EMULSION FOR ULTRASOUND ABLATION SURGERY

Abstract

The present description relates to an emulsion comprising an amphiphilic compound, a first phase comprising droplets including at least one perfluorocarbon compound and a second phase, which is aqueous. The droplets have a diameter d.sub.4,3 of between 0.5 pm and 5.5 pm, and the at least one perfluorocarbon compound has a boiling point above 100° C. The present description also relates to such an emulsion for use as an improving agent in ultrasound ablation surgery (FIG. 3).

Claims

1. An injectable emulsion comprising: a first phase comprising droplets including at least one perfluorocarbon compound; a second phase, which is aqueous; and an amphiphilic compound; wherein the droplets have a diameter d.sub.4,3 of between 0.5 μm and 5.5 μm; and wherein the at least one perfluorocarbon compound has a boiling point above 100° C.

2. The injectable emulsion of claim 1, wherein a concentration by volume of the first phase in the second phase is between 0.001% v/v and 10% v/v.

3. The injectable emulsion of claim 1, wherein the at least one perfluorocarbon compound has a boiling point above 100° C. and below 160° C.

4. The injectable emulsion of claim 1, wherein the at least one perfluorocarbon compound is selected from the group consisting of perfluorooctane, perfluorononane, perfluorodecalin, perfluorooctyl bromide (PFOB) and perfluoro-15-crown-5-ether (PFCE).

5. The injectable emulsion of claim 1, wherein the amphiphilic compound comprises a dendrimer of Dendri-TAC type or an oligomer of F.sub.iTAC.sub.n type.

6. The injectable emulsion of claim 5, wherein the amphiphilic compound is selected from the group consisting of F.sub.6TAC.sub.7, F.sub.6TAC.sub.12, F.sub.6TAC.sub.29, F.sub.8TAC.sub.7, F.sub.8TAC.sub.13, and F.sub.8TAC.sub.17.

7. A method for ultrasound ablation surgery, comprising: administering the injectable emulsion of claim 1 by intravenous or intra-arterial injection or direct injection into a tissue to be treated; transmitting a focused ultrasound beam; and applying the focused ultrasound beam on at least one zone of the tissue.

8. A method of treating a cancer affecting an organ selected from the group consisting of the liver, spleen, kidneys, prostate, breasts and pancreas, the method comprising administering the injectable emulsion of claim 1 by intravenous or intra-arterial injection or direct injection into a tissue to be treated and performing ultrasound ablation thereafter.

9. A process for producing the injectable emulsion of claim 1, comprising: providing an amphiphilic compound, a first phase comprising a perfluorocarbon compound, and an aqueous second phase; mixing the amphiphilic compound, the first phase and the second phase; cooling the mixture obtained; and homogenizing the mixture at low energy so as to obtain the injectable emulsion of claim 1.

10. The method of claim 7, wherein the at least one zone of the tissue comprises focusing points of the focused ultrasound beam.

11. The method of claim 7, wherein transmitting is performed in pulsed mode and for a predetermined insonification time.

12. The method of claim 11, wherein applying is performed with a duty cycle corresponding to insonification of each focusing point of the at least one zone and comprised between 0 and 100%.

13. The method of claim 7, wherein the focused ultrasound beam has a frequency between 500 kHz and 2 MHz.

14. The method of claim 7, wherein the focused ultrasound beam has an intensity of between 0.05 W/cm.sup.2 and 10 000 W/cm.sup.2.

15. The method of claim 7, wherein applying is performed on highly vascularized organs.

16. The injectable emulsion of claim 2, wherein the at least one perfluorocarbon compound has a boiling point above 100° C. and below 160° C.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0080] FIG. 1 is a representation of a molecular structure of a first and of a second example of amphiphilic compounds that may be used in emulsions according to embodiments of the present description.

[0081] FIG. 2 shows a .sup.19F MRI image of a model tissue treated with emulsions according to embodiments according to the present description.

[0082] FIG. 3 shows a frontal view of a superimposition between an MM image of a model tissue loaded with emulsions according to embodiments of the present description, and a temperature chart obtained by thermometry during the exposure of the model tissue to an ultrasound beam, for various concentrations by volume of emulsions (a) (b) and (c), that is to say, respectively, 0% v/v, 0.1% v/v, 0.5% v/v.

[0083] FIG. 4 shows the heating factor differences measured on model tissues loaded with emulsions according to embodiments of the present description, at different concentrations by volume and at two different duty cycles. Cases a) 0.1% v/v and 0% v/v; b) 0.5% v/v and 0.1% v/v are represented.

