Use of androgen receptor blockade as a novel liver regeneration treatment

Abstract

Methods and uses of increasing or accelerating liver regeneration in a subject in need thereof are provided. In some aspects, such methods and uses comprise administering an effective amount of an androgen receptor inhibitor to a subject in need thereof, including, but not limited to, a subject suffering from one or more risk factors of post-hepatectomy liver failure. Any androgen receptor is contemplated for use in the methods described herein including, but not limited to, enzalutamide, apalutamide, bicalutamide, darolutamide, flutamide, nilutamide, or proxalutamide.

Description

USE OF ANDROGEN RECEPTOR BLOCKADE AS A NOVEL LIVER REGENERATION TREATMENTFIELD

[0001] The disclosure relates to the field of androgen receptor blockade for increasing or accelerating liver regeneration in the treatment and / or prevention of liver disease. More particularly, the disclosure provides products and methods for increasing or accelerating liver regeneration in a subject in need thereof. More particularly, the disclosure provides methods and uses for treating a subject in need thereof, wherein the subject is suffering from liver disease, a subject planning to undergo or having undergone a partial hepatectomy, and / or a subject who is suffering from liver failure or one or more risk factors for post-hepatectomy liver failure.BACKGROUND

[0002] Surgical removal of the liver, termed partial hepatectomy, is required for successful treatment of multiple benign and malignant processes. The indications, number, and extent of resections has increased over time across the world as diagnostic, operative, and perioperative modalities have improved. For example, at Mayo Clinic’s Rochester campus approximately 400 partial hepatectomies are performed annually. The ability to perform partial hepatectomy safely is dependent on the ability of the liver to regenerate and return to normal physiologic function. In cases where liver regeneration is impaired or absent, liver dysfunction and / or failure can develop (Starlinger et al., JHEP Rep, 5(4): 100683, 2023). Supportive treatments exist for liver disfunction and failure, but in the worst cases impaired liver function leads to death. Predictive models have been developed to attempt to avoid liver dysfunction following partial hepatectomy, however these are not accurate in all patients. Critically, there are no currently approved medications to either avoid or treat post- hepatectomy liver dysfunction and / or failure (PHLF). This difficult clinical situation is further compounded by the fact most patients undergo these operations for malignant indications, and as such are not candidates for salvage liver transplantation. Hence substantial morbidity, resource utilization, and mortality are associated with this condition.

[0003] Risk factors have been identified for the development of PHLF, which include Metabolic Associated Fatty Liver Disease (MAFLD) and Metabolic Associated Steatohepatitis (MASH) (Kauffmann et al., Hepatobiliary Surg Nutr, 3(5): 238-46, 2014). Obesity remains a significant risk factor for MAFLD / MASH and the rate of obesity in the United States continues to rise, now surpassing 30% of the population overall (Quek et al., Lancet Gastroenterol Hepatol, 8(1 ): 20-30, 2023). Thus, a large proportion of patients undergoing partial hepatectomy are at risk of developing post-hepatectomy liverdysfunction / failu re, and there are no medications to prevent or treat this condition. Thus, there exists a need for compositions and methods to accelerate liver regeneration in a subject in need thereof.SUMMARY

[0004] The disclosure provides products, methods and uses of androgen receptor (AR) inhibition for increasing or accelerating liver regeneration in a subject in need thereof. More particularly, the disclosure provides products, methods and uses for treating a subject in need thereof, wherein the subject is suffering from liver disease, a subject planning to undergo or having undergone a partial hepatectomy, and / or a subject who is suffering from one or more risk factors for post-hepatectomy liver failure.

[0005] In various embodiments of the disclosure, the method disclosed herein involves bringing about, increasing, or accelerating liver regeneration in a subject in need thereof comprising administering an effective amount of an androgen receptor inhibitor to the subject. In an embodiment of the disclosure, the subject is suffering from liver cancer, a benign tumor of the liver, hepatitis, cirrhosis, fatty liver disease, hemochromatosis, or liver failure. In various aspects, the fatty liver disease is metabolic dysfunction-associated steatohepatitis (MASH) or metabolic dysfunction-associated fatty liver disease (MAFLD). In various aspects, the benign tumor is a hemangioma, an adenoma, or a focal nodular hyperplasia.

[0006] In some embodiments, the subject is planning to undergo or has undergone a partial hepatectomy. In various aspects, the partial hepatectomy is about a 50% hepatectomy. In various aspects, the partial hepatectomy is about a 70% hepatectomy.

[0007] In some embodiments, the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF). In various aspects, the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF) after receiving a partial hepatectomy. In various aspects, the subject exhibits multiple risk factors for the development of post-hepatectomy liver failure. In various embodiments, the risk factors for the development of post-hepatectomy liver failure include, but are not limited to, metabolic fatty liver disease (MAFLD), metabolic associated steatohepatitis (MASH), obesity, cirrhosis, diabetes mellitus, chemotherapy-associated steatohepatitis, Hepatitis B or C, malnutrition, renal insufficiency, hyperbilirubinemia, thrombocytopenia, lung disease, or a subject age of 65 years old or greater.

[0008] In some embodiments, the androgen receptor inhibitor is enzalutamide, apalutamide, bicalutamide, darolutamide, flutamide, nilutamide, or proxalutamide. In aspects of the disclosure, the androgen receptor is enzalutamide.

[0009] In some embodiments, the androgen receptor inhibitor is administered to the subject prior to receiving a partial hepatectomy. In various aspects, the androgen receptor inhibitor is administered to the subject at least about 24 hours prior to receiving a partial hepatectomy. In various aspects, the androgen receptor inhibitor is administered to the subject at least about 60 hours prior to receiving a partial hepatectomy. In various aspects, the androgen receptor inhibitor is administered at least about 14 days prior to receiving a partial hepatectomy. In various aspects, the androgen receptor inhibitor is continuously administered to the subject for at least several days after receiving a partial hepatectomy.

[0010] In some embodiments, the androgen receptor inhibitor does not affect the tumor growth kinetics in a subject comprising a tumor.

[0011] In some embodiments, the liver function of the subject increases after administering the androgen receptor inhibitor to the subject.

[0012] In some embodiments, the androgen receptor inhibitor is administered intravascularly, intravenously, subcutaneously, parenterally, pulmonarily, orally, topically, intramuscularly, buccally, sublingually, intranasally, transdermally, vaginally, rectally, or via subcutaneous implant.

[0013] In addition to the aforementioned methods of the disclosure, the disclosure provided compositions for use and uses of an androgen receptor inhibitor to practice the methods disclosed herein are contemplated. For example, the disclosure provides the use of an androgen receptor inhibitor for increasing or accelerating liver regeneration in a subject in need thereof. The embodiments provided herein above for the methods of the disclosure equally apply to the uses of the disclosure as described herein.

