Notice of Pre-AIA or AIA Status
The present application, filed on or after 16 Mar 2013, is being examined under the first inventor to file provisions of the AIA .
Claim Status
Amendment to claims filed 10 Jun 2024 are acknowledged. Claims 1-14 are canceled. Claims 15-28 are newly presented and pending.
Claim Objections
Claim 21 is objected to because of the following informalities:
Gene or protein names should recite the standardized gene/protein name to avoid confusion with other genes and proteins sharing the same name. In the present instance, p21 should be amended to recite CDKN1A and p16 should be amended to recite CDKN2A.
Furthermore, the feed composition does not increase expression of the genes in the liver. Hepatic steatosis brought on by feeding the composition induces expression of the inflammation- and fibrosis-associated genes. The following correction is suggested:
"The method of claim 15, wherein the method leads to increased expression of at least one gene..."
Appropriate correction is required.
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.
Claims 21, 24, and 28 are rejected under 35 U.S.C. § 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention.
The phrase " increases expression of at least one gene " in claim 21 is a relative phrase that renders the claim indefinite. The claim nor the specification set forth a standard for ascertaining the requisite degree of increased expression or a standard by which to compare expression. The specification provides that,
"the phrase 'increase the expression' means that the expression level of a protein or gene that is a marker of cause of inflammation, fibrosis, or senescence increases so that pathological inflammation, fibrosis, and senescence in the liver tissue progress accelerate, or worsen or the risk thereof increases. Specifically, the phrase may mean that the expression level increases by least 20%, more specifically at least 30%, even more specifically at least 40%, compared to that in a control group." (par. 31).
This teaching in the specification is not a limiting definition as it provides three different values for ascertaining the requisite degree of increased expression and as it provides no characteristics required in a control, such as a comparison diet or comparison between time points within a single animal. Without a standard of comparison, one of ordinary skill in the art would not be reasonably apprised of the scope of the invention.
A broad claim limitation together with a narrow limitation that falls within the broad limitation within the same claim may be considered indefinite if the resulting claim does not clearly set forth the metes and bounds of the patent protection desired. See MPEP § 2173.05(c). In the present instance, claim 24 recites the broad recitation "producing an animal" and the claim also recites "administering the feed composition of claim 15 to a mammal" which is the narrower statement of the limitation. The claim is considered indefinite because there is a question or doubt as to whether the feature introduced by such narrower language is (a) merely exemplary of the remainder of the claim, and therefore not required, or (b) a required feature of the claim.
The phrase "exhibits continuous disease progression from steatohepatitis to liver cancer" in claim 28 renders the claim indefinite. It is known the art that in patients with metabolic-associated fatty liver disease (MAFLD), and animal models recapitulating the disease, follow a progression of pathologic steps of hepatic steatosis, steatohepatitis, hepatic fibrosis, cirrhosis, and hepatocellular carcinoma (liver cancer) (M. Eslam, et al. J Hepatol, 2020). Other pathological states showing a progression from steatohepatitis to liver cancer are not known in the art. In the present case, it is unclear whether steatohepatitis and liver cancer must be observed in the same mammal at the same time, whether the mammal must progress from steatohepatitis to liver cancer, whether the animal may merely exhibit liver cancer as an end stage disease after resolution of the preceding pathological states, or whether the intermediate steps of hepatic fibrosis and cirrhosis must also be observed. Therefore, a skilled artisan would be unable to recognize the metes and bounds of the claimed invention, rendering the claim indefinite.
Claim Rejections - 35 USC § 112(a)
The following is a quotation of the first paragraph of 35 U.S.C. § 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
Claims 15-18 and 20-28 are rejected under 35 U.S.C. 112(a) because the specification, while being enabling for,
a method of inducing in a mammal a liver disease selected from the group consisting of steatohepatitis and fibrosis, the method comprising a step of administering a feed composition comprising proteins, carbohydrates, and fats accounting for 13 to 17%, 38 to 42%, and 43 to 47%, respectively, of total calories in the feed composition,
does not reasonably provide enablement for induction of cirrhosis or liver cancer in a mammal. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make or use the invention commensurate in scope with these claims.
