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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 04/01/2026 has been entered.
Claim Objections
Claims 1 and 4 are objected to because of the following informalities:
Claim 1 recites the phrase “seeding the pellet into a culture dish with a medium, and culturing the dermal papilla cells” in step (b3). This should be corrected to “seeding the pellet comprising dermal papilla cells into a culture dish with a medium, and culturing the dermal papilla cells” because a pellet obtained from hair bulbs contain cells other than dermal papilla cells.
Claim 4 recites the phrase “37° C.” in steps (c2) and (c5), which should be corrected to “37° C” without the period after C and the extra spaces between 37° and C.
Claim 4 recites the phrase “trypsin/EDTA” in step (c2), which should be corrected to “trypsin-EDTA” for the sake of clarity.
Appropriate correction is required.
Claim Interpretation
The term “hair bulb” is defined in the specification as “a thick club-like structure which forms the lower part of the hair root surrounded by the hair follicle and in which capillaries, dermal papilla cells, keratinocytes, etc. are located” (p 6, para 21).
Claim 1 recites the phrase “chopping the hair bulbs,” which not defined in the specification. For the purpose of examination, this phrase is be interpreted to mean “dissecting the hair bulbs.”
Claim 4 recites the phrase “next-step culture dish” (step (c5)), which is not a term recognized in the art. For the purpose of examination, given the context of this phrase in Example 3 on page 11, para 45 of the specification, “next-step culture dish” is interpreted to mean “a new culture dish,” which is distinct from the culture dish used in a previous step of the protocol.
Claim Rejections - 35 USC § 112(b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 2 and 4 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 2 recites the limitation “the MEM alpha medium” in line 3. There is insufficient antecedent basis for this limitation in the claim. This should be corrected to “MEM alpha” 1) to overcome the lack of antecedent basis, and 2) because MEM is an acronym for Minimum Essential Medium, so “MEM alpha medium” is redundant.
Furthermore, claim 2 recites the limitation “the scalp tissue collected from a subject” in line 3. There is insufficient antecedent basis for this limitation in the claim.
Claim 4 recites the limitation “(c1) determining a culture dish to be passaged depending on the number of the dermal papilla cells cultured in the culture dish of the step (b3)” (lines 3-4), which is indefinite for the following two reasons:
First, it is unclear what is meant by “depending on the number of the dermal papilla cells cultured in the culture dish.” The limitation “the number of dermal papilla cells” is unclear, as it could refer to the seeded number of cells or the number of cells at confluence, among other things. Moreover, is unclear whether “the number” must reach a particular threshold to make the determination, or whether any number, including one, is sufficient to satisfy this limitation.
Second, claim 1 recites “wherein the step (C) of passaging is performed three times.” It is unclear how “the number of the dermal papilla cells cultured in the culture dish of the step (b3)” is to be used to determine when a culture dish should be passaged for the second and third passages.
Furthermore, claim 4 recites the limitation “the MEM alpha medium containing 1% FBS” in line 1 of step (c3), which is indefinite for the following two reasons:
First, there is insufficient antecedent basis for this limitation in the claim. This should be corrected to “MEM alpha containing 1% FBS” 1) to overcome the lack of antecedent basis, and 2) because MEM is an acronym for Minimum Essential Medium, so “MEM alpha medium” is redundant.
Second, claim 1 recites “wherein the step (C) of passaging is performed three times using medium comprising Minimum Essential Medium (MEM) alpha, basic FGF, 10% fetal bovine serum, penicillin-streptomycin and amphotericin B.” The limitation of 1% FBS in claim 4, which is drawn to steps within step (C) of claim 1, directly contradicts the limitation of 10% FBS set forth in claim 1.
Furthermore, claim 4 recites the limitation “the supernatant” in line 1 of step (c4). There is insufficient antecedent basis for this limitation in the claim.
Furthermore, claim 4 recites the limitation “until the next-step culture dish is full of cells” (step (c5), line 3). The term “full” is a relative term which renders the claim indefinite. The term “full” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. For the purpose of examination, this phrase is interpreted to mean “until the next-step culture dish reaches confluence.”
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.
