Design, synthesis and biological evaluation of deuterated Vismodegib for improving pharmacokinetic properties

Article abstract of DOI:10.1016/j.bmcl.2018.06.025

Vismodegib is an oral and high selective hedgehog (Hh) inhibitor used for the treatment of basal cell carcinoma (BCC). In this work, analogs of Vismodegib with deuterium-for-hydrogen replacement at certain metabolically active sites were prepared and found to have a better pharmacokinetic properties in mice. In particular, deuterated compound SKLB-C2211 obviously altered the blood circulation behavior compared to its prototype, which was demonstrated by significantly prolonged blood circulation half-life time (t_1/2) and increased AUC_0→∞. These results suggested SKLB-C2211 had the potential to be a long-acting inhibitor against Hh signaling pathway, and laid the foundation for the further research of its druggability.

Full text of DOI:10.1016/j.bmcl.2018.06.025

Accepted Manuscript
Design, synthesis and biological evaluation of deuterated Vismodegib for im-
proving pharmacokinetic properties
Fangying Wang, Hongxia Jiang, Yufang Deng, Jiang Yu, Miao Zhan, Lifeng
Zhao, Yuanwei Chen
PII:
S0960-894X(18)30514-6
https://doi.org/10.1016/j.bmcl.2018.06.025
BMCL 25906
DOI:
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
10 April 2018
27 May 2018
13 June 2018
Please cite this article as: Wang, F., Jiang, H., Deng, Y., Yu, J., Zhan, M., Zhao, L., Chen, Y., Design, synthesis
and biological evaluation of deuterated Vismodegib for improving pharmacokinetic properties, Bioorganic &
Medicinal Chemistry Letters (2018), doi: https://doi.org/10.1016/j.bmcl.2018.06.025
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers
we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and
review of the resulting proof before it is published in its final form. Please note that during the production process
errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Graphical Abstract
To create your abstract, type over the instructions in the template box below.
F‘’onts or abstract dimensions should not be changed or altered.
Design, synthesis and biological evaluation of
deuterated Vismodegib for improving
pharmacokinetic properties
Leave this area blank for abstract info.
Fangying Wanga† , Hongxia Jianga† , Yufang Denga , Jiang Yua , Miao Zhand* , Lifeng Zhaoc and Yuanwei
Chena,b

Bioorganic & Medicinal Chemistry Letters
Design, synthesis and biological evaluation of deuterated Vismodegib for
improving pharmacokinetic properties
Fangying Wanga† , Hongxia Jianga† , Yufang Denga , Jiang Yua , Miao Zhand* , Lifeng Zhaoc and Yuanwei
Chena,b
a. State Key Laboratory of Biotherapy and Cancer Center/Collaborative Innovation Center of Biotherapy, West China Hospital, West China Medical School of
Sichuan University, Chengdu 610041, China.
b. Hinova Pharmaceuticals Inc., Suite 402, Building B, #5 South KeYuan Road, Chengdu, 610041, China.
c. Chengdu University, Sichuan Industrial Institute of Antibiotics, Chengdu 610052, China.
d. Frontier Institute of Science and Technology, Xi’an Jiaotong University, Xi’an, 710054, China
ARTICLE INFO
ABSTRACT
Vismodegib is an oral and high selective hedgehog (Hh) inhibitor used for the treatment of basal
cell carcinoma (BCC). In this work, analogs of Vismodegib with deuterium-for-hydrogen
replacement at certain metabolically active sites were prepared and found to have a better
pharmacokinetic properties in mice. In particular, deuterated compound SKLB-C2211 obviously
altered the blood circulation behavior compared to its prototype, which was demonstrated by
significantly prolonged blood circulation half-life time (t1/2) and increased AUC0→∞. These
results suggested SKLB-C2211 had the potential to be a long-acting inhibitor against Hh
signaling pathway, and laid the foundation for the further research of its druggability.
2009 Elsevier Ltd. All rights reserved.
Article history:
Received
Revised
Accepted
Available online
Keywords:
Vismodegib
Hedgehog signaling pathway
Basal cell carcinoma
Deuterated drug
Deuterium-hydrogen replacement
Pharmacokinetic properties
 Corresponding author. Tel.: +86 186 0283 1330; fax: +86 2885058465; e-mail: ywchen@scu.edu.cn.
 Corresponding author. Tel.: +86 158 8241 1095; fax: +86 2885058465; e-mail: lifengzhao@scu.edu.cn; mzhan90@xjtu.edu.cn
†These authors contributed equally