[0084] FIG. 5 shows the differences in heat deposit increase registered with respect to a control model tissue (without emulsion), for two different concentrations by volume of emulsions according to embodiments of the present description, and at two different duty cycles.

[0085] FIG. 6 shows the change in the temperature in model tissues loaded with emulsions according to embodiments of the present description, at various concentrations by volume of emulsions during the exposure of the model tissue to an ultrasound beam.

[0086] FIG. 7 shows the impact of the repetition of an exposure to ultrasound beams on the increase in temperature of the model tissue.

[0087] FIG. 8 shows a sagittal view of a superimposition between the MRI image of a model tissue loaded with emulsion according to one embodiment of the present description, at a concentration by volume of 0.5% v/v, and exposed to an ultrasound beam, and the temperature chart determined by thermometry.

DETAILED DESCRIPTION

[0088] In the following detailed description of the embodiments of the present invention, numerous specific details are disclosed in order to provide a fuller understanding of the present description. However, it will become apparent to those skilled in the art that the present description may be implemented without these specific details. In other cases, well-known characteristics have not been described in detail in order to avoid needlessly complicating the description.

[0089] Examples of emulsions 1-6 (see Table 2) according to embodiments of the present description are described below. Emulsions 1-6 use exemplary amphiphilic compounds, of F-TAC type, an example of a perfluorocarbon compound, namely perfluorooctyl bromide (PFOB), and also an example of a second phase, namely water. Emulsion 2, used as an example of an emulsion for use as an improving agent according to the present description, was introduced into tissues modeling target tissues in ultrasound ablation surgery. The results of the exposure of the model tissues loaded with emulsion 2 at various concentrations by volume are detailed below.

[0090] The description also gives an example of a process for producing emulsions 1-6.

1. Example of Synthesis of Amphiphilic Compounds

[0091] The surfactants of F-TAC type, used as examples of amphiphilic compounds, are synthesized by one-step radical polymerization according to a protocol described in the literature [Contino-Pepin et al., 2002, “Amphiphilic Oligomers: A New Kind of Macromolecular Carrier of Antimitotic Drugs. Curr. Med. Chem.—Anti-Cancer Agents”, 2, 645-665]. Two types of perfluoroalkanethiols (C.sub.6F.sub.13C.sub.2H.sub.4SH or C.sub.8F.sub.17C.sub.2H.sub.4SH, telogenic agents) and also azobisisobutyronitrile (AIBN) were used, respectively as transfer reagents and as radical initiator. 10 mL of solvent are used per gram of Tris-derived polymerizable monomer, namely tris(hydroxymethyl)acrylamidomethane (THAM; C=0.57 mol/L).

[0092] In a Schlenk tube under inert atmosphere, THAM (C=0.57 mol/L), a telogenic agent (introduced with a molar ratio Ro as defined in Table 1 below) and AIBN (0.5 molar equivalent relative to the telogenic agent) are dissolved in freshly distilled methanol or in a mixture of methanol and water (9/1, v/v) for the preparation of F-TAC having high DPn values (DPn=n=degree of polymerization). After 3 freeze-thaw cycles under reduced pressure, the mixture is heated at 90° C. for 4 hours with stirring until the monomer has completely disappeared, followed by thin layer chromatography (TLC). The reaction crude is then precipitated twice from diethyl ether and filtered. The filtrate is then vacuum-dried and corresponds to the expected product (white powder, 32-84%, see Table 1 below). The DPn is evaluated by .sup.19F NMR, as previously described in Astafyeva, K. et al., 2015, “Perfluorocarbon nanodroplets stabilized by fluorinated surfactants: characterization and potentiality as theranostic agents” J. Mater. Chem. B, 3, 2892-2907.

TABLE-US-00001 TABLE 1 Polymerization conditions for various F-TACs (where R.sub.0 is the (telogenic agent)/(THAM) molar ratio) F.sub.6TAC.sub.7 F.sub.6TAC.sub.12 F.sub.6TAC.sub.29 F.sub.8TAC.sub.7 F.sub.8TAC.sub.12 F.sub.8TAC.sub.18 1/R.sub.0 4 12 20 4 8 12 AIBN 0.5 0.5 0.5 0.5 0.5 0.5 Yield 63.4% 81.3% 31.8% 65.2% 84.0% 65.1%

2. Example of Process for Producing Emulsions 1-6

[0093] Exemplary emulsions 1-6, comprising the F-TACs of Table 1 and also an example of a PFC compound, namely PFOB, and also an example of a second phase, namely water, may be formed according to the general protocol below. The characteristics of these emulsions 1-6, such as the d.sub.4,3 or the F-TAC used, are grouped together in Table 2.

