DS0404 - Innovation biomédicale

Development of a Real Time Monitoring non invasive In Vivo Mass Spectrometry System for Diagnostic: REALITY'MS – REALITY'MS

SpiderMass a new tool for guided surgery

In oncology, improved patients uptake is tightly related to the quality of surgery in most cases. Currently there is no instrument allowing to retrieve molecular data in-real time under intraoperative conditions. There is, thus, a need for guiding surgeon in defining with precision excision margins and giving a preliminary diagnostic. SpiderMass corresponds to the development of a novel instrument design for guided surgery and diagnostic in the surgery room.

Developing SpiderMass a novel instrument based on Mass Spectrometry for in-vivo real-time analysis in the surgery room

The challenge of the SpiderMass project is to develop an instrument for allowing the analysis in-vivo and in real-time in the surgical room. This is a main issue with regards to the importance of the quality of the surgery on patient’s outcome and survival. Currently, none of the existing technologies is able to collect such data which require the acquisition in real-time and in-vivo of non-targeted data. The overall objective of REALITY’MS is to develop an instrument for real-time in-vivo analysis based on mass spectrometry because of the ability of this technology to discriminate different cellular phenotypes on the basis of their molecular profiles. The projects is, thus, focused on the development of the instrument and to make the proof on concept of operating on canine cancers in the veterinary surgery room.

The developed SpiderMass instrument is based on an in-vivo micro-sampling of the biological tissues based on laser ablation and then the transport in real-time of the gas phase ions generated by this mean, through a transfer tubing and a dedicated interface, to the mass spectrometer instrument that analyses the sample. The microprobe is a laser source that is composed of a pump laser followed by an OPO allowing for generating a wavelength tunable beam. The microprobe is set to be in resonance with the most intense absorption band of water molecules that is a major component of biological tissues. This microprobe promotes laser ablation and gas phase ions creation with a minimal damage to the analyzed surface. Formed ions can then be transferred to the Mass Spectrometer instrument through a tube which length can be of few meters allowing the microprobe to stand remotely from the spectrometer. The ions are analyzed in real-time by the mass analyzer which ion source is removed to equip the instrument with a dedicated interface. The generated molecular profiles acquired and followed are specific to a tissue type and physiological or physiopathological context (cell type and phenotype). Databanks can thus be realized with the instrument for a pathology and use to establish a classification model using bioinformatics tools. These models must then be used for real-time interrogation during the instrument operation. SpiderMass is then foreseen as a new tool within the surgical room to help the surgeon to define cancerous regions, find its excision margins and give pathology grading.

Starting from an initial prototype which allowed to demonstrate the ability of our system to obtain real-time analysis from ex-vivo tissue pieces, we first were able to show the possible use of the SpiderMass for in-vivo real-time analysis in man. Results demonstrate that SpiderMass is of low invasiveness, since only a white trace is observed from the skin which disappears a few minutes after the analysis, and painless. The system was then optimized to reach a second version of the prototype (SpiderMass v2.0) for which the parameters of the microprobe, the transfer tubing, the interface to the spectrometer and the mass spectrometer itself are improved. This prototype is equipped with a fibered laser source on which the transfer tube is directly fixed allowing for screening the surface of the tissues with the microprobe. The transfer tubing has been reduced in diameter down to 3mm internal diameter and is composed of a biocompatible material certified for medical applications. This is now connected to an IonMobilityQTOF mass spectrometer on which has been installed a dedicated interface for that application. This instrument demonstrates much more important performances in terms of sensitivity, mass accuracy, calibration stability, spectral resolution, resolving power and dynamic mass range than the previous instrument used for Prototype v1.0. SpiderMass 2.0 can thus use the positive and negative mode since we show that discriminating molecular profiles can be generated in these two modes from distinct phenotypes. This Prototype is now use to create the human and canine databanks which will be used as base to establish the classification model.

We have finished a second prototype version of our instrument and passed milestones including the possible development of fibred probe and the possible discrimination of pathological classes of tissue samples. We have demonstrate that SpiderMass can be use on man and is painless. The next step is focused on the realization of the databanks and the establishment of the classification model as well as the development of the interface for the real-time interrogation of the model. Databanks are currently under construction. First bioinformatics studies on data collected with Prototype 1.0 help to find the best appropriated algorithm to be used for the classification according the type of data. A longer term, the last objective will be to transfer the prototype and adapt it to the real surgery room conditions. This will be achieved by moving the prototype to the veterinary surgery room of OCR. The ultimate stage will be to make proof of concept of the instrument abilities using it during a surgery for guiding. Further, after the end of the ANR project several perspectives are envisaged. These include the miniaturization and realization of more integrated system for the microprobe as well as the development of a more compact mass spectrometer easily transportable from one room to another. We also plan to study the possibly coupling of our system to high resolution endoscopic systems or surgical robots as Da Vinci or Rosa. Finally, the addition to the probe of a second laser for treatment is also a possibility.