[0014] Other features and advantages of the present disclosure will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating preferred embodiments of the disclosure, are given by way of illustration only, because various changes and modifications within the spirit and scope of the disclosure will become apparent to those skilled in the art from this detailed description, including the drawings and the examples.BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Fig.1 provides a schematic of competitive Yes-associated protein 1 (YAP) YAP- Androgen-TEAD signaling. Transcriptional coactivator with PZD-binding motifs (TEADs) are the primary transcription factors for the YAP / TAZ transcription coactivators of the Hippo pathway and the TEAD family has been implicated in processes, such as development, cell growth and proliferation, tissue homeostasis, and regeneration.

[0016] Fig. 2 shows the fold of expression change of mRNA abundance as evaluated via RT-PCR for Yes-associated protein 1 (YAP) target genes CYR61 , CTGF, and NUAK. Hu1545 cells exposed to vehicle or enzalutamide for 6 hours. Exposure of immortalized human male cell line Hu1545 to the AR inhibitor enzalutamide was associated with a dramatic increase in YAP transcriptional activity, as indicated by increases in the expression of YAP target genes.

[0017] Fig. 3 provides the liver to body weight ratio of vehicle (VEH)- or enzalutamide (ENZA)-treated mice, ENZA-treated mice received 30mg / kg every 24 hours at 40 hours post-hepatectomy. Accelerated liver regeneration was observed with ENZA treatment, as denoted by increased liver to body weight ratio at 40 hours post-hepatectomy.

[0018] Fig. 4A-B shows KI-67 staining and quantification after ENZA treatment in mice. Fig. 4A shows Ki-67 staining in vehicle (VEH) or enzalutamide (ENZA) treated mice 30mg / kg q 24 hours at time of hepatectomy (0 timepoint) and at 40 hours. Fig. 4B provides a quantification of Ki-67 staining, with positive cells per 10 high powered fields (HPF). Ki-67 staining demonstrated a marked increase in proliferation at the 40-hour timepoint.DETAILED DESCRIPTION

[0019] The products, methods and uses described herein are used for increasing or accelerating liver regeneration in a subject in need thereof. The disclosure therefore provides androgen receptor inhibitors for increasing or accelerating liver regeneration in a subject in need thereof. In some aspects, the subject is suffering from any ailment which impairs the liver or liver function. In some aspects, such ailment is liver cancer, a benign tumor of the liver, hepatitis, cirrhosis, fatty liver disease, hemochromatosis, or liver failure.

[0020] As used herein, the term “androgen receptor inhibitor” refers to a therapeutic or therapeutic agent that antagonizes androgen receptor signaling. Such therapeutics include, but are not limited to, pharmacological agents, small molecule inhibitors, antigen binding domains, and other compounds or compositions that prevent androgen receptor dimerization, androgen receptor translocation to the nucleus, or androgen receptor binding to androgen response elements (AREs) in the promoters of target genes.

[0021] As used herein, the term “treating” or “treatment” refers to administering an effective amount or a therapeutically effective amount of androgen receptor inhibitor. Treatment, in certain aspects, requires administration of a single dose or multiple doses at regular intervals to alter the course of the condition.

[0022] As used herein, “administering” means providing a therapeutic or “a therapeutic agent”, or “a pharmaceutical agent” to a subject. In some aspects the subject is a mammalor an animal, including but not limited to a human patient, and includes, but is not limited to administering by a medical professional and / or self-administering.

[0023] As used herein, the terms “therapeutically effective amount” and “effective amount” are used interchangeably to refer to an amount of a therapeutic or pharmaceutical agent that provides a therapeutic benefit to an animal, including a human patient.

[0024] As used herein, the term “risk factors” refers to the underlying disease or diseases that necessitated a partial hepatectomy. Such underlying disease risk factors include, but are not limited to, metabolic fatty liver disease (MAFLD), metabolic associated steatohepatitis (MASH), obesity, cirrhosis, diabetes mellitus, chemotherapy-associated steatohepatitis, Hepatitis B or C, malnutrition, renal insufficiency, hyperbilirubinemia, thrombocytopenia, lung disease, or a subject age of 65 years old or greater.

[0025] As used herein, the term “multiple”, in the context of “multiple risk factors”, refers to a subject presenting with two or more risk factors.

[0026] The disclosure provides a novel strategy for increasing or accelerating liver regeneration via methods comprising inhibiting androgen receptor (AR) signaling. The disclosure is based, at least in part, on the discovery that treatment with an AR inhibitor increases the regenerative capacity of hepatic cells. These surprising findings highlight the utility of AR inhibition to promote liver regeneration a subject in need thereof, wherein the subject, for example, has received a partial hepatectomy, or wherein the subject exhibits a malignant indication, such as Metabolic Associated Fatty Liver Disease (MAFLD) and Metabolic Associated Steatohepatitis (MASH). Dysfunctional liver regeneration following surgical resection remains a major cause of postoperative mortality and has no therapeutic options. Without targeted therapies, the current treatment paradigm relies on supportive therapy until homeostasis can be achieved. Pharmacologic acceleration of regeneration represents an alternative therapeutic avenue (Watkins et al., PNAS Nexus, 3: 1-12, 2024). Disclosed herein is the first report concerning the utility of AR inhibition for inducing or accelerating liver regeneration. In exemplary embodiments, the methods comprise administering an effective amount of an androgen receptor inhibitor to a subject in need thereof, wherein said method increases or accelerates liver regeneration in the subject.

[0027] The disclosure provides methods and uses involving androgen receptor inhibitors in the treatment of liver diseases and for liver regeneration.

[0028] The liver functions as a filter for detoxifying various metabolites, and serves as a biochemical defense against foreign and toxic chemicals, antigens, bacteria and cells originating from food composition or in blood circulation. The liver is connected directly to the large and small intestine, the spleen, and the pancreas via the portal vein. The liverregulates protein levels, lipid and carbohydrate concentrations, ammonia blood levels and bile production (Monte et al., World J Gastroenterol, 15(7): 804-16, 2009). These wide arrays of functions make the liver a critical organ for maintaining homeostasis, and this key role is safeguarded by its unique regenerative ability.

[0029] Liver cells possess remarkable regenerative potential. It is well documented that the liver can regenerate its original volume even if 70% of its original mass is resected, with the caveat that the remaining tissue be normal and disease-free (Dong et al., Cell Mol Immunol, 4(4): 241 -52, 2007). The cells responsible for liver regeneration and tissue repair are mature hepatocytes and biliary epithelial cells, although other parts of the liver parenchyma contain and activate stem cells (Cantz et al., Cell Tissue Res, 331 (1): 241 -52, 2007). Such regenerative capacity allows for extensive surgical treatments of both benign and malignant diseases via hepatectomy, but in the context of preexisting liver disease liver resection may exceed the remaining hepatic tissues’ regenerative reserve. Intracellular signaling pathways in hepatocytes are very rapidly (within minutes) activated after partial hepatectomy (Michalopoulos et al., Nat Rev Gastroenterol Hepatol, 18(1): 40-55, 2021 ). Successful liver regeneration relies on a highly regulated response to maintain homeostasis after surgical resection. Disruption of this process, which occurs in various liver diseases including metabolic dysfunction-associated steatotic liver disease (MASLD), can result in subject morbidity and mortality. While individual signaling events associated with liver regeneration are well studied, the mechanisms underlying coordination of these complex processes are not fully understood (Michalopoulos et al., Nat Rev Gastroenterol Hepatol, 18(1 ): 40-55, 2021). Consequently, no pharmacological agents are approved for use to augment liver regeneration or prevent liver dysfunction after surgical resection.

