Sealant foam compositions for lung applications

Abstract

The present invention is directed to tissue sealant compositions comprising: a multi-arm reactive polyethylene glycol polymer having at least 3 electrophilic groups; albumin; a buffer; water; and entrained gas as bubbles; wherein concentration of albumin in a liquid component of the sealant is within range of 50-200 mg/ml; and wherein concentration of multi-arm PEG in said liquid component of the sealant is within range of 25-100 mg/mL.

Claims

1. A tissue sealant comprising: a) a multi-arm PEG having at least 3 electrophilic groups; b) albumin; c) a buffer; d) water; and e) entrained gas as bubbles; wherein when components (a-e) of the tissue sealant are combined produce a compliant soft set foam with closed cell; wherein concentration of albumin in a liquid component of the sealant is within range of 50-200 mg/ml; and wherein concentration of multi-arm PEG in said liquid component of the sealant is within range of 25-100 mg/mL, wherein gas comprises 65% to 75% of the foam by volume, and wherein E modulus of the foam in a cured state is above 8.5 kPa and below 110 kPa.

2. The tissue sealant of claim 1, wherein said gas is air, nitrogen, argon, carbon dioxide, or mixtures thereof.

3. The tissue sealant of claim 1, wherein said multi-arm PEG has molecular weight from about 5 kD to about 20 kD.

4. The tissue sealant of claim 1, wherein said multi-arm PEG comprises diester linkages.

5. The tissue sealant of claim 1, wherein said electrophilic groups are hydroxysuccinimide (NHS) or succinimidyl glutarate ester (SG).

6. The tissue sealant of claim 1, wherein said sealant comprises a foam and is bioresorbable.

7. The tissue sealant of claim 6, wherein foam flowability is such that the foam requires at least 7 seconds to travel 6 inches on a 30% inclined panel.

8. The tissue sealant of claim 6, wherein albumin and PEG-SG concentrations in mg/mL of liquid portion of the foam, are selected so that an equation for E Modulus yields values from 8.5 kPa to 110 kPa:
E Modulus (kPa)=−2.34−0.168Albumin+0.197PEG+0.000393Albumin{circumflex over ( )}2−0.00462PEG{circumflex over ( )}2+0.008318Albumin*PEG.

9. A method of making the tissue sealant of claim 1, comprising: mixing and foaming a composition comprising multi-arm PEG-SG, albumin, buffer, water, and gas.

10. A method of using the tissue sealant of claim 1, comprising: rapidly mixing and foaming a composition comprising multi-arm PEG-SG, albumin, buffer, water, and gas, forming a foam; immediately thereafter applying the foam onto a tissue; allowing the foam to cure on said tissue.

Description

BRIEF DESCRIPTION OF FIGURES

(1) FIG. 1 shows micrographs of cross-sections of the cured foam presented for foams with variable air (% by volume) content.

(2) FIG. 2 shows chart of the thickness of the liquid phase layer vs. % air content in foam and analysis of phase separation or liquid phase thickness.

(3) FIG. 3 shows results of foam flowability test as time (in seconds) to travel 6 inches vs. the air content of the foams.

(4) FIG. 4 shows contour plot of foam elasticity (E modulus; kPa) vs. PEG (mg/ml) and albumin (protein) (mg/ml).

(5) FIG. 5 shows testing of foam in a lung ex-vivo defect model.

(6) FIGS. 6A-B show rigidity and poor compliance of high modulus foams applied

(7) FIGS. 7A-D show application of the foam sealant to an air leak defect model and testing of the performance.

(8) FIG. 8 shows the same contour plot of foam elasticity as shown in FIG. 4, but with the desirable concentrations space defined by the dotted line.

