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Disclosures Personalized (Precision) I am an employed by Promega Corporation, a Madison Medicine based international biotechnology company Promega does not market any of the products mentioned in Wisconsin Association of Physician Assistants this presentation, although we do provide components to some of the manufacturers of these diagnostic assays October 10, 2014

Ashley G. Anderson Jr., MD, MS Chief Medical Officer Promega Corporation

Objectives Promega Corporation

Look into the future, and appreciate the growing role, and limitless potential, of molecular diagnostics

Understand the term “personalized medicine”

Be motivated to read, learn, and become an early adopter of precision diagnostics

Appreciate that these bill be big changes, but they may not come fast.

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Personalized medicine Precision medicine

“The molecular methods that make personalized medicine The tests themselves are not necessarily “personalized.” possible include testing for variations in genes, gene expression, They are standardized like most other tests proteins and metabolites as well as new treatments that target molecular mechanisms. Test results are correlated with clinical The results from these tests are increasingly more predictive factorsfactors--suchsuch as disease state, prediction of futurefuture disease states, of disease states drug response, and treatment prognosisprognosis--toto help physiciansphysicians Certain tests are more predictive of responses to specific individualize treatment for each patient” therapies Personalized Medicine Coalition The net result is a more precise method of assessing disease and response to therapy

Key developments in medicine

Sterility Antibiotics Eradication of smallpox, and vaccines for polio, diphtheria, rubella, chicken pox, hep A&B Blood banking and transfusion Insulin Medical imaging advances Heart surgery Minimally invasive surgery

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Physical exam /radiographic studies

Blood chemistry Biomarkers

A objectively measurable indicator of a biologic state or condition Physical exam Pulse, respiration, blood pressure Clinical chemistry Radiology finding

Precision medicine is the identification of more sensitive and specific biomarkers.

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A brief review of the genome Basic DNA structure

DNA is composed of 3 billion base pairs, AA--TT or C--GGC

Short sequences of DNA represent genes, each of which may code for one or more proteins

Each gene is located on one of the 23 paired chromosomes

The genetic information on an entire genome can be placed on one DVD

DNA toto RNA toto proteins Genes & alleles

Gene = A stretch of DNA that codes for a polypeptide or RNA chain that has a function in an organism

Gene = “A locatable region of genomic sequence, corresponding to a unit of inheritance, which is associated with regulatory regions or other functional sequence regions”

An “allele” is an area of variation in a gene responsible for variation in some trait.

Everyone has a gene for eye color, but different alleles for the specific color. An allele is a variant of, or within a gene.

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Genes as biomarkers Loci and chromosome rregionsegions

Locus = It is not a gene, but refers to a map position. “A •• There are 20,00020,000--25,00025,000 protein encoding genes locus is a point in the genome, identified by a marker, which can be mapped by some means. It could, for example, be an •• But it’s not all about genetics. anonymous nonnon--codingcoding DNA segment •• EpigeneticsEpigenetics--BaseBase pairs, genes are “turned on/turned off ” by Chromosome region : a genomic region which has been “epigenetic” factors associated with a specific syndrome or phenotype, •• So, is your genetic anatomy your destiny? Only partly. particularly when there is a possibility that several genes within it may be involved in the phonotype

Molecular biomarkers

Biomarker that gives an indication of risk

Diagnostic biomarkersbiomarkers--makemake the diagnosis of a specificspecific disease Drug development and

Prognostic biomarkersbiomarkers--showshow progression of disease therapeutics

Therapeutic predictive biomarkersbiomarkers--showshow the probableprobable effect of a specific treatment on a patient.

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Personalized medicine pparadigmaradigm One dose fits all vs. personalized medicine

Personalized medicine approach Future *Drugs targeted at small Evidence-based patient populations Medicine *Genomic profiles determine patient segmentation and therapy Improved therapeutic Genomic outcomes with fewer adverse Diagnosis events paradigm Current state *Blockbuster drugs targeted at broad patent diagnoses *Approximately 50% or more Today of patients to not have desired ‘Trial and error therapeutic outcomes medicine’ *Significant adverse events Diagnostic Broad Narrow- Targeted Population-based Patient specific Therapeutics

Source: Personalized Medicine©2009 Future Medicine, Ltd

Warfarin sensitivity Molecular diagnostics

Genetic analysis Antithrombotic effects of warfarinwarfarin——narrownarrow therapeutic index

2 Million people are placed on warfarin annually in the US. Some individuals are highly sensitive to warfarin

Risk of adverse drug events (bleeding) approaches 9.5%, mostly in the first month of therapy. Cost approaches $18.5M/10,000 patients/5 years

Many of these individual have polymorphisms in CYP2C9 and/or VKORC1 genetic loci Cancer Infectious disease So…we should be testing for these allelic variants, but few institutions are. Why?

