Biotech & Pharma

The Science, The Jobs, & Skills for Success

Neil Stahl Ph.D. Regeneron Pharmaceuticals

Overview
Science at a Company vs. Academia Attributes for Success at a Company Biotech vs. Big Pharma Biotech : Innovation and Risk Drug Development 101 Job Opportunities Outside of Research Getting Hired

My Experience
BS Zoology Duke 1978 Duke Marine Lab 1977, 78, 79 brought me to Science PhD Biochemistry Brandeis 1979-1985 Quantitative fundamentals of equilibria, kinetics, & how to make a conclusion Post-Doc at UCSF with Stan Prusiner - Scrapie Prions Learned many fields ranging from protein chemistry, cell biology, transgenics, human genetics Regeneron Discovery, 1991 (65 employees) Explored mechanisms of how cytokine receptors are activated and activate cytoplasmic signaling pathways Figured out a way to make tight binding cytokine blockers based on the mechanism of cytokine activation, using multiple cytokine receptor components in a recombinant fusion protein = Cytokine Trap Regeneron Drug Development (1999-2005; now 565 employees) Established preclinical development group - put 3 Traps into clinical trials Joined Senior Management - reorganized program management, clinical project teams, jointly manage all aspects of Research and Clinical Development, present to investors, analysts, potential partners

Science at a Company
Scientific endeavor on a project can be carried out at a scale that is very rare in a University setting Teams of competent people aligned toward a common goal can accomplish more than any individual scientist

Discoveries can be translated into therapeutic opportunities with the potential to create new drugs and technologies
Understand molecular and cellular pathways defining a particular biology and how it goes wrong in disease Create a drug to impact those pathways Explore how that drug works in animals and humans Design Clinical Program to prove that the drug is safe & effective

Register the drug with the FDA and Rest of World

Differences Between Academia & Industry

You will have access to far more resources, equipment, core facilities, and collaborative colleagues to advance your project

You will be required to work on projects of the companys choosing

You may be asked to switch to (or add on) new projects Although you will report to one person, you will interact with many Scientists instead of a single PI Participate and present in cross-functional meetings where data is vetted and the future directions of a project are established by discussion and consensus More heads are better than 1! You are likely to publish and attend scientific conferences

Some Myths of Industry

You have failed if you dont pursue an academic position Thats what some told me, but there are many, many incredibly competent people doing Science & Drug Discovery in Industry The working day is 9-5 Hard, effective work is expected and rewarded! Compensation is dramatically better than academia Entry level scientist positions (3-5 year postdoc) are compensated similarly to Assistant Professors, but much better than post-docs, and there are stock options! However, opportunity for advancement is more frequent and more rapid than Academia

You never get to publish

I published more rapidly at Regeneron than anywhere else! Also, compensation is based on contributions beyond publishing

You cant move from Industry to Academia More and more, Universities value Industry experience and perspective, making a reverse move more likely

Attributes for Success at a Company

Team player who can collaborate effectively with others Ability to become interested in a wide variety of different scientific areas - learning is a continuous Life-long experience! Superb analytical, communication, and presentation skills All of us have particular skills that make us good Scientists, although my exact skill set may not be the same as yours Contribute your particular talent and expertise toward the common goal

Success means that your project grows so that hundreds of people work on it!

Biotech
Often more innovative, high-risk scientific approaches More informal working environment, with a were all in this together spirit. A do what it takes to get the job done attitude that may provide more variety

vs

Big Pharma

Typically more traditional small molecule Drug Discovery, unless partnered with Biotech Typically more hierarchical Employees can become pigeon-holed in a particular function.