Enablement is considered in view of the Wands factors (MPEP 2164.01(a)). The court in Wands states, “Enablement is not precluded by the necessity for some experimentation such as routine screening. However, experimentation needed to practice the invention must not be undue experimentation. The key word ‘undue’, not ‘experimentation.’” (Wands, 8 USPQ2d 1404). Clearly, enablement of a claimed invention cannot be predicated on the basis of quantity of experimentation required to make or use the invention. "Whether undue experimentation is needed is not a single, simple factual determination, but rather is a conclusion reached by weighing many factual considerations." (Wands, 8 USPQ2d 1404). When determining whether a specification meets the enablement requirement, some of the factors to be analyzed are: (1) the breadth of the claims, (2) the nature of the invention, (3) the state of the prior art, (4) the level of one of ordinary skill in the art, (5) the level of predictability in the art, (6) the amount of direction provided by the inventor, (7) the existence of working examples, and (8) whether the quantity of any necessary experimentation to make and use the invention based on the content of the disclosure is undue (Wands). While all of these factors are considered, those sufficient for establishing a prima facie case are discussed below.
Nature of the invention
The claimed invention pertains to a diet-induced model of liver disease, and the claimed group of diseases, steatohepatitis, hepatic fibrosis, cirrhosis, and liver cancer, are stages in the progression of metabolic-associated fatty liver disease (MAFLD, previously referred in the art as non-alcoholic fatty liver disease or NAFLD). The feed composition recited in claim 15 is commonly referred to in the art as a Western diet.
Breadth of the claims
The claims are broad with respect to which diseases can be induced by feeding a composition comprising proteins, carbohydrates, and fats accounting for 13 to 17%, 38 to 42%, and 43 to 47%, respectively, of total calories in the feed composition.
Existence of working examples
The specification provides one species of diet as a working example for inducing liver disease in mice (par. 67-111). The feed composition is referred to as "New diet", and is compared to an amylin liver non-alcoholic steatohepatitis (NASH) diet (AMLN) and a choline-deficient high fat diet (CD-HFD) (Table 1 par. 69). Mice fed the New diet exhibit symptoms consistent with NASH, hepatic fibrosis, and intrahepatic tumors. No example is provided for induction of cirrhosis.
Amount of direction provided by the inventor
Regarding induction of cirrhosis by feeding a mammal the claimed feed composition, the specification provides that "hepatic fibrosis and cirrhosis are independent diseases with separate etiologies, but are continuous diseases progressing from chronic hepatitis through fibrosis to cirrhosis" (par. 20). However, the specification does not further teaching for which feed compositions could result in hepatic fibrosis without cirrhosis or which feed compositions can result in hepatic fibrosis severe enough to be considered cirrhosis.
Regarding induction of liver cancer by feeding a mammal the claimed feed composition, the specification provides that "that present inventors have found that, when a diet adjusted to have protein, carbohydrate, and fat contents accounting for 13 to 17%, 38 to 42%, and 43 to 47%, respectively, of the total calories of the diet is fed to a mammal, fatty liver, hepatic fibrosis, and insulin resistance are all efficiently induced, so that the progression of a series of liver disease stages from nonalcoholic steatohepatitis in humans through cirrhosis and hepatic fibrosis to liver cancer in humans is successfully reproduced in the mammal" (par. 13). However, this guidance is broad with respect to which diet compositions are capable of inducing each of the listed pathological features recited. The specification provides no further guidance regarding specific feed components required to induce cirrhosis or liver cancer.