Claims 1-2 and 4 remain rejected under 35 U.S.C. 103 as being unpatentable over Topouzi (Experimental Dermatology, 2017, 26(6): 491-496; cited in IDS dated 11/28/2024), in view of Freshney (Culture of Animal Cells: A Manual of Basic Technique and Specialized Applications, 2010), Li (CN 103497930, 2014; Clarivate Analytics English translation), and R&D Systems (Amphotericin B Product Datasheet, 2019), as evidenced by Miranda (British Journal of Dermatology, 2010, 163(2): 287-295).
Regarding claim 1: Topouzi teaches a method for the isolation and culture of dermal papilla cells from a skin biopsy, including scalp tissue (p 492, col 1, section 2.1; step (A)). Topouzi teaches isolating end bulbs (reads on hair bulbs) from the tissue (p 492, col 2, section 2.1.1; step (A)), isolating dermal papilla cells from the hair bulbs (p 493, col 1, section 2.1.2; step (B)), and culturing and passaging the dermal papilla cells 3-5 times (p 493, col 2, section 2.1.3.L; p 494, col 1, section 2.2; step (C)). Topouzi teaches placing tissue comprising the hair bulbs in a petri dish with DMEM (p 492, col 2, section 2.1.1.B), and using Noyles spring scissors (p 492, col 2, section 2.1.1.C) to transect the hair follicle to isolate the end bulb (p 493, col 1, section 2.1.1.D) and inverting the end bulb step to expose the dermal papilla (p 493, section 2.1.2), then further transecting the dermal papilla (p 493, col 2, section 2.1.3.J) (reads on chopping the hair bulb of (step (b1)).
Topouzi is silent regarding the size of the dissected hair bulbs in step (b1). Topouzi’s method of dissecting a hair bulb, which comprises dermal papilla, results in isolation of the dermal papilla (p 493, section 2.1.2; Fig 1 A-F). Thus, the dermal papilla isolated from a hair bulb (Fig 1F-G) reads on chopped hair bulb.
Miranda shows that the mean diameter of a terminal hair dermal papilla is 0.119 mm (119 µm), and the mean diameter of an intermediate hair dermal papilla is 0.063 mm (63 µm) (p 289, col 2, para 2; Fig 2). Therefore, the diameter of the dermal papilla (reads on chopped hair bulb) isolated following the method of Topouzi has a diameter between approximately 63 µm and 119 µm.
Topouzi does not teach collecting the dissected hair bulbs in a tube and centrifuging the hair bulbs to obtain a pellet (step (b2)).
Freshney teaches that centrifugation can increase the concentration of cells in cell suspensions (p 47, col 1, section 4.2.7). It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Topouzi by centrifuging the chopped hair bulbs before plating on a culture dish. One of ordinary skill in the art would have been motivated to make this modification because Freshney teaches that centrifugation can increase the concentration of cells in cell suspensions. One of ordinary skill in the art would have had a reasonable expectation of making this modification because Freshney teaches that cell suspensions, such as the hair bulbs in DMEM taught in the method of Topouzi, can be centrifuged.
Topouzi teaches using DMEM as the culture medium for dermal papilla cells in step (C) (Section 2.1.1.A). Topouzi does not teach the use of a medium comprising MEM alpha, basic FGF, 10 % fetal bovine serum, penicillin-streptomycin and amphotericin B.
Li teaches a method for culturing dermal papilla cells from hair follicles, wherein the culture medium comprises alpha-MEM, 10% fetal calf serum, penicillin-streptomycin, and basic fibroblast growth factor (p 1, Description, section C1).
Given the teachings of Topouzi and Li, there was a reasonable expectation that each of the media taught therein work equivalently as a culture medium for papilla cells. Therefore, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention have substituted the DMEM-based culture medium, as taught in Topouzi, with alpha-MEM based culture medium comprising basic FGF, 10 % fetal bovine serum, penicillin-streptomycin, as taught in Li, with predictable results. Substitution of one element for another known in the field, wherein the result of the substitution would have been predictable, is considered to be obvious. See KSR International Co. v Teleflex Inc 82 USPQ2d 1385 (US 2007) at page 1395.
The culture medium taught in Li does not comprise amphotericin B.
