DETAILED ACTION
Notice of AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Status of the Claims
The amendments filed 6/04/2026 have been entered.
Response to Arguments
Applicant’s arguments, filed 6/04/2026, have been fully considered.
Applicant first points out that “the cited reference to Polach bears the publication number US 2016/0331686, rather than WO 2016/0331686 as indicated” (Applicant Arguments, Page 4).
Applicant’s note is appreciated. The publication number has been updated in the instant Action and on the attached PTO-892 Notice of References Cited.
Applicant next traverses the rejection of claims under 35 U.S.C. 103(a) based primarily on Lin et al.
Applicant argues that “[n]either Lin nor Chao teaches or suggests exosome-based production or delivery systems for Ganoderma immunomodulatory proteins” while “Polach discloses only broad, non-specific approaches for concentrating extracellular vesicles, while failing to teach or suggest the specific exosome production and concentration processes realized by the present disclosure” (Applicant Arguments, Page 5). As further argued by Applicant, “Polach’s generic methods for vesicle handling cannot be assumed to apply to the specific production and concentration of exosomes” (Applicant Arguments, Page 5).
The argument is not found persuasive. Polach et al expressly teach in Example 1 that “[l]iquid cultures are grown to a final density optimal for exosome production for each yeast strain” (Paragraph 0124; see also Figures 11, 13, 14 and 18).
Applicant, however, further argues that, “[a]s demonstrated in Table 2 of the present disclosure, a working example successfully achieves the recited high concentration by delivering a remarkable 1000-fold increase. Because exosome concentration is a critical parameter for clinical efficacy... the cited prior art fails to achieve or suggest the enriched exosome concentration successfully realized by the present disclosure” (Applicant Arguments, Page 5; see also Applicant Arguments, Page 5: “a person of ordinary skill in the art would have had no reasonable expectation of success that the presently claimed high exosome concentrations could be achieved by modifying the teachings of Lin or Chao with the generalized vesicle technology of Polach” and “Yun Lin’s method [wherein the sampling concentration range is limited to 107 to 109 particles/mL] cannot achieve the high exosome concentration of the present claims”).
The argument is not found persuasive. Turning to the Specification, in Example 1 (Paragraphs 0078-0086), Applicant first transform KM71H yeast “to express the proteins of SEQ ID NOs. 1 to 4, as described in US Patent No. 7,601,808” (Paragraph 0079) wherein, of the transformed candidate yeast clones, the “[t]he strain designated as [Pichia pastoris strain] mm106... exhibited the highest production level [and] was selected as the production strain” (Paragraph 0083). Next, in Example 2 (Paragraphs 0087-0096), Applicant utilizes the mm106 strain from Example 1 to release vesicles comprising GMI (Paragraph 0089) which are isolated and concentrated using tangential flow filtration (Paragraphs 0090-0093), ultimately achieving high concentrations thereof (Table 2).
Yet, there is nothing unexpected or unpredictable in “the enriched exosome concentration successfully realized by the present disclosure” as asserted by Applicant. Indeed, Busatto et al (Cells 7:273, 2018) – provided only in response to Applicant’s arguments and not forming a basis of the rejection – “reports the use of tangential flow filtration (TFF) for the highly efficient isolation of EVs from large volumes of samples” (Abstract) and “highly efficient concentration of extracellular vesicles from large volumes of fluid” (Title).
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-2, 5-6 and 8-9 are rejected under 35 U.S.C. 103(a) as being unpatentable over Lin (US 2007/0207954; of record) in view of Polach et al (US 2016/0331686; of record), Yun Lin et al (Am J Physiol Heart Circ Physiol 319:H1162-1180, 2020; of record) and Bransby et al (WO 2019/227095; of record).
As amended, claim 1 is drawn to an exosome composition comprising a population of enriched exosomes of fungus (more specifically, wherein the fungus is Pichia pastoris (claim 6)) containing Ganoderma immunomodulatory protein, a recombinant thereof or a fragment thereof, or a mixture derived from the population of enriched exosomes (more specifically, wherein the Ganoderma immunomodulatory protein is GMI derived from Ganoderma microsporum (claim 2) and the fragment thereof comprises SEQ ID NOs 1 and 2 (claim 5)), wherein:
(a) the concentration of enriched exosomes in the exosome composition is greater than about 1 x 1010 exosomes/mL; and
(b) the exosome composition is obtained by performing tangential flow filtration (TFF) using a TFF membrane with a pore size of about 0.8 µm to 8.0 µm; performing TFF using a TFF membrane with a molecular weight cutoff of about 10 kDa to about 100 kDa; and performing depth filtration using a membrane with a pore size of about 0.22 µm to about 0.45 µm.
