Formation of extraterrestrial peptides and their derivatives.

The formation of protein precursors, due to the condensation of atomic carbon under the low-temperature conditions of the molecular phases of the interstellar medium, opens alternative pathways for the origin of life. We perform peptide synthesis under conditions prevailing in space and provide a comprehensive analytic characterization of its products. The application of 13C allowed us to confirm the suggested pathway of peptide formation that proceeds due to the polymerization of aminoketene molecules that are formed in the C + CO + NH3 reaction. Here, we address the question of how the efficiency of peptide production is modified by the presence of water molecules. We demonstrate that although water slightly reduces the efficiency of polymerization of aminoketene, it does not prevent the formation of peptides.

Ultrasound assisted extraction of amino acids and nucleobases from clay minerals and astrobiological samples.

The study of organic molecules in meteorite and return samples allows for the understanding of the chemistry that undergoes in our Solar System. The present work aims at studying ultrasound assisted extraction technique as effective extraction method for these molecules in extraterrestrial samples and analogs. Optimal conditions were selected from the investigation of ultrasonic frequency, irradiation duration and solvent effects on amino acids, nucleobases and dipeptides extraction yields from a model clay-rich mineral matrix. Optimal ultrasound-assisted extraction parameters were frequency of 20 kHz within 20 min irradiation time and methanol/water solvent ratio of 1. We then validated this protocol on Mukundpura and Tarda meteorite fragments and compared it to the reference extraction protocol used in astrobiology and based on 24 h extraction time at 100 °C in water We obtained similar quantitative results without any racemization with both methodologies.

Combination of Targeted Therapies for Colorectal Cancer Treatment.

The design of innovative therapeutic strategies enabling the selective destruction of tumor cells while sparing healthy tissues remains highly challenging in cancer therapy. Here, we show that the combination of two targeted therapies, including bevacizumab (Bev), and a ß-glucuronidase-responsive albumin-binding prodrug of monomethyl auristatin E (MMAE), is efficient for the treatment of colorectal cancer implanted in mice. This combined therapy produces a therapeutic activity superior to that of the association of FOLFOX and Bev currently used to treat patients with this pathology. The increased anticancer efficacy is due to either a synergistic or an additive effect between Bev and MMAE selectively released from the glucuronide prodrug in the tumor microenvironment. Since numerous drug delivery systems such as antibody-drug conjugates employ MMAE as a cytotoxic payload, this finding may be of great interest for improving their therapeutic index by combining them with Bev, particularly for the therapy of colorectal cancer.

VOC-Based Probes, a New Set of Analytical Tools to Monitor Patient Health from Blood Sample. Proof of Concept on Tracking COVID-19 Infection.

Induced volatolomics is an emerging field that holds promise for many biomedical applications including disease detection and prognosis. In this pilot study, we report the first use of a cocktail of volatile organic compounds (VOCs)-based probes to highlight new metabolic markers allowing disease prognosis. In this pilot study, we specifically targeted a set of circulating glycosidases whose activities could be associated with critical COVID-19 illness. Starting from blood sample collection, our approach relies on the incubation of VOC-based probes in plasma samples. Once activated, the probes released a set of VOCs in the sample headspace. The dynamic monitoring of the signals of VOC tracers enabled the identification of three dysregulated glycosidases in the initial phase after infection, for which preliminary machine learning analyses suggested an ability to anticipate critical disease development. This study demonstrates that our VOC-based probes are a new set of analytical tools that can provide access to biological signals until now unavailable to biologists and clinicians and which could be included in biomedical research to properly construct multifactorial therapy algorithms, necessary for personalized medicine.

Induced-volatolomics for the design of tumour activated therapy.

The discovery of tumour-associated markers is of major interest for the development of selective cancer chemotherapy. Within this framework, we introduced the concept of induced-volatolomics enabling to monitor simultaneously the dysregulation of several tumour-associated enzymes in living mice or biopsies. This approach relies on the use of a cocktail of volatile organic compound (VOC)-based probes that are activated enzymatically for releasing the corresponding VOCs. Exogenous VOCs can then be detected in the breath of mice or in the headspace above solid biopsies as specific tracers of enzyme activities. Our induced-volatolomics modality highlighted that the up-regulation of N-acetylglucosaminidase was a hallmark of several solid tumours. Having identified this glycosidase as a potential target for cancer therapy, we designed an enzyme-responsive albumin-binding prodrug of the potent monomethyl auristatin E programmed for the selective release of the drug in the tumour microenvironment. This tumour activated therapy produced a remarkable therapeutic efficacy on orthotopic triple-negative mammary xenografts in mice, leading to the disappearance of tumours in 66% of treated animals. Thus, this study shows the potential of induced-volatolomics for the exploration of biological processes as well as the discovery of novel therapeutic strategies.