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Sequencing the human genome Next gen sequencing

Began in 19901990--2003.2003. The sequence of last chromosomechromosome was Definition: published in Nature in 2006. Goal was to determine the Developed approximately 5 years ago sequence of chemical base pairs that make up human DNA, Low cost sequencingsequencing--approachingapproaching the $1000 genome and identify and map all the genes Very rapid results at significant cost reductions

Human Genome Project (HGP) = $3 Billion dollar project Ion Torrent Illumina (Solexa) sequencing Celera Corporation, using some data from the HGP, sequenced the genome for $300M with commercial Exome sequencingsequencing--lookinglooking only at the protein encodingencoding genes in aspirations, which caused controversy the genome Select for the subset of genes that encode proteins (180,000), or 1% Very slow, painstaking, expensive of the genome or 30 million base pairs Mutations in these are much more likely to have severe consequences

Next generation ssequencingequencing Next generation ssequencingequencing

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Genome WWideide Association The $1000 dollar genome Study (GWAS)

Collect genomic information

Analyze information for common genetic variants (alleles) in different individuals to see if they are associated with a trait.

Perform “SNP analysis” to determine the SNPs associated with the disease. SNP= sequence variation within population consisting of a single nucleotide (A,T,C or G) different in individuals

First GWA study was performed in 2005, for macular degeneration. Found 2 SNPs with altered allele frequency as contrasted with controls.

Genetic testing

Carrier screeningscreening--screensscreens for one copy of a gene mutationmutation

PrePre--implantationimplantation genetic diagnosis (PGD) Avoids selective pregnancy termination Sex discrimination? Ethical issues?

Forensic testing Crime Paternity

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Prenatal genomic sscreeningcreening (PGS)

Circulating fetal DNA (cfDNA) fragments are harvested from the maternal blood stream

Costs range from $200$200--$2500$2500 (or more)

Multiple companies, notably Sequenom, Illumina False positive rates for detection of trisomies 21 and 18 Illumina’s Verifi test has false positive rate of only 0.03% were significantly less than other methods of screening (0.03%) (Test is very specific) What are the implications of a diagnosis, and how might this impact parent’s decisions Use of cfDNA detected all cases of aneuploidy (100% sensitivity)

DNA Deletions in chromosome 22 associated with DeGeorge Synd

Molecular diagnostics

Genetic analysis

Cancer Infectious disease

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Phenotype to Breast cancer phenotype + genotype Myriad Bracanalysis® assay for BRCA1 and BRCA2 gene mutations

Patients with either mutation have up to an 87% chance of developing breast cancer, and a 44% chance of developing ovarian cancer prior to age 7070

Benefits: Increased surveillance for test positive individuals Enhanced customization Earlier diagnosis and treatment for specific individuals Cost = $4000.00 and tumors

Competitors tried to offer for less resulting in legal action ultimately confirming Myriad’s right to “own” genes

Companion diagnostics Herceptin® (trastuzumab)

Companion diagnostics are assays (a test or Monoclonal antibody that interferes with the Her2/neu measurement) intended to assist physicians in making receptor treatment decisions for their patients. They do so by HercepTest™ elucidating the efficacy and/or safety of a specific drug Approved by FDA in 2010 as companion diagnostic or class of drugs for a targeted patient group or subub-- groups. Identify tumors with over expression of HER2 protein Immunohistochemistry test Types of diagnostics Dako Corporation manufactures test, while Genentech Test developed after drug comes to market manufactures the therapeutic Test developed in conjunction with drug Numerous other companies now have companion diagnostic assays for HER2 (Ventana, LTI, Abbott, Biogenex)

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HER2 Oncotype DX

Tests surgical samples from breast CA patients for 21 genes to predict probability of recurrence.