[0094] Emulsions 1-6 are prepared with a Polytron® system PT 3100 homogenization device of the Kinematica brand.

[0095] The inventors carried out the following procedure in order to prepare an emulsion of droplets including PFOB and an F-TAC at 10% v/v: 835 mg (12.8 mg/ml of emulsion) of F-TAC surfactant are dissolved in 58.5 ml of distilled water using an ultrasound bath. 6.5 ml of PFOB are then added; the resulting mixture is cooled using an ice bath and the process for producing the emulsion is then initiated for three times 15 min using the Polytron® system PT 3100 at 22 500 RPM, with a pause of 30 min between each cycle in order to observe total disappearance of foam. The emulsion is then stored at 4° C. and diluted to the desired concentration before its use in ultrasound ablation surgery.

[0096] Determination of the Volume Fraction of PFC for Each Emulsion

[0097] The volume fraction of PFC for each emulsion 1-6 was evaluated after each preparation, using a known methodology based on .sup.19F NMR [Astafyeva, K. et al., 2015, “Perfluorocarbon nanodroplets stabilized by fluorinated surfactants: characterization and potentiality as theranostic agents” J. Mater. Chem. B, 3, 2892-2907]. The method was modified for the purpose of obtaining a totally homogeneous solution containing both the PFC compound and water. A mixture containing methanol and diethyl ether (50/50, v/v) was used for this purpose, in order to enable complete solubilization of the PFC compound.

[0098] Measurement of the d.sub.4,3 of the Droplets for Each Emulsion

[0099] The size and also the size distribution among a population of droplets of each emulsion 1-6 are evaluated using a Mastersizer 2000 laser diffraction particle size analyzer (Malvern Instruments, Orsay, France) equipped with a Hydro2000S as sample dispersion unit (A), using a dynamic light scattering instrument. The refractive indices used for the PFOB and the dispersant (water) are respectively 1.305 and 1.333. Depending on the sample, a variable number of drops of emulsion is added to the sample dispersion unit (stirring 500 RPM) and the volume-weighted mean diameter d.sub.4,3 (De Brouckère mean diameter) is determined by the Mie theory.

TABLE-US-00002 TABLE 2 Size and polydispersity of the droplets of emulsions 1-6 Amphiphilic compound F.sub.8TAC.sub.7 F.sub.6TAC.sub.7 F.sub.6TAC.sub.12 F.sub.6TAC.sub.29 F.sub.8TAC.sub.13 F.sub.8TAC.sub.17 Emulsion 1 2 3 4 5 6 d.sub.4,3 4.07 ± 3.67 ± 1.48 ± 1.47 ± 0.62 ± 0.62 ± in μm 0.12 0.17 0.22 0.09 0.02 0.09 (D90/ 4.84 4.00 2.90 2.00 3.97 3.30 D10)

3. Example of Ultrasound Beam Exposure of Model Tissues Loaded with Emulsion 2

[0100] Emulsion 2 comprising PFOB as first phase, water as second phase and the biocompatible amphiphilic compound F.sub.6TAC.sub.7 was used as an example of emulsion for use as an improving agent for ultrasound ablation surgery.

[0101] The examples of use of emulsion 2 were carried out on model tissues mimicking the acoustic properties of live soft tissues. The model tissues were uniformly loaded with emulsion 2, used at various concentrations by volume, and exposed to ultrasound beams.

[0102] The temperature increase of the model tissues exposed to ultrasound beams could be verified and compared to a control tissue not containing emulsion 2. The temperature increase through the model tissues was measured in real time, by PRSF (proton resonance shift frequency) magnetic resonance thermometry. This method makes it possible to obtain a precise return from the temperature chart at the focal point at high image number per second, and with millimetric resolution. The .sup.19F MRI imaging confirmed that the droplets of emulsion 2 were uniformly distributed through the model tissue for the two concentrations by volume tested, and also the absence of air bubbles (see FIG. 2).

[0103] Model Tissue Used

[0104] The model tissue, acoustically absorbent and composed in particular of agar-agar gel, was used for its capacity to reproduce acoustic properties very close to those of soft tissues, as for example attested to by Ramnarine, K., Anderson, T., Hoskins, P., 2001, Construction and geometric stability of physiological flow rate wall-less stenosis phantoms, ULTRASOUND Med. Biol. 27, 245-250. The composition of the agar gel used as model tissue is detailed in Table 3. It was in particular adjusted to the need to make it compatible with thermometry measurements, based on magnetic resonance imaging.

