Source: (cell) http://www.uvm.edu/~inquiryb/webquest/fa06/mvogenbe/ (Tissue) http://en.wikipedia.org/wiki/File:Oral_cancer_%281%29_squamous_cell_carcinoma_histopathology.jpg (Red lungs) http://en.wikipedia.org/wiki/File:Respiratory_system_complete_en.svg Living tissue is made up of cells. There are many different types of cells, but all have the same basic structure. Tissues are layers of similar cells that perform a specific function. The various kinds of tissues group together to form organs.

Inside (AND outside) of cells are molecules --which are composites of two or more elements (atoms). http://www.sfgate.com/cgi-bin/blogs/mbauer/category?blogid=26&cat=622&o=60 http://images.google.com/imgres?imgurl=http://www.sfgate.com/blogs/images/sfgate/mbauer/2006/10/13/waiter_242x350.JPG&imgrefurl=http://www.sfgate.com/cgi-bin/blogs/mbauer/category%3Fblogid%3D26%26cat%3D622%26o%3D60&usg=__UQjmrwJ2jKHFNtW_U_tmuEOP798=&h=350&w=242&sz=18&hl=en&start=16&um=1&itbs=1&tbnid=AlrBmpCPC4VZyM:&tbnh=120&tbnw=83&prev=/images%3Fq%3Dwaiter%26um%3D1%26hl%3Den%26tbs%3Disch:1 http://www.brooklyn.cuny.edu/bc/ahp/SDgraphics/PSgraphics/SD.PS.LG.Water.html

Two or more atoms put together. Everything is composed of atoms.

What you’re doing when you only look at DNA is “SEQUENCING” – which only looks at the molecules composing the DNA – not methylation, RNA, proteins, etcetera!

Source: http://www.noikhomes.com/content/design.html http://www.keima.com/ http://www.you-are-here.com/location/the_grove.html (The Grove)

End-conformation/expression is like a construction worker’s end result (i.e., installing a bathroom), not the master plan (akin to the design for the bathroom). An error in the DNA that doesn’t result in a change in expression or conformation/activity is like a mistake in the MP that is meaningless. Fixing a problem that is in the Master Plan (i.e., the size for the tub is doesn’t work) in his efforts (i.e., buying a different sized tub), and not fixing the mistake in the plan, is analogous to giving someone a monoclonal Antibody or siRNA. Fixing a problem that is in the MP, in the master plan, is like repairing the DNA when an error occurs.

Source: http://www.chemistry.wustl.edu/~edudev/LabTutorials/Vision/Vision.html

Source: http://www.chemistry.wustl.edu/~edudev/LabTutorials/Vision/Vision.html

“Everything psychological is biological.” “Technology is not only changing our lives, it’s changing our BRAINS.” – Dr Gary Small, UCLA - Director of the UCLA Center on Aging & Professor of Psychiatry and Behavioral Sciences

Bonnie Addario Lung Cancer Medical Institute http://www.alcmi.net/ Watch 6.50-8 in the video

Vs ALTERNATE: Expression levels and the sequence of molecules—which constantly are changing—are key to understanding the underlying molecular mechanisms describing our actions, at any given moment in time.

“prohibits group health plans and health insurers from denying coverage to a healthy individual or charging that person higher premiums based solely on a genetic predisposition to developing a disease in the future.” “The legislation also bars employers from using individuals' genetic information when making hiring, firing, job placement, or promotion decisions.” Wiki, “Genetic Information Nondiscrimination Act” http://www.ama-assn.org/ama/pub/physician-resources/medical-science/genetics-molecular-medicine/news.shtml

http://www.youtube.com/watch?v=aKBZbxBnpGM&feature=related http://www.youtube.com/watch?v=xbJ0nbzt5Kw Courtesy of GeneGo http://www.genego.com/map_373.php

It is important to remember: everything you see in the microscope and feel in your body is a result of molecularly driven processes. Happiness is associated with serotonin secretion, cancer metastasis is associated with various proteins and their interactions with surrounding cells. Source: http://en.wikipedia.org/wiki/File:Optical_microscope_nikon_alphaphot_%2B.jpg; http://en.wikipedia.org/wiki/File:Structural_MRI_animation.ogv

Giving you something with a known molecular mechanism of action, and giving it to you not knowing if it will work or not. It is a hypothesis and guess! And quite often, this guestimate is also based on the ideology that everyone’s differences are random and yield no predictive value on an Rx’s effectiveness or safety.

