1) The study used 11C-erlotinib PET imaging to determine if tumors with different EGFR mutations would specifically bind the tyrosine kinase inhibitor erlotinib. 2) They found that tumors with kinase domain mutations of EGFR demonstrated specific binding of 11C-erlotinib, while tumors with extracellular domain mutations did not. 3) The results indicate that 11C-erlotinib PET can identify tumors that are likely to be sensitive to treatment with erlotinib based on their level of drug binding, as determined by kinetic modeling of the PET data.

1. PET determination of specific uptake
of 11C-erlotinib by different tumor types
expressing EGFR, in vivo, through
kinetic modeling
J. Ryan Petrulli1,4, Jenna M. Sullivan1,4, Ming-Qiang Zheng2,4,
Yiyun Huang2,4, Joseph N. Contessa3, Evan D. Morris1,2,4
1. Biomedical Engineering
2. Diagnostic Radiology
3. Therapeutic Radiology
4. Yale PET Center
Yale University, New Haven, CT, USA

2. CLINICAL SIGNIFICANCE
• Epidermal growth factor receptor (EGFR)
– Extracellular domain mutations: 40% of glioma patients1
– Tyrosine kinase domain mutations: 10%-15% of non-
small cell lung cancer (NSCLC) patients2,3
WT EGFR
“ACTIVATION”
Cell growth
Apoptosis resistance
Cell proliferation
Kinase Domain
Mutant EGFR
Extracellular Domain
Mutant EGFR
Ligand Binding
[1] Pelloski CE et al. (2007). J Clin Oncol 25, 2288-2294. [2] Sequist LV et al. (2008).
J Clin Oncol 26, 2442-2449. [3] Yang CH et al. (2008). J Clin Oncol 26, 2745-2753.

3. CLINICAL SIGNIFICANCE
• EGFR-targeted treatment
– Diagnostic method: biopsy + mutation analysis
– Tyrosine kinase inhibitors (TKIs): erlotinib, gefitinib,
afatinib
ACTIVATION
TKIs INACTIVEMUTANT
EGFR
BIOPSY
SAMPLE
OR OR
MUTATION ANALYSIS
[4] Lynch, TJ et al. (2004).
N Engl J Med 21, 2129-39.

4. QUESTION: Can we use PET to identify
erlotinib-sensitive tumors?

5. EGFR PET
• Direct assessment of drug interaction with EGFR
• Target engagement
– TKI tracer binds specifically to EGFR kinase domain
– Excess cold drug scans: saturate system with
unlabeled TKI
• Kinetic modeling
– Simplified Reference Tissue Model (SRTM)5: compares
ROIs to a “control” (no target engagement) ROI
– Chosen outcome metric: binding potential (BP, index of
specific binding)
[5] Lammertsma A and Hume S (1996). Neuroimage 4, 153-158.

6. APPROACH: Perform 11C-erlotinib PET
on tumors of varying EGFR status with or
without excess cold erlotinib and
determine specific binding with kinetic
analysis (SRTM)

7. METHODS
• Subjects: nude mice implanted with 2-3 tumor xenografts
• Human cancer cell lines: SW620, U87, HCC827, PC9, and U87∆
• Scans: Siemens Focus 220; 11C-erlotinib injections with or without
excess erlotinib
• Analysis:
– Regions of interest (ROI) drawn on summed images; regional time-
activity curves
– Kinetic modeling with SRTM to produce BP
– Statistical comparison between BP in each xenograft and drug condition
HCC827, PC9
Kinase Domain+
Active
U87∆
Extracellular Domain+
Active
U87
WT EGFR
Inactive
Cell Line:
Mutation:
Status:
SW620
No EGFR
n/a

8. KINASE DOMAIN MUTANT TUMORS
0
0.5
1
1.5
2
0 50 100
SUV
Time (min)
0
0.5
1
1.5
2
0 50 100
SUV
Time (min)
Tracer Experiment Excess Cold Drug
SW620 (▲)
Muscle (○)
HCC827 (■)
PC9 (♦)
HCC827, PC9
KD Mutant
Activated EGFR
no EGFR
KD Mutant
KD Mutant

9. EXTRACELLULAR DOMAIN MUTANT TUMORS
Tracer Experiment Excess Cold Drug
Muscle (○)
U87 (*)
U87∆ (●)
0
0.5
1
1.5
2
0 50 100
SUV
Time (min)
0
0.5
1
1.5
2
0 50 100
SUV
Time (min)
U87
WT EGFR
Inactivated EGFR
U87∆
ECD Mutant
Activated EGFR
ECD Mutant
WT EGFR

10. -1
0
1
2
3
4
5
6
7
8
9
HCC827 PC9 SW620 U87 U87∆
MeanBP
tracer experiment
excess cold drug
SPECIFIC BINDING
** p<0.05 * p=0.06
**
*
NS NS
NS
n= 3 3 3 3 1 1 3 2 3 2
KD+
Active
ECD+
Active
WT EGFR
Inactive
Mutation:
Status:
No EGFR
n/a
KD+
Active

11. -1
0
1
2
3
4
5
6
7
8
9
HCC827 PC9 SW620 U87 U87∆
MeanBP
tracer experiment
excess cold drug
ACTIVATION ≠ TARGET ENGAGEMENT
** p<0.05 * p=0.06
NS
KD+
Active
ECD+
Active
WT EGFR
Inactive
Mutation:
Status:
KD+
Active
n= 3 3 3 3 1 1 3 2 3 2
No EGFR
n/a

12. -1
0
1
2
3
4
5
6
7
8
9
HCC827 PC9 SW620 U87 U87∆
MeanBP
tracer experiment
excess cold drug
SENSITIVITY OF KINETIC MODELING
** p<0.05 * p=0.06
**
KD+
Active
ECD+
Active
WT EGFR
Inactive
Mutation:
Status:
KD+
Active
n= 3 3 3 3 1 1 3 2 3 2
No EGFR
n/a

13. SUMMARY
FINDINGS:
• Not all tumors with activating EGFR mutations bind erlotinib
• 11C-erlotinib PET can image EGFR with kinase domain
mutations
• 11C-erlotinib PET cannot image EGFR with extracellular domain
mutations
INTERPRETATION:
• Sensitivity to erlotinib among EGFR mutant tumors requires
erlotinib binding and can be demonstrated by 11C-erlotinib PET
imaging
IMPLICATIONS:
• Binding potential is a sensitive index which can measure the
degree of drug binding

14. FUTURE DIRECTIONS
• Do all radiolabeled TKIs behave comparably?
• How does specific binding relate to TKI efficacy?
• How do tumors with acquired resistance to TKIs
interact with 11C-erlotinib?

15. SPECIAL THANKS
Funding: Yale Clinical Center for Investigation Scholar Award (UL1RR024139 and
KL2RR024138), the Kalimeris fund, Yale PET Center, NSF GRFP Grant DGE-1122492.
Yale PET Center
Sarah Goldberg, MD, MS

16. SUPPLEMENTAL INFORMATION
Presenting Author:
Joseph “Ryan” Petrulli
joseph.petrulli@yale.edu
Supporting Material:
Petrulli JR, Sullivan JM, Zheng MQ, Bennett DC, Charest J,
Huang Y, Morris ED, Contessa JN. (2013). Quantitative
analysis of [11C]-erlotinib PET demonstrates specific binding
for activating mutations of the EGFR kinase domain.
Neoplasia 15(12) 1347-53.