Lin teaches “[a]n immunomodulatory protein... cloned from Ganoderma microsporum” (Abstract) comprising “the amino acid sequences of sequence listings, SEQ ID NO 2 and SEQ ID NO 3” (Paragraph 0010) wherein:
SEQ ID NO 2 is Leu-Ala-Trp-Asn-Val-Lys (LAWNVK) (Page 7, Sequence Listing) which is identical to instantly claimed SEQ ID NO 1; and
SEQ ID NO 3 is Asp-Leu-Gly-Val-Arg-Pro-Ser-Tyr-Ala-Val (DLGVRPSYAV) which is identical to instantly claimed SEQ ID NO 2.
Specifically, Lin teaches that the “the immunomodulatory protein gene, gmi gene, cloned from G. microsporum, [was] used... to obtain [the] expression vector... pPGMI”, i.e., pichia plasmid Ganoderma microsporum immunomodulatory protein, (Paragraph 0045), and said construct was, in turn, introduced into Pichia pastoris to express and secrete reGMI, i.e., recombinant GMI (Paragraph 0045) for subsequent purification (Paragraph 0048). As further taught by Lin, “the immunomodulatory protein reGMI according to the present invention not only had the ability to promote the activation of all kinds of cells [but] also had better immunomodulatory efficiency than that of reLZ-8” (Paragraph 0063).
As such, Lin teaches a composition comprising GMI (Ganoderma microsporum immunomodulatory protein) derived from Ganoderma microsporum, a recombinant thereof or fragment thereof, comprising amino acid SEQ ID NOs 1 and/or 2, having an immunomodulator effect in target cells.
However, Lin does not teach the composition as an exosome composition as claimed, in particular an exosome composition having the recited concentration and obtained according to the recited method.
Yet, Polach et al, recognizing that “[t]he delivery of biologically active macromolecules to cells and tissues in vivo remains a challenge to the development of new biological drugs” (Paragraph 0004), teaches “compositions and methods for the use of yeast extracellular vesicles [in particular, exosomes (see Paragraph 0124 and Figures 11, 13, 14 and 18)] to deliver biologically active molecules to target cells” (Paragraph 0007) wherein “[e]xtracellular vesicles pre-loaded with a biological agent by the yeast cells can be collected from the growth media and purified for use as a drug delivery system for mammalian target cells” (Paragraph 0008). In particular, Polach et al teach that “[y]east cells [wherein “the yeast cell is a commensal yeast strain [and]... the yeast strain is Pichia pastoris” (Paragraph 0021)] are transformed with a circular DNA plasmid encoding the biologically active protein of interest... and cultures are grown to a level of confluence optimized for extracellular vesicle production” wherein “[t]he RNA is expressed in the nucleus of the yeast cell, exported to the cytoplasm, translated into the biologically active protein, which is then taken into yeast extracellular vesicles through cytoplasmic sampling. Cells are removed from the growth media by centrifugation and filtering to yield a media fraction containing extracellular vesicles and free extracellular protein” from which the extracellular vesicles are isolated and “then added to mammalian target cells, which take up the vesicles and biologically active protein” wherein “[r]elease of the protein to the cytoplasm of the target cell then allows for biological activity” (Paragraph 0040; see also Paragraph 0079: “the biologically active molecules are produced within the yeast cell itself and loaded during formation of vesicles, such that they emerge from the yeast cells ready to deliver their cargo to target cells”; see also Figure 4).
Based further on Polach et al, it would have been prima facie obvious to modify Lin such that the composition comprising GMI derived from Ganoderma microsporum, a recombinant thereof or fragment thereof, comprising amino acid SEQ ID NOs 1 and/or 2, is an exosome composition. Considering that “[t]he delivery of biologically active macromolecules to cells and tissues in vivo remains a challenge to the development of new biological drugs” (as taught by Polach et al), it would have been prima facie obvious to synthesize the composition comprising GMI derived from Ganoderma microsporum, a recombinant thereof or fragment thereof, comprising amino acid SEQ ID NOs 1 and/or 2, as an exosome composition in order “to deliver biologically active molecules to target cells” with a reasonable expectation of success.
