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AACR Annual Meeting · April 18–22, 2015 · Philadelphia, PA
Insulin-like growth factor 1 (IGF1R)/insulin receptor (INSR) inhibitory activity
of rociletinib (CO-1686) and its metabolites in nonclinical models
P 793
Andrew D. Simmons, Sarah Jaw-Tsai, Henry J. Haringsma, Minh Nguyen, Andrew Allen, Thomas C. Harding
Clovis Oncology Inc., San Francisco, CA
The parent CO-1686 molecule does not increase glucose or
insulin levels in a rat oral glucose tolerance test
BACKGROUND
• Rociletinib (CO-1686) is a novel, oral, irreversible tyrosine kinase inhibitor for
the treatment of patients with mutant epidermal growth factor receptor (EGFR)
non-small cell lung cancer (NSCLC) that has demonstrated efficacy against
the activating mutations (L858R and del19) and the dominant acquired
resistance mutation (T790M), while sparing wild-type (WT) EGFR.
Figure 1. Chemical structure
of rociletinib (CO-1686)
• Interim data from a phase 1/2 study demonstrated that heavily pretreated
patients with T790M+ NSCLC receiving rociletinib (500 or 625 mg BID)
had a 67% objective response rate.1
• An oral glucose tolerance testing (OGTT) was carried out in female Sprague-Dawley rats to directly evaluate
the role of CO-1686 on glucose homeostasis. The study design is shown in Table 3.
Table 3. Study design to evaluate CO-1686 in an OGTT in female Sprague-Dawley rats
Group
No. of females
Test article
Dose (mg/kg)
Dosing schedule
1
2
3
4
4
4
4
4
Vehicle
CO-1686
CO-1686
BMS-754807
NA
1000
500
25
PO x 1
PO x 1
PO x 4 days BID dosing
PO x 1
Rociletinib Safety Profile
• Interim data from a phase 1/2 study demonstrate that rociletinib is well tolerated, with no evidence of systemic
WT EGFR inhibition.1
• The most common treatment-related adverse events reported in ≥15 percent of all patients included
hyperglycemia, diarrhea, nausea, and reduced appetite.1
• Hyperglycemia was unexpected in the clinic since it was not observed in toxicology studies performed in rats
and dogs.
BID = twice daily; NA = not applicable; PO = oral
• The peak concentration (Cmax) of CO-1686 achieved in the OGTT was 3 – 6-fold greater than the Cmax
achieved in patients dosed with rociletinib at 500 mg BID (Table 4).
Table 4. Comparison of the Cmax of CO-1686 observed in rats and humans
Study
Rat OGTT
Human patients
IGF1R/INSR Pathway
• One possible explanation for the finding of hyperglycemia in clinical trials is inhibition of the insulin-like growth
factor 1 receptor (IGF1R) and/or the insulin receptor (INSR).
• IGF1R inhibition has been observed with other EGFR inhibitors, highlighting the structural similarity in the
kinase domain between these proteins.2
• IGF1R/INSR have established roles in glucose homeostasis and are part of a complex signaling pathway.3, 4
Total plasma Cmax (ng/mL)
Unbound plasma Cmax (ng/mL)
9680 – 15,100a
2690b
126 – 196
35.0
aMean
plasma Cmax following 4 days of 500 mg/kg BID (first value in range) or a single oral dose of 1000 mg/kg (second value) during the 2-hour OGTT
experiment period; bMean Day 1 levels in humans following repeated oral dosing of rociletinib at 500 mg BID
• Increased glucose and insulin levels were not observed with CO-1686 dosing in an OGTT (Figure 2).
Figure 2. (A) Blood glucose and (B) insulin levels over time in an OGTT in Sprague-Dawley rats
A
B
The goal of the studies reported here were to explore the mechanism(s) underlying hyperglycemia
observed in a subset of patients treated with rociletinib.
These data suggest M460 and M502 may be involved in the hyperglycemia observed in patients
Metabolites M460 and M502 are observed at greater levels
in humans and have increased potency against IGF1R/INSR
Preclinical studies in NSCLC models have demonstrated a role for IGF1R/INSR signaling in mediating
resistance to EGFR inhibitors; for example:
Table 5. Comparison of Cmax of metabolites M460, M502, and M544 in rats and humans
Metabolite and study
Total plasma Cmax (ng/mL)
Unbound plasma Cmax (ng/mL)
416 – 479a
4460b
3.33 – 3.83
250
4020 – 4570a
853b
80.4 – 91.4
43.5
7.72 – 11.1a
562b
0.80 – 1.14
39.9
M502
Rat OGTT
Human patients
M544
Rat OGTT
Human patients
M460
Rat OGTT
Human patients
• IGF1R/INSR signaling has been shown to be required for the emergence of drug tolerant persistor cells.5, 6
• IGF1R/INSR inhibitors can overcome resistance in mutant EGFR patient-derived cell lines.7
• Several in vitro studies were performed to evaluate the role of IGF1R and M502 in EGFR inhibitor resistance.