R&D Systems teaches that amphotericin B is a strong antimycotic agent that can effectively inhibit or eradicate fungus and/or yeast contamination in cell cultures, and is especially useful in establishing contamination-free primary cell cultures (p 1, Product Description, para 2). It would have been prima facie obvious for a person of ordinary skill in the art before the effective filing date of the claimed invention to have added amphotericin B to the culture medium taught in Li, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to make this modification because R&D Systems teaches that amphotericin B can inhibit or eradicate fungus and/or yeast contamination in cell cultures. One of ordinary skill in the art would have had a reasonable expectation of successfully making this modification because R&D Systems teaches that amphotericin B can be added to cell culture media.
Regarding claim 2: Following the discussion of claim 1, Topouzi teaches washing the skin biopsy in a petri dish containing Dulbecco’s minimal essential medium (DMEM), then transferring the skin biopsy to a new petri dish containing DMEM (p 492, col 2, section 2.1.1.A; step (a1)).
Topuozi teaches using Noyles spring scissors and forceps to trim adherent adipose or connective tissue surrounding the lower section of the follicle to expose and isolate the end bulb (reads on hair bulb) located at the base of the follicle (p 492, col 2, section 2.1.1.C), then transferring the end bulb to a small drop of DMEM formed on the petri dish lid (p 493, col 1, section 2.1.1.D; Fig 1B; steps (a2), (a3)). Topouzi teaches that the dermal papilla can be identified using a stereomicroscope at a high magnification (p 492, col 1, section 2.1.1.C; step (a3)).
Topouzi does not teach the use of a MEM alpha medium in step (A).
Li teaches a method for culturing dermal papilla cells from hair follicles, wherein the culture medium comprises alpha-MEM, 10% fetal calf serum, penicillin-streptomycin, and basic fibroblast growth factor (p 1, Description, section C1).
Given the teachings of Topouzi and Li, there was a reasonable expectation that each of the media taught therein would work equivalently as a culture medium for papilla cells. Therefore, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention have substituted the DMEM-based culture medium, as taught in Topouzi, with alpha-MEM based culture medium, as taught in Li, with predictable results. Substitution of one element for another known in the field, wherein the result of the substitution would have been predictable, is considered to be obvious. See KSR International Co. v Teleflex Inc 82 USPQ2d 1385 (US 2007) at page 1395.
Regarding claim 4: Topouzi teaches that dermal papilla cells are maintained in an incubator set at 37℃ and 5% CO2 (p 493, 2.1.2.L; steps (c1), (c5)). Topouzi teaches that the dermal papilla cells can be passaged using standard cell culture techniques, using a culture medium comprising DMEM supplemented with 10% FBS and 1% penicillin-streptomycin (p 493-494, section 2.1.3.L). Topouzi teaches that passaging comprises removing the media and washing cells with PBS (p 494, section 2.2.1.A, section 2.2.2.C; step (c1)); treating the cells with 0.5% trypsin-EDTA, followed by incubation for approximately 5 minutes in a 37℃ incubator (p 494, section 2.2.2.C; step (c2)); adding DMEM containing 10% FBS to inhibit the trypsin, then transferring the cells in suspension to a 50-mL centrifuge tube and centrifuging for 4 minutes to pellet the cells (p 494, section 2.2.2.C; step (c3)); removing the supernatant and resuspending the pellet in 10mL DMEM/10% FBS, counting the cells, then resuspending the cells for passaging (p 494, section 2.2.2.D; step (c4)). Topouzi teaches that the cells can be passaged using standard cell culture techniques (p 493, section 2.1.3.L).
Topouzi, in view of Li, teaches the use of MEM alpha in step (C), as set forth above in the discussion for claim 1.
Topouzi, in view of Li and R&D systems, teaches the use of “expansion medium 2” in step (c4), as set forth above in the discussion for claim 1.
Topouzi does not teach 1) 0.25% trypsin-EDTA in step (c2); 2) 1% FBS in step (c3); or 3) seeding the cells at a density of 1,500 cells/cm2, then replacing the medium every 3 days while the cells grow to confluency in step (c5).
Regarding differences 1-2: Freshney teaches a method for subculturing (reads on passaging) monolayer cells (p 196-204). Freshney teaches that the procedure for dissociating an adherent cell line varies depending on cell type (p 196, section 12.4), and teaches variations of the protocol, including one wherein 0.25% EDTA in trypsin is used (p 198, Table 12.5; p 199, section 12.4.2).