[0030] The disclosure relates to increasing or accelerating liver regeneration in a subject in need thereof. In various embodiments, the subject presents with a liver disease. Although the etiologies of liver diseases vary, many exhibit dysfunction in the same post-damage healing pathways. One such etiology is steatosis, the abnormal retention of lipids within hepatic cells where excess lipid accumulation in vesicles displaces the cytoplasm (Hashizume et al., Eur J Gastroenterol Hepatol, 19: 827-34, 2007). The most common risk factors associated with steatosis are diabetes mellitus, hypertension, obesity, and alcoholism (Sparks et al., J Clin Invest, 118: 2012-15, 2008). Liver damage can result in the accumulation of matrix proteins, the formation of scars, and the alteration of tissue structure and function. As fibrosis develops, chronic compensatory scar-healing processes in the liver commence. Once irreversible distortion of the hepatic architecture and vascular structure occurs, the cirrhotic liver begins to replace functional hepatocytes (Ma et al., Endocr Relat Cancer, 21 (3): R165-82, 2014). Cirrhosis is one etiological factor contributing tohepatocarcinogenesis, but a significant number of patients without cirrhosis develop hepatocellular carcinoma (HCC), indicating a disease process that involves both oncogenic events and virus-related factors. Hepatitis B virus (HBV) is a well-known etiological factor contributing to cirrhotic liver progression and early HCC development. Anti-viral therapeutics have been contemplated in this context, but there is evidence that such anti-viral therapeutics have little effect on survival of patients with advanced-stage disease (Shin et al., J Gastroenterol Hepatol, 27: 1528-34, 2012) or on disease recurrence after hepatectomy for HCC (Chan et al., Arch Surg, 146: 675-81 , 2011 ).

[0031] In various embodiments, the subject in need thereof is suffering from a disease whose symptoms or progression would be mitigated or alleviated by increasing liver regeneration. Exemplary diseases the subject may suffering from include, but are not limited to, liver cancer, a benign tumor of the liver, hepatitis, cirrhosis, fatty liver disease, hemochromatosis, or liver failure. Subjects suffering from these diseases rely on the regenerative capacity of any remaining healthy liver tissue, with the possibility of resection of the diseased tissue. While solely relying on the regenerative capacity of the subject’s liver would be preferable to undergoing invasive surgery, especially when the subject suffers from an underlying liver disease, there are currently no pharmaceutical interventions available for agonizing post-damage healing pathways and improving liver regeneration.

[0032] In various aspects, the fatty liver disease includes metabolic dysfunction- associated steatohepatitis (MASH) or metabolic dysfunction-associated fatty liver disease (MAFLD). Metabolic dysfunction-associated steatotic liver disease (MASLD) represents a clinical spectrum of disease from simple steatosis to metabolic dysfunction-associated steatohepatitis (MASH) (Hardy et al., Annu Rev Pathol, 11 : 451-96, 2016). MASLD, as a risk factor for liver injury and PHLF, is particularly important as the prevalence of MASLD is estimated to be 30% and rising in the United States (Younossi et al., Hepatology, 64(1): 73- 84, 2016). Consistent with clinical outcomes in patients with MASH, preclinical models of MASH also demonstrate significant risk for mortality following partial hepatectomy (Calthorpe et al., Ann Surg, 277: e592-96, 2021). The ability to augment or accelerate liver regeneration pharmacologically, in patients with or without MASH, may provide an avenue for prevention or treatment of PHLF. Furthermore, with the limited availability of organs suitable for transplantation (Vodkin et al., Clin Liver Dis, 21 (2): 289-301 , 2017), pharmacologic enhancement of liver regeneration could provide therapeutic options even for patients who are candidates for salvage liver transplantation.

[0033] MAFLD is a condition of fat accumulation in the liver in combination with metabolic dysfunction in the form of obesity and insulin resistance. It is also associated with an increased cardiovascular disease risk, including hypertension and atherosclerosis. Hepaticlipid metabolism is regulated by a combination of the uptake and export of fatty acids, de novo lipogenesis, and fat utilization via p-oxidation. When the balance between these pathways is altered, hepatic lipid accumulation commences, and long-term activation of inflammatory and fibrotic pathways can progress to worsen the liver disease. There are currently no FDA-approved therapeutics for management of MAFLD, and MAFLD surgical interventions include sleeve gastrectomies and bariatric endoscopies for weight loss, rather than liver resection (Jeeyavudeen et al., World J Gastroenterol, 29(1): 126-43, 2023).

[0034] In various aspects, the benign tumor is a hemangioma, an adenoma, or a focal nodular hyperplasia. Hepatic hemangiomas (HH) are the most common benign tumor of the liver, occurring in approximately 0.4-20% of primary hepatic lesions (Brouwers et al., Br J Surg, 84:314-16, 1997), while focal nodular hyperplasia (FHA) is the second most common, accounting for approximately 8% of primary hepatic lesions (Fukukura et al., J Hepatol, 29: 470-75, 1998). In contrast, hepatic adenoma (HA) rarely occurs. HHs consist of largely asymptomatic clusters of blood-filled cavities. Typical HHs range from a few mm to 3cm and do not increase in size over time. Small (mm to 3cm) and medium (3cm-10cm) HHs are well- defined lesions, requiring no active treatment aside from regular follow-ups. However, giant liver HHs range from 10-20cm in size, which will usually develop symptoms and complications that will require prompt surgical or therapeutic intervention (Bajenaru et al., J Med Life, 8:4-11 , 2015). FNHs are benign hyperplasic lesions that arise in response to preexisting arterial malformations, which are asymptomatic and have no malignant potential, but can cause hemorrhage or rupture in rare cases (LeGout et al., Radiographics, 42(4): 1043-61 , 2022). HAs are rare benign lesions, but they pose a risk of hemorrhage and malignant transformation makes identification and treatment of these lesions critical (Barthelmes et al., HPB, 7: 186-96, 2005). Hepatectomy is contemplated as a treatment modality for subjects presenting with benign tumors of the liver, to increase or accelerate liver regeneration while mitigating the risk of rupture via resection of the benign tumor tissue.

[0035] The disclosure provides methods and uses for treating a patient about to undergo or having undergone a partial hepatectomy.

