DETAILED ACTION
Notice of Pre-AIA or 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 .
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 11/03/2025 has been considered by the Examiner.
Status of Claims
Claims 1, 3, 11, 14, 16, 18, 29, 30, 38, 47, 66, 72, 89, 91, 107, 130, 134, and 138–140, filed on 11/03/2025, are under consideration. Claims 2, 4–10, 12–13, 15, 17, 19–28, 31–37, 39–46, 48–65, 67–71, 73 88, 90, 92–106, 108–129, 131–133, 135–137, and 141–160 are cancelled.
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 1, 3, 91, 107, and 130 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by CN 105367370 A (Zhikai et al.)
For claim 1, Zhikai discloses a method of isolating squalene from a squalene source,
comprising:
1. Extracting squalene from the squalene source: Zhikai discloses taking plant oil waste residue (natural vitamin E production foot oil containing squalene, vitamin E, and plant sterols), performing saponification with alkali in lower alcohol, and solvent-extracting the unsaponifiable squalene fraction with an organic solvent (n-hexane, n-heptane, or petroleum ether).
2. Evaporating the squalene resulting from (a): Zhikai discloses subjecting the unsaponifiable squalene extract to multi-stage vacuum fractional distillation (1st stage at 200 220°C and 10–100 Pa; 2nd stage at 220–260°C and 10–100 Pa) to distill and isolate a squalene concentrate.
3. Purifying the squalene resulting from (b) by way of chromatography: Zhikai discloses loading the distilled squalene concentrate onto a silica gel chromatography column and eluting with organic solvents to separate and collect high-purity squalene.
For claim 3, Zhikai explicitly extracts squalene from a plant source (vegetable oil / soybean oil deodorizer distillate waste residue), directly meeting this limitation.
For claim 91, Zhikai discloses that the evaporation step comprises using fractional distillation under vacuum: Operating pressures of 10–100 Pa (0.075 to 0.75 torr) directly anticipate and overlap the claimed range of 0.5 torr to 5 torr. Distillation heating temperatures of 200–220°C and 220–260°C fall squarely within the claimed range of 150°C to 300°C.
For claim 107, Zhikai discloses running the distilled squalene concentrate over a polar resin comprising silica (silica gel column chromatography): Zhikai explicitly specifies silica particle sizes of 60–100 mesh (~150–250 m), 100–200 mesh (~75–150 m), and 200–300 mesh (~48–75 m), which anticipate the claimed average particle size range of 50 m to 700 m. Zhikai performs chromatography at room/ambient temperature, falling within the claimed temperature range of 5°C to 70°C.
Claim 130 is a product-by-process claim reciting “A composition comprising squalene, wherein the composition is produced by the method of claim 1, optionally wherein the squalene has a purity of from about 90% (w/w) to about 100% (w/w)…”. Please see MPEP § 2113: the patentability of a product-by-process claim rests solely on the product itself, not on the process of making it. If the product in the prior art is the same as or Patent Claim Anticipation and Obviousness Analysis obvious over the claimed product, the claim is unpatentable even if the prior art product was made by a different process. Zhikai discloses a composition comprising squalene isolated with high purity. Specifically, Zhikai's Examples 6–12 disclose isolating squalene having purities of 91.1%, 95.3%, 96.1%, 96.5%, 97.3%, 97.7%, and 98.7% by weight. These purities fall squarely within the claimed purity range of 90% to 100% (w/w). Because the squalene composition produced by Zhikai is structurally identical to the composition of Claim 130.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 11, 14, 16, 18, 29, 30, 38, 47, 66, 72, 89, 134 and 138-140 are rejected under 35 U.S.C. 103 as being unpatentable over Thompson et al. (cited in the IDS; i-ACES Vol. 1 No. 1, 2014) in view of Zhikai et al. (CN 105367370 A) and Xynos et al. (“A single-step isolation of squalene from olive oil deodorizer distillates by using centrifugal partition chromatography”, SEPARATION SCIENCE AND TECHNOLOGY 2016, VOL. 51, NO. 5, 830–835).
Regarding claim 11, Thompson discloses culturing a population of host cells capable of producing squalene in a culture medium and under conditions suitable for the host cells to produce squalene (culturing engineered Saccharomyces cerevisiae SK2 overexpressing tHmg1 in glucose-containing synthetic complete medium) and extracting the squalene from the fermentation composition using an alcohol/KOH lysis and hexane extraction.
Thompson does not explicitly disclose evaporating the squalene or purifying the squalene by chromatography to isolate a purified squalene product (Thompson utilizes a C18 HPLC column strictly as an analytical detector to quantify squalene content, rather than as a downstream recovery process).
However, Zhikai discloses in the analogous art of lipid separation that crude squalene extract obtained by alkaline extraction is effectively separated and purified to high purity by fractional vacuum distillation followed by polar resin (silica gel) chromatography. Also, Xynos discloses chromatography separation of sequalene from plant oil (olive).
Therefore, and before the effective filing date of the instant invention, it would have been obvious to one of ordinary skill in the art to apply the downstream evaporation (vacuum distillation) and chromatographic purification steps of Zhikai to the crude squalene extract obtained from the yeast fermentation of Thompson with the motivation to combine these teachings because Thompson's crude alcohol/KOH cell lysate contains unreacted cellular debris, lipids, and pigments; employing Zhikai's proven sequence of vacuum distillation to remove volatiles/heavies and polar chromatography to adsorb residual sterols predictably isolates squalene at commercial purity levels (see MPEP § 2143).
Regarding Claim 14, the claim recites that extracting comprises "one or more of: (i) homogenizing..., (ii) separating... into sediment and supernatant by way of centrifugation; (iii) demulsifying...; and (iv) separating the demulsified supernatant... into an aqueous component and an oil component." Thompson explicitly teaches centrifuging harvested yeast cells at 4,000 rpm to separate cell pellets from supernatant (meeting step (ii)). Because Claim 14 only requires one or more of (i)–(iv), the combination of Thompson and Zhikai renders Claim 14 obvious.
Regarding Claim 89, the claim is directed to purifying squalene from an extraction composition previously obtained from fermentation host cells, comprising optionally evaporating and purifying by chromatography. As established above for Claim 11, modifying Thompson's fermentation extraction composition with Zhikai's distillation and chromatographic purification renders this claim obvious.
Claims 16, 18, 29, 30, 38, 47, 66, and 72 recite upstream recovery limitations for yeast fermentation broths, including dilution of whole-cell broth to 20%–40% solid material and high-pressure homogenization at 400–1,200 bar, chemical demulsification of whole-cell broth supernatant using specific surfactants (e.g., DOWFAX® 2A1, DOWFAX® 3B2) at elevated temperature (50–90°C) and pH 6 8; and an intermediate aqueous caustic base wash (e.g., 1 M NaOH) between two liquid-liquid centrifugation stages to remove free fatty acid impurities prior to evaporation. These steps/details are assumed part of the purification, handling and processing techniques and are obvious to one of ordinary skill in the art.
Claim 134 recites a pharmaceutical composition requiring an ultra-high squalene purity of 99.5% to 100% (w/w) with impurities ≤0.5% (w/w). Zhikai's shows purity of 98.7% which is close to and approaching the claimed range; this overlap/touch establishes a prima facie case of obviousness—see MPEP 2144.05.I.
Regarding claims 138–140, Zhikai suggest the use of the squalene for cancer (adjuvant formulations and vaccines comprising the high-purity squalene).
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ALI Z FADHEL whose telephone number is (571)270-0267. The examiner can normally be reached M-F 9am-6pm PST.
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/ALI Z FADHEL/Primary Examiner, Art Unit 1772
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