Trial Information

Targeted Demethylation to Enhance Response or Overcome Resistance to EGFR Blocking Agents in KRAS Wild-type Metastatic Colorectal Cancer Patients Using Sequential Decitabine and Panitumumab

Patients with metastatic colorectal cancer are living longer and running out of therapeutic
options due to disease resistance. Epidermal growth factor receptor (EGFR) has been
validated as a therapeutic target in colorectal cancer (CRC). Ligand binding to EGFR
activates the RAS/RAF/MAPK, STAT, and PI3K/AKT signaling pathways, which together modulate
cellular proliferation, adhesion, migration, and survival. Anti-EGFR targeted antibodies
cetuximab and panitumumab administered as monotherapy in CRC have shown response rates of
approximately 9% and 17% respectively (Amado et al., 2008; Saltz et al., 2004). Single
agent panitumumab has been approved for use in third line colorectal cancer and has been
shown improve progression free survival over supportive care. Further subset analysis showed
the response rate of 17% was confined to patients with KRAS wild type tumors only and that
this group (approximately 60-70% of all CRC patients) should be considered for further study
(Amado et al., 2008). According to the Huntsman Cancer Hospital registry, colorectal cancer
patients are the largest disease group within our gastrointestinal cancer group and many
have or eventually will progress on available therapy or are or will become intolerant to
the side effects of second line therapies such as oxaliplatin neuropathy or irinotecan
induced diarrhea, yet still are candidates for treatment.

In the lab through translational research studies, we hope to identify re-expression or a
reduction in promoter methylation of genes involved in tumor suppressor pathways known to be
important in colorectal cancer (CRC) or involved in EGFR signaling pathway. Candidate genes
we will evaluate will include genes described in prior studies as associated with the CpG
island methylator phenotype (CIMP) as well as genes previously reported to be
hypermethylated in association with colorectal neoplasia. These will include APC, SFRP
family members, CDH-1 (e-cadherin) and p16 (Belshaw et al., 2008; Lind et al., 2004; Suehiro
et al., 2008). Other genes more specific to EGFR or KRAS signaling that will be assessed
include: RASSF1A , a tumor suppressor gene know to be hypermethylated in several human
cancers including CRC, is occasionally associated with KRAS wild type and when silenced by
methylation allows for RAS activation (Kang et al., 2006; Oliveira et al., 2005); SOX17, a
member of the transcription factor superfamily know to be hypermethylated in CRC and lead to
disrupted Wnt signaling (Zhang et al., 2008); SOCS-1 a negative regulator of STAT3 an
activating ligand for EGFR that has been shown to be silenced by hypermethylation and allow
for constitutive signaling via EGFR (Lee et al., 2006); and PTEN, a tumor suppressor that
antagonizes the PI3K- AKT/PKB signaling pathway by dephosphorylating phosphoinositides (Noro
et al., 2007). Further candidate genes may be discovered or added based on preliminary data
and ongoing research. Methylation analysis and gene expression pattern changes will be
done using methylation specific PCR and bisulfite sequencing of genes known to be involved
in EGFR signaling pathways and colorectal neoplasia as described above. We have prior data
from our own work as well as others to suggest the use of a hypomethylating agent can
resensitize colon cancer cells to therapeutic agents (Karpf et al., 1999; Morita et al.,
2006). The translational component of this research will be supported by institutional
translational grant awarded to the PI, Kimberly Jones, as of July 1st, 2008. This
information may help identify other important targets and allow for the design of better
combination therapies. We plan to do these assays on weekly blood and buccal samples while
patients are on therapy, on epithelial cells swabbed from panitumumab associated skin rash,
and on archived or biopsied tumor specimens when available (from KRAS testing (required) and
optional end-of treatment biopsy).

The pharmacokinetic profile for decitabine has been well described and offers several
possible dosing schedules feasible for clinical practice and combination with other agents.
Decitabine is currently being tested in combination with standard cytotoxic agents. It has
shown some activity in solid tumors, however, myelosuppression is a common side effect,
especially when given concurrently with other myelosuppressive therapy (carboplatin)
(Appleton et al., 2007; Plimack et al., 2007). We propose a novel study using decitabine in
combination with a non-myelosuppressive targeted biological agent as well as giving it
sequentially rather than concurrently to try to maximize the effect of the second drug by
dosing it during the demethylation window. In the dose-finding study reported by Appleton
et al., they recommended a dose of 90 mg/m2 over 6 hours every 28 days, but went up as high
as 135 mg/m2 and combined this with carboplatin (Appleton et al., 2007). We have chosen a
dose of 45 mg/m2 decitabine every 14 days based on its reported safety and biological
equivalence from this study. There was no grade 3 or 4 hematological toxicities observed
with 3 patients infused with 45 mg/m2 of decitabine followed by 5 AUC carboplatin; there was
1 episode of grade 3 leukopenia and 1 of grade 3 neutropenia in 4 patients infused with 45
mg/m2 of decitabine followed by 6 AUC carboplatin (Appleton et al., 2007). At the higher
dose of 90 mg/m2 with 5 AUC carboplatin in 13 patients, 5 episodes of grade 4 leukopenia or
neutropenia were observed; in 10 patients with 90 mg/m2 and 6 AUC carboplatin, there were 4
episodes of grade 4 leukopenia or neutropenia. The proposed infusion over 2 hours should
not create any infusion rate toxicities and will be a schedule patients will tolerate.
Daily lower dose infusions used in hematological disorders maybe be efficacious but daily
intravenous chemotherapy impacts patient's quality of life significantly. Decitabine also
has an elimination half-life of 30 minutes, so will clear the body rapidly. The proposed
administration of the decitabine twice, at half the dose, in a 28-day period should be
better tolerated than the 90 mg/m2 combined with carboplatin. The myelosuppressive toxicity
which is the main toxicity of decitabine should be less significant as it is not being
combined with a second myelosuppressive agent as it was with carboplatin. Low dose
decitabine (45 mg/m2) has nearly equivalent hypomethylating effects to 90 mg/m2 in both
blood and buccal cells. The more frequent dosing (every 2 weeks versus every 4 weeks)
should maintain the hypomethylating effect at a lower dose. Additionally, we propose a
novel dosing schedule where our second agent, panitumumab, will be given on alternating
weeks and targeting the EGFR pathway when it is hypomethylated from the prior decitabine
treatment. Our work as well as others, has demonstrated that specific promoter
hypomethylation is observed by 8-14 days after the start of treatment and genomic DNA
reverts to baseline levels by 28 to 35 days after the start of treatment (Appleton et al.,
2007; Samlowski et al., 2005, Kantarjian, 2007 #23).

We will also be assessing clinical response and progression free survival (PFS) and
comparing it to historical controls of patients treated with panitumumab monotherapy. If
the combination can be given safely and responses are seen that are equal to or better than
the single agent panitumumab data, we would plan a multi-center larger phase II trial.