[0036] Partial hepatectomy is the surgical removal of a portion of the liver. Post- hepatectomy, both death and regeneration of remaining hepatocytes occurs. Under normal conditions regeneration outweighs hepatocyte death, and both liver mass and function are restored rapidly (Sue et al., Br J Surg, 79: 39-42, 1999). In humans, restoration of liver mass is complete around 3 months following partial hepatectomy, with a peak in DNA synthesis at days 7-10 after the operation (Fausto et al., J Hepatol, 57: 692-94, 2012). Regeneration is triggered by an increased metabolic demand placed upon remnant hepatocytes (Michalopoulos et al., Science, 274: 60-66, 1997). Considerable hemodynamic changesoccur following hepatectomy, even in the absence of blood extravasation. The arterial supply through the hepatic artery appears to remain unaffected, although the portal supply per unit of liver tissue triples; the portal vein flow in the hepatic sinusoids elevates the blood pressure and causes decreased oxygen levels within the liver. Similarly, the concentration of intestinal and pancreatic nutrients, including amino acids, carbohydrates, lipids, insulin, toxins, growth factors and numerous cytokines are increased three-fold in a two-thirds hepatectomy (Michalopoulos, J Cell Phys, 2013(2): 286-300, 2007). When a hepatectomy is performed, more than 100 genes, inactivated in normal liver, are expressed rapidly leading to activation of multiple biological factors (Taub, FASEB J, 10(4): 413-27, 1996; Taub, Nat Rev, 5(10): 836-47, 2004). For example, HGF, which is stored in liver matrix in large quantities, is rapidly diminished 1-2 hours after partial hepatectomy, while its plasma levels substantially increase by 10- to 20-fold. There is no agreement in the field that a single agent alone can lead to liver regeneration, although HGF has been considered a primary candidate for single agent liver regeneration as its receptor (cMet) is activated early in the regenerative process, exerting significant mitotic action on hepatocytes and triggering multiple events during regeneration including massive liver enlargement (Pediaditakis et al., Hepatology, 34(4 pt 1): 688-93, 2001 ; Huh et al., Proc Natl Acad Sci, 101 (13): 4477-82, 2004). Disclosed herein is the first report that androgen receptor inhibition can induce liver regeneration as a single agent modality.

[0037] The disclosure provides methods and uses involving the delivery of androgen receptor inhibitors for treating liver failure including, but not limited to, post-hepatectomy liver failure. Liver failure is a severe postoperative complication of hepatectomy. Inefficient or absent regeneration of hepatic cells post-hepatectomy can lead to PHLF, a condition in which the liver cannot functionally meet a subject’s physiologic requirements (Kauffmann et al., Hepatobiliary Surg Nutr, 3: 238-46, 2014). Incidence of liver failure post-hepatectomy is reportedly between 0.7% and 33.83% (Riberio et al., Eur J Surg Oncol, 39: 380-85, 2013). However, PHLF is the number one cause of postoperative mortality, and there are no effective treatment options (Calthorpe et al., Ann Surg, 277: e592-96, 2021 ). Instead, PHLF is managed with postoperative supplementation of albumin, fibrinogen or prothrombin complex, intravenous nutrition, and blood transfusion (Jin et al., World J Gastroenterol, 19(44): 7983-91 , 2013). The regenerative capacity of the liver is the single most relevant factor determining postoperative outcome after hepatic resection, as PHLF remains the most frequent cause of mortality after partial hepatectomy (Sparrelid et al, BJS Open, 6(6): 142, 2022). Consequently, increasing the regenerative capacity of a subject’s liver may decrease the risk of liver failure post-hepatectomy.

[0038] PHLF represents a spectrum of diseases ranging from limited hepatic insufficiency to acute fulminant hepatic failure following liver resection (Ray et al., Ann Med Surg (Lond), 34: 4-10, 2018). PHLF manifests with progressive multi-system organ failure, including renal insufficiency, encephalopathy, need for ventilator support, and need for pressor support. As hepatic function deteriorates, patients often develop persistent hyperbilirubinemia and coagulopathy (Roberts et al., HPB (Oxford), 15: 345-51 , 2008). Up to 10% of major hepatectomy patients develop PHLF (Vibert et al., Surgery, 155:94-105, 2014), and development of this condition represents the major source of morbidity and mortality after liver resection. Liver resection outcomes have improved due to refinements in operative technique and advances in critical care (Jaeck et al., Liver Transpl, 10: S58-63, 2004), but even then resection is still burdened by relatively high rates of postoperative morbidity (4.09% to 47.7%) and mortality (0.24% to 9.7%) (Sato et al., J Gastroenterol, 47: 1125-33, 2012).To date there are no approved therapeutics for prophylaxis or treatment of PHLF (Soreide et al., Eur J Surg Oncol, 47: 216-24, 2021). The most effective treatment for PHLF is liver transplantation, but patients receiving a partial hepatectomy due to underlying liver disease are often too medically fragile to qualify as candidates for salvage liver transplantation (Ray et al., Ann Med Surg (Lond), 34: 4-10, 2018), which is further reserved for patients who have failed all other supportive therapies (Chan et al., Liver Cancer, 2: 332- 37, 2013). Disclosed herein are methods of treatment that increase or accelerate liver regeneration, wherein liver function is maintained and treated subjects are less likely to develop and / or succumb to physiological deterioration associated with PHLF.

[0039] In various embodiments, the subject is planning to undergo a partial hepatectomy, and prophylactic treatment with an AR inhibitor serves to increase the regenerative capacity of the healthy tissue prior to resection of the diseased tissue. In various embodiments, the subject has already had a partial hepatectomy, and therapeutic treatment with an AR inhibitor serves to increase the regenerative capacity of the remaining hepatic tissue and reduce risk of the subject developing PHLF. In exemplary aspects, the partial hepatectomy is a 30%, 40%, 50%, 60%, or 70% partial hepatectomy. In various aspects the partial hepatectomy is a 50% or 70% partial hepatectomy.

[0040] The disclosure, in some aspects, includes various means of observing whether a subject is suffering from PHLF or exhibits any of the risk factors for PHLF.

[0041] For example, there are multiple clinical measures used to determine whether a subject is experiencing PHLF, including small remnant liver volume (RLV), excessive intraoperative blood loss and need for blood transfusion, pre-operative hypoalbuminemia, prolonged operating time, and male gender (Ray et al., Ann Med Surg (Lond), 34: 4-10, 2018). However, the underlying liver disease that necessitated the partial hepatectomygreatly increases the risk of PHLF (Wakai et al., J Gastrointest Surg, 15(8): 1450-58, 2011). Risk factors, as defined in the instant application, are these underlying liver diseases that predispose the subject to liver failure post-hepatectomy. These underlying liver diseases are often associated with inflammation, fibrosis, and cirrhosis (viral hepatitis, chemotherapy associated steatohepatitis (CASH), non-alcoholic steatohepatitis (NAHS), and alcoholic steatohepatitis, amongst others), are known to increase the risk of post-operative dysfunctional liver regeneration (DLR), and are frequently present in subjects requiring hepatic resection (Forbes et al., Nat Rev Gastroenterol Hepatol, 13: 473-85, 2016). In exemplary embodiments, the risk factors a subject may exhibit for the development of PHLF include metabolic fatty liver disease (MAFLD), metabolic associated steatohepatitis (MASH), obesity, cirrhosis, diabetes mellitus, chemotherapy-associated steatohepatitis, Hepatitis B or C, malnutrition, renal insufficiency, hyperbilirubinemia, thrombocytopenia, lung disease, or a subject age of 65 years old or greater.