Regarding (a) that the concentration of enriched exosomes in the exosome composition is greater than about 1 x 1010 exosomes/mL:
As stated by MPEP 2144.05, “[g]enerally, differences in concentration or temperature will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical” (see also In re Aller (220 F.2d 454 (CCPA): “where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation…” Indeed, as further discussed by the court, “[s]uch experimentation is no more than the application of the expected skill of the [ordinarily skilled artisan] and failure to perform such experiments would, in our opinion, show a want of the expected skill”; see also In re Peterson, 315 F.3d at 1325 (Fed. Cir. 2005): “[t]he normal desire of scientists or artisans to improve upon what is already generally known provides the motivation to determine where in a disclosed set of percentage ranges is the optimum combination of percentages” and “[o]nly if the ‘results of optimizing a variable’ are ‘unexpectedly good’ can a patent be obtained for the claimed critical range” (quoting In re Antonie (559 F.2d 618 (CCPA 1977))).
In the instant case, the concentration of active molecules in a composition is clearly a result-effective variable. As taught by Polach et al, “the high culture densities achievable with yeast and the relatively low cost of their growth media combine to yield vesicle production systems that are sufficiently concentrated for applications in vivo, scalable and cost effective” (Paragraph 0079) wherein “the number extracellular vesicles produced can vary with the strain of yeast and can be influenced by mutations that influence relevant pathways in the yeast cell [“or the addition of methanol to P. pastoris cultures” (Paragraph 0092)]... In some embodiments, optimized vesicle loading and/or production can be obtained through screens of various yeast strains to determine conditions where both the biologically active molecule and vesicle accumulation levels are the highest” (Paragraph 0080). Not surprisingly, as noted by the court in Ariosa Diagnostics, Inc. v. Sequenom, Inc., 809 F.3d 1282, 1293 (Fed. Cir. 2015), every ordinary artisan in medicine performs “merely routine optimization of drug dosage to maximize therapeutic effect.”
Accordingly, it would have been customary for an artisan of ordinary skill in the art to determine the optimal concentration of enriched exosomes comprising active molecules to include in the exosome composition in order to best achieve the desired results.
And, regarding (b) that the exosome composition is obtained by performing tangential flow filtration (TFF) using a TFF membrane with a pore size of about 0.8 µm to 8.0 µm; performing TFF using a TFF membrane with a molecular weight cutoff of about 10 kDa to about 100 kDa; and performing depth filtration using a membrane with a pore size of about 0.22 µm to about 0.45 µm:
As further taught by Polach et al, after obtaining “a media fraction containing extracellular vesicles... [t]he extracellular vesicles and protein are then precipitated from the media with polyethylene glycol (PEG, M.Wt. optimized for vesicle precipitation)” (Paragraph 0040).
Yet, as taught by Yun Lin et al, while “commercial polyethylene glycol (PEG)-based reagents have been developed to precipitate exosomes with a standard countertop centrifuge... the exosome-enriched pellet is contaminated with residual PEG polymer and other soluble proteins” (Page H1168, Column 1). As such, Yun Lin et al teach “[a]n alternate method to enrich exosomes using the countertop centrifuge is centrifugal ultrafiltration” wherein “[t]he purity of the exosome samples will depend on the choice of filter pore sizes” (Page H1168, Column 1), further teaching, “[i]n a similar fashion... tangential flow filtration (TFF)” wherein “[p]olyethersulfone membranes with a molecular mass cutoff of 100 kDa are most commonly used” and which “is fairly efficient at concentration EVs and allows for a buffer exchange to wash away residual medium contents” (Page H1168, Column 1).
As further taught by Bransby et al, however, “[d]espite their advantages, TFF systems filters may foul when filter flux limits are exceeded, and TFF systems have finite process capacities” (Paragraph 0005). As such, Bransby et al teaches “methods for TFF which employ non-laminar flow through tubular depth filters (TDFs)... referred to... as tangential flow depth filtration systems or TFDF systems” which provide “improved process capacities by reducing fouling characteristics while increasing flux” (Paragraph 0006; see also Paragraph 0034: “[t]he present disclosure overcomes many of these hurdles by combining the advantages of tangential flow filtration with the advantages of depth filtration”) utilizing filters comprising “a mean pore size... typically ranging from 0.2 to 5 microns” (Paragraph 0037).
As such, it would have been prima facie obvious to obtain the exosome by ultrafiltration, more specifically by performing at least one step of tangential flow filtration (TFF) using a TFF membrane with a pore size of about 0.8 µm to 8.0 µm and a TFF membrane with a molecular weight cutoff of about 10 kDa to about 100 kDa, and further performing depth filtration using a membrane with a pore size of about 0.22 µm to about 0.45 µm. It would have been obvious to do so in order to avoid contamination “with residual PEG polymer and other soluble proteins” associated with the method of isolation disclosed by Lin, while also avoiding “foul[ing] when filter flux limits are exceeded, and... finite process capacities” associated with isolation by TFF processes, with a reasonable expectation of success.