Traceless Staudinger Ligation to Access Stable Aminoacyl- or Peptidyl-Dinucleotide.

Aminoacyl- and peptidyl-tRNA are specific biomolecules involved in many biological processes, from ribosomal protein synthesis to the synthesis of peptidoglycan precursors. Here, we report a post-synthetic approach based on traceless Staudinger ligation for the synthesis of a stable amide bond to access aminoacyl- or peptidyl-di-nucleotide. A series of amino-acid and peptide ester phenyl phosphines were synthetized, and their reactivity was studied on a 2'-N3 di-nucleotide. The corresponding 2'-amide di-nucleotides were obtained and characterized by LC-HRMS, and mechanistic interpretations of the influence of the amino acid phenyl ester phosphine were proposed. We also demonstrated that enzymatic 5'-OH phosphorylation is compatible with the acylated di-nucleotide, allowing the possibility to access stable aminoacylated-tRNA.

Stimuli-Responsive Catenane-Based Catalysts.

Rotaxanes and molecular knots exhibit particular properties resulting from the presence of a mechanical bond within their structure that maintains the molecular components interlocked in a permanent manner. On the other hand, the disassembly of the interlocked architecture through the breakdown of the mechanical bond can activate properties which are masked in the parent compound. Herein, we present the development of stimuli-responsive CuI -complexed [2]catenanes as OFF/ON catalysts for the copper-catalyzed alkyne-azide cycloaddition (CuAAC) reaction. The encapsulation of the CuI ion inside the [2]catenanes inhibits its ability to catalyze the formation of triazoles. In contrast, the controlled opening of the two macrocycles induces the breaking of the mechanical bond, thereby restoring the catalytic activity of the CuI ion for the CuAAC reaction. Such OFF/ON catalysts can be involved in signal amplification processes with various potential applications.

A ß-Cyclodextrin-Albumin Conjugate for Enhancing Therapeutic Efficacy of Cytotoxic Drugs.

Enhancing the selectivity of anticancer drugs currently used in the clinic is of great interest in order to propose more efficient chemotherapies with fewer side effects for patients. In this context, we developed a ß-cyclodextrin trimer that binds to circulating albumin to form the corresponding bioconjugate in the bloodstream. This latter can then entrap doxorubicin following its i.v. administration via the formation of a host-guest inclusion complex and deliver the drug in tumors. In this study, we demonstrate that the ß-cyclodextrin trimer improves the therapeutic efficacy of doxorubicin for the treatment of a subcutaneous murine Lewis lung carcinoma (LLC) implanted in C57BL/6 mice. This outcome is associated with an increased deposition of doxorubicin in malignant tissues when used in combination with the ß-cyclodextrin trimer compared to the administration of the drug alone.

Focused ultrasound extraction versus microwave-assisted extraction for extraterrestrial objects analysis.

Search for organic bioindicators in the solar system is a fundamental challenge for the space research community. If tremendous improvements have been achieved in detection, little or no research has been dedicated to extraction of the targets from the studied mineral matrices. Apart from thermodesorption, no extraction step was ever performed in situ within the context of biomarker detection experiments. This work presents an extraction protocol compatible with in situ space constraints. Two extraction methods, i.e., microwave-assisted extraction (MAE) and focused ultrasonic extraction (FUSE), were optimized with the aim of extracting molecules having an astrobiological interest (amino acids, nucleobases, polyaromatic carboxylic acids) and that are included in mineral matrices representative of the Martian soil. Higher efficiency was obtained with the FUSE method (20 kHz, amplitude 80%, pulse and relaxation 1 s each, for 10 min) with yields ranging from 30 to 95%. It was then applied on an Atacama Desert soil sample and Aguas Zarcas meteorite fragment. Both water-soluble and organic-soluble compounds present at trace levels were extracted using this short extraction time, and small amounts of sample and solvent compliant with in situ requirements (50 mg, 500 µL). This unique FUSE/derivatization-GC-MS approach gave similar yields to usual 24 h hot water extraction and increased the recovery of the target molecules compared to the derivatization-GC-MS method already used for in situ space experiments by a factor from 2 to 8. The data highlighted the suitability of a focused ultrasonic method for the extraction of trace organic compounds from extraterrestrial samples.

Integrative analytical workflow to enhance comprehensive analysis of organic molecules in extraterrestrial objects.