(9) FIG. 9 shows the results of testing of 3 levels of albumin and PEG-SG4 concentration compositions in liquid sealant form, with foamed compositions also included as a control

DETAILED DESCRIPTION

(10) The inventors have discovered sealing and hemostatic materials and process for making thereof, the sealant materials having surprising and highly beneficial properties for tissue sealing and hemostasis. More specifically, described are absorbable or bioresorbable tissue sealants comprising a polymerizable or curable foam comprising multi-arm PEG-NHS (where NHS is N-hydroxysuccinimide) (or PEG-SG, where SG is a succinimidyl glutarate ester), water, albumin, buffer, and gas, with the foam being able to seal lung air leaks, does not limit lung expansion, has acceptable runniness or viscosity. The gas component preferably comprises 55% to 75% of the foam by volume. The albumin/PEG-SG content is preferably defined by a) equation b) contour plot as will be shown below. The E modulus of the cured foam is preferably above 8.5 kPa and below 110 kPa. The multi-arm PEG-SG is also referred to as 2, 3, 4, 6,8 arm polyethylene glycol succinimidyl glutarate ester; PEG-SG4, PEG-SG2, 4-PEG-SG, PEG-SG 4-ARM, PEG-NHS, Succinimidyl PEG NHS, PEG-SUCCINIMIDYL GLUTARATE ESTER, tetra functional poly (ethylene glycol) succinimidyl glutarate, Multiple Arm PEG Polymer, PEG succinate-NHS, PEG-N-hydroxysuccinimide.

(11) According to one embodiment of the present invention, the inventive composition comprises an absorbable compliant foam, comprising a) a protein, such as albumin; b) a polyethylene glycol (PEG) with at least 2 functionalized groups that can form a covalent bond with protein, such as NHS group; c) a gas which is used to form the foam and is present as bubbles trapped or encapsulated in the foam, with the gas component preferably comprises 55% to 75% of the foam volume; d) a fluid, such as water or saline; and e) an optional buffer which regulates pH. The foam is formed by mixing the above components and is applied to the tissue whereby the foam cures or cross-links while simultaneously bonding to the tissue. The foam is preferably characterized as compliant soft set foam with closed cell, a pliable liquid foam that is flowable, spreadable and/or a compliant soft set foam with closed cell.

(12) The preferred tissue application of the foam is lung tissue, i.e. when the foam is brought into contact with several types of tissue, at least one of these is preferably lung tissue. The foam, even after curing needs to have good adhesion to tissue and simultaneously good compliance and compressibility to accommodate expansion and contraction of the lung. The present sealant can be applied to any tissue in vivo.

(13) The inventive foam is a flowable foam that rapidly polymerizes (within a few minutes such as within interval from 0.5 min to about 30 min, more preferably 1 minutes-15 minutes, most preferably 1 minute to 10 minutes) to a soft or compliance and pliable cured or set foam.

(14) In some embodiments, the inventive foams are prepared as a mixture of three components, 2 liquid components and gas or air. The liquid components can preferably include 2, 3, 4, 6, 8, etc. arm polyethylene glycol succinimidyl glutarate, such as 4-arm polyethylene glycol succinimidyl glutarate (PEG-SG4) and albumin dissolved in a buffered carbonate solution. Other aqueous solutions and solvents can be utilized as well. Albumin can be of any source, including animal derived, e.g. bovine, porcine, human derived, recombinant, etc.

(15) One formulation embodiment includes 1 part liquid components (50 mg/ml PEG-SG4 in carbonate buffer and 100 mg/ml albumin in water) and 2 parts air or gas. To create an efficacious foam, ie, foam capable of sealing an air leak in the lung in an animal, the two liquid components and gas/air are combined and processed by passive or active techniques to generate a homogenous foam. Passive refers to methods such as passing components through beads, meshes, small orifices, while active methods require energy and include propellers and agitators. Upon mixing of the components, polymerization is initiated and by 30 seconds visual evidence of foam solidification occurs. Preferably, within about 2 minutes, the foam polymerization has progressed enough to seal an air leak in a preclinical air leak model.

(16) Advantageously, the inventive foams have several benefits as flowable polymerizable soft set foams, including strong adherence with ability to stretch; conformability; targeted controlled application without runoff; ability to get into holes and fissures; visual confirmation of application without additives; greater coverage with less material, due to large volume to mass ratio limiting the amount of implanted material; ability to occupy space and bridge surfaces.