State of the prior art and Level of predictability in the art
Regarding induction of cirrhosis, while cirrhosis progresses from hepatic fibrosis and shares common etiology, cirrhosis is a distinct pathology that does not always follow from hepatic fibrosis. Nevzorova (Y.A. Nevzorova, et al., J Hepatol, 2020) teaches that cirrhosis is a feature of advanced chronic liver disease (ACLD) that progresses from alcoholic-related- or non-alcoholic steatohepatitis (ASH and NASH, respectively) (Introduction p. 423). Nevzorova further teaches that cirrhosis is characterized by "excessive deposition of ECM in the liver, leading to formation of regenerative nodules surrounded by fibrous bands, parenchymal extinction, and vascular occlusion" and is strongly associated with portal vein hypertension (p. 432). These features are distinct from hepatic fibrosis, which is simply excess deposition of ECM components without the additional pathological consequences. Nevzorova reviews diet models for inducing NASH and notes that high fat and high-fat/high-fructose models are insufficient to induce cirrhosis (Table 1 and p. 430). Nevzorova further teaches that such diets require additional modifications, such as choline deficiency, addition of trans fats, or chemical insults (pp. 430-431).
Teachings from the art are in disagreement with the recitation from par. 13 of the specification quoted above. Eng (J.E. Eng and J.E. Estall, Cells, 2021, cited in the IDS filed 17 May 2024) teaches that the progression of non-alcoholic steatohepatitis (NASH), which comprises hepatic steatosis, inflammation, and fibrosis, to cirrhosis and hepatocellular carcinoma (HCC, liver cancer or tumor formation) in the liver is a widely recognized progression, but achieving the "transition in mice solely through diet is an ongoing challenge" (§ 12, p. 10). Eng further states that "promoting late-stage NASH is possible using dietary methods (cirrhosis is rare), it requires long time periods and has unreliable transition from NASH to HCC" (§ 12, p. 10). Eng further states that "HCC models relying solely on diet have yet to be widely used because of the extensive time required to detect tumors (>12 months) and overall low tumor accompanying these models" (§ 12, p. 11). Eng further teaches that induction of HCC typically requires additional insults, such as adding 0.2% w/w or more cholesterol (§ 7, p. 6) or directly inducing transformation by "genetic modification, chemotoxins, or implantations" (§ 12, p. 10). Taken together, the art teaches that a Western-style diet, such the feed composition recited in claim 15, is insufficient to induce cirrhosis or liver cancer in a predictable manner.
Conclusions
The specification and the prior art provide working examples that narrowly are enabling for the invention as claimed. However, the breadth of the claims encompasses numerous non-enabled embodiments and neither the specification nor the prior art provide sufficient direction for a skilled artisan to make and/or use the full scope of the claimed invention. The specification and the art recite specific embodiments of feed compositions that result in cirrhosis or liver cancer when fed to a non-human mammal, but the art further teaches that without specific insults, induction of cirrhosis or liver cancer using a high-fat/high-sugar diet alone is rare and unpredictable. Therefore, in order to make and/or use the full scope of the claimed invention, a skilled artisan would have to overcome limits of knowledge in the art to identify enabled embodiments. This represents an undue burden of experimentation to a skilled artisan attempting to make and/ or use the invention as claimed.
Claim 25 is rejected under 35 U.S.C. 112(a) as failing to comply with the enablement requirement. The claim contains subject matter which was not described in the specification in such a way as to enable one skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention.
Enablement is considered in view of the Wands factors (MPEP 2164.01(a)). The court in Wands states, “Enablement is not precluded by the necessity for some experimentation such as routine screening. However, experimentation needed to practice the invention must not be undue experimentation. The key word ‘undue’, not ‘experimentation.’” (Wands, 8 USPQ2d 1404). Clearly, enablement of a claimed invention cannot be predicated on the basis of quantity of experimentation required to make or use the invention. "Whether undue experimentation is needed is not a single, simple factual determination, but rather is a conclusion reached by weighing many factual considerations." (Wands, 8 USPQ2d 1404). When determining whether a specification meets the enablement requirement, some of the factors to be analyzed are: (1) the breadth of the claims, (2) the nature of the invention, (3) the state of the prior art, (4) the level of one of ordinary skill in the art, (5) the level of predictability in the art, (6) the amount of direction provided by the inventor, (7) the existence of working examples, and (8) whether the quantity of any necessary experimentation to make and use the invention based on the content of the disclosure is undue (Wands). While all of these factors are considered, those sufficient for establishing a prima facie case are discussed below.