Larger organizations usually have more More likely to participate in decision-making rules! process Much larger experience base More resources than Academia, but often partners with Pharma for expensive late stage clinical programs Can be acquired, have layoffs, or slowly go out of business More opportunities for advancement than Academia or Pharma if company grows Stock options can provide financial windfall if company successful Can bring huge resources to bear on a project, although there is always internal competition for resources Can be acquired, or have periodic layoffs

Base compensation often higher than Biotech, but usually doesnt have as large a stock option upside Promotion may occur more slowly

Biotech : Innovation & Risk

Biotech companies have traditionally been founded to exploit cutting edge ideas and technology. Examples include: Using our own cytokines, growth factors, and enzymes as drugs

Engineering human fusion proteins, combining functionalities to achieve new properties

Creating Humanized and Human Monoclonals as drugs Transcriptional control siRNA Ribozymes Aptamers Gene Therapy Many Biotech ventures are unsuccessful, often because there is not a realistic business plan of how to create an income-generating product before their ability to raise money runs out You need to assess whether the companys scientific and business plan makes sense, their history and future potential of raising capital, partnering deals they have closed, and how soon they will generate revenue

The Promise of the Human Genome Sequence

The Hype: Drug Development will be revolutionized following the identification of novel genes in druggable classes The Fact: Identifying novel genes is the first baby step. Understanding their biology and creating therapeutics against them is the difficult step that, in many instances, can take decades Accelerating these steps is the key to creating novel therapeutic opportunities

Target Validation : Velocigene Allows Rapid Creation of Mutant Mice, and Detailed Visualization of Expression

1 Nature Biotechnology paper described 10% of KOs ever made!

Huge Opportunities in Protein-Based Therapeutics

Good Drug Targets are hard to come by Many companies make Me Too drugs against targets for which drugs already exist Many Interesting Targets are large proteins (eg Cytokines and Growth Factors) that drive broad biological responses These pathways cause disease if inappropriately stimulated These targets are usually not amenable to small molecule approaches Current successful approaches include monoclonal antibodies that block cytokine action, and receptor - Fc fusion proteins As we learn more about Biology, we will uncover an ever growing number of Targets that will require protein-based interventional approaches

Protein Therapeutics - Examples

Success Stories Insulin - First administered to humans in 1922
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Interferons
Erythropoietin - 1989 Growth Hormone Enbrel - a receptor-Fc fusion protein Antibodies - eg Herceptin, Rituxan, Remicade, Humira, Avastin Mouse immunoglobulins - antigenicity Thrombopoietin (Tpo)- efficacy, immunogenicity Lenercept - Receptor-Fc fusion - immunogenicity Leptin - misunderstood mechanism - efficacy

Less Successful Stories

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Protein Therapeutics Strengths & Weaknesses

Strengths high specificity compared to small molecules

Little off-target toxicity - less likely to fail in early trials

Block Targets not amenable to small molecules - eg growth factors & receptors

Weaknesses

More difficult to manufacture

Potential immunogenicity even from fully human proteins Low abundance proteins that dont circulate Protein variants (aggregates, oxidation, deamidation) that break tolerance

More difficult to evaluate toxicology injectable

Regeneron Technology: Heteromeric Soluble Receptors Form Tight-Binding Traps

Antibody Structure Heavy Chain

Ra

Rb

Light Chain

Fc
(Drives Dimerization)

Fc

Cytokine
Kd = 5 nM Kd = 10 pM

Ra

Rb

In-Line Heteromeric Traps

Ra Ra Ra Rb Rb Rb

Must Overexpress 2 cDNAs Must Purify Heterodimer away from Homodimers Waste Cells Production Capacity with Unwanted Homodimers

In-Line fusion of 2 receptors without intervening linkers creates simple homodimer with very high affinity

Making a Proof of Concept into a Drug

Make T-shirts Do Preclinical Work File Investigational New Drug Application with FDA Clinical Trials

File Biologics License Application

PreClinical Development Checklist

BioMolecular Engineering Cell line Development - FASTR

Process Development
Formulation Assay Development Pharmacology Pharmacokinetics Toxicology Regulatory - IND = Investigational New Drug Application