Hedgehog (Hh) signaling pathway is highly conservative
which plays a very important role in cell differentiation,
development of embryo, organ formation, and keeping internal
environment of mature organs.[1-5] This signaling pathway is
mainly composed of Hh protein, two transmembrane receptors
Patched (Ptc) and Smoothened (Smo), and downstream
transcription factors (Gli family).[3] Under specific time or
position-dependent stimuli, cells secrete the Hh family ligands
(sonic, desert, and indian),[4] whose binding associates with Ptc
on target cells. The role of Ptc, in the absence of Hh ligands, is to
inhibit the activity of Smo, thus blocking Hh signal transduction.
Binding of Hh proteins to Ptc relieves this inhibition and initiates
activation of Smo. The increase in Smo activity leads to an
augment in activated forms of Gli, which serves to regulate the
expression of Hh target genes.[5,6,7] In adults the Hh pathway is
mainly quiescent, activation of the Hh pathway has been
implicated in the development of cancers in various organs,
including brain, liver, prostate, mammary gland, lung and
skin.[8,9,10,11] Basal cell carcinoma, the most common form of
cancerous malignancy, has the closest association with Hh
signaling pathway.[12]
colorectal cancer, small-cell lung cancer, advanced stomach
cancer,
pancreatic
cancer,
medulloblastoma
and
chondrosarcoma.[15] This drug acts as a cyclopamine-competitive
antagonist of the Smo, Smo inhibition causes the transcription
factors Gli1 and Gli2 to remain inactive, which prevents the
expression of tumor mediating genes within the Hh pathway,
which is pathogenetically relevant in more than 90% BCC[13,16]
Preclinical
.
pharmacokinetic
studies[17,18,19]
shows
that
Vismodegib was metabolically stable in mouse, rat, dog, and
human hepatocytes and had a more rapid turnover in monkey
hepatocytes. The metabolites were identified by using various
LC-MS/MS techniques.[19] The major metabolites and the
metabolic pathways were illustrated in Fig. 1, three oxidative
metabolites (M1, M2 and M3)[17,18] were formed by oxidation of
the pyridine or the middle benzene ring. Two glucuronide
conjugates (M4, M5) and an oxidative sulfate metabolite (M6)
were also produced subsequently. In addition, three major
pyridine ring-opened metabolites (M7, M8, and M9)[19] have
been found.
On the basis of this information, we considered that the
blockade of metabolically active sites may slow down the
metabolism rate. Replacing a hydrogen atom with fluorine has
been extensively used to achieve this goal. Recently, the use of
Vismodegib (GDC-0449) is the first oral and high selective
Hh signaling pathway targeting agent for the treatment of
BCC,[14] and it is also undergoing clinical trials for metastatic
Fig 1. The structures of metabolites and major metabolic pathways of Vismodegib. UGT, UDP-glucuronosyltransferase; Gluc,
glucuronic acid.
Fig 2. Recent examples of deuterated drug analogues. Deuterium used as a bioisostere of hydrogen to improve pharmacokinetic
properties.