Nature of the invention
The claimed invention pertains to induction of a liver disease by feeding 2.5 to 4.0 g/kg/day of a composition comprising proteins, carbohydrates, and fats accounting for 13 to 17%, 38 to 42%, and 43 to 47%, respectively, of total calories in the feed composition.
Existence of working examples and Amount of direction provided by the inventor
The specification provides one species of diet as a working example for inducing liver disease in mice (par. 67-111). The feed composition is referred to as "New diet". The specification provides that mice were fed starting at 8 weeks of age for 20 weeks (par. 72 and Fig. 1). Fig. 1 shows that mice fed New diet had an average starting weight of about 25 g and an average ending weight of about 45 g. Feeding 2.5 to 4.0 g/kg/day to mice with average weight of 35 g (average of start and end weight) for 140 days equates to 12.25 g to 19.60 g total feed. The specification provides that mice were fed starting at 8 weeks of age for 20 weeks and that "obesity, one pathological characteristics of NASH, was induced by ... 45.32 g of the diet of the present invention" (par. 72). Therefore, Example 1 appears to provide 2.3 to 3.7 times more feed than the claimed range. No other example is provided for feeding a mammal the claimed composition, and thus no working example is provided in the specification for the claimed invention. No teachings are found in the specification regarding the minimum amount of feed is required to induce steatohepatitis, hepatic fibrosis, cirrhosis, or liver cancer.
State of the art
Panasevich (M.R. Panasevich, et al. Am J Physiol Endocrinol Metab, 2018, cited in the IDS filed 17 May 2024) teaches a method of inducing non-alcoholic steatohepatitis (NASH) in Ossabaw swine by feeding a high-fat, high-fructose corn syrup, and high-cholesterol diet (HFHFHC diet) (Abstract and Methods p. E79) (described in detail in the rejection of claims 15, 21-22, 24 and 26-27 under 35 U.S.C. § 102 below). Panasevich is silent with regard to the amount of feed provided to the swine on a per weight basis, but teaches that "[l]ean pigs consumed an average of 550 g per day (~1,1500 kcal/day), while obese pigs averaged 1,200 g/day (~5000 kcal/day)" (Results p. E81). Lee (L. Lee, et al., Hepatol, 2009) teaches the same method in the same species and diet (Ossabaw fed Purina® TestDiet catalog no. 5B4L) (Abstract and Materials and Methods p. 58). The swine fed this diet consumed 6000 kcal/day, 20% higher than taught by Panasevich, equating to 1,440 g feed/day. The swine in the study taught by Lee averaged 68.35 kg (Table 2), and thus consumed 21.1 g/kg/day, which is 5.3 to 8.4 times more food than the claimed range of 2.5 to 4.0 g/kg/day.
Heinrichs (S.C. Heinrichs, Behav Brain Res, 2001) teaches that mice eat about 4.5 g food per day. Using the average weight of 35 g per mouse from Example 1 above, this equates to 128.6 g/kg/day, which is 32.2 to 51.4 times higher than the claimed range. Heinrich further teaches that obesogenic diets, which includes the claimed feed composition, induce hyperphagia in mice, which further increases the amount of feed consumed.
Conclusions
Using both swine and mice as example from the art, the claimed range of 2.5 to 4.0 g/kg/day is far lower than the amount of feed consumed by animals on obesogenic diets in the art. It is, therefore, unclear how the claimed amount of feed is sufficient to support mortality of the animals fed the diet, much less be sufficient to induce the cited liver diseases, which require steatosis for induction, which therefore requires a caloric excess.
Claim Rejections - 35 USC § 102
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 15, 21-22, 24 and 26-27 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Panasevich (M.R. Panasevich, et al. Am J Physiol Endocrinol Metab, 2018, cited in the IDS filed 17 May 2024).