For all the foregoing reasons, claims 1-2 and 5-6 are rejected as prima facie obvious.
Claim 8 is drawn to the exosome composition of claim 1, wherein the exosome composition comprises a population of enriched exosomes having a mean particle size from about 150 nm to about 250 nm.
As taught by Polach et al, “[i]n some embodiments, the vesicles have particle diameters in the hundreds of nanometers” (Paragraph 0092), further demonstrating exosomes of approximately 100-200 nm (Figure 12).
As such, claim 8 is also rejected as prima facie obvious.
Claim 9 is drawn to the exosome composition of claim 1, wherein the fungus is engineered with a nucleotide encoding the Ganoderma immunomodulatory protein, a recombinant thereof or a fragment thereof.
As discussed above, Lin teaches that the “the immunomodulatory protein gene, gmi gene, cloned from G. microsporum, [was] used... to obtain [the] expression vector... pPGMI”, i.e., pichia plasmid Ganoderma microsporum immunomodulatory protein, (Paragraph 0045), and said construct was, in turn, introduced into Pichia pastoris to express and secrete reGMI, i.e., recombinant GMI (Paragraph 0045).
Similarly, Polach et al teach that “[y]east cells are transformed with a circular DNA plasmid encoding the biologically active protein of interest” (Paragraph 0040).
As such, claim 9 is also rejected as prima facie obvious.
Claims 3-4 are rejected under 35 U.S.C. 103(a) as being unpatentable over Lin (US 2007/0207954; of record) in view of Polach et al (US 2016/0331686; of record), Yun Lin et al (Am J Physiol Heart Circ Physiol 319:H1162-1180, 2020; of record) and Bransby et al (WO 2019/227095; of record) as applied to claims 1-2, 5-6 and 8-9 above, in further view of Chao (US 2019/0298794; of record).
Claims 3-4 are drawn to the method of claim 1, wherein the Ganoderma immunomodulatory protein comprises an amino acid sequence SEQ ID NO. 3 (claim 3) and/or wherein the recombinant of Ganoderma immunomodulatory protein comprises an amino acid sequence of SEQ ID NO. 4 (claim 4).
The facts of Lin and Polach et al have been set forth above.
Chao teaches an “immunomodulatory protein derived from Ganoderma microsporum” (Paragraph 0001) wherein “[c]ertain embodiments of the immunomodulatory protein of Ganoderma or a recombinant thereof include an amino acid sequence selected from the group consisting of...” (Paragraph 0008):
SEQ ID NO 1 is Leu-Ala-Trp-Asn-Val-Lys (LAWNVK) which is identical to instantly claimed SEQ ID NO 1;
SEQ ID NO 2 is Asp-Leu-Gly-Val-Arg-Pro-Ser-Tyr-Ala-Val (DLGVRPSYAV) which is identical to instantly claimed SEQ ID NO 2;
SEQ ID NO 3 is MSDTALIFTLAWNVKQLAFDYTPNWGRGRPSSFIDTVTFPTVLTDKAYTY RVVVSGKDLGVRPSYAVESDGSQKINFLEYNSGYGIADTNTIQVYVIDPD TGNNFIVAQWN which is identical to instantly claimed SEQ ID NO3; or
SEQ ID NO 4 is EAEAEFMSDTALIFTLAWNVKQLAFDYTPNWGRGRPSSFIDTVTFPTVLT DKAYTYRVVVSGKDLGVRPSYAVESDGSQKINFLEYNSGYGIADTNTIQV YVIDPDTGNNFIVAQWNYLEQKLISEEDLNSAVDHHHHHH which is identical to instantly claimed SEQ ID NO 4.
Accordingly, for the same reasons as discussed above, it would have been prima facie obvious to synthesize an exosome composition comprising GMI derived from Ganoderma microsporum, a recombinant thereof or fragment thereof, comprising amino acid SEQ ID NOs 1, 2, 3 and/or 4. Considering that “[t]he delivery of biologically active macromolecules to cells and tissues in vivo remains a challenge to the development of new biological drugs” (as taught by Polach et al), it would have been prima facie obvious to synthesize the composition comprising GMI derived from Ganoderma microsporum, a recombinant thereof or fragment thereof, comprising amino acid SEQ ID NOs 1, 2, 3 and/or 4, as an exosome composition in order “to deliver biologically active molecules to target cells” with a reasonable expectation of success.
As such, claims 3-4 are also rejected as prima facie obvious.
Conclusion
Any new ground(s) of rejection presented in this Office action are necessitated by Applicant’s amendments to the claims. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any extension fee pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the date of this final action.
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/CRAIG D RICCI/Primary Examiner, Art Unit 1611
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