One-step radiosynthesis of18F-IRS: A novel radiotracer targeting mutant EGFR in NSCLC for PET/CT imaging

Article abstract of DOI:10.1016/j.bmcl.2016.10.084

EGFR (epidermal growth factor receptor) targeted therapy has shown great success in clinical comparing with chemotherapy in EGFR mutation NSCLCs. Such as, gefitinib, first generation EGFR TKI, has obviously prolonged the FPS (progression free survival) of the subgroup of patients, but to those who did not get a certain mutation in EGFR kinase domain, the outcome is poor. In view of this situation, scientists have synthesized many radiotracers for selecting the right people by PET/CT imaging to NSCLC TKI therapy. In this study, we developed a novel PET radiotracer¹⁸F-IRS in one-step with a radio yield 20% (non-corrected), radiochemistry > 98.5%, specific activity > 105G Bq/μmol, the pharmacokinetics and capacity of the tracer binding to mutant EGFR were evaluated both in vitro and in vivo.

Full text of DOI:10.1016/j.bmcl.2016.10.084

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
One-step radiosynthesis of 18F-IRS: A novel radiotracer targeting mutant
EGFR in NSCLC for PET/CT imaging
⇑
⇑
Zunyu Xiao, Yan Song, Kai Wang, Xilin Sun , Baozhong Shen
a TOF-PET/CT/MR Center, The Fourth Hospital of Harbin Medical University, China
b Molecular Imaging Research Center, Harbin Medical University, China
a r t i c l e i n f o
a b s t r a c t
Article history:
EGFR (epidermal growth factor receptor) targeted therapy has shown great success in clinical comparing
with chemotherapy in EGFR mutation NSCLCs. Such as, gefitinib, first generation EGFR TKI, has obviously
prolonged the FPS (progression free survival) of the subgroup of patients, but to those who did not get a
certain mutation in EGFR kinase domain, the outcome is poor. In view of this situation, scientists have
synthesized many radiotracers for selecting the right people by PET/CT imaging to NSCLC TKI therapy.
In this study, we developed a novel PET radiotracer 18F-IRS in one-step with a radio yield 20% (non-cor-
Received 28 July 2016
Revised 21 October 2016
Accepted 27 October 2016
Available online xxxx
Keywords:
18F-IRS
EGFR
rected), radiochemistry > 98.5%, specific activity > 105G Bq/
lmol, the pharmacokinetics and capacity of
the tracer binding to mutant EGFR were evaluated both in vitro and in vivo.
Ó 2016 Published by Elsevier Ltd.
TKIs
PET/CT
Imaging
For the last decade, non small cell lung carcinoma (NSCLC) is
still the leading causes of cancer death [1]. Conventional treatment
method, like chemotherapy, has only lead to marginal effect, but
great toxicity. With the development of genomics, the epidermal
growth factor receptor (EGFR) has been known widely by its sig-
naling pathway which is concerned with the growth, proliferation,
differentiation of advanced non small cell lung cancer [2,3]. It has
become a crucial molecular target for NSCLC TKIs targeted therapy
[4–6], especially for those patients who have got a certain muta-
tion in EGFR kinase domain. Gefitinib, first generation EGFR-TKI,
has significantly prolonged the progression free survival (PFS) of
patients who have got a certain mutation (EGFR 19 deletion or
L858R) in EGFR kianse domain compared with chemotherapy,
but to EGFR wild type, secondary mutation T790M, its outcomes
were not good [7–10]. So, despite its promising effect in the treat-
ment of advanced NSCLC, detection of EGFR mutation status is still
needed before treatment.
of treatment. Molecular imaging with PET tracer may provide a
noninvasive method that could be repeatedly operated and reflect
receptor status in real-time [15–19]. In view of gefitinib has shown
great response in targeted therapy of NSCLC patients who have got
a certain mutation in EGFR kinase domain, to detect EGFR status by
molecular imaging is evaluable. So, scientists have developed
many PET tracers which could specific bind to EGFR kinase domain,
such like 18F-gefitinib, 11C-gefitinib, 11C-erlotinib [20–22], but they
have a common insufficient, high lipophilicity results in too much
tracer accumulation in liver, which lead to a high background
noise, and will affect the quality of imaging pictures. Also, with
the short half-life 20 min, radionuclide 11C was not ideal and suit-
able for PET imaging, in the other hand, for 18F labeling, it need
multi steps to label the TKIs. So, we suppose that whether chemical
modification using polyethyleneglycol will reduce lipophilicity of
the tracer, lower its accumulation in liver, and improve the quality
of PET imaging pictures. Here we designed and synthesized a
p-toluene sulfonic acid (TSO) precursor IRST in multi steps, and
radiolabeled in one step. The process of IRST (compound 10)
synthesizing was shown as below (Scheme 1), and the detailed
method was described in Supplemental information.
The major method of EGFR status detection gene sequencing
analysis needs an invasive puncture, which has many limitations
[11,12]. It can not reflect EGFR status of all tumor regions due to
the heterogeneity of cancer [13,14], and repeating invasive punc-
ture will make patient suffer much more pain during the process
At the end of synthesis, the chemical purity of IRST was ana-
lyzed by HPLC (Agilent 1200) with the elunt (methanol:wa-
ter = 80:20), UV detector wavelength 254 nm, and the chemical
purity was 99.80 0.31% (Fig. 1 and Table 1).
⇑
Corresponding authors at: TOF-PET/CT/MR Center, The Fourth Hospital of
Harbin Medical University, China.
The process of radiosynthesis was performed in GE Health
E-mail addresses: sunxilin@aliyun.com (X. Sun), shenbzh@vip.sina.com
(B. Shen).
tracerlab FX-Fn synthesizer, shown in Scheme 2, and 18F-IRS was
http://dx.doi.org/10.1016/j.bmcl.2016.10.084
0960-894X/Ó 2016 Published by Elsevier Ltd.
Please cite this article in press as: Z. Xiao, et al.. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.10.084