Regarding claims 15, 24, and 27, Panasevich discloses a method of inducing non-alcoholic steatohepatitis (NASH) in Ossabaw swine by feeding a high-fat, high-fructose corn syrup, and high-cholesterol diet (HFHFHC diet) (Abstract and Methods p. E79). The swine (a mammal) exhibit obesity, liver steatosis and inflammation (steatohepatitis), and hepatic fibrosis (Results p.E81 and Fig. 1). The diet fed is LabDiet® catalog no. 5B4L, which comprises, by %kcal with respect to total calories, 16.2% protein, 40.8% carbohydrates, and 43.0% fat (within each of the claimed ranges of 13 to 17%, 38 to 42%, and 43 to 47, respectively) (Methods p. E79).
Regarding claim 21, livers of the HFHFHC-fed swine have elevated protein expression of α smooth muscle actin (αSMA) and tumor necrosis factor α (TNFα) in liver tissue compared to swine fed a diet of normal chow (p. E81 and Fig. 3).
Regarding claim 22, feeding swine the HFHFHC diet induces symptoms of non-alcoholic steatohepatitis consistent with human disease (Discussion p. E84).
Regarding claim 26, swine were fed the HFHFHC diet for 16 weeks (112 days) or 36 weeks (252 days) (both embodiments within the claimed range of 30 to 400 days) to induce NASH (Methods p. E79).
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 15, 20, 22-24, and 26-28 are rejected under 35 U.S.C. § 103 as being unpatentable over Malehmir (M. Malehmir, et al. Nat Med, 2019) in view of Sanches (S.C.L. Sanches, et al., Biomed Res Int, 2015) and as evidenced by Research Diets® 1 (Research Diets®, D05010402 Rodent Diet [Product data sheet], 2026), Bao (L. Bao, et al. Brit J Pharmacol, 2018), Research Diets® 2 (Research Diets®, D09100301 Rodent Diet [Product data sheet], 2026), and Kannt (A. Kannt, et al., Brit J Pharmacol, 2021).
Regarding claims 15, 24, and 27, Malehmir teaches a method of inducing non-alcoholic steatohepatitis (NASH) in C57BL/6JOldHsd mice by feeding a choline-deficient high fat diet (CD-HFD) or a Western diet with trans fats (WD-HTF) (Abstract and Methods p. 657). The mice (a mammal) fed CD-HFD or WD-HTF exhibit non-alcoholic steatohepatitis (NASH) and hepatic fibrosis (Results p.643 and Fig. 1-2).
The CD-HFD feed is Research Diets® catalog no. D05010402 (RD D05010402), which comprises, by %kcal with respect to total calories, 20% protein, 35.1% carbohydrates, and 44.9% fat (within the claimed range for fat, 43 to 47%) (Malehmir Methods p. 657, Research Diets® 1, and Bao Methods p. 3381). The WD-HTF feed is Research Diets® catalog no. D09100301 (RD D09100301), which comprises, by %kcal with respect to total calories, 20% protein, 40% carbohydrates, and 40% fat (within the claimed ranges for carbohydrates and fat, 38 to 42%, and 43 to 47%, respectively) (Malehmir Methods p. 657, Research Diets® 2, and Kannt Methods p. 2413).
Regarding claim 20, the CD-HFD feed taught by Malehmir is a choline-deficient formulation (does not comprise choline) (Malehmir Methods p. 657).
Regarding claim 22, Malehmir teaches that mice fed CD-HFD or WD-HTF exhibit non-alcoholic steatohepatitis (NASH) (Abstract).
Regarding claim 23, Malehmir teaches induction of NASH in mice (rodents) (Abstract).
Regarding claim 26, Malehmir teaches feeding CD-HFD diet for 6 or 12 mo. (about 180 days or 356 days, respectively) or WD-HTF for 6 or 7.5 months (about 180 days or about 227 days, respectively) (each within the claimed range of 30 to 400 days).