TABLE-US-00003 TABLE 3 Composition of the first model tissue SiO.sub.2 SiO.sub.2 Material Glycerol BAL Agar (1.5 μm) (0.5 μm) Water Mass (g) 33.6 0.27 9.0 2.85 2.64 251.6

[0105] The model tissue loaded with droplets has the same composition as the first model tissue described above, with the difference that the volume of emulsion introduced into the preparation replaces one and the same volume of water in order to achieve a constant final gel volume.

[0106] Influence of the Concentration by Volume of the Emulsion on the Thermal Improvement Effect

[0107] The impact of the concentration of the example of emulsion 2 on the thermal improvement effect under exposure to ultrasound beams was measured. Two different concentrations of emulsion 2, namely 0.1% v/v and 0.5% v/v, were tested.

[0108] FIG. 3 represents the superimposition of an MRI image of a model tissue and of the temperature chart at different concentrations by volume: 0.0% v/v, 0.1% v/v and 0.5% v/v. FIG. 3 makes it possible to observe that the surface area affected by the heat deposit is greater for the model tissue loaded with emulsion 2 than for the control model tissue (without emulsion 2), and that this surface area increases with the concentration by volume of PFOB droplets of emulsion 2.

[0109] Insonification Parameters

[0110] Focused Ultrasound Shot:

[0111] The ultrasound beam is transmitted to a zone of the target tissue of the ultrasound shot. The zone is delimited by 16 points at which the ultrasound beam is applied, all of the 16 points forming a circle 4 mm in diameter. The insonification of this zone lasts 1.65 s and covers the insonification of each point. Each point is insonified in pulsed mode over a period of 100 ms with a duty cycle of 70% or 90%. The insonification of the 16 points, corresponding to one complete turn of the circle, is repeated 20 times per shot, resulting in a total shot time of 33 s. The ultrasound beam has a frequency centered about 1 MHz and the acoustic power was adjusted to 94 W over the total ultrasound irradiation time, which corresponds to an intensity of 748 W/cm.sup.2 of the ultrasound beam applied to the zone. Two duty cycles were tested, 70% and 90%, corresponding to pulse durations of 70 ms and 90 ms, respectively, over the range of 100 ms of insonification allotted to each point of the zone, per complete turn of the circle. This corresponds to a difference in delivered energy of 32% between the two assemblies, and to an equivalence of total insonification time for each point of 1.40 s and 1.80 s respectively, with a duty cycle over the total duration of the shot of 4.24% and 5.45%, respectively.

[0112] As shown in FIG. 5, under the two conditions, the results are similar with a heat efficiency compared to the shot performed on the control model tissue (without emulsion), of 9° C. and 15° C. for volume fractions of droplets of 0.1% v/v and 0.5% v/v, respectively.

[0113] Heat Efficiency Measurement:

[0114] The differential heating factor reports, for each measurement, the increase in temperature relative to the control model tissue per unit of delivered energy (in kJ), and is calculated according to Formula 2, according to the respective heating factors of each gel (loaded or not loaded with droplets), calculated according to Formula 1:

Heating factor(° C./kJ)=[(temperature increase in ° C.)÷((total insonification time in seconds))×(Power in Watts))]  Formula (1):

Differential heating factor(° C./kJ)=Heating factor.sub.loaded gel−Heating factor.sub.blank gel  Formula (2):

[0115] The measurements of differential heating factor in the model tissues uniformly loaded with emulsion 2 according to the present description showed that the relationship between the concentration by volume of emulsion 2 and the generation of heat induced by exposure of the model tissue to an ultrasound beam is not linear. FIG. 4 clearly shows that the maximum increase in heating factor lies, for low concentrations by volume, between 0% v/v and 0.1% v/v.

[0116] Evidence of the Possibility of Control in Surgery Provided by the Present Emulsions

[0117] For each droplet concentration, the temperature continuously increases as long as the insonification lasts, but as soon as it is stopped, the temperature begins to decrease, as illustrated by FIG. 6. This is evidence of a possibility of additional control in ultrasound ablation surgery, provided by the emulsion according to the present description.

[0118] Likewise, FIG. 7 illustrates the fact that, contrary to the results of other research teams who observe a loss in heat efficiency of the droplets under exposure to ultrasound beams, the experimental results demonstrated the possibility of repeating the exposure to ultrasound beams with a minimal loss of 5% of gain in temperature. This droplet behavior confirms, if vaporization takes place, that the liquid to gas transition is reversible once the ultrasound irradiation is stopped.

[0119] Absence of Pre-Focal or Post-Focal Heating:

[0120] As shown in FIG. 8 by the cigar shape obtained after exposure of a model tissue to ultrasound beams according to the present description, the inventors noted that the use of an emulsion according to the present description did not result in pre-focal or post-focal heating.