BioMolecular Engineering
IL1 Receptor Complex Create Trap candidates with different receptor order (eg: ab-Fc, ba-Fc, a-Fc-b, b-Fc-a), different fusion IL1R Type 1 IL1R-AcP IL-1 position in receptor sequence + linkers to increase flexibility IL1 Single Chain Trap

IL1R-AcP Extr ace llular Doma ins IL1R Type 1 s s s s

Evaluate for bioactivity, high Ce ll M e mbrane expression level from CHO cells, Cytoplasmic clean folding Doma ins a-Fc with no extraneous linkers

Extr ace llular Doma ins

CH2

hIgG Fc

CH3

Something Old, Something New Ways to Isolate Over-Expressing Cell Lines

How to isolate clones after transfection:

Traditional:
GOI

Random isolation of clones

Pick clones ELISA Dilution clone ELISA

FASTR :

Isolation based on expression / characteristic of secreted protein

GOI FACS ELISA

FASTR Cell Line Selection

Flow cytometry-based Autologous Secretion Trap

CHO Parental cell with doxycylineinducible expression of FcR

Binds Trap internally and displays on cell surface Whole population of transfected cells can be sorted by FACS with fluorescent anti-Fc Allows selection of highest expresser from amongst millions of transfected cells Turn off expression of FcR after selection to allow unhindered secretion for manufacturing

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Process Development
Goal is to have protein secreted from CHO (Chinese Hamster Ovary cells) which have low viral burden and make human carbohydrate structures Batch-Fed Bioreactor Process yield is 1-3 g/L after 10-12 days of culture Start at 2L scale, eventually to 10,000 L, which yields 10 kg at expression of 1 g/L 3 step purification process (Protein A, ion exchange, hydrophobic interaction chromatography) with up to 70% yield

Formulation
Desire high concentration with adequate stability to give > 2 year shelflife

Add GRAS (Generally Regarded as Safe) excipients to stabilize protein from aggregation, deamidation, oxidation, fragmentation
Polysorbate, sucrose, amino acids, PEG

IV formulations generally <10 mg/ml

Subcutaneous (SC) - 25-100 mg/ml

IL1 Trap: liquid at 50 mg/ml or lyophilized at 80 mg/ml

Assay Development/Pharmacokinetics
Immunoassays to measure Trap and their complexes with target cytokines in plasma Assays to detect formation of antibodies against the Trap
100000 10000

IL-1 Trap (ng/mL)

1000 100 10 1 0 0 50 100 150 Time (hr) 200 250 Group 4: 3 mg/kg IV Group 5: 3 mg/kg SC

Use to measure PK - how the blood levels change over time, which often guides dosing frequency and active dose levels

IV & SC pharmacokinetics in Monkeys

Pharmacology: Murine Model of Collagen-Induced Arthritis

Arthritis Severity Index
CIA model in dba-1 mice is the most widely accepted model of rheumatoid arthritis
4

Vehicle

Injection of bovine collagen II induces immune response that results in progressive autoimmune joint destruction
Injection of zymosan IP at day 30 gives more robust and synchronous arthritis response Arthritis severity index grades inflammation, swelling, and deformity IL1 Trap blocks cartilage erosion, as well as joint swelling and deformity

Trap 10 mg/kg

Trap 31 mg/kg
30 34 38 Day 42 46 50

0 26

Trap

Vehicle

Toxicology
Usually, new drugs are tested at high doses in 2 animal species to identify NOAEL (No Adverse Event Level) and MTD (Maximum Tolerated Dose) Test drugs at >10x higher doses than expected human dose Many protein therapeutics have strict species specificity, and can only be tested in primates, but often KO data in animals is predictive of safety issues IL1RI KO shows no adverse phenotype except increased susceptibility to some types of bacterial infections Moreover, human proteins are often immunogenic in animals IL1 Trap only binds primate IL1 6 week toxicology study in monkeys showed no evidence of toxicity, but an antibody response was observed after a few weeks that resulted in clearance of Trap from circulation No MTD observed, adequate safety to proceed to clinical trials!