Fig 3. Chemical structure of deuterated analogues of Vismodegib.
Scheme 1. Synthesis of intermediates 3 and 4. Reagents and conditions: (a) HCl/H2O, NaNO2, KI, 0 ℃to rt; (b) Fe, AcOH, EtOH, 60 ℃
to 70 ℃to 23 ℃; (c) HCl, D2O, microwave, 180 ℃.
Scheme 2. Synthesis of intermediates 7a and 7b. Reagents and conditions: (d) HBr, Br2, NaNO2, -2 ℃; (e) n-BuLi, THF, -78 ℃,
chlorotributyltin, -78 ℃to rt.
Scheme 3. Synthesis of Vismodegib analogs. Reagents and conditions: (f) SOCl2, reflux; (g) DMAc, rt; (h) Pd(PPh3)2Cl2, NMP,
microwave, 130 ℃.
deuterium has also served a similar effect.[20,21,22] In fact, well-
known applications using deuterium exist within every
subdiscipline in pharmaceutical discovery and development.[23,24]
Because of the isotope effect, deuterium-hydrogen replacement
can have a major impact on improving pharmacokinetic
properties (ADMET),[25,26] mainly were: (1) Reducing the
metabolic rate, thus gaining a lower drug dose and improving the
tolerance of drugs. (2) Decreasing the clearance rate and
increasing the biological half-life of the drugs. These clear
benefits result from a slower rate of C-D bond cleavage
compared to C-H bond, making the deuterium-labeled drugs
more resistant to chemical or enzymatic degradation.[27] As a
result, many pharmacists devoted to the research of deuterium-
hydrogen replacement at metabolically active sites expecting to
achieve better drug candidates. Clinical trials of some known
drugs have been initiated,[28,29,30] including D-venlafaxine (SD-
254), D-atazanavir (CTP-518),[25] D1-telaprevir, D4-paroxetine,
D6-nifedipine.[22] On April 3rd, 2017, FDA approved the first
deuterated drug-Deutetrabenazine.[31] (Fig. 2)
studies of their pharmacokinetic profiles. Therefore, we designed
deuterated analogues of Vismodegib on the pyridine and/or the
middle benzene ring, namely SKLB-C2209, SKLB-C2210,
SKLB-C2211 (Fig. 3).
The designed deuterated analogues together with their
intermediates (compounds 3, 4, 7) were prepared according to
Schemes 1-3.
The synthesis of key intermediates 3 and 4 commenced from
commercially available 2-chloro-5-nitroaniline (1). As illustrated
in Scheme 1, 1 was first transformed into 2 via a diazotizating
iodination followed by a reduction reaction of nitro group using
Iron to obtain 3. Then 3 was efficiently deuterated at the ortho
position of amino group in the presence of 1 equiv HCl in D2O
under microwave irradiation at 180 ℃, receiving compound 4
with nearly total deuterium incorporation.
As described in Scheme 2, intermediate 7a was obtained by a
two-step procedure. Bromo-substitution of 2-aminepyridine (5a)
with HBr and Br2 afforded compound 6a, followed by
nucleophilic substitution with chlorotributyltin to give compound
7a. Corresponding deuterated compounds 5b, 6b, and 7b were
Here we wish to report the deuterium-for-hydrogen
replacement at the metabolically active sites of Vismodegib to
generate the corresponding deuterium-labeled drugs and the