Given the teachings of Freshney, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to have optimized the conditions for dissociating cells, as taught by Freshney, to arrive at the claimed invention of 25% trypsin-EDTA in step (c2) and 1% FBS in step (c3). See MPEP 2144.05(II)(A). As noted in In re Aller, 105 USPQ 233 at 235, 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.
Regarding difference 3: Freshney teaches that after counting, cell suspensions should be diluted to the appropriate seeding concentration (p 200, steps 12-13), then placed in a 5% CO2 incubator (p 201, steps 14-16). Freshney further teaches that a cell culture, whether it is a primary culture or a subculture, needs a periodic medium change, followed by subculture if the cells are proliferating (reads on until the culture dish is full of cells) (p 193, section 12.3). Freshney teaches that the intervals between medium changes vary among cell lines, depending on the rate of growth on metabolism (p 193, section 12.3). Freshney teaches that some cell lines require media change every 24 or 48 hours, while other require a change every 4 or 7 days (p 193, section 12.3 – p 194, section 12.3.2).
Given the teachings of Freshney, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to have optimized the seeding concentration and intervals between media changes, as taught by Freshney, to arrive at the claimed invention. See MPEP 2144.05(II)(A). As noted in In re Aller, 105 USPQ 233 at 235, 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.
Response to Arguments
RE: Claim Rejections - 35 USC § 112(b)
Applicant argues: Step (c1) of claim 4 has been amended to recite "determining a culture dish to be passaged depending on the number of the dermal papilla cells cultured in the culture dish of the step (b3)." This amendment unambiguously identifies the culture dish of step (b3) as the one whose cell count forms the basis for the passaging determination, and clarifies that the relevant cells are those that have grown in that culture dish over the course of the primary culture established in step (b3).
In response: Applicant’s arguments have been fully considered, but are not persuasive. The limitation “number of cells” is still unclear, as it could refer to the seeded number of cells or the number of cells at confluence, among other things. The language in claim 4 does not clarify “that the relevant cells are those that have grown in that culture dish over the course of the primary culture established in step (b3)” (emphasis added), as asserted by Applicant. Moreover, the language “over the course of the primary culture” leaves room for multiple interpretations, as it does not set forth a definite time period or number of counted cells.
RE: Claim Rejections - 35 USC § 103
1. The Claimed Size Limitation Pertains to the Chopped Hair Bulb Tissue Fragments, Not to the Intact Dermal Papilla Structure Within the Hair Bulb
Applicant argues: “Claim 1, in part, recites, "wherein the hair bulbs chopped in the step (b1) have a diameter of 15 μm to 120 μm." The subject of this size limitation is unambiguously the chopped hair bulb tissue fragments, discrete tissue pieces generated by mechanical dissection of hair bulbs using precision microscissors, not the intact dermal papilla structure residing within the hair bulb” (p 2).
In brief, Applicant argues on pages 2-3 of the Remarks that the Examiner erroneously applied the evidence of Miranda by conflating the diameter of hair bulbs and dermal papilla. Applicant argues that the Examiner relies on the evidence of Miranda to show that the hair bulbs chopped in step (b1) of claim have a diameter of 15 μm to 120 μm, but that these are values for the diameter of intact dermal papilla from terminal and intermediate hairs, and that Miranda in fact shows that the diameter of a hair bulb is 308 μm for terminal follicles and 191 μm for intermediate follicles.
Applicant argues, “Because the examiner's rejection conflates the claimed chopped hair bulb fragment size with the intact dermal papilla diameter taught by Miranda, the overlap in numerical ranges identified by the examiner is between two entirely different parameters describing two entirely different objects. The prima facie case of obviousness based on overlapping numerical ranges under MPEP §2144.05(1) can only arise where the overlapping ranges describe the same parameter in the claimed invention and the prior art” (p 3).
