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Lessons Learned in T1 Research: mouse to human. Jean Y. Tang MD PhD Assistant Professor Department of Dermatology. Mechanism of disease. In vitro experiments. Animal studies. Human clinical trials. New drugs. Epidemiological studies. Lessons learned: challenges.

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lessons learned in t1 research mouse to human

Lessons Learned in T1 Research: mouse to human

Jean Y. Tang MD PhD

Assistant Professor

Department of Dermatology

mechanism of disease
Mechanism of disease

In vitro experiments

Animal studies

Human clinical trials

New drugs

Epidemiological studies

lessons learned challenges
Lessons learned: challenges
  • Jack of all trades, Master of none
    • Journals
    • Conferences
    • Students and trainees
  • Not that many role models

– find a true believer and the experts

Slower time to publication

Grants: enthusiasm from NIH

slide4
KL2/K23 Mentors:
  • Ervin Epstein, Children’s Hospital Oakland Research Institute
  • Mary-Margaret Chren, Dept of Dermatology, UCSF
  • Charles McCulloch, Steve Cummings, Dept of Epidemiology and Biostatistics, UCSF
slide5

Translational Research in BCC

Screen for drugs in cell lines and

Ptch1+/- mice

Clinical trials

in PTCH1 +/-

BCNS patients

Observational studies

at Kaiser/UCSF

epidemiology of bcc
Epidemiology of BCC
  • 1 million BCC cases per yr in US
  • Estimated annual incidence of 0.1% to 0.5%
  • Rare risk of metastasis: < 0.5%
  • 5th most costly cancer for Medicare
  • The age-adjusted incidence per 100,000 white individuals: 475 cases in men, 250 cases in women
  • The estimated lifetime risk of BCC in the white population is 33-39% in men and 23-28% in women.
  • Risk of second BCC: 44% in 3 yr
slide9
BCC basic science:
  • Almost all BCCs have mutations in PTCH1 tumor suppressor gene
  • All BCCs have increased Hedgehog signaling
hedgehog signaling pathway regulates cell proliferation and growth

HH

HH

HH

Ptch

Ptch

Ptch

Ptch

Ptch

Ptch

Mutant

Mutant

Mutant

Ptch

Ptch

Ptch

CPN

Smo

Smo

Smo

Smo

Smo

Smo

Smo

Smo

Smo

Smo

Smo

Gli

Gli

Gli

off

off

off

Gli

Gli

Gli

on

on

on

Gli

Gli

Gli

on

on

on

Gli

Gli

off

Hedgehog signaling pathway regulates cell proliferation and growth
basal cell nevus syndrome
Basal cell nevus syndrome

Basal cell nevus syndrome are PTCH1+/-

slide12

(A) Photo of multiple circled BCCs on the back of a patient with Basal Cell Nevus Syndrome. (B) Photo of Ptch1+/- K14-Cre-ER2 p53 fl/fl mice with multiple BCCs

Ptch1+/- mice mimic BCNS phenotype: develop BCC tumors after IR or UV treatmentGoodrich and Scott, Science 1997Aszterbaum, Oro, Scott, Epstein Nature Medicine 1999

mechanism of disease13

Roadmap for finding new therapeutics

Mechanism of disease

Hedgehog pathway

BCC cell lines

In vitro experiments

Ptch1 +/- mice

Animal models

PTCH1+/- Basal cell nevus syndrome patients

Human clinical trials

Patients with sporadic BCCs

Epidemiological studies

slide15

Genetic deletion of Cox1 or Cox2 decreases microscopic BCCs in Ptch1+/- mice

IR-treated Ptch1+/- mice wild type (n=24), deleted for Cox1 (n=12) or for Cox2 (n=6). Mean and SEM. p<0.05

Cox1 and Cox 2 KO: Smithies Cell 1995

*

*

slide17
Study design: Phase II randomized, double-blinded, placebo controlled trial
  • Subjects: 60 patients with Basal Cell Nevus Syndrome (BCNS)
  • Treatment: oral Celecoxib at 200mg BID versus placebo for 24 months followed by 12 months of observation
  • Primary endpoint: change in BCC numbers during study periods
slide18

Baseline characteristics of study participants

are similar in two groups (mean and SD)

how to analyze bcc development
How to analyze BCC development
  • Regession technique: Linear mixed models
  • Calcuates a slope or rate (number of BCCs/yr) for each patient
  • Compare percent change in rate of BCCs in placebo and celecoxib groups
  • Accounts for drop-outs
  • Adjust for age, gender, BCC at baseline
lessons learned importance of finding a reliable mouse model
Lessons learned: importance of finding a reliable mouse model
  • Ptch1+/- mice are a reliable model for testing new anti-BCC agents in humans
    • Moderate effect of celecoxib on BCCs in mice and in BCNS patients
    • Greater effect on tumor size rather than number in both mice and BCNS patients
lessons learned statistics
Lessons learned: statistics
  • Statistics – linear mixed models for determining slope of BCCs in RCT
  • Regression models for tumors in mice
    • Go to class
  • Building a database (and managing)
    • Go to class
lessons learned
Lessons learned:
  • Long time to publication
  • Journals and co-authors disagree on whether to present data from mice and clinical trial study together
  • Cancer Prevention Research
new and relatively safe agents that decrease hedgehog signaling

Vitamin D3, Statins

New and relatively safe agents that decrease Hedgehog signaling

Cyclopamine

Smo

Gli

Corcoran and Scott, Proceedings of the Natl Acad Sci 2006

Bijlsma and Peppelenbosch, PLOS Biology 2007

mechanism of disease29
Mechanism of disease

Statins blocks Hedgehog pathway

BCC cell lines

In vitro experiments

Ptch1 +/- mice

Animal models

Human clinical trials

Statin therapy and risk of subsequent BCCs in Kaiser cohort

Epidemiological studies

lessons learned31
Lessons learned
  • Have a good biomarker or target gene (Gli1 mRNA)
  • Have a good way to measure bioavailability (24OHase)
  • Have a reliable cell line
slide34

Ptch1+/- K14-Cre-ER2 p53 fl/fl mice treated with vehicle control (Left) versus topical vitamin D3 (right) at 7 months of age.