Example 1. Foams Preparation

(17) Compliant foams comprising albumin at a concentration of 100 mg/ml of the liquid and 50 mg/ml of PEG-SG-4 having molecular weight of 10 kDa and varying volumes of air (0%, 15%, 25%, 35%, 45%, 55%, 65%, 75%, 85% of total foam volume) were prepared by a syringe exchange method as described below. The starting albumin concentration of the standard foam is 200 mg/ml, but after combining with the PEG-SG-4 in carbonate buffer, the final concentration of albumin is 100 mg/ml.

(18) In one series of tests, the following materials were used: 200 mg/ml albumin (bovine serum albumin, Sigma) solution; PEG-SG-4 (Jenkem, China.) having molecular weight of 10 kDa; Buffer Carbonate at 100 mM concentration at pH 8; and Air—variable amounts as noted above and described below.

(19) PEG-SG-4 powder was dissolved in 100 mM carbonate buffer (pH=8.0) to form 100 mg/mL solution. Albumin was dissolved in water to form 20% solution (w/v). 1.25 mL of 20% albumin solution and 1.25 mL of 100 mg/mL PEG-SG-4 in buffer were aspirated into separate 20 mL syringes. The volume of each syringe was adjusted to yield air content in the foams between 0% to 85% (air volume/total volume). The syringes were connected using a female-to-female or luer connector. The solutions were then rapidly passed back and forth or moved from one syringe to another and back for 20 times to create foams, with such mixing foaming performed over about 10-30 seconds. Such movement resulted in thorough mixing and formation of air-filled foam within the syringes. After forming the foam, the foam was finally transferred into the first syringe, the second syringe disconnected, and the foam expressed from the first syringe onto a substrate or onto a tissue for further evaluation and characterization.

(20) The resulting foams were assessed for their physical attributes using quantitative methods as described below.

Example 2. Air Bubbles Distribution (at the Tissue-Foam Interface)

(21) For the foam to be an effective lung sealant, the presence of air bubbles at the tissue-foam interface is important as it accommodates the change in lung surface and volume during lung inflation and deflation. Minimizing phase separation between the air and liquid is desirable. Foams with various air content (15% to 85% air volume/total volume) were made as described above and deposited on a horizontal flat polymeric surface and allowed to cure at least 10 minutes at room temperature 20-25 C. Once cured, the foams were processed in paraffin using standard histological techniques, sectioned and stained with hematoxylin and eosin (H&E). Referring to FIG. 1, micrographs of cross-sections of the cured foam are presented for foams with variable air (% by volume) content. Representative images of foams including the liquid or non-foamed phase as a function of air content are shown with the arrow indicating the liquid phase. The liquid phase refers to a portion of the expressed material that was non-foamed during or immediately collapsed after application. The liquid phase then cured and solidified as well as the foamed portion of the material.

(22) As can be seen in FIG. 1, H&E stained cross-section images show a liquid phase having no air bubbles but only cured sealant at the interface between the foam sealant and the surface (bottom of foam, or lower area of the micrographs) representing collapsed foam. Such collapsed foam having no air bubbles can be seen to be significantly reduced as the air content in the foam increased, with very little liquid phase, if any as indicated by FIG. 1 at 45%-85%, and practically none at 65%-85%.

(23) Analysis of phase separation or liquid phase thickness was performed in four cross-sections per foam and the results are shown in FIG. 2 as chart of the thickness of the liquid phase layer vs. % air content in foam. The results indicate that in 15% to 35% air content foams, relatively high thickness of liquid phase was present. In foams with 45% air content and above the liquid phase thickness are not statistically different to the negligibly low value observed for 85% air content foams (a thickness mean of 65 μm for 85% versus a thickness mean of 640 μm for 35%. ANOVA (analysis of variation) is a statistical test used to determine if there are statistically significant differences between the mean values of any groups in the study. Based on this evaluation, ˜45% air content and above in the foam is preferred.

Example 3. Foam Flowability

(24) For the foam to be useful in a lung sealant application, it is desirable to have a sealant that coats the tissue surface without running off while undergoing curing or cross-linking or polymerization. Low viscosity sealant will not stay in place long enough to cure and will migrate due to gravitational and other forces present. Foams made with the variable air content as described above were also prepared with similar compositions but having non-reactive PEG components (i.e. PEG with the same molecular weight but having no reactive SG or NHS groups). Such non-curing formulations were evaluated in order to eliminate the curing aspect of the foam for the flowability analysis. The viscosity and flowability of these non-curing foam formulations is expected to closely mimic these of the inventive curing formulations.