Table 1. Pharmacokinetic parameters after oral compound administration to mice
GDC-0449
SKLB-C2209
SKLB-C2210
SKLB-C2211
Dose (mg/kg)
tmax (h)
5
5
5
5
1
1
1
1
t1/2 (h)
5.617
854
2.556
972.3
6106.538
6113.538
2.82
8.547
Cmax (ng/mL)
AUC0→t (μg/L*h)
AUC0→∞ (μg/L*h)
1606.7
6284.838
8290.712
1870
4049.8
7657.678
6511.863
17147.309
prepared from the original material 5a, which was deuterated
using a Pd/C-H2-D2O system, details are described in Supporting
Information.
Fig 4. Mean plasma concentration of 5mg/kg Vismodegib
analogs after oral administration in mice (n=3/group)
SKLB-C2211) in female CD-1 mouse were performed to
evaluate the alteration of blood circulation behaviors by
deuterium replacement at metabolically active sites of
Vismodegib. (Fig. 4) Vismodegib and its analogs were
suspended in vehicle (0.5% methyl cellulose with 0.2% Tween
80 in distilled water) and the suspension was administered orally
to the mice within a day of preparation with dose of 5 mg/kg.
Blood samples were taken at 0.25, 0.5, 1.0, 3.0, 6.0 and 24 h, and
drug plasma concentration were quantified by LC-MS/MS. As
shown in Table 1, deuterated compounds generally exhibited
better PK properties as compared to their prototype Vismodegib
at the same dose. Among all deuterated compounds, SKLB-
C2211 displayed the best than others. More specifically, SKLB-
C2211 exhibited an almost 2.24-fold increase in AUC0→∞
(μg/L*h) when compared with Vismodegib (17147.309 vs.
7657.678). Meanwhile, the half-time (t1/2) of SKLB-C2211 was
also increased to 8.547 hours by comparison with Vismodegib
(5.617 hours), indicating that SKLB-C2211 acted prolonged
blood circulation time and longer duration of action. In addition,
its peak concentration Cmax (1870 ng/mL) was observed higher
than Vismodegib (854 ng/mL), which suggested the dosage of
SKLB-C2211 may be decreased when used in clinic. By contrast,
the similar peak time (tmax) indicated that SKLB-C2211 did not
change the time of taking effect. These results implied that
deuterated compound SKLB-C2211 successfully improved the
Deuterium-labeled analogs SKLB-C2209, SKLB-C2210, and
SKLB-C2211 were synthesized according to Scheme 3.
Compound 8 was first transformed into 9 via reaction with
thionyl chloride followed by an acylation reaction using
compound 3 or 4. The target compounds were obtained via Still
Coupling reaction using 10a / 10b with 7a/ 7b.
The in vivo pharmacokinetic profile studies of Vismodegib
and its deuterated analogues (SKLB-C2209, SKLB-C2210 and
blood circulation behavior compared to its prototype
Vismodegib, and had the potential to be a long-acting inhibitor
against Hh pathway with decreased dosage. These characters are
important for clinical use because lowered dosage administration
can significantly improve the compliance of patients and reduce
side effects.
3. Ruiz-Gómez, A.; Molnar, C.; Holguín, H.; Mayor, F.; de Celis, J. F.
Biochim. Biophys. Acta. 2007, 1768, 901.
4. Rubin, L. L.; de Sauvage, F. J. Nat. Rev. Drug Discov. 2006, 5,
1026.
5. Varjosalo, M.; Taipale, J. Genes. Dev. 2008, 22, 2454.
6. Ma, G.; Xiao, Y.; He, L. J. Genet. Genomics. 2008, 35, 129.
7. Corbit, K. C.; Aanstad, P.; Singla, V.; Norman, A. R.; Stainier, D.
Y.; Reiter, J. F. Nature 2005, 437, 1018.
8. Sahebjam, S.; Siu, L. L.; Razak, A. A. Oncologist 2012, 17, 1090.
9. Ma, HY.; Zhang, F.; Li, J.; Mo, M. L.; Cheng, Z.; Liu, L.; Zhou, H.
M.; Sheng, Q. Oncol. Lett. 2011, 2, 1291.
10. Riobo, N. A. Curr. Opin. Pharmacol. 2012, 12, 736.
11. Pan, J. Y.; Zhou, S. H. Pharmazie 2012, 67, 475.
12. Wong, H.; Alicke, B.; West, K. A.; Pacheco, P.; La, H.; Januario,
T.; Yauch, R.; de Sauvage, F.; Gould, S. E. Clin. Cancer Res. 2011,
17, 4682.
In conclusion, compound SKLB-C2211 with deuterium-for-
hydrogen replacement in a metabolically active site significantly
exhibited better pharmacokinetics profiles as compared to its
prototype Vismodegib at the same dose and had a better plasma
concentration which could prolong its half-life and improve the
potency with a lower dosage. Thus, SKLB-C2211 may be a novel
long-acting inhibitor of Hh signaling pathway.
13. LoRusso, P. M.; Rudin, C. M.; Borad, M. J.; Vernillet, L.;
Darbonne, W. C.; Mackey, H.; Dimartino, J. F.; de Sauvage, F.;
Low, J. A.; VonHoff, D. D. J. Clin. Oncol. 2008, 26, 3516.
14. Geeraert, P.; Williams, J. S.; Brownell, I. J. Drugs. Dermatol. 2013,
12, 519.
15. Rudin, C. M.; Hann, C. L.; Laterra, J.; Yauch, R. L.; Callahan, C.
A.; Fu, L.; Holcomb, T.; Stinson, J.; Gould, S. E.; Coleman, B.;
LoRusso, P. M.; VonHoff, D. D.; de Sauvage, F. J.; Low, J. A. N.
Engl. J. Med. 2009, 361, 1173.
Acknowledgments
This work was supported by the National Natural Science
Foundation of China (No.81472418).
References and notes
1. Ruiz, i. Altaba. A.; Sánchez, P.; Dahmane, N. Nat. Rev. Cancer
2002, 2, 361.
2. Lum, L.; Beachy, P. A. Science 2004, 304, 1755.
16. Robarge, K. D.; Brunton, S. A.; Castanedo, G. M.; Cui, Y.; Dina,
M. S.; Goldsmith, R.; Gould, S. E.; Guichert, O.; Gunzner, J.
L.; Halladay, J.; Jia, W.; Khojasteh, C.; Koehler, M. F.; Kotkow,