Figure 5. (A) Exogenous insulin-like growth factor 1 (IGF-1) increases p-IGF1R and p-INSR levels in PC-9 cells
(del19 EGFR). (B) Exogenous IGF-1 decreases CO-1686 sensitivity in PC-9 cells, and combining CO-1686 with
a selective IGF1R inhibitor (OSI-906) or M502 reverses IGF-1 induced resistance to CO-1686.
aMean
plasma Cmax following 4 days of 500 mg/kg BID (first value in range) or a single oral dose of 1000 mg/kg (second value) during the 2-hour OGTT
experiment period; bMean steady state levels in humans following repeated oral dosing of rociletinib at 500 mg BID
A
B
PC-9
pIGF1R
• We explored the potency of the metabolites against IGF1R and the INSR (Table 6). Metabolites M460 and
M502 have increased potency against IGF1R and INSR.
pEGFR
• We also explored the potency of the metabolites against WT and mutant EGFR (Table 7). CO-1686 and its
metabolites do not inhibit WT or mutant EGFR.
pINSR
PC-9 + 100 ng/mL IGF1
Table 6. Potency of metabolites against IGF1R and INSR
Assay
CO-1686 IC50 (nM)
M460 IC50 (nM)
pIGF-1R
M502 IC50 (nM)
M544 IC50 (nM)
BMS-754807 IC50 (nM)
INSR kinase
166
27
23
3
133
INSR cellular
162
74
51
7
167
IGF1R kinase
477
52
57
1
401
IGF1R cellular
732
210
101
12
662
Kinase IC50 profiling was performed using functional biochemical radiometric assays (Reaction Biology). Cellular profiling was performed using Ba/F3
INSR and IGF1R engineered cell lines (Advanced Cellular Dynamics). IC50 = half maximal inhibitory concentration
Table 7. Biochemical and cellular profiling of rociletinib metabolites
Assay
EGFR kinase
EGFR kinase
A431 cellular
NCI-H1975 cellular
PC-9 cellular
HCC827 cellular
The parent CO-1686 molecule shows limited activity against
IGF1R and INSR in biochemical and cellular assays
IGF1R inhibition may play a role in EGFR TKI resistance
Genotype
CO-1686 IC50 (nM)
M460 IC50 (nM)
M502 IC50 (nM)
M544 IC50 (nM)
WT
T790M
WT
L858R T790M
Del19
Del19
83
5
952 ± 40
38 ± 5
75 ± 4
23 ± 4
>1000
901
737 ± 67
1094 ± 97
1468 ± 50
1334 ± 147
>1000
>1000
1128 ± 155
811 ± 68
1596 ± 121
697 ± 64
>1000
>1000
4688 ± 907
3496 ± 217
3713 ± 115
3243 ± 204
pEGFR
pINSR
(A) Serum starved PC-9 cells were incubated with exogenous 100 ng/mL IGF-1 for 1 hour and evaluated using receptor tyrosine kinase antibody arrays (R&D
Systems). (B) Cell viability in PC-9 cells was determined by CellTiterGlo 72 hours after ligand and compound addition. Exogenous IGF-1 was added at 100
ng/mL, OSI-906 at 0.5 µM, and M502 at 1 µM. DMSO = dimethyl sulfoxide
Figure 6. In both first- and second-line EGFR mutant models, the addition of M502 or OSI-906 delayed the time
to CO-1686 resistance.
A
B
Kinase IC50 profiling was performed using functional biochemical radiometric assays (Reaction Biology). Cellular viability is reported as average IC50 ± SEM
in nM (cellular). IC50 = half maximal inhibitory concentration
SEM = standard error of the mean
• Whole kinome profiling of CO-1686 in functional
biochemical showed limited potency against IGF1R
and INSR (Table 1).