(25) The non-curing foams were prepared using the syringe exchange method as explained in Example 1, but using non-reactive PEG. After forming the foam, 2 mL of the foam was dispensed vertically over the course of two seconds on a glass plate on an inclined plane set at 30 degrees. 30 degrees is the angle between the horizontal plane and the surface. The timer was started at the beginning of the dispense and the time at which the foam traveled 6 inches was recorded. Each group was tested twice from each preparation using two separate preparations for a total of n=4 samples. The flowability of the foams was thus measured as time required for the bead of foam to travel 6 inches on the inclined surface. Flowability is measured by applying the foam to an incline plane and measuring how long it takes (in seconds) to travel 6 inches. A foam that is not too runny is preferred so that is stays at the application site. It is preferred that foam takes longer than 7 seconds to travel the 6 inches on the incline plane.

(26) Referring now to FIG. 3, the results of the flowability test are presented as the plot of the time (in seconds) to travel 6 inches vs. the air content of the foams.

(27) As seen in FIG. 3, the travel time did not significantly increase from 0% to 45% air (p=0, one-way ANOVA). Advantageously, between 55% and 75% there was a significant increase in travel time compared with lower air content foams reflecting an optimal flowability of the foams, ie, a foam that will not migrate from the application site. 65% and 75% air content foams were statistically equivalent and were the least runny foams tested. There is a sharp increase in foam flowability when air content is increased from 75% to 85% resulting in the 85% air foam being statistically equivalent to the flowability observed for foams made with air content ranging between 0% and 45%. Therefore, in terms of foam flowability, the air content preferred range is between 55% and 75%, most preferred 65%-75% (air content/total foam volume). Bars show standard deviation.

(28) The results demonstrate that the preferred air content in the inventive foams is in the range of 55% to 75% air content (air volume/total volume) as at these compositions the flowability is lowest, resulting in longest travel times which are above about 10 seconds or longer.

(29) Based on the studied attributes of the inventive foams, specifically the air bubbles distribution and the flowability, the preferred range for air content in the inventive foams (% volume in total foam volume) is 55% to 75%.

Example 4. Foam Strength and Compliance

(30) The inventors have further discovered specific advantageous properties of the inventive foams achieved at specific concentration ranges. Compliant foams were developed, the foams comprising albumin, polyethylene glycol (PEG) such as linear PEG or multi-arm (dendrimer) PEG with 4 arms or 8 arms or similar, with at least two or more functional groups (such as SG or NHS) that can form a covalent bond with proteins; gas such as air; and optional buffer. PEG-SG and protein concentrations are defined by an experimentally derived regression equation of “E modulus (kPa)”, where the concentrations yield optimal E modulus (kPa) value that is above 8.5 kPa and below 110 kpa.

(31) Compliant foams were prepared, comprising 66% of air (% volume); variable concentration of albumin (ranging between 50 and 200 mg/ml of the liquid portion of the foam); and variable concentration of PEG-SG4-10K (ranging between 25 and 100 mg/ml of the liquid portion).

(32) The compositions were prepared as described above and foamed by syringe exchange method. The foams were allowed to cure inside a fixture having a hollow cylindrical opening (height of 18.5 mm and cross sectional surface area of 194.8 mm{circumflex over ( )}2) for 10 minutes. The cured foams inside the cylindrical opening were then characterized for their biomechanical integrity by measuring the foam's elasticity under cyclical normal to surface mechanical load and record the E modulus (kPa) using an Instron electromechanical testing apparatus.

(33) The compositions of the tested foams and the E modulus are shown in Table 1.

