch2.gif 43

44. Plasmid-based non-viral vectors or vehicles used to transfer DNA or RNA: DNA/RNA phospholipid (liposome) plasmids Liposomes – aggregate of fat (lipo) molecules in the body (soma) to transfer DNA Antisense (complementary strands of RNA or DNA) oligonucleotides. Access: http://en.wikipedia.org/wiki/Oligonucleotide http://en.wikipedia.org/wiki/Oligonucleotide May be used to turn off cancer genes. siRNA (short-interfering RNA) to facilitate entry into the cell End result is a nucleic acid (DNA or RNA)-lipid complex Which enters cell by fusion or by endocytosis 44

45. Gene therapies and/or research applications today Gene therapy in oncology Addition of wild type tumor suppressor gene to complement a mutant tumor suppressor gene to slow cancer growth (may not cause tumor size regression) Antisense RNA therapies to turn off expression of cancer gene Transfer of a gene to enhance expression of an immunomodulator gene or of a cytokine (IL) gene Transfer of gene coding for a prodrug which produces cytotoxic metabolite Inhibition of tumor angiogenesis Transfer of chemoprotective genes to decrease chemotoxicity or genes to decrease radioresistance to increase radiosensitivity of cells. 45

46. Gene therapies and/or research applications today Infections disease Prevention: Vaccine gun discussed http://www.scientificamerican.com/ media/inline/000052A5-43F6- 1CBC- B4A8809EC588EEDF_arch2.gif http://www.scientificamerican.com/ media/inline/000052A5-43F6- 1CBC- B4A8809EC588EEDF_arch2.gif 46

47. Gene therapies and/or research applications today 47 Fischer, A., Hacein-Bey-Abina, S. & Cavazzana-Calvo, M. (2010). 20 years of gene therapy for SCID Nature Immunology,11: 457–460 http://www.nature.com/ni/journal /v11/n6/full/ni0610-457.html http://www.nature.com/ni/journal /v11/n6/full/ni0610-457.html

48. Gene therapies and/or research applications today 48 New gene therapy makes T-cells resistant to HIV, completely blocking proliferations in culture: http://www.liebertpub.com/contentframe.as px?code=MDoqsls2tWBQ0c%2bZWT0nPtZ 1zyJ60HXWr8kmqZBHbUfkxIBDwf3RDtPb RDToWe2xI0rLqyT2GBmC5ra%2f9NOebnro VhFTf4S%2fZOpVpXsV2OI%3d http://www.liebertpub.com/contentframe.as px?code=MDoqsls2tWBQ0c%2bZWT0nPtZ 1zyJ60HXWr8kmqZBHbUfkxIBDwf3RDtPb RDToWe2xI0rLqyT2GBmC5ra%2f9NOebnro VhFTf4S%2fZOpVpXsV2OI%3d

49. Pharmacology related laboratory tests and CLIA requirements Antiretroviral (ARV) resistance testing Warfarin resistance Clopidrogel resistance 49

50. Antiretroviral resistance (ARV) Addition of an antiviral drug fails to halt growth/multiplication or fails to kill microbe. Antiviral drug resistance results from a genetic change or mutation that occurs in the virus after being introduced to a drug. Viruses that mutate survive; viruses that don’t mutate die. 50

51. Antiretroviral resistance (ARV) 51 Surviving, mutant (now resistant) viruses are passed on to other people. Antiviral drugs are ineffective in either in first patient and in those to whom the virus is passed, because both now possess the mutant and resistant virus Rx: Place patient on drugs that remain sensitive to virus or treat patient from drugs from two categories known to be highly active (remember HAART), i.e., drugs from two different classes that attack same microbe.

52. HIV genotypic resistance testing AKA: Antiretroviral drug resistance testing, ARV resistance testing Formal name: Human immunodeficiency virus genotype resistance testing Related tests HIV viral load CD4 and CD8 counts HIV antibody titers HIV phenotypic resistance testing 52

53. HIV genotypic resistance testing Blood sample Need viral load of at least 1,000 copies/ml If viral load is very low, resistance to all AVRs may not be detected. Consider testing when viral load reaches 500 – 1000 copies/ml Source: www.labtestsonline.org 53

54. Warfarin resistance Warfarin: narrow therapeutic range 20 fold inter-patient therapeutic range Dosing is difficult Dose too high yields high INR yields increased risk for bleeding Dose too low yields low INR yields increased risk for thromboembolism Death from warfarin related hemorrhage is a lead cause of drug related mortality in Western world. 54

55. Warfarin background Warfarin uses: Antithrombotic (e.g., AF, valve prostheses, DVT/PE prophylaxis) External factors that affect warfarin dosing Weight BMI Age Diet Medications 55

56. Warfarin MOA 56 Interrupts the VKOR complex (Vitamin K oxidase reductase complex) yielding less clotting But, there is a genetic subunit to VKOR It is called VKORC1. Why is the subunit C1 important? A polymorphism in the E element -1639 of the base of VKORC1 explains 44% in the variance of dosing requirements 1639G allele is less active than 1639A allele. Chinese require a much lower dose of warfarin than Caucasians. Therefore, the 1639A allele is responsible for increased sensitivity to warfarin, requiring a lower dose. One more time….