Taken together, these data suggest that the parent CO-1686
molecule does not directly play a role in hyperglycemia
Table 1. Wild-type kinome profiling of CO-1686
Kinase
1 µM
0.1 µM
EGFR (T790M)
FAK/PTK2
CHK2
LRRK2
ERBB4/HER4
JAK3
FES/FPS
FLT3
STK22D/TSSK1
BMX/ETK
ROS/ROS1
ACK1
TNK1
ALK
IRR/INSRR
FER
TYK1/LTK
PYK2
ERBB2/HER2
BTK
IGF1R
EGFR
IR
98
96
96
94
96
98
91
92
90
95
91
96
89
87
94
89
81
75
91
83
80
89
79
94
77
70
69
68
64
61
60
57
56
55
53
51
45
42
40
39
31
31
30
29
26
25
CO-1686 selectivity was tested against 434 kinases in duplicate at concentrations
of 1 μM and 0.1 μM using an ATP concentration of 10 μM (Reaction Biology).
Target kinases are sorted by percentage inhibition at 0.1 µM.
Metabolites M460 and M502 increase glucose and insulin
levels in an OGTT
• In cellular assays, CO-1686 showed limited
potency against IGF1R and INSR (Table 2).
Figure 3. Rociletinib metabolites identified in human plasma
• BMS-754807 is a potent inhibitor of IGF1R
and INSR that was used as a positive control
for these studies.
• To directly evaluate the potential role of these metabolites in hyperglycemia, M460 and M502 were
administered to Sprague-Dawley rats in an OGTT.
Table 2. Biochemical and cellular profiling of CO1686 and metabolites against IGF1R and INSR
Assay
INSR kinase
INSR cellular
IGF1R kinase
IGF1R cellular
CO-1686 IC50
(nM)
BMS-754807 IC50
(nM)
166
162
477
732
3
7
1
12
Kinase IC50 profiling was performed using functional biochemical
radiometric assays (Reaction Biology). Cellular profiling was
performed using Ba/F3 IGF1R and INSR engineered cell lines
(Advanced Cellular Dynamics). IC50 = half maximal inhibitory
concentration
The increased exposure to M460 and M502 in humans, combined with the greater binding potency of
these 2 metabolites for IGF1R and INSR, suggests that M460 and M502 may play a role in the
hyperglycemia observed in patients.
Metabolite M502 delayed the time to CO-1686 resistance in (A) PC-9 and (B) NCI-H1975 cells. Cells (1 × 105) were plated on day 0 in the presence of CO1686, M502, and/or OSI-906 at a concentration of 1 µM, and cell number was assessed over time.
• Doses of M460 and M502 were selected to achieve plasma Cmax levels comparable to those observed in
patients dosed at 500 mg BID.
CONCLUSIONS
• Metabolites M460 and M502 increase glucose and insulin levels in an OGTT (Figure 4).
•
Figure 4. (A) Blood glucose and (B) insulin levels over time in rats administered M460 and M502
A
•
B
•
The hyperglycemia observed in patients may result from metabolite(s) at higher exposure in humans as
compared with rats and dogs. Rociletinib metabolites identified in human plasma are shown in Figure 3.
• To explore this hypothesis, metabolite levels were determined in plasma samples collected from the OGTT
study and compared with metabolite levels observed from dosing rociletinib in patients at 500 mg BID (Table
5). Metabolites M460 and M502 were observed at higher levels in humans than in rats.
• Metabolites M460 and M502 were also observed at higher levels in humans as compared to the levels
observed in rat and dog toxicology studies (data not shown).
The parent CO-1686 molecule does not appear to play a role in the hyperglycemia observed in a subset of
patients treated with rociletinib.
The metabolite M502 is likely to be the main driver of hyperglycemia observed in humans, through inhibition
of the IGF1R and INSR kinases.
• These data suggests that hyperglycemia may be most effectively managed by oral agents that target
insulin resistance (e.g. metformin, thiazolidinediones, SGLT2 inhibitors), rather than by administration of
exogenous insulin or by drugs that stimulate insulin production
IGF1R inhibition may contribute to the efficacy of rociletinib and play a role in delaying the onset of
resistance.
1. Soria J et al. Presented at ENA; November 18–21, 2014; Barcelona, Spain. 10LBA.
2. Pollack M Nat Rev Cancer 2012;12:159-169.
3. Chen HX and E Sharon Chin J Cancer 2013;32:242-252.
4. Finlay MR et al. J Med Chem. 2014;57:8249-8267.
5. Sharma SV et al. Cell. 2010;141:69-80.
6. Cortot AB et al. Cancer Res. 2013;73:834-843.
7. Crystal AS et al. Science. 2014;346:1480-1486.