(34) TABLE-US-00001 TABLE 1 Foam E modulus as a function of PEG- SG4-10K and Albumin concentrations. PEG-SG4-10K Protein (mg/ml) (Albumin mg/ml) E (kPa) 25 50 3.09 25 50 3.80 50 100 19.23 50 100 21.61 50 100 25.64 100 200 116.34 100 50 6.89 100 50 6.65 100 50 6.17 25 200 18.52 25 200 18.76 50 200 73.13 50 200 61.97 25 100 10.68 25 100 9.26 25 100 10.68 100 100 36.09 100 100 46.53 50 50 5.70 50 50 5.70 50 50 11.40

(35) The experimental data in Table 1 was processed using Minitab statistical (version 17) to yield the contour plot shown in FIG. 4. FIG. 4 shows a contour plot of foam elasticity (E modulus; kPa) vs. PEG (mg/ml) and albumin (protein) (mg/ml). The ranges for each band of the contour plot are shown as well. The average value for each of the 9 recorded concentration combinations are indicated on the plot outlined in squares. The concentration scale presented in FIG. 4 does not start at the value of zero. Foams made with PEG concentration below 25 mg/ml and albumin concentration below 50 mg/ml in the liquid portion of the foam yielded poor foam biomechanical integrity and therefore, were not further evaluated.

Example 5. Testing of Critical Biomechanical Requirements for the Foam in Terms of E Modulus

(36) The critical biomechanical requirements for the foam in terms of E modulus (kPa) were determined based on ex-vivo studies where various concentrations from Table 1 were tested on ventilated/expanding lung tissue and the feasibility of the foam as a sealant was scored by the pre-clinical surgeon as acceptable or not-acceptable.

(37) The foams were prepared by the dual syringe exchange method using 2.5 mL of solution of the specific concentration of PEG-SG4-10K, 2.5 mL of solution of the specific concentration of albumin and 10 mL of air. The components were rapidly passed 20 times between syringes to achieve a homogenous foam, which was immediately applied to the lung tissue.

(38) Example of failure at the lower end of the E modulus range. Foam made by a syringe exchange method with 66% air content (air volume/total volume) and with 25 mg/ml of PEG-SG-4-10K and 50 mg/ml albumin in the liquid portion of the foam, which exhibited E modulus of ˜3 kPa was applied to a ventilated cadaver porcine lung ex-vivo model. After being applied to lung tissue, this foam demonstrated a mechanical failure as shown in FIG. 5, due to low mechanical integrity, and was scored as un-acceptable to be used as a sealant by the surgeon. FIG. 5 shows testing of foam in a cadaver porcine lung ex-vivo defect model. Arrow indicates the foam failure location as evidenced by the bulging of the foam over the defect due to air pressure from the lung. As a consequence of the reduced concentrations of albumin and PEG crosslinker, the foam's low biomechanical integrity shows inability to serve as a sealant barrier during ventilations due to a pocket of air formed over the defect under the foam.

(39) Example of failure at the higher end of the E modulus range. Conversely, foam made with 100 mg/ml of PEG-SG-4-10K and 200 mg/ml albumin in liquid portion of the foam, which exhibited E modulus of ˜116 kPa was tested and found to be too rigid and thus was restricting lung expansion as indicated by the deformation of the lung tissue as it expands. After being applied to lung tissue, this foam demonstrated unacceptable rigidity and lack of compliance as shown in FIG. 6. FIG. 6A shows the high modulus foam applied to deflated porcine lung surface in an ex-vivo model. The lung maintains physiological structure. FIG. 6B shows that once the same lung is ventilated and expands, the areas in proximity to the high modulus foam deform into an unnatural, non-physiological shape and it appears that the natural expansion of the lung is constricted by the high E modulus foam. This phenomenon was not observed in lower E modulus foams tested. The performance of the high E modulus foam was deemed unacceptable.

(40) The high E modulus foam was prepared by the dual syringe exchange method using 2.5 mL of 200 mg/mL PEG-SG4-10K, 2.5 mL of 40% albumin and 10 mL of air. The concentrations represent the starting concentrations of the components before mixing in a 1:1 ratio. After the mixing of albumin and PEG, the concentrations are 50% lower.

(41) Example of successful performance in the middle range of the E modulus range. Foams prepared with amounts of PEG crosslinker and albumin within the optimal sealing range were tested in the same model (foam made with 50 mg/ml of PEG-SG-4-10K and 100 mg/ml albumin in liquid portion of the foam having a 1:2 liquid to air ratio). We have tested the optimal foam in the ex vivo model and it adhered well and complied with the lung during ventilations. The foam did not delaminate i.e. did not separate from the tissue, and the foam maintained its cohesive properties, i.e. the foam did not crack or rupture.