[0042] In various aspects, the subject exhibits a risk factor for the development of PHLF prior to receiving the partial hepatectomy. In various embodiments, the subject exhibits a risk factor for the development of PHLF after receiving the partial hepatectomy. In various embodiments, the subject exhibits multiple risk factors for the development of PHLF, wherein “multiple” is defined as two or more risk factors.

[0043] Thus, the disclosure provides a method of treating subjects suffering from a disease of the liver by increasing or accelerating liver regeneration in the subject, optionally wherein the patient has undergone a partial hepatectomy.

[0044] An understanding of YAP-TEAD signaling could be relevant to understanding the role of AR blockade in liver regeneration.

[0045] Yes-associated protein (YAP), a transcriptional coactivator, was initially identified as an oncogene due to YAP’s upregulation of pro-proliferative and survival-enhancing gene programs. YAP and transcriptional co-activator TAZ are regulated by the Hippo tumor suppressor signaling pathway, which controls organ size via regulating cellular proliferation, survival, and differentiation (Coulev et al., Cur Biol, 18: 435-41 , 2008). Hippo pathway activation results in the inactivating phosphorylation and cytoplasmic retention of YAP. Conversely, upon loss of Hippo function unphosphorylated YAP migrates to the nucleus and associates with various DNA binding proteins, such as TEAD, which controls gene transcription. Several genes upregulated by YAP-TEAD include connective tissue growth factor 28 (Ctgf28), CYR61, NUAK, Jagged 1 (Jag 1), and Notch receptor 2 (Notch 2) (Lai et al, Cancer Res, 71 : 2728-38, 2011 ; Totaro et al., Trends Cell Biol, 28(7): 560-73, 2018). YAP is amplified, and its protein level and nuclear localization are elevated, in many types ofhuman cancer including liver, lung, colon, and ovarian cancers. While antagonizing YAP signaling is a potential strategy for cancer treatment, agonizing YAP to promote proliferation and differentiation could serve the purposes of organ regeneration in the context of hepatectomy.

[0046] YAP also plays a role in liver regeneration (Fausto et al., J Hepatol, 57(3): 692-94, 2012), as activating YAP can augment or accelerate liver regeneration (Watkins et al., JCI Insight, 7(15): e159930). Preclinical studies have demonstrated a central role of YAP in regulating liver regeneration; however, the mechanisms both upstream and downstream of YAP remain unresolved and contradictory (Fan et al., Hepatology, 75(1 ): 74-88, 2022; Fan et al., Sci Transl Med, 8(352): 352ra108, 2016; Verboven et al., Gastroenterology, 160(3): 847- 62, 2021 ; Tharehalli et al., Int J Mol Sci, 19(12): 3801 , 2018). For example, TGF-p has been suggested to be both required for YAP-mediated regenerative signaling (Zhang et al., Theranostics, 11 (10): 4743-58, 2021 ), and to be detrimental (Oh et al., J Hepatol, 69(2):359- 67, 2018). Thus, context-specific signaling leading to differences in YAP-mediated signaling effects. In the field of YAP activation in both cancer and regeneration, research in prostate cancer models suggests that AR is directly competing with YAP for occupancy of the TEAD transcription factors such that YAP activation could inhibit growth of AR-responsive prostate cancer (Li et al., EMBO J, 42(4): e112184, 2023). Conversely, AR inhibition may enhance a YAP-TEAD transcriptional program by eliminating AR’s competition for TEAD binding, allowing enhanced YAP binding to TEAD and preferentially upregulating YAP pro- proliferative and survival-enhancing downstream gene programs.

[0047] Based on the work involving the regeneration-associated Hippo / YAP pathway, an interplay between the pro-regenerative molecule YAP and the androgen receptor has been identified. Preliminary studies suggest a direct competition between YAP and androgen receptor for transcriptional activity, such that androgen receptor blockade increases YAP activation in human hepatocytes and liver regeneration in mice. These preliminary data suggest that androgen receptor blockade may be a viable treatment option to augment liver regeneration. If confirmed, this would be a highly impactful finding because androgen receptor blockers are FDA approved and could easily be repurposed in a clinical trial assessing the efficacy of such an approach; providing a therapeutic option for patients where no options currently exist.

[0048] Androgens and steroid hormones bind to the Androgen Receptor (AR), which subsequently dimerizes and translocates to the nucleus to bind genomic androgen response elements (AREs) in the promoters of androgen-responsive genes (Bolton et al., Genes Dev, 21 (16): 2005-17, 2007). AR is differentially expressed in various tissues, including male and female reproductive organs, bones, muscles, the brain, the cardiovascular system, neuraltissues, as well as the immune and hematopoietic systems (Naamenh et al., Best Pract Res Clin Endocrinol Metab, 36(4): 101665, 2022). To date, AR does not have a reported role in liver regeneration or the wound healing response. However, AR signaling does play a role in the development and progression of several liver diseases, including hepatocellular carcinoma (HCC) and nonalcoholic fatty liver diseases. AR activation has also been implicated in obesity, altered lipid metabolism, development of insulin resistance, and hepatic steatosis (Lin et al., Hepatology, 47(6): 1924-35, 2008). Disclosed herein are methods of using AR inhibition to prevent AR competition for TEAD binding, inducing expression of YAP downstream genes and increasing or accelerating liver regeneration in a subject in need thereof.

[0049] Androgen receptor inhibitors contemplated for use with the methods described herein include, but are not limited to, enzalutamide (Xtandi, Astellas Pharma USA, Inc.), apalutamide (Erleada, Janssen Biotech, Inc.), bicalutamide (Casodex, Astrazeneca), darolutamide (Nubeqa, Bayer Healthcare), flutamide (Eulexin, Waylis Therapeutics), nilutamide (Nilandron, ANI Pharmaceuticals), or proxalutamide (Kintor Pharmaceuticals). Disclosed herein is the first report of AR inhibitor use in the context of organ regeneration broadly, and specifically liver regeneration.

[0050] Enzalutamide is an orally bioavailable, nonsteroidal, second-generation antiandrogen that acts at multiple points in the AR signaling pathway (Tran et al., Science, 324(5928): 787-90, 2009). It competitively inhibits androgen binding to AR, nuclear translocation of AR, DNA binding, and coactivator recruitment (Sternberg, Future Oncology, 15(13): 1437-57, 2019). Enzalutamide is indicated for the treatment of patients with castration-resistant prostate cancer and, in some regions, metastatic hormone sensitive prostate cancer.