In response: Applicant’s arguments have been fully considered, but are not persuasive. As set forth above in Claim Interpretation, absent a definition of “chopping” in the specification, the broadest reasonable interpretation of the limitation is “dissecting.” As set forth above in the rejection for claim 1, Topouzi teaches a method of inverting a hair bulb to expose the dermal papilla (p 493, section 2.1.2), then further transecting the dermal papilla (p 493, col 2, section 2.1.3.J); that is, a hair bulb, which comprises the dermal papilla, is dissected (i.e., chopped) to expose the dermal papilla therein, which is further transected (i.e., chopped). Thus, the method of Topuzi reads on the limitation chopping the hair bulb of step (b1) of claim 1, and the exposed dermal papilla, as shown in Fig 1F-G of Topouzi, reads on the hair bulbs chopped in step (b1). Therefore, the evidentiary evidence from Miranda regarding the diameter of the dermal papilla, rather than the diameter of the undissected whole hair bulb, is relied upon for the limitation regarding the diameter of the hair bulbs chopped in step (b1) in claim 1.
Applicant argues: The size of the intact dermal papilla in no way dictates, predicts, or constrains the size of the tissue fragments that result from mechanically chopping the hair bulb, as the latter depends on the chopping technique, the number of chopping steps, and the physical characteristics of the tissue being chopped, none of which are related to the dimensions of the intact dermal papilla residing within the hair bulb.
In response: Applicant’s arguments have been fully considered, but are not persuasive. In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., chopping technique, the number of chopping steps, and the physical characteristics of the tissue being chopped) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
Claim 1 does not require a specific chopping technique or the number of chopping steps (the latter of which is emphasized in the specification), as asserted by Applicant. Claim 1 merely requires that the hair bulbs chopped in the step (b1) have a diameter of 15 μm to 120 μm, which is a condition satisfied by the dermal papilla isolated by incising a hair bulb, as taught in Topouzi. Furthermore, although there is a difference between extracting dermal papilla from a hair bulb, as taught in Topouzi, and chopping an intact hair bulb comprising a dermal papilla, such that the dermal papilla is chopped into multiple fragments, the method taught in Topouzi reads on the broadest reasonable interpretation of claim 1 as currently drafted.
2. Topouzi's Disclosure of Three Passages Is Not Applicable to the Fundamentally Different Chopping-Based Method of the Claimed Invention
Applicant argues: The examiner alleges that Topouzi's disclosure of passaging cells three times renders the claimed limitation of three passages obvious. This argument is not persuasive because it fails to account for the fundamental difference between Topouzi's method and the claimed invention.
Topouzi's method involves precise surgical manipulation to isolate individual intact dermal papillae from hair follicles, which are then directly plated onto culture dishes as intact structures. In Topouzi's method, the starting material for culture is therefore an intact, surgically isolated dermal papilla, fundamentally different from the chopped hair bulb fragments used in the claimed invention.
The claimed invention, by contrast, employs an entirely different approach. Hair bulbs are mechanically chopped into tissue fragments having a diameter of 15 μm to 120 μm, and the resulting pellet is seeded into a culture dish in step (b3) to establish a primary culture, from which dermal papilla cells migrate and proliferate over the course of culture. The starting material for the passaging step of the claimed invention is therefore cells that have migrated and expanded from chopped hair bulb fragments, the cells that are fundamentally different in its origin, composition, and growth characteristics from the intact dermal papilla directly plated in Topouzi's method.
Because the cells obtained by the two methods are fundamentally different, the passaging conditions described in Topouzi's method, including the number of passages, cannot be applied to the claimed invention. A person of ordinary skill in the art would have had no basis to conclude, from Topouzi's disclosure alone, that three passages would be appropriate or optimal for the chopping-based primary culture of the claimed invention.
In response: Applicant’s arguments have been fully considered, but are not persuasive. In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., the difference between the method taught in Topouzi and the method allegedly claimed by the Applicant) are not recited in the rejected claim(s). As set forth above, under the broadest reasonable interpretation of instant claim 1, the dissecting method of Topuzi reads on step (b1) of claim 1. Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
Regarding the pellet in step (b3), this limitation is taught by Freshney, as set forth above in the rejection. In response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Risa Takenaka whose telephone number is (571)272-0149. The examiner can normally be reached M-F, 12-7 EST.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Peter Paras can be reached at (571) 272-4517. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/RISA TAKENAKA/ Examiner, Art Unit 1632
/KARA D JOHNSON/ Primary Examiner, Art Unit 1632
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