Regarding claims 15 and 28, Malehmir teaches that the CD-HFD diet induces a NASH to hepatocellular carcinoma (HCC, liver cancer) transition in 25% of mice fed the diet (exhibit continuous disease progression from steatohepatitis to liver cancer) (Results pp. 647-648 and Fig. 3e-g).
Malehmir does not teach a diet comprising 13 to 17% protein, 38 to 42% carbohydrates, and 43 to 47% fat, %kcal respective to total calories, as required by claims 15, 20, 22-24, and 26-28.
However, Malehmir teaches two different formulations with protein, carbohydrate, and fat composition close to, or overlapping with, the claimed ranges of macronutrients. A prima facie case of obviousness exists where the claimed ranges or amounts do not overlap with the prior art but are merely close. In re Becket, 88 F.2d 684 (CCPA 1937) ("Where the component elements of alloys are the same, and where they approach so closely the same range of quantities as is here the case, it seems that there ought to be some noticeable difference in the qualities of the respective alloys."). In re Dreyfus, 73 F.2d 931, 934, 24 USPQ 52, 55 (CCPA 1934)(the prior art, which taught about 0.7:1 of alkali to water, renders unpatentable a claim that increased the proportion to at least 1:1 because there was no showing that the claimed proportions were critical). Where the proportions of components in a feed composition can be varied and result in the same phenotype, the claimed range(s) are patentably indistinct from the prior art. Furthermore, it is routine procedure to optimize component amounts to arrive at an optimal product that is superior for its intended use, since it has been held where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. See MPEP § 2144.05.
Furthermore, Sanches teaches that NASH can be induced in mouse models to recapitulate features of the disease observed in humans by manipulation of diet alone (section 4, p. 6). Sanches reviews the art regarding nutritional models of NASH and teaches that variations in the proportion of total carbohydrates and fats impact which features of the disease are observed, such as hepatic steatosis, inflammation, and fibrosis (Sec. 4, pp. 6-8). Specifically, high fat diet with the majority of total energy derived from fats induces mild steatosis and inflammation without fibrosis (pp. 6-7). Shifting the proportion of calories from fat to carbohydrates using the simple sugars fructose and sucrose results in hepatic fibrosis, which is not observed in models using high fat and/or high fructose alone (p. 8). Therefore, a skilled artisan would recognize that varying the relative proportion of macronutrients of a feed composition results in different phenotypes relevant to producing an animal model of NASH. Further, alteration in the proportion of fat and carbohydrates would also shift the proportion of protein in the composition as a percent of the total composition.
Therefore, it would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to modify the composition of the CD-HFD or WD-HTF diets taught by Malehmir by optimizing the proportion of protein, carbohydrates, and fat as taught by Sanches to arrive at the claimed invention. One would be motivated to make such a modification as Sanches teaches that varying the macronutrient composition of feeds for inducing NASH in animal models is routine in the art. One would have a reasonable expectation of success in making the modification as Malehmir teaches two different compositions that are very close to, and partially overlap with, the claimed ranges of protein, carbohydrate, and fat proportions that both result in the claimed phenotypes of steatohepatitis and hepatic fibrosis.
Claims 16-19 are rejected under 35 U.S.C. 103 as being unpatentable over Malehmir in view of Sanches as applied to claims 15, 20, 22-24, and 26-28 above and in further view of Yin (J. Yin, et al., Mol Nutr Food Res, 2016) and as evidenced by Research Diets® 1 (Research Diets®, D05010402 Rodent Diet [Product data sheet], 2026) and Research Diets® 2 (Research Diets®, D09100301 Rodent Diet [Product data sheet], 2026).
Malehmir and Sanches teach a method of inducing steatohepatitis, hepatic fibrosis, and liver cancer as discussed in the rejection of claims 15, 20, 22-24, and 26-28 under 35 U.S.C. § 103 above.
