drug studies using mouse models - national cancer institute
TRANSCRIPT
![Page 1: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/1.jpg)
Drug Studies Using Mouse Models
Melinda Hollingshead, D.V.M., Ph.D. Biological Testing Branch
Developmental Therapeutics Program Division of Cancer Treatment and Diagnosis
NCI
![Page 2: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/2.jpg)
Why should the taxpayers fund this type of work?
Cancer is the second leading cause of death in America. It accounts for one of every four deaths. More than 565,000 - that’s
more than 1,500 people a day - die annually from cancer. Close to 1.4 million new cases are diagnosed each year. This estimate does not include pre-invasive cancer or the more than 1 million cases of non-melanoma skin cancer expected to be diagnosed annually.
![Page 3: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/3.jpg)
Why do we use mouse models?
genetics progression prevention diagnostics therapeutics physiologically complex vs in vitro studies cost effective develop human clinical protocols
To prevent, diagnose and treat human disease
![Page 4: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/4.jpg)
What models are available?
Spontaneous Virus-induced Transgenic Knock-out/in Induced/carcinogens Transplanted
![Page 5: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/5.jpg)
Spontaneous Models
Natural history is most “normal” Random tumors Difficult to predict occurrence Monitor through lifespan ~ 2 yr in mice Large number of animals may be required
for a small experiment Tumors may be heterogeneous Significant animal holding space needed
These allow the study of the biological history of natural disease. They can be applied to many types of studies.
![Page 6: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/6.jpg)
Virus-Induced Models
Classic models of virus-induced leukemia: Rauscher Moloney LP-BM5 Friend
Non-leukemic tumors including: Mammary tumors due to MMTV Thymomas of AKR mice
High tumor occurrence rate Predictable time to tumor development Well-characterized disease states/natural history Do not accurately reflect the natural history of most human
tumors
Early studies of cancer, both as a disease state and for therapeutic intervention, used virus-induced models. These models continue to be applied
in a variety of studies.
![Page 7: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/7.jpg)
Transgenic Models
Many available through commercial and collaborative arrangements http://emice.nci.nih.gov/emice/mouse_models http://mouse.ncifcrf.gov/ http://jaxmice.jax.org/query/f?p=205:1:1510299228434659165
Not all patents have expired - check with OTT regarding legalities Remember MTAs are needed to receive or ship material that may
have patent or licensing rights. This also applies to mice. Rate of tumor occurrence may be low Time to tumor occurrence may be difficult to predict Breeding schemes may be complex e.g., 3, 4 or more
intermediate genetic crosses Genetic error(s) are generally well characterized Disease may follow a more natural course e.g., time to
significant tumor burden may more accurately mimic the human; lesions likely relevant to their site of occurrence
Transgenic models aid our understanding of the genetics of cancer and are being pursued as models for intervention.
![Page 8: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/8.jpg)
Induced/Carcinogens
Epithelial tumorigenesis 7,12-dimethylbenz[a]anthracene [initiator mutagen] followed by multiple
applications of 12-O-tetradecanoylphorbol-13-acetate [pro-inflammatory]
GI tumorigenesis 1,2 dimethlhydrazine-2-HCl azoxymethane
Sarcoma induction Methylcholanthrene
Lung tumorigenesis nitrosamine 4- (methyl-nitrosamino)-1-(3-pyridyl)-1-butanone
These models aid studies of the local, systemic, and environmental factors that influence tumor susceptibility, growth, and progression.
![Page 9: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/9.jpg)
Induced/Carcinogens
Time to tumor development may be variable
Mimics some human diseases very well Requires technical work with known
carcinogenic agents thus special handling/equipment/facilities/training required
Provides the entire natural history of tumor development to study
![Page 10: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/10.jpg)
Transplanted Models
Tissue Source Syngeneic - same inbred strain – e.g., B16 tumors
in C57Bl/6 mice Allogeneic - same species, different strain
(genetically diverse) – e.g., M5076 sarcomas in athymic mice
Xenogeneic - different species e.g., human, rat or dog tumors grown in immunocompromised mice
Implant Site Orthotopic Heterotopic
Endpoints
These models are commonly applied in studies of genetics, diagnostics, and medical interventions.
