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Phosphate‐Based Self‐Immolative Linkers for Tuneable Double Cargo Release

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Chemistry - A European Journal
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Petr Šimon
Petr Šimon
Petr Šimon
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Markéta Tichotová
Markéta Tichotová
Markéta Tichotová
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María García Gallardo
María García Gallardo
María García Gallardo
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Eliška Procházková at Czech Academy of Sciences, Institute of Organic Chemistry and Biochemistry
Eliška Procházková
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Abstract and Figures

Phosphorus‐based self‐immolative (SI) linkers offer a wide range of applications, such as smart materials and drug‐delivery systems. Phosphorus SI linkers are ideal candidates for double‐cargo delivery platforms because they have a higher valency than carbon. A series of substituted phosphate linkers was designed for releasing two phenolic cargos through SI followed by chemical hydrolysis. Suitable modifications of the lactate spacer increased the cargo release rate significantly, from 1 day to 2 hours or 5 minutes, as shown for linkers containing p‐fluoro phenol. In turn, double cargo linkers bearing p‐methyl phenol released their cargo more slowly (4 days, 4 hours, and 15 minutes) than their p‐fluoro analogues. The α‐hydroxyisobutyrate linker released both cargos in 25 minutes. Our study expands the current portfolio of SI constructs by providing a double cargo delivery option, which is crucial to develop universal SI platforms.
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Phosphate-Based Self-Immolative Linkers for Tuneable
Double Cargo Release
Petr Šimon+,[a] Markéta Tichotová+,[a, b] María García Gallardo,[a] Eliška Procházková,*[b] and
Ondřej Baszczyňski*[a, b]
We dedicate this paper to the memory of Dr. John C. Martin (1951–2021), head of Gilead Sciences, to commemorate his achievements in
the development of anti-HIV drugs (acyclic nucleoside phosphonates, “Holy Trinity”: A. Holý, E. De Clercq, J. C. Martin).
Abstract: Phosphorus-based self-immolative (SI) linkers offer a
wide range of applications, such as smart materials and drug-
delivery systems. Phosphorus SI linkers are ideal candidates
for double-cargo delivery platforms because they have a
higher valency than carbon. A series of substituted phosphate
linkers was designed for releasing two phenolic cargos
through SI followed by chemical hydrolysis. Suitable modifica-
tions of the lactate spacer increased the cargo release rate
significantly, from 1 day to 2 hours or 5 minutes, as shown for
linkers containing p-fluoro phenol. In turn, double cargo
linkers bearing p-methyl phenol released their cargo more
slowly (4 days, 4 hours, and 15 minutes) than their p-fluoro
analogues. The α-hydroxyisobutyrate linker released both
cargos in 25 minutes. Our study expands the current portfolio
of SI constructs by providing a double cargo delivery option,
which is crucial to develop universal SI platforms.
Introduction
Self-immolative (SI) linkers are chemical constructs subjected to
irreversible fragmentation triggered by external stimuli.[1] After
external activation, SI linkers disassemble (e.g., by cyclisation or
electronic cascade), thereby releasing a leaving group (cargo).
Triggered cargo release can be used in drug delivery[2]
(prodrugs,[3] antibody-drug conjugates,[4] and chemosensors[5]),
smart materials[6] (stimuli-responsive SI dendrimers[7] and
polymers[8]), or in vivo cell labelling,[9] thus highlighting the
wide range of applications of SI linkers. In turn, different
structural motifs, including carbamate[3,6,10] or phosphate,[11] can
be used to attach chemically variable cargos to the SI linker.
Phosphorus-based SI linkers can offer higher versatility than
their carbamate analogues because they allow us to attach an
additional substituent (a second cargo), which significantly
broadens their potential applications. For example, a double
cargo linker can be used to simultaneously tether a drug
(warhead, first cargo) and a reporter molecule (chromogenic
molecule, second cargo).[12] Systems combining double (multi)
cargo release have been studied[13] to pursue effective cancer
chemotherapy using complementary drug combination,[14] anti-
biotic drug resistance[15] or novel fluorescent reporters/signal
amplification chemosensors.[5a,16] In addition, phosphorus linkers
enable us to monitor the reaction pathway by 31P NMR
spectroscopy in real time,[17] providing detailed structural and
kinetic information, and to even detect the cyclic intermediates
in some cases,[18] confirming cargo release by self-immolation,
more specifically intramolecular cyclisation, and not by chemical
hydrolysis.
Self-immolation, and consequently the rate of the cargo
release reaction, can be tuned up by varying substituents on
the phosphorus atom and is mainly directed by two effects: 1)
the Thorpe-Ingold[19] effect and 2) the pKaeffect of the leaving
groups. The Thorpe-Ingold effect refers to the spacer – sterically
demanding substituents in the α-position accelerate intra-
molecular cyclisation.[20] Conversely, the pKaeffect is related to
the cargo – the more acidic the cargo is (lower pKa), the faster
the cargo will be released upon external activation.[18] Fast
cargo release systems are usually designed for biological
applications. However, slow SI may also be advantageous when
constructing slowly reacting (decomposing) polymers[8,21] or
developing prodrugs with delayed cargo release (e.g.,
antibiotics).[22] Notwithstanding these advances and the poten-
tial of SI linkers, no phosphate-based, double cargo release
system has been reported yet.
Considering the above, we designed a novel set of
phosphate linkers bearing a lactate spacer (Figure 1) to attach
two cargos (double cargo option). We also fine-tuned the
release of both cargos by introducing a suitable modification in
the lactate spacer and by adjusting the pKaof the leaving
groups. SI was triggered by UV light (365 nm) and monitored
by 31P NMR spectroscopy. The structures of intermediates and
products were determined in situ by combining 13C and 31P
[a] Dr. P. Šimon,+M. Tichotová,+M. García Gallardo, Dr. O. Baszczyňski
Faculty of Science, Charles University
Prague 128 43 (Czech Republic)
E-mail: baszczyo@natur.cuni.cz
[b] M. Tichotová,+Dr. E. Procházková, Dr. O. Baszczyňski
Institute of Organic Chemistry and Biochemistry
The Czech Academy of Sciences
Prague 166 10 (Czech Republic)
E-mail: prochazkova@uochb.cas.cz
[+]These authors contributed equally to this work.
Supporting information for this article is available on the WWW under
https://doi.org/10.1002/chem.202101805
Chemistry—A European Journal
Full Paper
doi.org/10.1002/chem.202101805
12763Chem. Eur. J. 2021,27, 12763 –12775 © 2021 Wiley-VCH GmbH
Wiley VCH Mittwoch, 25.08.2021
2150 / 210680 [S. 12763/12775] 1
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