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showed higher cytotoxicity in vitro. In particular, the clusterization of Notes and references
the nanobody further enhanced the anti-tumor effect of NDC.
1
(a) V. Chudasama, A. Maruani and S. Caddick, Nat. Chem., 2016, 8,
14–119; (b) P. Gotwals, S. Cameron, D. Cipolletta, V. Cremasco,
A. Crystal, B. Hewes, B. Mueller, S. Quaratino, C. Sabatos-Peyton and
L. Petruzzelli, Nat. Rev. Cancer, 2017, 17, 286.
H. D. Herce, D. Schumacher, A. F. Schneider, A. K. Ludwig, F. A.
Mann, M. Fillies, M.-A. Kasper, S. Reinke, E. Krause and H. Leonhardt,
Nat. Chem., 2017, 9, 762.
C. McMahon, A. S. Baier, R. Pascolutti, M. Wegrecki, S. Zheng,
J. X. Ong, S. C. Erlandson, D. Hilger, S. G. Rasmussen and
A. M. Ring, Nat. Struct. Mol. Biol., 2018, 25, 289.
The toxicity of various formulations was analyzed using a
TUNEL assay. The results confirmed that cNDC@PEG markedly
promoted the apoptosis of tumor cells (Fig. 3C). On the other
hand, cisplatin led to a continuous decrease in body weight.
NDC@PEG decreased the body weight in the first 7 days and
then increased afterward, showing decreased toxicity in com-
parison to cisplatin. The cNDC@PEG group almost reached the
level of the PBS group, showing minimal toxicity among all
treatments (Fig. 3B). This result reveals that NDCs possess
higher anti-tumor efficacy and lower systemic toxicity, while
the clustered cNDC@PEG demonstrated an improved effect
compared to the monomeric NDC@PEG.
1
2
3
4 K. R. Schmitz, A. Bagchi, R. C. Roovers, P. M. V. B. Henegouwen and
K. M. Ferguson, Structure, 2013, 21, 1214–1224.
5
(a) H. Huang, T. Wu, H. Shi, Y. Wu, H. Yang, K. Zhong, Y. Wang and
Y. Liu, Chem. Commun., 2019, 55, 5175–5178; (b) T. Fang, J. N.
Duarte, J. Ling, Z. Li, J. S. Guzman and H. L. Ploegh, Angew. Chem.,
Int. Ed., 2016, 55, 2416–2420.
D. Virant, B. Traenkle, J. Maier, P. D. Kaiser, M. Bodenh ¨o fer, C. Schmees,
I. Vojnovic, B. Pisak-Luk ´a ts, U. Endesfelder and U. Rothbauer, Nat.
Commun., 2018, 9, 930.
6
In addition to tumor targeting, anti-EGFR Nb can also
2
3
downregulate the EGFR level by inducing internalization,
1
8a,24
7 B. J. Stenton, B. L. Oliveira, M. J. Matos, L. Sinatra and G. J. Bernardes,
and further blocking the EGFR signalling pathway.
This
Chem. Sci., 2018, 9, 4185–4189.
F. Zhang, H. Wei, X. Wang, Y. Bai, P. Wang, J. Wu, X. Jiang, Y. Wang,
H. Cai and T. Xu, Cell Discovery, 2017, 3, 17004.
E. Pardon, C. Betti, T. Laeremans, F. Chevillard, K. Guillemyn,
P. Kolb, S. Ballet and J. Steyaert, Angew. Chem., Int. Ed., 2018, 57,
alteration can suppress the proliferation and metastasis of tumors
8
9
18a
since the EGFR signal is important for these processes. Here the
effect of NDC on the EGFR level has been measured using
immunohistochemical analysis. The result confirmed that a high
5292–5295.
expression of EGFR occurred in the region of the A431(++) 10 (a) X. Wang, X. Wang, S. Jin, N. Muhammad and Z. Guo, Chem. Rev.,
2
018, 119, 1138–1192; (b) S. Dhar, N. Kolishetti, S. J. Lippard and
tumor (Fig. S16, ESI†). The treatment of cisplatin did not change
the EGFR level. Interestingly, a clearly reduced EGFR level was
observed with the treatment of NDCs. A more significant change
was observed with the treatment of cNDC@PEG, which suppressed
the EGFR expression level down to 29% relative to the PBS group.
In conclusion, a tetravalent NDC was designed using den-
drimeric poly-lysine as a cross-linking agent. The site-specific
protein PEGylation was achieved by the mTGase mediated
O. C. Farokhzad, Proc. Natl. Acad. Sci. U. S. A., 2011, 108, 1850–1855.
1 H. Wang, M. Johnston and R. D. Mitra, Genome Res., 2007, 17,
1202–1209.
2 (a) Q. Cheng, H. Shi, H. Wang, Y. Min, J. Wang and Y. Liu, Chem.