57. Vitamin K cycle (http://www.mayomedicallaboratories.com/articles/c ommunique/2008/08.html)http://www.mayomedicallaboratories.com/articles/c ommunique/2008/08.html 57 Vit K → reductase → Vit K epoxide → reductase → Vit K Warfarin inhibits reductase, interrupting Vitamin K cycles, and thereby decreasing or inhibiting production of Vitamin K But, there are polymorphisms/mutations in the 1639 gene that controls Vitamin K metabolism 1639 A allele decreases or inhibits metabolism of Vitamin K epoxide to Vitamin K, yielding less Vitamin K and increased tendency to bleeding. Such persons are hypersensitive to warfarin and will need less warfarin, 1639G allele is less active. It doesn’t inhibit metabolism of Vitamin K epoxide to Vitamin K. Such persons require a more normal dose of warfarin.

58. Distribution of 1639 alleles among Caucasians and Chinese, with inputs for Sconce algorithm A/A (VKORC1 Input “3”) A/G (VKORC1 Input “2”) G/G (VKORC1 Input “1) Caucasians14%47%39% Chinese82% 58

59. But, not finished yet…. It is not just the Chinese that are highly sensitive to warfarin, we do have Caucasians as well who require very small doses to achieve a therapeutic effect. What accounts for the genetic variation among Caucasians? 59

60. Warfarin and the P450 genetic connection Metabolized by CYP2C9 CYP2C9*1 is the normal or wild type allele CYP2C9*2 decreases P450 activity yielding increased plasma concentration, yielding need for decreased dose. This is called increased sensitivity to warfarin. CYP2C9*3 decreases P450 activity, yielding increased plasma concentration, yielding need for decreased dose. This is called increased sensitivity to warfarin. CYP2C9*1/*2 and CYP2C9*1/*3 occur in the Caucasian population 20% and 12%, respectively. Usually CYP2C9*2/*3 are completely absent in the Chinese population. 60

61. Who requires the lower dose of warfarin? AgeHeig ht CYP 2C9*1 Input 0 CYP 2C9*2 Input 1 (inhi- bits P450) CYP 2C9*3 Input 2 (inhi- bits P450) VKO R AA Input 3 VKO R GA Input 2 VKO R GG Input 1 Patient 1 90 years 170 cm (5’8”) Yes Patient 2 30 years 170 cm (5’8”) Yes 61

62. Answer: Patient 1 will require a much lesser (six times lesser by algorithm) than Patient 2. Why? 62

63. Sconce Reference Sconce, EA, Khan, Ti, Wynne, H.A., et al. (2005). Impact of CYP2C9 and VKORC1 genetic polymorphism and patient characteristics upon warfarin dose requirements: proposal for a new dosing regimen. Blood, 106(7):2329-2333. For an update on work by Sconce, access MyNCBI ID: SLUSONgenepharm Password: search 63

64. Latest: University of Utah, McMillan et al., 2010 64 229 adults having orthopedic surgery Assigned to: Gene-based warfarin algorithm (Sconce, 2005 alternating with Pendleton, 2008), post determination of CYP2C9 and VKORC1 variants Standard care supervised by dedicated warfarin team, with only change being going slower on dose increases when variants were present. Results When variants were present, gene based algorithms led to earlier attainment of INR goal with less adverse events. When no variants were present, gene-based algorithms underpredicted dosing. In other words, standard care methods were superior.

65. The future and warfarin dosing? Note: Research is underway to include in the algorithm The multidrug resistance gene ApoE genotype Drugs that interact with warfarin Foods that interact with warfarin 65

66. Clopidrogel resistance Clopidrogel metabolizes in the P450 system to an active metabolite But, esterases can capture the prodrug and inactivate it. That accounts for about 85% of the prodrug produced. But, the remaining 15% gets metabolized by the P450 system to the active metabolite that inhibits platelet production. 66

67. Clopidrogel resistance (con’t) 67 The isoenzyme is CYP 2C19. But, it has one genotype (*2) that inactivates CYP2C19. If 2C19 *2 is inactivated, then clopidrogel is not metabolized to its active metabolite and platelets are not inhibited and patient is at increased risk of death, MI, CVA, etc. VerifyNow (Accumetrics, San Diego): Point of care measure of response to clopidrogel

68. Population Frequency of Cytochrome P450 (CYP) 2C19 Metabolizer Types http://www.healthanddna.com/drug-safety-dna- testing/clopidogrel.html 68 Poor metabolizers No isoenzyme No active metabolite production Reduced metabolizers Reduced isoenzyme Reduced active metatolite production Extensive metabolizers Normal isoenzyme production CYP2C193-21%24-36%43-73%

69. Treatment options and issues 69 Increase clopidrogel dose: Issue increased risk of bleeding, joint pain. Increase clopidrogel per algorithm allows for successful treatment of 80% - 85% of patients per a German trial (Neubauer, et al, 2011). Prescribe IIb/IIIA inhibitors, e.g., abciximab, tirofiban Prescribe a thienopyridine class inhibitor of platelet activation and aggregation (mediated by the P2Y12 ADP receptor), e.g., Prasugrel. Issue: Increased risk of bleeding. Too, there may be P2y12 ADP polymorphisms that will affect response and subsequent dosing. Ticlopidine (Ticlid), but it is metabolized by CYP2C19 as well and converted to an active metabolite. Its polymorphisms? Also, omeprazole issue – same with clopidrogel. Omeprazole inhibits P450 system, yielding less active metabolite.