Immunogenicity in animals not predictive of Ab response in humans

Regulatory
FDA regulates testing of experimental drugs in people

Must submit IND - Investigational New Drug Application

Usually takes us ~1 year to complete, and may involve ~100 people

Describes everything you know about the manufacturing and structure, PK, pharmacology, formulation, stability, toxicology, proposed clinical plan for Phase I trials
FDA gets 30 days to respond, allowing you to go forward, or request more information, or to tweak your clinical trial design

Clinical Trial Overview

Phase I Safety Dose Escalation in Volunteers or Patients Dose Ranging Efficacy Studies to decide on dose and interval Proof of Efficacy Treat larger number and broader range of patients to evaluate overall safety and look for less frequent adverse events (AEs)

Phase II
Phase III

As few as 4 clinical studies (each one a single experiment) could suffice to get a drug approved for use in humans!!

Entry Level Positions in Biotech

Research Post-Doctoral Scientist Analogous to Academia, except more resources and mentoring available As in academic post-doc, a good publication record should allow return to Assistant Professor route Pharmaceutical Post-Doctoral Scientist Contribute to Clinical Development Projects or Core Technologies in ways that may not result in high profile publications Would lead to a career in Biotech/Pharma

Scientist
2-5 years post-doctoral experience Staff Scientist 3 years experience following Post-Doc

Career Opportunities Outside of Research

Preclinical Development Immunoassays & Sample Analysis from Human Clinical Trials Formulation Development Pharmacology - Assessing Drugs in Animal Models Protein Sciences Cell line generation to overexpress recombinant proteins Protein characterization

New technology and assay development

Protein Manufacturing Process Development Program Coordination & Management Core Facilities

Methodology Oriented (DNA, in situ, FACS, Mass Spec, Biacore)

Clinical Regulatory - understand FDA Guidance, liason for company to FDA, EU Scientific Writing Quality Control Business Development

Getting Hired

Application & Hiring Process

Typically, job descriptions are posted, applications solicited Human Resource personnel (non-scientists) review applications, winnowing down to those that match job description, and pass on to Hiring Scientists

Unsolicited applications to HR and Hiring Scientists can sometimes hit paydirt and find an opening before its even listed

CV & Cover Letter Essentials

Must communicate to multiple audiences Scientists - trying to figure out if you have the raw materials that they can mold into a productive scientist and useful contributor Human Resources - non-scientists checking for a match between your CV and a job description Usually your First & Only Chance to make a positive impression Should convey your Intelligence & ability to communicate (Clear Writing = Clear Mind!) Perspective of your field beyond your own project Accomplishments - aimed at a non-expert and placed in context of the open questions in your field

Skill set - techniques that you really know as well as those for which you may have a passing knowledge and vocabulary
Enthusiasm!

CV
Same CV can be used for all applications Need not be 1 page - can be 3-4 or longer

Research summary
explain in 1 paragraph your projects and conclusions aimed at someone who is not in your field Can also briefly describe rotation & graduate research

Clearly identify core skill sets

Dont exaggerate - youll get busted just because you have seen a mass spectrometer doesnt mean you should list it as a core competency!!!

Presentations
Awards/Grants Initiatives that youve undertaken outside your core requirements Publications - including submitted / in preparation

Supervisory & Collaborative experiences

Summary & Technical Skills

Cover Letter
Ideally should be customized for each application Should connect your skill set and experience to the job you are applying for so that its easy for HR to understand and pass on to hiring scientist Should describe your project and findings in the broad context of your field - often the best way to convey to the Hiring Scientist that you were not just a skilled set of hands directed by your PI Rarely is an applicant perfect for the job - often we look for someone that appears to be smart, communicates well, and can grow into a job Therefore, its usually a stretch to say that you can make Regeneron a success More reasonable to emphasize your flexibility and ability to learn quickly