Commun., 2014, 50, 7427–7430; (b) N. Adarsh, C. Frias, T. P.
Lohidakshan, J. Lorenzo, F. Novio, J. Garcia-Pardo and D. Ruiz-
Molina, Chem. Eng. J., 2018, 340, 94–102.
3 L. O. Gainkam, L. Huang, V. Caveliers, M. Keyaerts, S. Hernot,
I. Vaneycken, C. Vanhove, H. Revets, P. De Baetselier and
T. Lahoutte, J. Nucl. Med., 2008, 49, 788–795.
1
1
1
ligation, and the Pt(IV) prodrug was site-specifically conjugated 14 T. Wu, H. Huang, Y. Sheng, H. Shi, Y. Min and Y. Liu, J. Mater. Chem. B,
2
018, 6, 1011–1017.
by an amide reaction. This clustered cNDC@PEG demonstrated
excellent targeting to EGFR positive cancer cells. Interestingly,
the clusterization of Nbs further increased the drug retention
time in vivo, and enhanced the drug efficacy and reduced the
systemic side effects. Meanwhile, the clustered cNDC@PEG can
efficiently penetrate tumor tissues. This work provides a novel
clusterization strategy to design multi-valent NDCs by taking
advantage of the unique properties of Nbs and the enzyme-
1
5 H. P. Patil, D. Freches, L. Karmani, G. A. Duncan, B. Ucakar,
J. S. Suk, J. Hanes, B. Gallez and R. Vanbever, J. Controlled Release,
2018, 272, 62–71.
6 S. Hu, H. Wang, A. A. Knisely, S. Reddy, D. Kovacevic, Z. Liu and
S. M. Hoffman, Genetica, 2008, 133, 215–226.
7 (a) D. Rusnak, K. Alligood, R. Mullin, G. Spehar, C. Arenas-Elliott,
A. M. Martin, Y. Degenhardt, S. Rudolph, T. Haws and B. Hudson-
Curtis, Cell Proliferation, 2007, 40, 580–594; (b) S. Ganta, A. Singh,
N. R. Patel, J. Cacaccio, Y. H. Rawal, B. J. Davis, M. M. Amiji and
T. P. Coleman, Pharm. Res., 2014, 31, 2490–2502.
1
1
mediated protein ligation. This new generation of NDCs 18 (a) R. C. Roovers, M. J. W. D. Vosjan, T. Laeremans, R. el Khoulati,
R. C. G. de Bruin, K. M. Ferguson, A. J. Verkleij, G. A. M. S. van
showed remarkable targetability, higher antitumor efficacy
Dongen and P. M. P. V. E. Henegouwen, Int. J. Cancer, 2011, 129,
and low systemic side-effects in vivo.
2013–2024; (b) K. Uchibori, N. Inase, M. Araki, M. Kamada, S. Sato,
This work was supported by the National Science Foundation
of China (21877103 and 51873218), the National Key R&D
Program of China (2017YFA0505400) and the Major Program
Y. Okuno, N. Fujita and R. Katayama, Nat. Commun., 2017, 8, 14768.
9 H. X. Lu and M. H. Stenzel, Small, 2018, 14, 1702858.
0 J. Friedrich, C. Seidel, R. Ebner and L. A. Kunz-Schughart, Nat.
Protoc., 2009, 4, 309.
1
2
of Development Foundation of Hefei Center for Physical Science 21 (a) A. Rondon, N. Ty, J. B. Bequignat, M. Quintana, A. Briat,
T. Witkowski, B. Bouchon, C. Boucheix, E. Miot-Noirault, J. P.
and Technology (2018ZYFX004). A portion of this work was
Pouget, J. M. Chezal, I. Navarro-Teulon, E. Moreau and F. Degoul,
performed on the Steady High Magnetic Field Facilities, High
Magnetic Field Laboratory, CAS.
Sci. Rep., 2017, 7, 14918; (b) H. T. Pan, J. Y. Liu, W. T. Deng,
J. Y. Xing, Q. Li and Z. Wang, Int. J. Nanomed., 2018, 13, 3189–3201.
2 B. Yang, S. I. Lim, J. C. Kim, G. Tae and I. Kwon, Biomacromolecules,
2
2
2016, 17, 1811–1817.
3 S. Oliveira, R. M. Schiffelers, J. van der Veeken, R. van der Meel,
R. Vongpromek, P. M. P. V. E. Henegouwen, G. Storm and R. C.
Roovers, J. Controlled Release, 2010, 145, 165–175.
Conflicts of interest
2
4 A. Kol, A. T. van Scheltinga, M. Pool, C. Gerdes, E. de Vries and S. de
There are no conflicts to declare.
Jong, Oncotarget, 2017, 8, 45432–45446.
Chem. Commun.
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