(42) Example of successful performance in the middle range of the E modulus range. Foam prepared with amounts of PEG crosslinker and albumin within the optimal sealing range were shown to seal air leaks in the ex-vivo model above as well as in a canine prolonged air leak model in acute and survival pre-clinical studies. The foam formulation used consisted of 50 mg/ml PEG-SG4 and 100 mg/ml albumin. Referring to FIG. 7, the data indicates that the foam sealant can successfully seal an air leak in the canine model. FIG. 7A shows application of the foam sealant to an air leak defect model. When the foam was applied, it flowed over the surface and coated the defect. After 2 minutes, the foam polymerized to form an adhesive and cohesive barrier that sealed the air leak (FIG. 7B). The leak was tested by a bubble leak test at 20 cm water pressure in the lung to ensure the leak was sealed (FIG. 7C). After confirmation of the sealed leak, the foam was peeled away to demonstrate that the leak had not spontaneously resolved and therefore scored as acceptable by the surgeon (FIG. 7D showing air leak still present after foam removal). This indicates that a foam with a biomechanical integrity of ˜22 kPa provided was rigid but also pliable/complaint enough to prevent air leak without constricting the lung natural expansion.

(43) Referring to Table 2, comparison of different foams and their properties is presented. The foams with low E modulus (3.44) are compared to foams with intermediate E modulus (22.16) and to foams with high E modulus (116.34). It can be seen that only foams with an intermediate E modulus are able to meet all criteria, including sealing air leaks, not limiting lung expansion, and having acceptable runniness.

(44) TABLE-US-00002 TABLE 2 Comparison of different formulations of foams and their properties Formulations 25 mg/ml of 50 mg/ml of 100 mg/ml of PEG-SG-4-10K PEG-SG-4-10K PEG-SG-4-10K and 50 mg/ml and 100 mg/ml and 200 mg/ml Criteria albumin albumin albumin E modulus (kPa) 3.44 22.16 116.34 Sealing air leaks No Yes Yes Does not limit lung Yes Yes No expansion Acceptable Runniness No Yes No

(45) A regression model was developed based on the experimental data to represent the relationship between foam biomechanical properties and the PEG and albumin concentrations. Regression Analysis was performed for E Modulus (kPa) versus Albumin, PEG concentrations and the results are shown in Tables 3-6.

(46) TABLE-US-00003 TABLE 3 Analysis of Variance Source DF Adj SS Adj MS F-Value P-Value Regression 5 15922.3 3184.46 136.19 0 Albumin 1 44.1 44.11 1.89 0.19 PEG 1 15 15 0.64 0.436 Albumin*Albumin 1 18.6 18.57 0.79 0.387 PEG*PEG 1 158.6 158.63 6.78 0.02 Albumin*PEG 1 3841.6 3841.63 164.29 0 Error 15 350.7 23.38 Lack-of-Fit 3 189.4 63.14 4.7 0.022 Pure Error 12 161.3 13.44 Total 20 16273.1

(47) TABLE-US-00004 TABLE 4 Model Summary S R-sq R-sq(adj) R-sq(pred) 4.83559 97.84% 97.13% 93.93%

(48) TABLE-US-00005 TABLE 5 Coefficients Term Coef SE Coef T-Value P-Value VIF Constant −2.34 9.89 −0.24 0.816 Albumin −0.168 0.123 −1.37 0.19 44.62 PEG 0.197 0.246 0.8 0.436 48.21 Albumin*Albumin 0.00039 0.00044 0.89 0.387 37.91 PEG*PEG −0.00462 0.00177 −2.6 0.02 42.13 Albumin*PEG 0.00832 0.00065 12.82 0 6.9
E Modulus (kPa)=−2.34-0.168 Albumin+0.197 PEG+0.000393 Albumin*Albumin−0.00462 PEG*PEG+0.008318 Albumin*PEG  Regression Equation

(49) TABLE-US-00006 TABLE 6 Fits and Diagnostics for Unusual Observations E Modulud Obs (kPa) Fit Resid Std Resid 6 73.13 61.21 11.92 2.9 R R Large residual