[0051] The methods contemplated herein comprise treating a subject in need thereof with an effective amount or “therapeutically effective amount” or “effective dose” of AR inhibitor, wherein AR inhibition increases or accelerates liver regeneration. Thus, in various aspects, such terms as “effective amount” or “therapeutically effective amount” or “effective dose” are used interchangeably herein. In embodiments of the disclosure, an effective dose or a therapeutically effective dose is a dose that alleviates (eliminates or reduces) at least one symptom associated with the disease or disorder being treated, that slows or prevents progression of the disease or disorder, that slows or prevents progression of the disease or disorder, that diminishes the extent of the disease or disorder, that results in remission (partial or total) of the disease or disorder, and / or that prolongs survival. The effective amount or effective dose may vary depending on a variety of factors and conditions related to the patient being treated and the severity of the disorder. In some aspects, the effectivedose is a therapeutically effective dose which is determined by the clinician and the dose will depend upon age, weight, and condition of the subject or patient. In some embodiments, the dose or effective dose of AR inhibitor is about 10mg to about 800mg and the dose is administered once or twice daily. In some embodiments, the dose or effective dose of AR inhibitor is about 20mg to about 400mg. In some aspects, the dose is about 10mg, about 20mg, about 30mg, about 40mg, about 50mg, about 60mg, about 70mg, about 80mg, about 90mg, about 100mg, about 110mg, about 120mg, about 130mg, about 140mg, about 150mg, about 120mg, about 130mg, about 140mg, about 150mg, about 160mg, about 170mg, about 180mg, about 190mg, about 200mg, about 21 Omg, about 220mg, about 230mg, about 240mg, about 250mg, about 260mg, about 270mg, about 280mg, about 290mg, about 300mg, about 325mg, about 350mg, about 375mg, about 400mg, about 425mg, about 450mg, about 450mg, about 475mg, about 500mg, about 550mg, about 600mg, about 650mg, about 700mg, about 750mg, or about 800mg. In some aspects, the dose is provided as a higher initial dose given twice daily and then the dose is tapered to a lower dose provided once or twice daily. In an embodiment, the effective dose of AR inhibitor is about 160mg daily. In some aspects, the effective dose of AR inhibitor is administered to the subject in need thereof daily, or in some aspects it is provided every other day or every week.

[0052] In some aspects, the AR inhibitor is administered to the subject at least about 24 hours prior to receiving a partial hepatectomy. In some aspects, the AR inhibitor is administered to the subject at least about 60 hours prior to receiving a partial hepatectomy. In some aspects, the AR inhibitor is administered to the subject at least about 14 days prior to receiving a partial hepatectomy. Thus, in some aspects, the AR inhibitor is administered to the subject one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, 10 days, 11 days, 12 days, 13 days, or 14 days prior to receiving a partial hepatectomy. In some aspects, the AR inhibitor may delivered more than 14 days prior to receiving a partial hepatectomy. In some aspects, the AR inhibitor is continuously administered to the subject for at least several days after receiving a partial hepatectomy. In some aspects, the AR inhibitor is administered to the subject continuously for up to several weeks, or even up to several months or more after receiving a partial hepatectomy.

[0053] In some aspects, the AR inhibitor is administered orally, including in the form of a capsule, tablet, or pill. In some aspects, the AR inhibitor is administered intravascularly, intravenously, subcutaneously, parenterally, pulmonarily, orally, topically, intramuscularly, buccally, sublingually, intranasally, transdermally, vaginally, rectally, or via subcutaneous implant.

[0054] AR inhibitors including, but not limited to, enzalutamide, apalutamide, and darolutamide, are FDA approved for use in multiple types of prostate cancer (Chen et al., Cell Death Dis, 13(7): 632, 2022). However, their approved use has not been reported yet in liver cancer. Nevertheless, the disclosure provides for the use of AR inhibitors in the methods and uses of the disclosure. Such AR inhibitors include, but are not limited to, enzalutamide, apalutamide, bicalutamide, darolutamide, flutamide, nilutamide, or proxalutamide. In various embodiments, wherein the subject in need thereof is suffering from cancer, administration of the AR inhibitor does not affect tumor growth kinetics.

[0055] In various embodiments, administration of an AR inhibitor to a subject in need thereof via the methods disclosed herein increases the subject’s liver function. In various embodiments, the subject’s liver function is determined by the assay of the level or activity of total cholesterol, triglyceride, LDL-cholesterol, aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyl transferase (GGT), alkaline phosphatase (ALP), albumin, total protein, total bilirubin, globulin, creatine kinase (CK), and / or lactate dehydrogenase (LDH) in the subject.

[0056] Throughout this disclosure, and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integer or step. When used herein the term "comprising" can be substituted with the term "containing" or "including" or sometimes when used herein with the term "having."

[0057] When used herein, "consisting of" excludes any element, step, or ingredient not specified in the claim element. When used herein, "consisting essentially of" does not exclude materials or steps that do not materially affect the basic and novel characteristics of the claim.

[0058] In each instance herein any of the terms "comprising", "consisting essentially of" and "consisting of" may be replaced with either of the other two terms.

[0059] It should be understood that this disclosure is not limited to the particular methodology, protocols, material, reagents, and substances, etc., described herein and as such can vary. The terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the subject matter of the disclosure, which is defined solely by the claims.

[0060] All publications and patents cited throughout the text of this disclosure (including all patents, patent applications, scientific publications, manufacturer's specifications, instructions, etc.), whether supra or infra, are hereby incorporated by reference in theirentirety. To the extent the material incorporated by reference contradicts or is inconsistent with this disclosure, this disclosure will supersede any such material.

[0061] A better understanding of the disclosure and of its advantages will be obtained from the following examples, offered for illustrative purposes only. The examples are not intended to limit the scope of the disclosure.EXAMPLES

[0062] Aspects and embodiments of the disclosure are illustrated by the following examples.Example 1Treatment with an AR inhibitor increased YAP transcription and accelerated liver regeneration

[0063] The role of YAP, and its paralog PDZ-binding motif (TAZ), in contributing to tissue growth and hence regeneration has been evaluated in multiple systems. For example, it has been previously demonstrated that YAP activation can accelerate liver regeneration in a murine partial hepatectomy model, wherein two experimental strategies were employed to improve YAP activation; inhibition of a phosphatase responsible for deactivating YAP, and use of a small-molecule inhibitor for mammalian Ste20-like kinases 1 / 2 (MST1 / 2), which is a kinase that can negatively regulate YAP activity (Watkins et al., JCI Insights, 7(15): e159930, 2022; Watkins et al., PNAS Nexus, 3(3): pgae096, 2024). In these studies, liver regeneration was accelerated in wild-type mice, and survival improved in MASH mice following a 70% hepatectomy. These effects were likely reliant on YAP activity, as they were eliminated in YAP / TAZ double knockout mice. Thus, YAP activation appeared to be a potential therapeutic strategy to augment liver regeneration.

[0064] In the study of YAP activation in both cancer and regeneration, research in prostate cancer models has suggested that the androgen receptor (AR) was directly competing with YAP for occupancy of the TEAD transcription factors such that YAP activation (through MST1 / 2 inhibition) could inhibit growth of AR-responsive prostate cancer (Li et al., EMBO J, 42(4): e112184, 2023). Chromatin immunoprecipitation studies demonstrated overlap of the binding sites. Similarly, it has been noted that utilizing YAP- TEAD inhibitor, CA3 in multiple adenocarcinoma models, including both cholangiocarcinoma and gastric cancer models, was associated with upregulation of an AR-regulated transcriptional program (data not shown). Based on this intriguing data, the potential for AR signaling to limit YAP transcriptional activity was investigated in multiple tissues, including hepatocytes (Fig. 1). This research became of particular interest, as the role of estrogen inaltering liver regeneration has been explored, but the role of androgen receptor signaling in altering liver regeneration has not.