Malehmir does not teach a single composition comprising 15 to 19% w/w casein and 0.2 to 0.3 % w/w L-cysteine as required by claims 16-19, 21 to 25% w/w fructose and 10 to 12% w/w sucrose as required by claims 17-19, 16 to 20% w/w lard as required by claims 18-19, or 0.5 to 0.7% w/w cholesterol as required by claim 19.
However, regarding claim 16, Malehmir further teaches feeding a choline-deficient high fat diet (CD-HFD, Research Diets® cat. no. D05010402), which comprises 200.0 g casein and 3.0 g L-cystine out of a total feed composition of 856.15 g (23.4% w/w and 0.35% w/w, respectively) (Malehmir Methods p. 657 and Research Diets® 1). Malehmir further teaches feeding a Western diet with trans fats (WD-HTF, Research Diets® cat. no. D09100301), which comprises 200.0 g casein and 3.0 g L-cystine out of a total feed composition of 904.05 g (22.1% w/w and 0.33% w/w, respectively) (Malehmir Methods p. 657 and Research Diets® 2).
Regarding claim 17, Research Diets® D05010402 comprises 100.0 g sucrose out of a total feed composition of 856.15 g (20.7% w/w) (Malehmir Methods p. 657 and Research Diets® 1). Research Diets® D09100301 comprises 200.0 g fructose and 100.0 g sucrose out of a total feed composition of 904.05 g (22.1% and 11.1% w/w, respectively) (each within the claimed ranges of 21 to 25 %w/w fructose and 10 to 12 %w/w sucrose) (Malehmir Methods p. 657 and Research Diets 2).
Regarding claim 18, Research Diets® D05010402 comprises 177.5 g lard (20.7% w/w) (Malehmir Methods p. 657 and Research Diets® 1). Research Diets® D09100301 comprises 135.0 g Primex® shortening, 25.0 g soybean oil, and 20.0 g lard out of a total feed composition of 904.05 g (14.90%, 2.77%, and 2.21% w/w, respectively, for a total of 19.88% w/w fat) (Malehmir Methods p. 657 and Research Diets 2).
Regarding claim 19, Research Diets® D05010402 does not comprise cholesterol (Malehmir Methods p. 657 and Research Diets® 1). Research Diets® D09100301 comprises 18.0 g cholesterol out of a total feed composition of 904.05 g (1.99% w/w) (Malehmir Methods p. 657 and Research Diets 2).
Regarding claim 16, L-cystine (CAS Reg No. 56-89-3, MW = 240.30 g/mol) is the oxidized and dimerized form of L-cysteine (CAS Reg No. 52-90-4, MW = 121.16 g/mol). Yin teaches that in vivo, that L-cystine and L-cysteine are interconverted by oxidizing/reducing conditions based on cellular needs, and that L-cystine is the predominant form found in food and outside of cells due to rapid oxidation of L-cysteine (Introduction p. 134). Thus, a skilled artisan would recognize that in a feed composition, L-cystine is an obvious substitute of L-cysteine.
Furthermore, Sanches teaches that NASH can be induced in mouse models to recapitulate features of the disease observed in humans by manipulation of diet alone (section 4, p. 6). Sanches reviews the art regarding nutritional models of NASH and teaches that variations in the proportion of total carbohydrates and fats impact which features of the disease are observed, such as hepatic steatosis, inflammation, and fibrosis (Sec. 4, pp. 6-8). Specifically, high fat diet with the majority of total energy derived from fats induces mild steatosis and inflammation without fibrosis (pp. 6-7). Shifting the proportion of calories from fat to carbohydrates using the simple sugars fructose and sucrose results in hepatic fibrosis, which is not observed in models using high fat and/or high fructose alone (p. 8). Therefore, a skilled artisan would recognize that varying the relative proportion of macronutrients of a feed composition results in different phenotypes relevant to producing an animal model of NASH. Sanches further teaches that multiple sources of fat for high fat diets to induce NASH are used in the art, including lard, fats containing trans fats such as partially hydrogenated vegetable oil (also referred to as shortening), and corn oil, which all result in the same phenotype of obesity, insulin resistance, hepatic steatosis, elevated serum ALT, and other related markers of steatosis and mild inflammation (pp. 6-8). Sanches further teaches that addition of cholesterol to a feed for inducing NASH enhances liver inflammation and expression of collagen, a necessary step in progression of fibrosis, compared to high fat diet alone (p. 7).