(50) Regression Equation Obtained (Inserting the Albumin and PEG Concentration in mg/ml) is as Follows:
E Modulus (kPa)==−2.34−0.168Albumin+0.197PEG+0.000393Albumin{circumflex over ( )}2−0.00462PEG{circumflex over ( )}2+0.008318Albumin*PEG

(51) Based on the regression model 95% confidence interval (IC) at the data points showed biomechanical integrity insufficiency, a range of “E modulus (kPa)” of above 8.5 kPa and below 110 kPa was determined to be critical to provide acceptable foam sealant performance.

(52) The PEG and albumin concentrations that define this critical “E modulus (kPa)” biomechanical desired range can be obtained by setting the “E modulus (kPa)” in the equation above to be above 8.5 kPa and below 110 kPa. This range is illustrated as the inner surface of the dashed, closed geometric shape in FIG. 8. FIG. 8 is the same contour plot of foam elasticity as shown in FIG. 4, but with the desirable concentrations space defined by the dotted line. The PEG and Albumin Concentrations that intersect within this shape are the preferred range of concentrations to be used in the foam. The surface inside the dashed, closed geometric shape represents the biomechanical acceptable region for foam sealants (Above 8.5 and below 110 kPa E modulus).

Example 6. Comparative Data for Non-Foam Liquid-Only Sealant

(53) Modulus data for comparing foamed and liquid-only compositions was obtained for the compression modulus. The liquid sealant formulations were tested at 3 levels in exactly the same way as in the foam testing, but using a liquid formulation instead (no air). The results of testing of 3 levels of albumin and PEG-SG4 concentration compositions in liquid sealant form, with foamed compositions also included as a control are presented in Table 7 and in FIG. 9. The results show that the “standard” concentration of liquid yielded results outside the preferred range identified for the foam (8.5-110 kPa modulus). The “standard” concentration of liquid represents the same concentration of albumin (100 mg/ml) and PEG-SG4 (50 mg/ml) as that found in the liquid portion of the foam having a 1:2 liquid to air ratio), which was found to be acceptable in the preclinical animal model. The foam performed similarly to the previously presented data and demonstrated advantageous properties as discussed above.

(54) TABLE-US-00007 TABLE 7 Compressive Modulus comparisons for foamed and liquid-only sealants Compressive Modulus (kPa) Formulation (Mixing Technique) Mean Low Concentration Liquid (50 mg/mL albumin 15.89 and 25 mg/mL PEG-SG4-10k, no air) Sealant Formulation Standard Concentration Liquid (100 mg/mL 118.00 albumin and 50 mg/mL PEG-SG4-10k, no air) Sealant Formulation High Concentration Liquid (200 mg/mL albumin 347.60 and 100 mg/mL PEG-SG4-10k, no air) Sealant Formulation Standard Titan Foam (100 mg/mL albumin and 22.57 50 mg/mL PEG-SG4-10k, 1:2 liquid to air ratio)

Example 7. Methods of Making and Treating

(55) According to embodiments of the present invention, there is provided a method of making a foam for lung sealing applications, comprising the steps of mixing the components and sequentially or simultaneously adding gas forming a foam, then immediately expressing the resulting foam onto lung tissue and allowing the foam to cure. The mixing and foaming steps are preferably performed rapidly, such as over 20 s-2 min, such as over 30 s-1 min. The mixing and foaming steps are performed so that no substantial curing can occur prior to expressing the foam onto tissue.

(56) The mixing and foaming steps can be performed in a variety of sequences and some are illustrated below. The foaming step can be performed by injecting gas, high speed mixing with gas, co-expression through static mixers, and other methods of making foams known to a skilled artisan. Examples of mixing sequences include:

(57) Foaming all-Component Mixture M3

(58) Two Liquid Precursors Method of Making Steps:

(59) Preparing a mixture M1 of reactive PEG and water and optional buffer;

(60) Preparing a mixture M2 of albumin and water and optional buffer;

(61) Rapidly intermixing M1 and M2 forming mixture M3;

(62) Adding a gas and rapidly foaming M3;

(63) Immediately thereafter expressing M3 onto lung tissue;

(64) Allowing M3 to cure on tissue.