[0065] The immortalized human male hepatocyte cell line Hu1545 was treated with the AR inhibitor enzalutamide. Treated cells showed a dramatic increase in YAP transcriptional activity, as indicated by increased in YAP target genes (Fig. 2).

[0066] Exposure of immortalized human male hepatocyte cell line Hu 1545 to AR inhibitor enzalutamide was associated with a dramatic increase in YAP transcriptional activity, as indicated by increases in YAP target genes (Fig. 2). To further develop these findings, male C57BL / 6 mice were treated with vehicle or enzalutamide (30mg / kg gavage once daily) starting 24 hours before a 70% partial hepatectomy. Accelerated liver regeneration was observed, as denoted by increased liver to body weight ratio at 40 hours post-hepatectomy (Fig. 3), a timepoint associated with peak proliferative activity. Ki-67 staining demonstrated a marked increase in proliferation at the 40-hour timepoint (Fig. 4A-B). This data supports a model where AR inhibition via enzalutamide treatment can accelerate liver regeneration, and provides a rationale for comprehensive evaluation of this treatment paradigm in relevant preclinical models.Example 2The effect of AR blockade on liver regeneration after partial hepatectomy in both male and female mice

[0067] Preliminary data indicates that AR blockade can activate YAP signaling in human hepatocytes and accelerate liver regeneration in a partial hepatectomy model. Additional experiments are being carried out to evaluate the effect of AR blockade in both wild-type male and female mice, in conjunction with partial hepatectomy utilizing both a pretreatment approach as well as rescue dosing protocol.

[0068] The efficacy of AR blockade in a partial hepatectomy model is further assessed based on liver to body weight ratio, overall survival, proliferation via Ki-67 staining and Brdll incorporation, YAP activation via immunoblot and RT-PCR, and toxicity based on serum biochemistries. In wild-type mice, the analyses are extended to include RNA sequence and spatial transcriptomics. Success is assessed by accelerated liver regeneration, indicated by increased liver to body weight ratio, and improved survival in the MASH model.

[0069] Both male and female mice experience liver regeneration after AR blockade.Example 3The effect of AR blockade on liver regeneration after partial hepatectomy in a validated MASH mouse model

[0070] Experiments are being carried out to evaluate the effect of AR blockade on liver regeneration after 70% hepatectomy in a validated MASH mouse model (i.e., mice fed a high fat, high fructose (FFC) diet for 6 months), and evaluate survival, with and without AR blockade, but with pretreatment and rescue dosing. For each sex, a minimum of 5 animals per time point is utilized. Mice, fed an FFC diet for 6 months, undergo treatment with enzalutamide (30mg / kg gavage once daily) or vehicle starting either 5 days before (pretreatment cohort) or 12 hours after 70% partial hepatectomy (rescue cohort). Dosing of enzalutamide continues every 24 hours until each mouse has reached humane survival endpoint or 7 days, and the mouse is euthanized.

[0071] Enzalutamide treatment as compared to vehicle treatment increases liver function and or liver regeneration and improves survival.Example 4Determining the role of YAP (and its paralog TAZ) by completing partial hepatectomy with and without AR blockade in conditional YAP / TAZ double knockout mice

[0072] Experiments are carried out to determine the role of YAP (and its paralog TAZ) by completing partial hepatectomy with and without AR blockade in conditional YAP / TAZ double knockout mice. These conditional knockout mice are developed by breeding the commercially available CK19Cre and YAPflox / flox / TAZflox / floxmice strains which were obtained from Jackson Labs (Watkins et al., JCI Insight, 7(15): e159930, 2022).

[0073] For each animal sex (i.e., male or female), a minimum of five animals per time point are used in the experiments. Homozygous CK19Cre-Yapflox / flox / TAZflox / floxmice undergo treatment with tamoxifen to induce YAP / TAZ knockout in cholangiocytes 5 days prior to initiation of treatment. A separate hepatocyte-specific conditional knockout is achieved by administering AAV8Cre to YAPflox / flox / TAZflox / floxmice 5 days prior to initiation of treatment. In these conditional knockout mice, enzalutamide (30mg / kg gavage once daily) or vehicle is administered, starting either 24 hours before (pretreatment cohort) or 12 hours after 70% partial hepatectomy (rescue cohort). Dosing of enzalutamide continues every 24 hours until each mouse has reached a human survival endpoint or 7 days and the mouse is euthanized. The efficacy of treatment is evaluated by overall mouse survival, liver to body weight ratio and Ki-67 staining.

[0074] Treatment with enzalutamide does not improve survival of treated mice as the mechanism of action is anticipated to require YAP / TAZ signaling.Example 5The effects of AR blockade on the growth kinetics of indwelling tumors

[0075] An important consideration of regenerative therapeutics is the effect of such therapeutics on cancer, since patients undergoing partial hepatectomy are typically doing so for treatment of cancer. A potential advantage of identifying a drug that can enhance liver regeneration is that dosing would be limited to the perioperative timeframe if used clinically.

[0076] Evaluation of the effects of AR blockade on the growth kinetics of indwelling tumors is conducted in several clinically relevant models, including syngeneic cholangiocarcinoma models, patient-derived xenograft cholangiocarcinoma models, and patient-derived colorectal cancer metastases models. Tumor size following vehicle versus enzalutamide treatment is recorded as the primary endpoint. YAP activation and proliferation are evaluated additionally.

[0077] Enzalutamide treatment does not have a significant effect on tumor growth kinetics.Example 6Additional testing for effects of AR blockade on the growth kinetics of indwelling tumors

[0078] Evaluation of the effects of AR blockade on the growth kinetics of indwelling tumors is conducted in several clinically relevant models, including syngeneic cholangiocarcinoma models, patient-derived xenograft cholangiocarcinoma models, and patient-derived colorectal cancer metastases models. Tumor size following vehicle versus enzalutamide treatment is recorded as the primary endpoint. YAP activation and proliferation are evaluated additionally. Treatment with AR blockade is limited to five total treatments to evaluate if an acute dosing regimen eliminates any effects.

[0079] Acute dosing of enzalutamide does not have a significant effect on tumor growth kinetics.Example 7Successful off-label usage of enzalutamide in treating liver failure in a human subject

[0080] A 60-year old male underwent extended right hepatectomy for locally advanced cholangiocarcinoma. Following resection, the male subject suffered from progressive postoperative liver failure with rising biochemical markers, including increased levels ofInternational Normalized Ration (INR) and bilirubin. By standard International Study Group of Liver Surgery (ISGLS) guidelines, the subject suffered from grade C liver failure. By postoperative day 15, the subject’s total bilirubin level had risen to 14 mg / dL (whereas normal is usually about 0.3 to 1 .2 mg / dL); INR level was 4.0 with plasma supplementation (whereas normal is usually about 0.8 to 1 .2), and creatine level was 2.5 mg / dL (whereas normal is usually about 0.7 to 1 .3 mg / dL). Based on the preclinical studies and the dire clinical situation without other treatment options, off-label enzalutamide (160 mg orally daily) was initiated for 10 days.