![Page 11: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/11.jpg)
Transplanted Models
Easy to control time of tumor occurrence Many tumor types/lines available Well accepted models Do not accurately recapitulate human disease Metastatic lesions are difficult to find Tumor growth rates may preclude multiple
treatment cycles
![Page 12: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/12.jpg)
production In vitro studies
efficacy trials
Discovery & Development
New therapy
Pharmacology & toxicology
![Page 13: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/13.jpg)
• Human Tumors – Hollow fiber – Subcutaneous – Intravenous – Intraperitoneal – Orthotopic
• Mammary fat pad • Intracranial • Intrarenal • Intrahepatic • Intracecal
• Rodent Tumors – Subcutaneous – Intravenous – Intraperitoneal – Orthotopic – Metastatic – Transgenic – Knock-in/out – Induced – Spontaneous
In Vivo Efficacy Models
![Page 14: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/14.jpg)
Issues faced during Efficacy Evaluations
Model Vehicle, formulation, stability Dose, route and schedule Experimental protocol Endpoints
![Page 15: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/15.jpg)
What are you assessing? Which type of model is most appropriate? Is the treatment designed to:
impact the tumor chemically, e.g., cytotoxic impact the tumor genetically, e.g., modulator impact the stroma e.g., vasculature impact the immune system act as an adjuvant synergize with known drugs interact with specific proteins
Model Selection
![Page 16: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/16.jpg)
Model Selection
A
0
200
400
600
800
1000
1200
1400
1600
1800
22 25 28 31 35 39 43 46 49 54 61
Day Post-Implantation
Med
ian
Tum
or W
eigh
t
B
-200
0
200
400
600
800
1000
1200
1400
1600
1800
13 19 22 26 29 33 36 39 46 50 53 56 60 63
Day Post-implantation
Med
ian
Tum
or W
eigh
t
![Page 17: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/17.jpg)
Experimental Protocol
When will treatment start? When will treatment end? Will samples be collected for ex vivo
evaluation? Will tumors be monitored visually, by
imaging techniques? What will terminate the experiment,
i.e., what are the endpoints?
![Page 18: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/18.jpg)
What is published/prior knowledge? What is proposed/expected mechanism? How much exposure is required for effect? Is the material soluble/stable in aqueous
solution? What routes of administration are technically
feasible? Options
What is the maximum tolerated dose (MTD)? IP, IV, SC, PO? QDx?; BIDx?; TIDx?
Dose, Route, Schedule
![Page 19: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/19.jpg)
A
0
500
1000
1500
2000
2500
19 22 27 29 32 34 40Day Post-implantation
med
ian tu
mor
weig
ht (m
g)
▬×▬ vehicle ▬□▬ temozolomide 400 mg/kg ▬Δ▬ temozolomide 200 mg/kg x 3 ▬●▬ topotecan 1 mg/kg QDx10
![Page 20: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/20.jpg)
Endpoints Tumor size Weight loss Time to sacrifice Imaging Pre-defined time of termination Time post-treatment
![Page 21: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/21.jpg)
MDA-MB-231-Luc
0
200
400
600
800
1000
1200
13 16 19 23 27 33 40 47
Tum
or W
eigh
t (m
g)10% Etoh/10% Cremaphor/80% saline
75 mg/kg cyclophosphamide Q3Dx4 IP
8 mg/kg paclitaxel Q7Dx3 IV
Saline Q3Dx4 IP
![Page 22: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/22.jpg)
0
10
20
30
40
50
60
70
80
90
100
1 6 11 16 21 26 31 36 41 46 51 56 61 66
Day Post-Implantation
% S
urvi
vors
PBS oligonucleotide in PBS
lipofectamine oligonucleotide in lipofectamine
![Page 23: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/23.jpg)
Prior to Treatment (Day 6)
7 days after third treatment (Day 21)
Vehicle BCNU
Measuring antitumor drug activity using bioluminescence vs. tumor weights
Average Luminescence and Median Tumor Weight
0.00E+00
5.00E+07
1.00E+08
1.50E+08
2.00E+08
2.50E+08
3.00E+08
3.50E+08
4.00E+08
4.50E+08
11 18 21 24/25 32 39 46Days from Implant
Pho
tons
/sec
0
500
1000
1500
2000
2500
3000
Tum
or W
eigh
t (m
g)
Average Lumin. ETOHAverage Lumin. Nsc 409962Median TW 2% ETOHMedian TW NSC 409962
![Page 24: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/24.jpg)
PC-3 Tumor
12
0
10
0
80
60
40
20
0 PS-341 (mg/m2)
0.9 1.8 Vehicle
20S
Act
ivity
(% V
ehicle)
120 100 80 60 40 20 0
0.3 0.9 3.0 9.0 VCTL PS-341 (mg/m2) 20
S Act
ivity
(% V
ehicle)
Mouse WBC