(65) One Liquid Precursor with Peg Method of Making Steps:

(66) Preparing a mixture M1 of reactive PEG and water and optional buffer;

(67) Rapidly intermixing M1 and powdered albumin forming mixture M3;

(68) Adding a gas and rapidly foaming M3;

(69) Immediately thereafter expressing M3 onto lung tissue;

(70) Allowing M3 to cure on tissue.

(71) One Liquid Precursor with Albumin Method of Making Steps:

(72) Preparing a mixture M2 of albumin and water and optional buffer;

(73) Rapidly intermixing M2 and powdered reactive PEG forming mixture M3;

(74) Adding a gas and rapidly foaming M3;

(75) Immediately thereafter expressing M3 onto lung tissue;

(76) Allowing M3 to cure on tissue.

(77) Dry Mixture Precursor Method of Making Steps:

(78) Preparing a dry mixture M4 of reactive PEG and albumin and optional buffer;

(79) Rapidly intermixing M4 with water and optional buffer forming mixture M3;

(80) Adding a gas and rapidly foaming M3;

(81) Immediately thereafter expressing M3 onto lung tissue;

(82) Allowing M3 to cure on tissue.

(83) Pre-Foaming Intermediate Compositions M1 or M2

(84) Two Liquid Precursors Method of Making Steps:

(85) Preparing a mixture M1 of reactive PEG and water and optional buffer;

(86) Adding a gas and rapidly foaming M1;

(87) Preparing a mixture M2 of albumin and water and optional buffer;

(88) Rapidly intermixing foamed M1 with M2 forming mixture M3;

(89) Immediately thereafter expressing M3 onto lung tissue;

(90) Allowing M3 to cure on tissue.

(91) Or

(92) Preparing a mixture M1 of reactive PEG and water and optional buffer;

(93) Preparing a mixture M2 of albumin and water and optional buffer;

(94) Adding a gas and rapidly foaming M2;

(95) Rapidly intermixing foamed M2 with M1 forming mixture M3;

(96) Immediately thereafter expressing M3 onto lung tissue;

(97) Allowing M3 to cure on tissue.

(98) One Liquid Precursor with Peg Method of Making Steps:

(99) Preparing a mixture M1 of reactive PEG and water and optional buffer;

(100) Adding a gas and rapidly foaming M1;

(101) Rapidly intermixing foamed M1 and powdered albumin forming mixture M3;

(102) Immediately thereafter expressing M3 onto lung tissue;

(103) Allowing M3 to cure on tissue.

(104) One Liquid Precursor with Albumin Method of Making Steps:

(105) Preparing a mixture M2 of albumin and water and optional buffer;

(106) Adding a gas and rapidly foaming M2;

(107) Rapidly intermixing foamed M2 and powdered reactive PEG forming mixture M3;

(108) Immediately thereafter expressing M3 onto lung tissue;

(109) Allowing M3 to cure on tissue.

(110) According to embodiments of the present invention, there is provided a method of treating a lung tissue by applying a compliant and curable foam prepared by the methods described above and comprising components described above. According to embodiments, a foam is prepared and immediately dispensed, prior to complete curing, more preferably prior to any substantial curing, such as prior to viscosity changing after mixing not more than 50%, such as not more than 40%, more preferably not more than 25%, such as not more than 10%, expressing the foam onto lung tissue and allowing the foam to cure.

(111) A number of additives may be delivered with/in the foam sealant, including chemotherapeutic agents, growth factors, cytokines, antimicrobials, procoagulant hemostatic agents, antifibrinolytics, etc.

(112) Having shown and described various versions in the present disclosure, further adaptations of the methods and systems described herein may be accomplished by appropriate modifications by one of ordinary skill in the art without departing from the scope of the present invention. Several of such potential modifications have been mentioned, and others will be apparent to those skilled in the art. For instance, the examples, versions, geometrics, materials, dimensions, ratios, steps, and the like discussed above are illustrative and are not required. Accordingly, the scope of the present invention should be considered in terms of the following claims and is understood not to be limited to the details of structure and operation shown and described in the specification and drawings.