2
Z. Xiao et al. / Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Scheme 1. The synthetic process of precursor IRST.
Fig. 1. HPLC analysis of precursor IRST.
Table 1
Chemical property of IRST.
chemistry was >98.5%. 18F-IRS is stable in both PBS and
serum, the radiochemistry of 18F-IRS incubation in PBS was
98.56 0.60% and 97.87 0.51% at 1 h, 2 h time point, respectively
(n = 3). The radiochemistry of 18F-IRS incubation in serum was
97.69 0.93% and 92.43 1.26% at 1 h, 2 h time point, respectively
(n = 3). The LogP value of 18F-IRS was À0.647 0.096 (n = 3), and
the low lipophilicity will lead to less accumulation of 18F-IRS in
liver which may be concerned with the chemical modification
using polyethyleneglycol.
Parameter
Bacterial
endotoxins
Appearance
Purity
Method
Specification
Result
LAL
<2.0 I.U./25 mg
<0.5 I.U./25 mg
Organoleptic
HPLC
Off-white to white solid
>95.0%
White solid
99.80%
763.30
ESI
Identity
LC-MS
[M+H]+, 763.25
Conformed
1H NMR
Conformed
To evaluate the pharmacokinetics of 18F-IRS, ex vivo biodistri-
bution was performed. The uptake of 18F-IRS in organs were
expressed as %ID/g (n = 3), the result was shown in Fig. 3. As the
data suggested, the highest level of accumulation of 18F-IRS in liver
was only 2.12 0.60%ID/g at 30 min post injection, which is lower
than reported before. The evidence provided by LogP value and
biodistribution assay has proved that the chemical modification
using polyethyleneglycol do decreased the lipophilicity of the
purified by HPLC (Fig. 2). The whole process was only about
30 min, the little time cost was due to the contribution of the p-
toluene sulfonic acid (TSO), which is active group that could be
substituted by halogen 18F in only one step, and also with the high
chemical yield >20%. At the end of synthesis, the radiochemistry
was determined by Thin-layer chromatography, and the radio-
Scheme 2. One-step radiolabeled of 18F-IRS.
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Z. Xiao et al. / Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
3
Fig. 2. Chromatogram from HPLC separation of 18F-IRS.
Fig. 3. Biodistribution of 18F-IRS in KM mice.
tracer, which will lead to more tracer circulation in the blood sys-
tem, and the tumor region may has more tracer accumulation. The
accumulation of 18F-IRS in kidney was as high as 6.86 2.81%ID/g,
which means 18F-IRS was mainly excreted from urinary. The
uptake of 18F-IRS in lung has reached the highest level, only
1.90 0.31%ID/g at 30 min time point, which will be good for
PET/CT imaging of NSCLC due to the low background. The activity
in blood at 240 min time point was 0.46 0.28%ID/g, which sug-
gested the clearance of 18F-IRS was rapid.
shown in Fig. 4. As the data suggested, the accumulation of
18F-IRS in HCC827 was much higher than that in H1975, H358,
H520 at all time points examined. At 2 h time point, the uptake
of 18F-IRS in HCC827 cells reached the highest level 19.07 0.70%
of total input radioactivity, which is about 17.66, 26.12 and 19.46
times higher than that in H1975, H358 and H520 cell lines. In
the cell efflux assay, 18F-IRS has shown good retention in HCC827
cells, even at 2 h time point, the accumulation of 18F-IRS in
HCC827 cells still remained 4.10 1.06% of total input radioactiv-
ity, which is much higher than H1975, H358 and H520 cells. The
cell uptake and efflux assay have proved 18F-IRS could specifically
bind to HCC827 cells, which may be concerned with EGFR 19 exon
deletion in HCC827 cells, so 18F-IRS has the potential to be used in
To test the capacity of 18F-IRS in binding mutant EGFR and
retention, cell uptake and efflux were performed in four NSCLC cell
lines, HCC827(19 exon deletion), H1975(L858R/T790M), H358
(EGFR wild type), H520(low EGFR expression). The result was
Fig. 4. Cell uptake and efflux of 18F-IRS in NSCLC cell lines.
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Z. Xiao et al. / Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
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In summary, PET/CT imaging with molecular tracer was an ideal
method to detect the EGFR status and monitor the response of TKIs
therapy. In this study, we have successfully developed a new PET
tracer 18F-IRS in one-step radio-labeling. With the chemical modi-
fication by polyethylene glycol, the lipophilicity of 18F-IRS was
decreased by detecting the LogP value, the tracer was also stable
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background. Due to the specificity of 18F-IRS to HCC827 cells, it
could be used to select the appropriate patient for TKIs therapy.
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was required.
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The costs of publication of this article were defrayed in part by
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A. Supplementary data
Supplementary data associated with this article can be found, in
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084.
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