  • Example of large BCC tumor on the dorsal skin these transgenic mice
  • Histological confirmation of BCC.

Topical Vitamin D3 decrease BCC development

by 50% (p<0.05)

slide35

Topical vitamin D3 decreases BCC development

in mice

*Adjusted for gender and coat color of mouse

lessons learned36
Lessons learned
  • Have a good biomarker or target gene (Gli1 mRNA)
  • Have a good way to measure bioavailability (24OHase)
  • Pilot trial of topical and oral vitamin D on human BCC
mechanism of disease37
Mechanism of disease

Vit D3 blocks Hedgehog pathway

BCC cell lines

In vitro experiments

Ptch1 +/- mice

Animal models

PTCH1+/- Basal cell nevus syndrome patients

Human clinical trials

Vit D3 levels in BCC pts

Epidemiological studies

lesson learned
Lesson Learned
  • Translating from mouse studies to epidemiologic studies (skip the human clinical trial/pilot)
  • Mouse to Epi (Ralph Gonzalez)
slide39
The association of serum vitamin D with skin cancer risk in elderly men

Jean Y. Tang1,3, Neeta Parimi2, Angela Wu1,3, John Boscardin1, Meg Chren1,Steven R. Cummings1,2, Ervin Epstein3, and Douglas C. Bauer1,2

1 University of California San Francisco, 2 San Francisco Coordinating Center, California Pacific Medical Center Research Institute, 3 Children’s Hospital Oakland Research Institute

table 3 association of increasing serum 25 oh d levels with non melanoma skin cancer
Table 3 Association of increasing serum 25(OH)D levels with non-        melanoma skin cancer  

* Adjusted for age (continuous variable), BMI (continuous variable), season of blood draw, and clinic site

† Adjusted for age, BMI, season of blood draw, clinic site, outdoor walking activity (continuous variable), and

cigarette smoking (yes/no)

slide44

Topical Cur-414 cream

Oral Hh antag

Cyclopamine

Smo

Gli

Williams and Wang , Proceedings of the Natl Acad Sci 2003

slide49

Hh Antagonist cream decreases Gli1 mRNA in BCC tumors

S

Tu

Tu

Tu

S

Tu

S

Tu

Re

S

Re

Re

Tu

S

Tu

Tu

Hh Antagonist cream

Placebo

Lesson: benefits of collaborator

lessons learned mechanism of disease
Lessons learned: mechanism of disease
  • Topical and oral HH antagonists significantly reduce murine BCCs by decreasing tumor proliferation and/or inducing follicular differentiation.
  • We know how to collect tumors, do these assays, get to mechanism of disease in future trials
investigator sponsored trial in bcc prevention
Investigator sponsored trial in BCC prevention
  • Genentech Hh antagonist GDC 0449: 150mg daily
  • Phase II, placebo controlled RCT in 41 Basal Cell Nevus Syndrome subjects
  • Primary endpoint: change in BCCs at 12 mo and 18 mo
new and relatively safe agents that decrease hedgehog signaling56

Itraconazole

New and relatively safe agents that decrease Hedgehog signaling

Cyclopamine

Smo

Gli

J Kim and P Beachy, Stanford

mechanism of disease57
Mechanism of disease

Small molecule library screen for inhibitors of Hedgehog pathway

Cell based assays

In vitro experiments

Ptch1 +/- mice

Animal models

Patients with sporadic BCCs

Human clinical trials

slide58

Day 4 – irritation and necrosis, 11 mm

Day 0, BCC A: 10mm

Day 11 – residual BCC, 3mm

Itraconazole#178

cyclodextrin 5460
Cyclodextrin#5460

Day 0 – BCC B (11 mm), BCC D (10 mm), BCC G (9mm)

Day 7 – BCC B (11 mm), BCC D (10 mm), BCC G (9mm)

lessons learned60
Lessons learned:
  • Collaboration with another basic lab
    • Post-doc fellows who are experts at specific assays (5 years) vs training someone new
  • Focus on getting the first pilot clinical trial of itraconazole on human BCCs
    • Measure Gli1mRNA in BCCs
    • Measure Ki67
    • Paired t-test (at biopsy and at excision)
  • New opportunities/markets
lessons learned61
Lessons learned:
  • Easier to translate FDA approved drug or a drug manufactured and tested by Pharma - already have paid the $$$
  • New agent (vitamin D) – investigator pays
    • Efficacy
    • Stability
    • GMP grade for human
    • IND
slide63
Children’s Hospital Oakland

Ervin Epstein

Po Lin So

Tony Zheng Xiao

Elana Shpall

Angela Wu

Kris Chang

Yefim Khaimsky

UCSF EpiBiostat

Charles McCulloch

Ralph Gonzalez

Steve Hulley

Steve Cummings

Doug Bauer

Neeta Parimi

John Boscardin

Michael Kohn

Kaiser Division of Research

Maryam Asgari

Genentech

Fred de Sauvage

Tracy Tang

Chris Callahan

UCSF Derm

Meg Chren

Dan Bikle

Loretta Chan

Funding Sources

NRCC – CTSA KL2

NIAMS – K23

Prevent Cancer Foundation

American Skin Association