As illustrated in previous slides, it’s important to know particular molecular expression levels of your cells to effectively treat illnesses and minimize side effects. What I did not mention was the Window of Opportunity this brings us. This Window is open at the point where we undergo these tests. At this moment, we know what molecular expression our cells have and can treat based off this knowledge. However, this window doesn’t stay open forever as I will illustrate in the next slide.

Genes (our DNA) code for proteins, RNA strands, and enzymes – which in turn act as the workhorses of our body.

Cancer refers to the uncontrollable growth of cells. This arises from an over-expression of what are called oncogenes, and an under-expression of what are called tumor suppressors. You can think of oncogenes as a GREEN light for cells to grow (their definition is any protein/enzyme that, when normally expressed, promotes cell growth), and tumor suppressors as a RED light (they are any enzyme/protein that, when normally expressed, suppresses or stops cell growth). There are many, many oncogenes and tumor suppressors in the cell, very often working at the same time. “EGFR” is a growth receptor / oncoprotein (remember genes code for proteins) that is often over-expressed in tumors. There is a therapy known as Tarceva that inhibits EGFR proteins from functioning by binding to that protein’s energy source – it’s ATP binding site. However, an over-expression of “MET” – another oncoprotein, is associated with RESISTANCE to Tarceva. This means that if you have an over-expression of MET and EGFR, then Tarceva will likely NOT be beneficial to you (even if you have an overexpression of EGFR). http://awurl.com/7urUWKWlv This dynamic concept illustrates why it is important to know the molecular profile of the diseased cells in the body to see if they express MET and EGFR or if they just over-express EGFR when deciding treatment options. Another Example: Xeloda + TP overexpression = good, but Xeloda + thymidylate synthase overexpression = BAD! http://www.biomedcentral.com/1471-2407/8/386; http://awurl.com/xODVcukKl

FUS1, MDM2, P53 Therapy: This is going to get very tough to follow so be careful. P53 is a “stopper” of cell growth. It is a very often knocked out (inactive or missing) in tumor cells. MDM2 is an inhibitory protein of p53 (so it’s a promoter of cell growth – an oncoprotein). FUS1 is an inhibitor of MDM2, which inhibits p53, thus FUS1 is a stopper of cell growth (tumor suppressor). Application Let’s say, you took a sample of someone’s tumor and it showed an overexpression of MDM2; however, you didn’t test for other mutations/expression levels. You then gave that person MDM2 antibodies / inactivators as a treatment. The problem here with this is, if this tumor cell also has a mutated, inactive p53 gene, then blocking the action of this inhibitory protein makes no difference because p53 will still be inactive. This illustrates the need to test for various proteins in these known signaling pathways to realize the most effective benefits of a therapy. It is important to note here that every protein has another promoter/inhibitor molecule regulating it. Each of these regulating proteins are coded by a cell’s DNA. The signaling pathways within cells has taken decades to understand and is still far from fully being understood. However, there are known signaling pathways that we can test patients for particular discrepancies in – enhancing the effectiveness and safety of therapeutic effects on patients. As well, if a database were to be collected and everyone were to undergo this “sequencing” and testing of proteins/genes, then we could isolate the causes of rare side effects.

Not just evident in cancer, but also in antibiotics, ALS, and every disease/therapy known. Every action is mediated by molecules driving it.

Statistics = A+B+x= Y, where A is disease histology; B is patient medical history; X is the therapy; and Y is likely outcome. A+B+c+d+e+f+g+x =Y is new equation.