70. CLIA Clinical Laboratory Improvement Amendment (1988) 70

71. CLIA’s good practices for molecular genetics test for inheritable conditions Link: http://www.cdc.gov/mmwr/pdf/rr/rr5806.pdfhttp://www.cdc.gov/mmwr/pdf/rr/rr5806.pdf Introduction DNA based diagnosis often crucial, plus… Newborn screening tests, plus… Pharmacogenetic and pharmacogenomic tests for SNPs, single nucleotide polymorphisms’ Haplotype markers Alterations in gene expression No genetic test is waived or of moderate complexity 71

72. Purpose of genetic testing Detect/confirm rare genetic diseases Detect mutations or genetic variations associated with Cancer Coagulation disorders CV diseases Diabetes Pharmacogenetics/pharmacokinetics 72

73. Laboratory test issues Validity (sensitivity and specificity) Benefits (the “so what” question), i.e., value of information versus cost Utility (or usefulness) Marfan’s If tested, so what? Aortic aneurysm and aortic valve problem is Marfan’s? Surgery? Is not Marfans? 73

74. Preanalytic phase: Labs provide info to clients (practices, clinical settings) regarding Molecular genetic tests available and information for selecting appropriate tests Intended use Indications Test method description in user friendly language Info on collection, handling, transport and specimen submission Patient information needed, if applications racial/ethnic info, family history, pedigree, and consent information meeting federal, state, and local requirements A statement indicating implication of results for the patient family. Availability of lab consultants for info on test selection/ordering, specimen submission, results interpretation. 74

75. Provider obtains informed consent from the patient for the genetic test, when indicated Not all molecular genetic tests for inheritable conditions require informed consent Provider obtains federal, state, local, institutions regs/policies Lab can help provide useful information, but the informed consent is responsibility of the patient care provider Document in medical record, lab requisition 75

76. Info needed on lab test requests Patient name/unique identifiers Patient DOB Indication for testing and relevant clinical or lab data Racial/ethnic data, if applicable Info on patient family history, pedigree (tree) or both that is disease pertinent ICD diagnosis(es) Checkbox indicated appropriate level of informed consent obtained. 76

77. Lab provides specimen handling info to clients (practices, clinical settings) Appropriate type and amount of specimen Collection container or devices Special timing of specimen being collected Special preparation and handling of specimen Specimen stability/integrity info Specimen transport Reasons for specimen rejection 77

78. Criteria for specimen rejection Improper handling/transport Exposure to temperatures that degrade stability/integrity of specimen Insufficient specimen volume or amount Use of inappropriate anticoagulants, media, degraded specimens, or inappropriate specimen types Comingled or contaminated specimens Mislabeled or insufficiently labeled specimens Mislabeled or insufficiently labeled lab request forms Lack of MD/NP order info to determine if test ordered and specimen submitted are appropriate to answer clinical question asked. 78

79. CLIA regs: Analytic phase in-lab 79

80. CLIA regs: Post analytic phase regarding reports Patient name/identifiers DOB Indication(s) for testing Date of specimen collection and arrival in lab Referring MD/NP Test method, including the nucleic acid targets of the test, e.g., targeted mutation detection or DNA sequence analysis Test results in “current recommended standard nomenclature” Interpretation of test results References to literature, if applicable Recommendations for genetic consultation, when appropriate 80

81. CLIA regs: Post analytic phase regarding reports Implications of test results for relatives/family members, who might benefit from results Statement indicating that results are based on current information and technology. Report retention on molecular genetic tests: 25 years by lab 81

82. De-identified lab results from Quest Clopidrogel 82

83. Handouts Pretest and posttest Slides Glossary Samples of lab results, available for reading here, upon request 83

84. Online CEUs 84 http://nursingworld.org/mods/mod484/gen eticsfull.htm http://nursingworld.org/mods/mod484/gen eticsfull.htm http://www.kumc.edu/gec/prof/genecour. html http://www.kumc.edu/gec/prof/genecour. html http://www.cdc.gov/mmwr/pdf/rr/rr5806. pdf http://www.cdc.gov/mmwr/pdf/rr/rr5806. pdf http://www.ornl.gov/sci/techresources/Hu man_Genome/medicine/medicine.shtml http://www.ornl.gov/sci/techresources/Hu man_Genome/medicine/medicine.shtml http://www.who.int/genomics/professional s/education/en/index.html http://www.who.int/genomics/professional s/education/en/index.html

85. 85