GENOMICS Tumor RNA
BIOMARKER
SURROGATE BIOMARKER
Retention Time (min)0 5 10 15 20 25
Inte
nsit
y (A
rbit
ray
Uni
ts)
0
100
200
300
400
500
600
FR (IS)Staurosporine (IS)
NC381
NC383
NC384
DRUG ANALYSIS
Plasma
In Vivo Studies
![Page 25: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/25.jpg)
IHC Original tumor and each passage Antibodies for specific gene products
Molecular analysis gene expression: microarray (LCM), RT-PCR proteins/phosphoproteins: reverse arrays, IHC biomarkers: serum proteomics
Analysis of host response immune system response (syngeneic v.s. immunosuppressed) Angiogenesis (MFP v.s. sc)
Stem Cell search OCT
IHC
Snap-Frozen (RNA)
Harvest and freeze cell suspension
Tools for Analysis
![Page 26: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/26.jpg)
Colo-829 SC Tumors Collected Post Dose 4
Drug Vehicle Q12hx5D PO
Topotecan 5mg/kg QDx5 IP
ABT-888 25mg/kg Q12hx5D PO
Poly-ADP-ribose (Trevigen)
Min 20.0 Max 400.0 4 min exp
Mou
se 5
Mou
se 9
Mou
se 1
1
Mou
se 1
3
Mou
se 1
5
Mou
se 1
8
AA
XR2
-7 #
1
Mou
se 3
Mou
se 7
Mou
se 1
6
Mou
se 1
7
Mou
se 1
9
Mou
se 2
1
AA
XR2
-7 #
1
Mou
se 1
Mou
se 2
Mou
se 4
Mou
se 6
Mou
se 8
Mou
se 1
0
AA
XR2
-7 #
1
200
140
100
80
513 517 521
b-Actin
Topotecan 5mg/kg QDx5 IP
ABT-888 25mg/kg Q12hx5D PO
Poly-ADP-Ribose (Trevigen) Min 20.0
Max 400.0 4 min exp
Mou
se 2
6
Mou
se 2
7
Mou
se 3
0
Mou
se 3
4
Mou
se 3
6
AA
XR2
-7 #
1
Mou
se 1
2
Mou
se 1
4
Mou
se 2
5
Mou
se 2
8
Mou
se 3
2
Mou
se 3
3
AA
XR2
-7 #
1
519 523
Drug Vehicle Q12hx5D PO
Mou
se 2
0
Mou
se 2
3
Mou
se 2
4
Mou
se 2
9
Mou
se 3
1
Mou
se 3
5
AA
XR2
-7 #
1
200
140
100
80
515
b-Actin
Colo-829 SC Tumors Collected Post Dose 10
![Page 27: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/27.jpg)
Many Models – Many Options
With few exceptions, every rodent model, even if conducted with hundreds of experimental mice, represents a single patient. Interpreting the
preclinical results and applying these outcomes to human clinical trials continues to prove challenging for those charged with translating the preclinical
experience into viable drug candidates.
Statistically valid model assessing relevant endpoints on an optimal schedule
with clinically appropriate doses.
![Page 28: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/28.jpg)
PROBLEMS
The appropriateness of animal models to identify, qualify and promote new therapies for cancer has been under review, and in some ways under attack, for many years. Continuing concerns about the failure rate of agents being sent to the clinic has led to a flurry of publications on the irreproducibility of published preclinical data and their over-prediction of activity.
![Page 29: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/29.jpg)
![Page 30: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/30.jpg)
![Page 31: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/31.jpg)
![Page 32: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/32.jpg)
![Page 33: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/33.jpg)
CAVEAT!
![Page 34: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/34.jpg)
![Page 35: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/35.jpg)
What can you do? 1. use well powered animal studies 2. reproduce your own data 3. have 2 separate operators generate the data 4. provide adequate details in publications 5. don’t over-interpret your data 6. stage tumor studies correctly 7. don’t selectively use/present your data 8. remember the clinical situation and what can be
assessed in man
![Page 36: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/36.jpg)
Clinical Endpoints In Man Toxicity - What harmful effects are induced? Tumor response - Does the cancer respond to the treatment?
Biomarker modulation as a measure of the effect of a treatment that may correlate with a traditional clinical endpoint (PFS; TR)
Progression-free survival (stable disease) Tumor regression Statistically significant improvement in survival
Survival - how long does the person live? Quality of life - how does the treatment affect a person's
overall enjoyment of life and sense of well being?
![Page 37: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/37.jpg)
Progression, Stable, Regression
![Page 38: Drug Studies Using Mouse Models - National Cancer Institute](https://reader031.vdocuments.site/reader031/viewer/2022012505/6180d50cf7fe7071e80fdc6d/html5/thumbnails/38.jpg)