Genomic HealthHealth--RedwoodRedwood City, CA

Marketing began in Jan 2004

Predicts likelihood of chemotherapy benefit as well as recurrence in invasive breast cancer, and has been shown to predict the likelihood of recurrence in ductal carcinoma in situsitu

Cologuard® test for colon Cologuard® ccancerancer

Madison based molecular diagnostics company How much will it cost? MultiMulti--targettarget test performed on stool samples How much is that cost spread over a population? KRAS mutations Aberrant NDRG4 and BMP3 methylation What will be the net effect on population health? BB--actinactin Hemoglobin immunoassay Does it do a good enough job of detecting adenomas? Does Approved by FDA in August 2014 that matter?

92% sensitivity to detection of colon CA, 42% sensitivity to Should it replace colonoscopy? If not, how should it be detection of prepre--cancerouscancerous polyps. 66% sensitivitysensitivity for polyps greater than 2 cm in size used? 87% specificity (true negative results)

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Erbitux (cetuximab) Biobanks

Indicated for certain head and neck cancers, along with some Repositories for human biological samples types of metastatic colorectal carcinoma In many cases, known therapeutic courses, with known Tumors with KRAS mutation negative gene (wild type), results, can be studied along with molecular characterization along with positive EGFR of diseases

Companion diagnostic: Therascreen® for KRAS, along with Used in genome wide association studies Dako EGFR assay Importantly, specimens can be shared across laboratories and Therascreen detects seven somatic mutations of the human disciplines KRAS oncogene, using DNA extracted from FFPE slides from colon cancer tissues. Discovery of many more SNPs associated with diseases

Retrieving DNA from sslideslides Molecular Diagnostics

Genetic analysis DNA can be recovered from pathology slides that are fixed in formalin & embedded in paraffin (FFPE slides = formalin fixed, paraffin embedded slides.

Implication #1: Tissue can be harvested anywhere, and DNA can be recovered from the sample at a later date.

Implication #2:= There are billions of slides from patients who were treated for various conditions, and in many cases, we know the outcomes! Cancer Infectious disease

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Point of care testing Identifying bacteria The Cephied GeneXpert®

Traditional methods = culture and gram stains Sample to answer “box,” that can be operated with minimal training Subjective analysis, sometimes difficult to grow bacteria Very rapid assessment of MRSA & c. difficile (66 minutes or New methods involve sequencing of bacteria (and viruses) less) for identification purposes

Flu virus from 1918 was sequenced from victim’s tissues

Standard technologies include PCR and microarrays, but mass spectrometry has also been used

Point of care testing

HPV testing FDA Considerations

Over 80% of women are infected with HPV between ages of 1515--25.25. Duration is usually less than 1 year, with spontaneous resolution The primary role of the FDA is to keep our population safe

Over 100 HPV types, but certain types are more pathologic (16,18,45). Vast majority of women with highhigh--riskrisk HPV types do not develop cancer Most molecular tests are laboratory developed tests (LDTs). The FDA does not regulate LDTsLDTs——theythey are regulated under HPV can’t be culturedcultured——therethere is are molecular tests Digene HC2 HighHigh--riskrisk HPV DNA Test (Qiagen) the Clinical Laboratory Improvement Act (CLIA). But Cervista HPV HR Test (Hologic)* (Uses Invader technology developed in Madison at Third Wave) ultimately, the FDA does have authority over them. Cobas 4800 HPV Test (Roche)

Implications for surveillance and treatmenttreatment——phenotypephenotype of cells vs. HPV status. Plenty of The industry is huge compared to the FDA. How can the controversy, but bottom line is that coco--testingtesting cancan result in increased screening intervals.intervals. FDA keep up with the scientific advances in molecular Vaccines for prevention of highhigh--riskrisk HPV medicine? Should they? Development of head and neck cancerscancers-- Increasing incidence of HPV related head and neck carcinoma. Why? Better prognosis

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23 and Me

Summary

Medicine is changing rapidlyrapidly——therethere is an explosion of research in the area of molecular diagnostics and targeted therapeutics

The way we look at diseases will change dramatically over the next decade

The rate of change will be tremendous, and we will need to work hard to keep up with new scientific developments

Hang on tight and enjoy the ride!

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Thank you!

Ashley G. Anderson Jr, MD, MS Chief Medical Officer Promega Corporation 2800 Woods Hollow Road Madison, WI 5371153711--53995399 608608--474474--43304330 [email protected]

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