Need to not only find different molecular patterns, but you have to be able to know the signaling patterns to properly predict and devise a therapeutic regiment!! For instance, an inhibitor to an oncogene may be under-expressed.. Hence attacking the over-expressed oncogene wont matter.

A person and society’s Window of Opportunity can be lost if a therapy becomes resistant by mutating or upregulating/downregulating various other proteins => Resistance to treatment at the personal level occurs because an organism adapts to these treatments by turning up the expression of certain proteins and turning down the expression of others. In doing this, they can adapt to therapies. Diseases and infections can also adapt when these disease cells have unstable genomes, hence the cells with DNA that cannot survive die and ones that can survive. This is why cancer and HIV are very hard to eliminate from the body. With this in mind, it is important to know the molecular profile of your target illness before you prescribe therapies and after to see these changes and effectively treat these ailments. As resistance grows throughout the population, when tumors or bacterial infections / viruses do adapt, the therapies that took decades to discover and test in clinical trial become ineffective and we must develop novel therapies based on novel technology and discoveries. These effects can be seen as new antibiotics are constantly being developed.

a) Prognoses refers to the likely outcome of your illness. b) Profile cells for future risk of getting a disease. (However, this is of little significance if “C” is realized because any genomic or proteomic “defect” can be fixed if therapies are out there and these pathways are known.) c) Knowing the molecular profile of illnesses and infections, one can effectively target these ailments and minimize side effects.

“prohibits group health plans and health insurers from denying coverage to a healthy individual or charging that person higher premiums based solely on a genetic predisposition to developing a disease in the future.” “The legislation also bars employers from using individuals' genetic information when making hiring, firing, job placement, or promotion decisions.” Wiki, “Genetic Information Nondiscrimination Act” http://www.ama-assn.org/ama/pub/physician-resources/medical-science/genetics-molecular-medicine/news.shtml

Molecular based (i.e, personalized) Medicine By I&C President Ryan Witt witt.rj@gmail.com

Schedule Bio 101 Molecular Medicine

Remember Ask Qs Always ask “ Why/how, What, and When” Try to understand EVERYTHING!

Our Bodies Made up of 10 to 100 trillion cells Cells make up Tissues Tissues make Organs (i.e., Skin, lungs, brain)

Molecules So… what and where are “molecules?” What: A group of two or more atoms bonded to one another. (Pauling, Linus (1970). General Chemistry. New York: Dover Publications, Inc.. ISBN 0-486-65622-5 ) Where: Everywhere Including inside AND outside of cells H20 Water

Now, let’s look at function…

The Driver Molecules are the drivers of your cells. Forming the structure of DNA, RNA, proteins, enzymes, and EVERYTHING pushing our body to do what it does!

DNA RNA Proteins Newly discovered / other components: Methylases (methylate DNA), mRNA, alternative splicing, micro-RNA, tRNA This is how cells work! Environment / extra-cellular signals

DNA Been called “the Master Plan” and “your destiny.” Is there merit? Codes for ALL players – proteins, enzymes, microRNA, RNA, etc. But, DNA is the code… not an actor / player itself

Reality 101 DNA isn’t everything! DNA mutations change our DNA throughout our lifetime + Expression and function is partially independent of DNA (i.e., epigenetics (methylation); post-translational modifications) Expression and “end-conformation” of molecules is what matters most -- that is, what players (i.e., RNA, glycoproteins, proteins) will be active, and how are they functioning.

3 fun examples of environment/our actions + DNA, and their results!

Example: Stubbing your Toe (DNA change) + Rubbing You can either rub or not! Depending on which affects the molecular characteristics describing you in a diff’t way!

Ex: form of Dieting (methylation change) “Excessive alcohol consumption has been shown to alter DNA methylation in the colon of humans.”