Molecular identification is a fundamental issue in astrobiology to investigate the routes of emergence of life on our planet involving in particular a potential seeding of extraterrestrial organic matter on the primitive Earth. However, this project encompasses major difficulties due to the low concentration of molecules present in bodies of the Solar System. This work proposes an integrative analytical workflow, no longer based on GC-MS instruments, to enhance comprehensive analysis of organic markers in these objects. Our strategy combines UPLC-HRMS and UPLC-MRM MS methods to bring both a broad molecular mapping and detailed data on indigenous compounds present in any extraterrestrial objects or laboratory analogs. Applied on water extracts from fresh meteorites, our workflow highlights a wide range of free molecules in the non-treated extracts and reveals the wide diversity of amino acid and nucleobase isomers that could lead to misinterpretation as far as the molecular composition of meteorite extracts cannot be anticipated. This strategy, never explored so far, would provide new clues for studying the organic matter in space and should offer new perspectives on its evolution and reactivity.

Induced volatolomics of pathologies.

Volatolomics allows us to elucidate cell metabolic processes in real time. In particular, a volatile organic compound (VOC) excreted from our bodies may be specific for a certain disease, such that measuring this VOC may afford a simple, fast, accessible and safe diagnostic approach. Yet, finding the optimal endogenous volatile marker specific to a pathology is non-trivial because of interlaboratory disparities in sample preparation and analysis, as well as high interindividual variability. These limit the sensitivity and specificity of volatolomics and its applications in biological and clinical fields but have motivated the development of induced volatolomics. This approach aims to overcome issues by measuring VOCs that result not from an endogenous metabolite but, rather, from the pathogen-specific or metabolic-specific enzymatic metabolism of an exogenous biological or chemical probe. In this Review, we introduce volatile-compound-based probes and discuss how they can be exploited to detect and discriminate pathogenic infections, to assess organ function and to diagnose and monitor cancers in real time. We focus on cases in which labelled probes have informed us about metabolic processes and consider the potential and drawbacks of the probes for clinical trials. Beyond diagnostics, VOC-based probes may also be effective tools to explore biological processes more generally.

A Novel Proteomics-Based Strategy for the Investigation of Peptide Sequences in Extraterrestrial Samples.

Method development is one of the objectives of the astrophysical community for characterizing the organic matter in objects of the solar system. In this context, we report on the development of an enzyme-catalyzed stereoselective hydrolysis, inspired by the proteomics discipline, which has enabled the indirect detection of peptide sequences in extraterrestrial samples. A proof of concept has been performed on a Murchison extract. We show that our approach can successfully highlight l- and d-amino acids involved in peptide bonds. While we show that some d-amino acids must have been involved in peptide bonds, we cannot at this stage conclude on the indigenous or exogenous nature of these biopolymers. However, our strategy constitutes the first step toward direct UPLC-MS evidence of peptide sequences in extraterrestrial samples. It should thus contribute to deepening knowledge on the molecules available in the solar system, hence providing new clues about their chemical history, especially on Earth.

Phosphine-Mediated Bioconjugation of the 3'-End of RNA.

Staudinger ligation is an attractive bio-orthogonal reaction that has been widely used to tag proteins, carbohydrates, and nucleic acids. Here, we explore the traceless variant of the Staudinger ligation for 3'-end modification of oligoribonucleotides. An azido-containing dinucleotide was used to study the ligation. Nine phosphines containing reactive groups, affinity purification tags, or photoswitch probes have been successfully obtained. The corresponding modified dinucleotides were synthesized and characterized by LC/MS. Mechanistic interpretations of the reaction are proposed, in particular, the unprecedented formation of an oxazaphospholane nucleotide derivative, which was favored by the vicinal position of 2'-N3 and 3'-OH functional groups on the terminal ribose has been observed. The post-functionalization of a 24-nt RNA with a photoactivable tag is also reported.

In vivo synthesis of triple-loaded albumin conjugate for efficient targeted cancer chemotherapy.

The development of selective anticancer drugs avoiding side effects met in the course of almost all current treatments is of major interest for cancer patients. Here, we report on a novel ß-glucuronidase-responsive drug delivery system allowing the in vivo synthesis of triple-loaded albumin conjugate. Following intravenous administration, the glucuronide prodrug reacts in the blood stream with the cysteine-34 residue of circulating albumin through thio-Michael addition, enabling the bioconjugation of three Monomethylauristatin E (MMAE) molecules to the plasmatic protein. The albumin conjugate then accumulates in malignant tissues where tumor-associated ß-glucuronidase triggers the selective release of the whole transported drugs. By operating this way, the trimeric glucuronide prodrug produces remarkable anticancer activity on orthotopic MIA PaCa-2 pancreatic tumors, leading to dramatic reduction or even remission of tumors (3/8 mice).

Volatile Organic Compound Based Probe for Induced Volatolomics of Cancers.

The development of efficient protocols for cancer diagnosis remains highly challenging. An emerging approach relies on the detection in exhaled breath of volatile organic compounds (VOC) produced by tumours. In this context, described here is a novel strategy in which a VOC-based probe is converted selectively in malignant tissues, by a tumour-associated enzyme, for releasing the corresponding VOC. The latter is then detected in the exhaled breath as a tumour marker for cancer diagnosis. This approach allows the detection of several different tumours in mice, the monitoring of tumour growth and tumour response to chemotherapy. Thus, the concept of "induced volatolomics" provides a new way to explore biological processes using VOC-based probes that could be adapted to many biomedical applications.

