Chemical Reactions in Living Systems

Abstract

The term "in vivo ("in the living") chemistry" refers to chemical reactions that take place in a complex living system such as cells, tissue, body liquids, or even in an entire organism. In contrast, reactions that occur generally outside living organisms in an artificial environment (e.g., in a test tube) are referred to as in vitro. Over the past decades, significant contributions have been made in this rapidly growing field of in vivo chemistry, but it is still not fully understood, which transformations proceed efficiently without the formation of by-products or how product formation in such complex environments can be characterized. Potential applications can be imagined that synthesize drug molecules directly within the cell or confer new cellular functions through controlled chemical transformations that will improve the understanding of living systems and develop new therapeutic strategies. The guiding principles of this contribution are twofold: 1) Which chemical reactions can be translated from the laboratory to the living system? 2) Which characterization methods are suitable for studying reactions and structure formation in complex living environments?

Keywords: bioconjugation; cellular compartments; click-chemistry; in vivo chemistry; monitoring reactions.

Figure 1

A) Overview of chemical reactions for controlled bond cleavage (left), covalent bond formation (middle), and formation and cleavage of several bonds by, that is, cascade reactions (right) proceeding in living organisms. B) Overview of different living systems from cells to large organisms used for “in vivo chemistry”, with increasing size and complexity.

Figure 2

A) Schematic illustration of a chemoselective reaction product. B) Simulation of second‐order reactions between 100 µm reactants (A + B) yielding product (C) as a function of the reaction rate constant. Rate constants of current chemoselective ligation reactions typically range from 10⁻³ to 10² m ⁻¹ s⁻¹. Reproduced with permission.^{[ 27 ]} Copyright 2008, American Chemical Society.

Figure 3

Schematic illustration of compartments in eukaryotic cells, specific chemical triggers of these compartments, and representative examples of small molecule targeting groups.

Figure 4

Concept of cleavable linkers by chemical triggers: A) Schematic overview of linker cleavage for selective release. B) Imine‐linkages, hydrazones and oximes, ketals, disulfides, boronate esters, and salicylhydroxamates.

Figure 5

Selected examples of prodrugs that are cleaved by chemical triggers (reactions) in cells.^{[ 63 ]}

Figure 6

Concept of covalent bioconjugation reactions: A) Schematic overview of covalent bond formation from reactive but also stable functional groups. B) Traceless bioconjugation reactions that do not require any additive or catalyst (Staudinger ligation, strain‐promoted azide‐alkyne cycloaddition (SPAAC), inverse electron‐demand Diels–Alder reaction (IEDDA)). C) Transition metal‐catalyzed bioconjugation reactions (copper(I)‐catalyzed azide‐alkyne cycloaddition (CuAAC), palladium‐catalyzed Suzuki‐Miyaura cross‐coupling, gold‐catalyzed amide bond formation).

Figure 7

Strain‐promoted click chemistry in mice. A) Mice were injected with N‐azidoacetylmannosamine‐tetraacylated (Ac4ManNAz) to allow for metabolic labeling of glycans with SiaNAz. They were then injected with a cyclooctyne‐FLAG conjugate for in vivo covalent labeling of azido glycans. B) Different FLAG conjugates used. Reproduced with permission.^{[ 119 ]} Copyright 2010, The Authors, published by National Academy of Sciences.

Figure 8

Pretargeting of SKBR3 cells with norbornene and tetramethylrhodamine co‐labeled trastuzumab, followed by tagging the live cells with tetrazine‐VT680. A) Rhodamine channel. B) Near‐IR channel (tetrazine‐VT680). C) Merged image of A and B. Adapted with permission.^{[ 127 ]} Copyright 2008, American Chemical Society.

Figure 9

General scheme of tumor pretargeting by using the inverse‐electron‐demand Diels–Alder reaction. Small animal single photon emission computed tomography of live mice bearing coloncarcinoma xenografts. Reproduced with permission.^{[ 128 ]} Copyright 2010, Wiley‐VCH.

Figure 10

Pd⁰‐mediated Suzuki–Miyaura cross‐coupling within HeLa cells. Pd⁰‐catalyzed intracellular cross‐coupling generates the mitochondria‐localized fluorescent compound. Reproduced with permission.^{[ 129 ]} Copyright 2011, Springer Nature.

Figure 11

Pd‐catalyzed activation and synthesis of two anticancer agents. Simultaneous decaging of the alkyne and Suzuki–Miyaura cross‐coupling reaction of aryl iodide and aryl boronic acid. Reproduced with permission.^{[ 135 ]} Copyright 2017, Wiley‐VCH.

Figure 12

Organ‐selective accumulation within live mice directed by disialo‐ and galactosyl‐linked glycoalbumins. Preparation of glycoalbumins as “transition‐metal carriers” to produce Glyco‐Au complexes. Glyco‐Au (Sia) and Glyco‐Au (Gal) were synthesized with (2‐6)‐disialoglycoalbumin and galactosylglycoalbumin, respectively. General scheme for liver‐ and intestine‐selective in vivo fluorescence labeling by Au(III)‐catalyzed amide bond formation between propargyl ester‐based imaging probes and surface amino groups of targeted tissues. Reproduced with permission.^{[ 136 ]} Copyright 2017, Wiley‐VCH.

Figure 13

A) Schematic overview of Intracellular co‐assembly of peptides; B) Chemical reactions leading to cellular uptake, peptide linearization, and peptide co‐assembly.^{[ 80 ]}

Figure 14

A) Oxidative polymerization of Te‐containing molecules by ROS;^{[ 149 ]} B) Condensation reaction between cyanobenzothiazole and cysteine upon reductive activation by GSH.^{[ 150 ]}

Figure 15

Schematic representation of self‐immolative polymers^{[ 151 ]} by A: 1,6‐elimination reaction, B: Disulfide reduction, or C: Acetal hydrolysis.

Figure 16

Overview of models of living organisms and suitable imaging techniques in research.

Figure 17

Selected examples of “turn‐on” fluorescents by Staudinger‐ligation (top) and tetrazine‐ligation (bottom).

Figure 18

Real‐time monitoring of hydrogen peroxide in rat brains by combining sensing strategy and a peroxide bond excited chemiluminescent scaffold. Reproduced with permission.^{[ 170 ]} Copyright 2020, Wiley‐VCH.

Figure 19

Pre‐targeting/labeling protocol for in vivo click reaction. B) 3D PET images (top) and transversal slides (bottom) of a U87 MG tumor‐bearing mouse injected with ω‐[18F]fluoro‐pentaethylene glycolic azide without pretargeting. C) 3D PET images (upper row) and transversal slides (lower row) of a U87 MG tumor‐bearing mouse injected with ω‐[18F]fluoro‐pentaethylene glycolic azide with pretargeting using DBCO‐PEG‐NPs. Reproduced with permission.^{[ 181 ]} Copyright 2013, Wiley‐VCH.