How We Turn Light into Sight

How We Turn Light into Sight

In our BRAINS too! “Technology is not only changing our lives, it’s changing our BRAINS.” - Dr Gary Small, Director of the UCLA Center on Aging & Professor of Psychiatry and Behavioral Sciences at UCLA

Coming back to the Q Is our DNA everything? No, our actions change our molecules - which changes our cells, organs, and bodies!

Reality Reality is… We are a product of environment / actions plus our “blueprint.” The blueprint is the start; everything from there is up to you!

Lastly Beware! ALWAYS ask, “What, Why, When and How” http://www.youtube.com/watch?v=R34Sguwxe2o

4-slide Recap

Molecules Two or more atoms bound together! Molecules make up the components of cells, tissue, organs, and us as a whole

How your body works

Our actions affect our bodies -- at the molecular level Rubbing

Take Home Points Our actions  the molecular attributes describing those actions. Blueprint / cards dealt (DNA) + choices and environment Expression levels, the sequences of molecules, among other factors—which constantly are changing—are the keys to understanding the underlying molecular mechanisms describing our actions, at any given moment. (DNA is code, not the actor!) 1 & 3 are VERY important for next lecture – know them well!

COOL! Woooooooooooooooo – done with bio! (next is Medicine!) Any questions?

Break Ask your doc about molecular testing if you have a disease, infection, or simply want to learn more about yourself. What to test -- DNA Expression levels, sequence, and functionality of molecules (DNA, micro-RNA, RNA, proteins, etc)

Break: Myths about Genetic Testing and Molecular Med Insurers can drop you based on results. GINA prohibits -- “Denying coverage to a healthy individual or charging that person higher premiums based solely on a genetic predisposition to developing a disease in the future.” “Employers from using individuals' genetic information when making hiring, firing, job placement, or promotion decisions.” “prohibits health insurers from using genetic information to determine eligibility or premiums, prohibits insurers and employers from requesting or requiring that a person undergo a genetic test, and prohibits employers from using genetic information to make employment decisions.” – AMA site If someone did happen to get a hold of my DNA, they could clone an identical copy of me!

Medicine (the standard, then innovation)

Key point There are – Our actions and characteristics the Molecules describing them & Rubbing Rubbing

Things can get very hazy Cancer

Medicine When it comes to medicine, challenge is Clinical Symptoms Where & What is molecular-Cause Stuffy nose Fever Headache Slurred speech Twitchy eye

Medical Diagnostics: standard / currently “Differential diagnosis” What is the most likely cause Physical Examination – checking your signs and symptoms Imaging Tools (MRIs, CT scans, fMRI, etc) Histopathology – looking under a microscope to distinguish cell / tissue characteristics

Treatment Treatment by “trial-and-error” and “one-size-fits all” AMA and medical terminology

Right now Many of the same diseases by modern classification have different molecular characteristics/drivers. Meaning: “NSCLC of squamous type” –> MDM2 mutation? p53 mutation? And/or EGFR mt? Met? Etc. Picture Source: http://www.iftheshoefitz.com/

Innovation (Currently, not the standard)

BUT Wait a minute! …What about the “molecular drivers?”

Molecular Profiling Now we have the capability to take a sample of diseased or infected cells (or a person’s “normal cells”), and examine it at the molecular level -- looking at RNA, protein structure, expression levels, and genetic makeup (DNA). The methods are there (immunohistochemistry, DNA arrays, RNA microarrays, real-time quantitative PCR) Which is best – unsure!

Cool! What’s the value? Predictive to suggest treatment? No…not by itself (if no clinical experiments are performed) I.e., MammaPrint, EGFR test, Tissue of Origin But it could be (predictive), if inferences are made by some person…(as you will see)

Profiling provides insights into the molecular mechanisms governing/describing your symptoms -- AKA Knowledge!!!!!