K.; La, H.; Lalonde, R. L.; Lau, K.; Lee, L.; Marshall, D.; Marsters,
J. C. Jr.; Murray, L. J.; Qian, C.; Rubin, L. L.; Salphati, L.; Stanley,
M. S.; Stibbard, J. H.; Sutherlin, D. P.; Ubhayaker, S.; Wang, S.;
Wong, S.; Xie, M. Bioorg. Med. Chem. Lett. 2009, 19, 5576.
17. Wong, H.; Chen, J. Z.; Chou, B.; Halladay, J. S.; Kenny, J. R.; La,
H.; Marsters, J. C. Jr.; Plise, E.; Rudewicz, P. J.; Robarge, K.; Shin,
Y.; Wong, S.; Zhang, C.; Khojasteh, S. C. Xenobiotica 2009, 39,
850.
18. Wong, H.; Theil, F. P.; Cui, Y.; Marsters, J. C. Jr.; Khojasteh, S. C.;
Vernillet, L.; La, H.; Song, X.; Wang, H.; Morinello, E. J.; Deng,
Y.; Hop, C. E. Drug. Metab. Dispos. 2010, 38, 1029.
19. Yue, Q.; Chen, Y. H.; Mulder, T.; Deese, A.; Takahashi, R.;
Rudewicz, P. J.; Reynolds, M.; Solon, E.; Hop, C. E.; Wong, H.;
Khojasteh, S. C. Drug. Metab. Dispos. 2011, 39, 952.
20. Meanwell, N. A. J. Med. Chem. 2011, 54, 2529.
21. Zhu, Y.; Zhou, J.; Jiao, B. ACS. Med. Chem. Lett. 2013, 4, 349.
22. Gant, T. G. J. Med. Chem. 2014, 57, 3595.
23. Xu, G.; Lv, B.; Roberge, J. Y.; Xu, B.; Du, J.; Dong, J.; Chen, Y.;
Peng, K.; Zhang, L.; Tang, X.; Feng, Y.; Xu, M.; Fu, W.; Zhang,
W.; Zhu, L.; Deng, Z.; Sheng, Z.; Welihinda, A.; Sun, X. J. Med.
Chem. 2014, 57, 1236.
25. Harbeson, S. L.; Tung, R. D. Annu. Rep. Med. Chem. 2011, 46,
403.
26. Katsnelson, A. Nat. Med. 2013, 19, 656.
27. Butear, K. C. J. High. Tech. Law. 2010, 10.
28. Yarnell, A. Chem. Eng. News. 2009, 87, 36.
29. Frank, S.; Moritz, H.; Rossen, K.; Rudolf-Giesbert, A. Arzneim.
Forsch. 2006, 56, 295.
30. Szostak, M.; Spain, M.; Procter, D. J. Org. Lett. 2014, 16, 5052.
31. Heo, Y. A.; Scott. L. J. Drugs. 2017, 77, 1857.
Supplementary Material
Supplementary material that may be helpful in the review
process should be prepared and provided as a separate electronic
file. That file can then be transformed into PDF format and
submitted along with the manuscript and graphic files to the
appropriate editorial office.
Click here to remove instruction text...
24. Sanderson, K. Nature 2009, 458, 269.

Highlights:



SKLB-C2211 has prolonged blood circulation half-
life time (t1/2) and increased AUC.
Three deuterated analogues produced greater AUC0
than Vismodegib.
→
t
SKLB-C2211 is potential to be a long-acting inhibitor
against Hh signaling pathway.