-Finalist Prix Foundation France Altran, Nov 2014
-Prix « Cutting Edge Technology », MATWIN, May 2015
- Maturation Program SATT Nord, July 2015
-Publication: Fatou, B., Saudemont, P., Leblanc, E., Vinatier, D., Mesdag, V., Wisztorski, M., Focsa, C., Salzet, M., Ziskind, M., and Fournier, I. (2016) In vivo Real-Time Mass Spectrometry for Guided Surgery Application. Scientific reports 6, 25919 (IF:5.578)
-Patent: Deposited France Sept 2014, Deposited Europe 2015, Published Mars 2016 ; I. Fournier, M. Salzet, B. Fatou, M. Wisztroski, C. Focsa, M. Ziskind. Device for Real-Time in-vivo Molecular Analysis WO 2016046748(A1)

Data collection for diagnosis and prognosis of pathologies provides critical information for the physicians and surgeons to make a decision for the subsequent step of treatment and follow-up of the patients with tumors. Currently, conventional diagnostics employs several different in vivo non-invasive technologies including MRI and PET-Scan. However, if these technologies give important information such as tumor volume they might be, to a certain extent, limited to give stage and grade of pathologies which, in turn requires molecular information. Nowadays, tumor grading is still largely performed by pathological examination of biopsies using morphological histology and immunohistochemistry. However, these tests remain difficult especially regarding data interpretation leading to significant inter-exam variability or are narrowly targeted to certain selected markers. (e.g. KI-67, cytokeratins or P53 for immunohistochemistry in ovarian cancers) More recently, diagnostic procedures have been improved by the introduction of quantitative PCR technologies targeting sets of genetic markers. But these tests remain time consuming and might require multiple biopsy samples. These tests become even more complicated if the surgeons need the information during the surgical intervention. Using conventional tools, it is still difficult for surgeons to determine the histological classification of tissues in-situ in order to establish clear excision margins. To provide a solution to this technology gap, there is a need to develop a completely novel instrumentation providing molecular information in real time, in the operating theatre. This is clearly the major goal of the REALITY’MS project; i.e. to develop a monitoring instrument for in vivo real time diagnostic of patients. Among the various technology platforms available we have chosen mass spectrometry due to the numerous advantageous features of this technique including sensitivity, rapidity, automation, reliability, robustness and ability to separate complex mixtures. The principle of the instrument is based on the collection of molecular profiles (molecular signatures) of the investigated tissues. Number of recent studies has already showed the ability of mass spectrometry to observe specific molecular signatures of tissues showing excellent correlation with the cellular types studied, and in particular to distinguish benign, cancerous and borderline regions. Indeed, cancerous cells show particular metabolic features that induce significant changes in the classes of biomolecules that they express. Thus, by creating a model of reference signatures for the different stages and grades of a given pathology, it becomes possible to associate a signature in real-time with the grade of the tumor that can be used for the physician to make decisions. REALITY’MS has the basic rationale to provide surgeons with fast and reliable diagnostic information, without biopsing, and to determine regions to be excised and examine excision margins. The instrument will be embedded into the current context of other imaging modalities routinely used in the operating rooms for guided surgery such as MRI or ultrasonography as well as innovative therapies focused in particular interstitial laser thermotherapy (LITT) and photodynamic therapy (PDT).

Project coordination

Isabelle Fournier (Laboratoire de Spectrométrie de Masse Biologique Fondamentale et Appliquée, Université Lille 1)

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

ICL Computational and Systems Medicine, Department of Surgery and Cancer, Faculty of Medicine, Imperial College London
OCR
PHLAM Laboratoire de Physique des Lasers, Atomes et Mole´cules - CNRS UMR 8523, Université Lille 1
PRISM Laboratoire de Spectrométrie de Masse Biologique Fondamentale et Appliquée, Université Lille 1

Help of the ANR 387,396 euros
Beginning and duration of the scientific project: September 2014 - 48 Months

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