What can be done with knowledge: Applying Molecular Profiling to Treatment CANCER Xeloda – Thymidine Phosphorylase overexpression EGFR & MET overexpression FUS1, MDM2, & P53 therapy

FUS1, MDM2, & P53 therapy

Application is to ALL diseases With every ailment, there are respective proteins, genes, and molecules driving its progression. Bacterial Infections Azithromycin – antibiotic; works on various strains of bacteria (inhibits 50S ribosomal subunit from acting) ALS SOD1 gene target -- Arimoclomol …Alzheimer’s, MS, Parkinson’s, Crohn’s, Lupus

Value By seeing a comprehensive picture of one’s expression levels, sequences, and so forth, one has a more clear view of the underlying molecules and structure of molecules, hence a more clear perspective on the likely outcomes of a therapeutic intervention.

Knowledge Nearly every drug approved on the market by a regulatory agency has a known molecular mode of action. If you distinguish the molecular characteristics of a disease from “normal cell” characteristics, then you can match drugs perfectly with those disease characteristics and known molecular signaling pathways within a disease! Hence, less of a trial-and-error / one-size-fits-all approach. Idea that fuels (i) “Molecular Medicine”, (ii) Personalized Med, (iii) Combination Therapies and (iii) “Drug Repositioning”

Without molecular knowledge, and continued trial-by-error Rx… W/o mDx, you have -- Different responses to the same treatments, in regards to the safety and effectiveness of that therapy. http://learn.genetics.utah.edu/content/health/pharma/intro/ Life? Death?

W/o molecular testing and treatment Resistance Development & your Window of Opportunity is lost! Personal level Society at large

Translation, Please W/o molecular profiling and continued trial-by-error treatment -- Therapies available are no longer effective; must find something else. Antibiotics Not everything is available now – avg new cancer therapy takes 14 yrs of testing before final approval. In the instance of antibiotic resistance and infectious diseases, this means spreading these adverse effects to society!

Questions Any questions? (ask: HOW, WHY, WHAT, WHEN!)

Benefits of m-based Medicine More specific detection and classification of existing diseases, leading to more accurate prognoses and earlier detection. Predictive/predisposition tests and actionable things you can do with such info Better idea of what—molecularly—to target and how to most effectively and efficiently treat a patient with a particular illness or infection. http://learn.genetics.utah.edu/content/health/pharma/intro/ Less adverse side effects from metabolism variants (PGx) via dosage variations for diff’t patients, combination drug adverse reactions, and side effects in general Better idea of what will work, and gives u additional ideas of what to use as an Rx

Increasing Trends in Molecular Testing “Learning about #lungcancer mgmt trends. EGFR mut'n testing is up from 2-3% to ~10-12% from early to late 2009; should rise more in next yr.” - Tweet from Oncologist in Seattle, WA. “The biology of lung cancer differs from one patient to the next, depending on age and sex, according to scientists at Duke University Medical Center. The findings may help explain why certain groups of patients do better than others, even though they appear to have the same disease.” - http://www.brightsurf.com/t/52409/

Myths about Genetic Testing and Molecular Med Insurers can drop you based on results. GINA prohibits -- “Denying coverage to a healthy individual or charging that person higher premiums based solely on a genetic predisposition to developing a disease in the future.” “Employers from using individuals' genetic information when making hiring, firing, job placement, or promotion decisions.” “prohibits health insurers from using genetic information to determine eligibility or premiums, prohibits insurers and employers from requesting or requiring that a person undergo a genetic test, and prohibits employers from using genetic information to make employment decisions.” – AMA site If someone did happen to get a hold of my DNA, they could clone an identical copy of me!

Where to go from here Talk with your doc about molecular testing if you have a disease, infection, or simply want to learn more about yourself and the underlying molecular mechanisms of your disease, infection or biological actions. “Molecular profiling”, “molecular testing”, “genetic testing” What to test -- DNA – “genetic testing” Expression levels, sequence, and functionality of molecules (DNA, micro-RNA, RNA, proteins, etc) -- “molecular profiling”

What Are We (I&C) Doing? Bringing innovation to you at the fastest rate possible is our goal. Community-based education and resource sharing Building an interface to empower you to make a difference Working to integrate these innovations and patient options into physicians’ workflow.

Thank you for listening!