Data-Driven UPLC-Orbitrap MS Analysis in Astrochemistry.

Meteorites have been found to be rich and highly diverse in organic compounds. Next to previous direct infusion high resolution mass spectrometry experiments (DI-HR-MS), we present here data-driven strategies to evaluate UPLC-Orbitrap MS analyses. This allows a comprehensive mining of structural isomers extending the level of information on the molecular diversity in astrochemical materials. As a proof-of-concept study, Murchison and Allende meteorites were analyzed. Both, global organic fingerprint and specific isomer analyses are discussed. Up to 31 different isomers per molecular composition are present in Murchison suggesting the presence of ≈440,000 different compounds detected therein. By means of this time-resolving high resolution mass spectrometric method, we go one step further toward the characterization of chemical structures within complex extraterrestrial mixtures, enabling a better understanding of organic chemical evolution, from interstellar ices toward small bodies in the Solar System.

Primary Step Towards In Situ Detection of Chemical Biomarkers in the UNIVERSE via Liquid-Based Analytical System: Development of an Automated Online Trapping/Liquid Chromatography System.

The search for biomarkers in our solar system is a fundamental challenge for the space research community. It encompasses major difficulties linked to their very low concentration levels, their ambiguous origins (biotic or abiotic), as well as their diversity and complexity. Even if, in 40 years' time, great improvements in sample pre-treatment, chromatographic separation and mass spectrometry detection have been achieved, there is still a need for new in situ scientific instrumentation. This work presents an original liquid chromatographic system with a trapping unit dedicated to the one-pot detection of a large set of non-volatile extra-terrestrial compounds. It is composed of two units, monitored by a single pump. The first unit is an online trapping unit able to trap polar, apolar, monomeric and polymeric organics. The second unit is an online analytical unit with a high-resolution Q-Orbitrap mass spectrometer. The designed single pump system was as efficient as a laboratory dual-trap LC system for the analysis of amino acids, nucleobases and oligopeptides. The overall setup significantly improves sensitivity, providing limits of detection ranging from ppb to ppt levels, thus meeting with in situ enquiries.

Controlled Release of a Micelle Payload via Sequential Enzymatic and Bioorthogonal Reactions in Living Systems.

A bioorthogonal approach is explored to release the content of nanoparticles on demand. Exploiting our recently described click-and-release technology, we developed a new generation of cleavable micelles able to disassemble through a sequential enzymatic and bioorthogonal activation process. Proof-of-concept experiments showed that this new approach could be successfully used to deliver the substances encapsulated into micelles in living cells as well as in mice by two complementary targeted strategies.

TCA precipitation and ethanol/HCl single-step purification evaluation: One-dimensional gel electrophoresis, bradford assays, spectrofluorometry and Raman spectroscopy data on HSA, Rnase, lysozyme - Mascots and Skyline data.

The data presented here are related to the research paper entitled "Study of a Novel Agent for TCA Precipitated Proteins Washing - Comprehensive Insights into the Role of Ethanol/HCl on Molten Globule State by Multi-Spectroscopic Analyses" (Eddhif et al., submitted for publication) [1]. The suitability of ethanol/HCl for the washing of TCA-precipitated proteins was first investigated on standard solution of HSA, cellulase, ribonuclease and lysozyme. Recoveries were assessed by one-dimensional gel electrophoresis, Bradford assays and UPLC-HRMS. The mechanistic that triggers protein conformational changes at each purification stage was then investigated by Raman spectroscopy and spectrofluorometry. Finally, the efficiency of the method was evaluated on three different complex samples (mouse liver, river biofilm, loamy soil surface). Proteins profiling was assessed by gel electrophoresis and by UPLC-HRMS.

Development of liquid chromatography high resolution mass spectrometry strategies for the screening of complex organic matter: Application to astrophysical simulated materials.

The present work aims at developing two LC-HRMS setups for the screening of organic matter in astrophysical samples. Their analytical development has been demonstrated on a 100-µg residue coming from the photo-thermo chemical processing of a cometary ice analog produced in laboratory. The first 1D-LC-HRMS setup combines a serially coupled columns configuration with HRMS detection. It has allowed to discriminate among different chemical families (amino acids, sugars, nucleobases and oligopeptides) in only one chromatographic run without neither a priori acid hydrolysis nor chemical derivatisation. The second setup is a dual-LC configuration which connects a series of trapping columns with analytical reverse-phase columns. By coupling on-line these two distinct LC units with a HRMS detection, high mass compounds (350