Furthermore, Malehmir teaches two different formulations with overlapping components overlapping with, or very close to, the claimed ranges of components. Research Diets® D05010402 comprises 23.4% w/w casein, 0.35% w/w L-cystine, 20.7% w/w sucrose, and 20.7% w/w lard. Research Diets® D09100301 comprises 22.1% w/w casein, 0.33% w/w L-cystine, 22.1% fructose, 11.1% w/w fructose, a mixture of lipids comprising 14.90% w/w Primex® shortening, 2.77% w/w soybean oil, and 2.21% w/w lard, and 1.99% w/w cholesterol. As discussed in the rejection of claims 15, 20, 22-24, and 26-28, both formulations induce steatohepatitis and hepatic fibrosis in mice. A prima facie case of obviousness exists where the claimed ranges or amounts do not overlap with the prior art but are merely close. In re Becket, 88 F.2d 684 (CCPA 1937) ("Where the component elements of alloys are the same, and where they approach so closely the same range of quantities as is here the case, it seems that there ought to be some noticeable difference in the qualities of the respective alloys."). In re Dreyfus, 73 F.2d 931, 934, 24 USPQ 52, 55 (CCPA 1934)(the prior art, which taught about 0.7:1 of alkali to water, renders unpatentable a claim that increased the proportion to at least 1:1 because there was no showing that the claimed proportions were critical). Where the proportions and identity of components in a feed composition can be varied or substituted and result in the same phenotype, the claimed range(s) and specific components are patentably indistinct from the prior art. Furthermore, it is routine procedure to optimize component amounts to arrive at an optimal product that is superior for its intended use, since it has been held where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. See MPEP § 2144.05.
Therefore, it would have been obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to modify the composition of the WD-HTF diet taught by Malehmir by substituting L-cystine for L-cysteine as taught by Yin, substituting the mixture of fats comprising shortening, soybean oil, and lard for lard alone as taught by Malehmir and Sanches, and optimizing the proportion of casein, L-cysteine, lard, and cholesterol as taught by Sanches to arrive at the claimed invention. One would be motivated to substitute L-cystine for L-cysteine as Yin teaches that the two molecules are interchangeable in vivo and as L-cysteine is usually already in its oxidized form, L-cystine, in feed. One would be motivated to substitute the fat mixture of the WD-HTF diet for lard alone as Malehmir teaches that both the WD-HTF diet and CD-HFD diet comprising lard alone as a source of energy from fat result in the same phenotype of steatohepatitis and hepatic fibrosis. One would be motivated to optimize the concentration of feed components as Sanches teaches variation in levels of feed components, particularly fats, sugars, and cholesterol, impacts the phenotype of the animal with NASH induced by diet. One would have a reasonable expectation of success in substituting L-cystine for L-cysteine as Yin teaches that the two molecules are interchangeable after feeding. One would have a reasonable expectation of success in substituting the fat mixture of WD-HTF for lard alone as Malehmir and Sanches both teach that fat source results in substantially the same phenotype in NASH models. One would have a reasonable expectation of success in optimizing feed component amounts as the two different compositions taught by Malehmir that are very close to, and partially overlap with, the claimed ranges of feed components both result in the claimed phenotypes of steatohepatitis and hepatic fibrosis and optimization of component composition ranges is routine in the art.
Conclusion
No claim is allowed.
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Eric B Wright, PhD
Examiner
Art Unit 1632
/Eric B Wright/Examiner, Art Unit 1632
/VALARIE E BERTOGLIO/Primary Examiner, Art Unit 1632
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