[0081] By day 4 of treatment (i.e., dosing with enzalutamide), the subject’s serum biochemistries stabilized, and by day 10 of treatment the subject’s blood biochemistry markers improved, e.g., creatine level went down to 2 mg / dL, total bilirubin level went down to 10 mg / dL, and INR level went down to 3.6 without plasma supplementation. Further, the subject’s clinical condition and serum biochemical markers, i.e., lab values, continued to improve for 10 days following the initial drug dosing, when his total bilirubin plateaued at 5 mg / dL.

[0082] Redosing of enzalutamide was initiated at 160mg daily for 7 additional days, with resumption of normalization of the subject’s lab values. At the end of the second dosing interval, the subject’s total bilirubin level decreased to 2.1 mg / dL and his INR level decreased to 1 .8 without any plasma supplementation.

[0083] After two rounds of dosing with enzalutamide, the subject’s liver function fully recovered, providing clinical proof that treatment with an androgen receptor inhibitor, e.g., enzalutamide, is successful in improving the function of the liver consistent with accelerating and promoting liver regeneration.

[0084] While the disclosure has been described in terms of specific embodiments, it is understood that variations and modifications will occur to those skilled in the art.Accordingly, only such limitations as appear in the claims should be placed on the disclosure.

[0085] All documents referred to in this application are hereby incorporated by reference in their entirety.

Claims

CLAIMSWHAT IS CLAIMED:1 . A method of increasing or accelerating liver regeneration in a subject in need thereof comprising administering an effective amount of an androgen receptor inhibitor to the subject.

2. The method of claim 1 , wherein the subject is suffering from liver cancer, a benign tumor of the liver, hepatitis, cirrhosis, fatty liver disease, hemochromatosis, or liver failure.

3. The method of claim 2, wherein the fatty liver disease is metabolic dysfunction-associated steatohepatitis (MASH) or metabolic dysfunction-associated fatty liver disease (MAFLD).

4. The method of claim 2, wherein the benign tumor is a hemangioma, an adenoma, or a focal nodular hyperplasia.

5. The method of any one of claims 1-4, wherein the subject is planning to undergo or has undergone a partial hepatectomy.

6. The method of claim 5, wherein the partial hepatectomy is about a 50% hepatectomy.

7. The method of claim 5, wherein the partial hepatectomy is about a 70% hepatectomy.

8. The method of any one of claims 1-7, wherein the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF).

9. The method of any one of claims 1-8, wherein the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF) after receiving a partial hepatectomy.

10. The method of claim 8 or claim 9, wherein the subject exhibits multiple risk factors for the development of post-hepatectomy liver failure (PHLF).11 . The method of claim 8 or claim 9, wherein the risk factor or factors for the development of post-hepatectomy liver failure (PHLF) is metabolic fatty liver disease (MAFLD), metabolic associated steatohepatitis (MASH), obesity, cirrhosis, diabetes mellitus, chemotherapy-associated steatohepatitis, Hepatitis B or C, malnutrition, renal insufficiency, hyperbilirubinemia, thrombocytopenia, lung disease, or a subject age of 65 years old or greater.

12. The method of any one of claims 1-11 , wherein the androgen receptor inhibitor is enzalutamide, apalutamide, bicalutamide, darolutamide, flutamide, nilutamide, or proxalutamide.

13. The method of claim 12, wherein the androgen receptor inhibitor is enzalutamide.

14. The method of any one of claims 1-13, wherein the androgen receptor inhibitor is administered to the subject prior to receiving a partial hepatectomy.

15. The method of any one of claims 1-14, wherein the androgen receptor inhibitor is administered to the subject at least about 24 hours prior to receiving a partial hepatectomy.

16. The method of any one of claims 1-15, wherein the androgen receptor inhibitor is administered to the subject at least about 60 hours prior to receiving a partial hepatectomy.

17. The method of any one of claims 1-16, wherein the androgen receptor inhibitor is administered at least about 14 days prior to receiving a partial hepatectomy.

18. The method of any one of claims 1-17, wherein the androgen receptor is continuously administered to the subject for at least several days after receiving a partial hepatectomy.

19. The method of any one of claims 2-18, wherein the androgen receptor inhibitor does not affect tumor growth kinetics in a subject comprising a tumor.

20. The method of any one of claims 1-19, wherein liver function of the subject increases after administering the androgen receptor inhibitor to the subject.21 . The method of any one of claims 1-20, wherein the androgen receptor is administered intravascularly, intravenously, subcutaneously, parenterally, pulmonarily, orally, topically, intramuscularly, buccally, sublingually, intranasally, transdermally, vaginally, rectally, or via subcutaneous implant.

22. Use of an androgen receptor inhibitor for increasing or accelerating liver regeneration in a subject in need thereof.

23. The use of claim 22, wherein the subject is suffering from liver cancer, a benign tumor of the liver, hepatitis, cirrhosis, fatty liver disease, hemochromatosis, or liver failure.

24. The use of claim 23, wherein the fatty liver disease is metabolic dysfunction- associated steatohepatitis (MASH) or metabolic dysfunction-associated fatty liver disease (MAFLD).

25. The use of claim 23, wherein the benign tumor is a hemangioma, an adenoma, or a focal nodular hyperplasia.

26. The use of any one of claims 22-25, wherein the subject is planning to undergo or has undergone a partial hepatectomy.

27. The use of claim 26, wherein the partial hepatectomy is about a 50% hepatectomy.

28. The use of claim 26, wherein the partial hepatectomy is about a 70% hepatectomy.

29. The use of any one of claims 22-28, wherein the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF) after receiving a partial hepatectomy.

30. The use of any one of claims 22-29, wherein the subject exhibits a risk factor for the development of post-hepatectomy liver failure (PHLF) after receiving a partial hepatectomy.31 . The use of claim 29 or claim 30, wherein the subject exhibits multiple risk factors for the development of post-hepatectomy liver failure (PHLF).

32. The use of claim 29 or claim 30, wherein the risk factor or factors for the development of post-hepatectomy liver failure (PHLF) is metabolic fatty liver disease (MAFLD), metabolic associated steatohepatitis (MASH), obesity, cirrhosis, diabetes mellitus, chemotherapy-associated steatohepatitis, Hepatitis B or C, malnutrition, renal insufficiency, hyperbilirubinemia, thrombocytopenia, lung disease, or a subject age of 65 years old or greater.

33. The use of any one of claims 22-32, wherein the androgen receptor inhibitor is enzalutamide, apalutamide, bicalutamide, darolutamide, flutamide, nilutamide, or proxalutamide.

34. The use of claim 33, wherein the androgen receptor inhibitor is enzalutamide.

35. The use of any one of claims 22-34, wherein the androgen receptor inhibitor is administered to the subject prior to receiving a partial hepatectomy.

36. The use of any one of claims 22-35, wherein the androgen receptor inhibitor is administered to the subject at least about 24 hours prior to receiving a partial hepatectomy.

37. The use of any one of claims 22-36, wherein the androgen receptor inhibitor is administered to the subject at least about 60 hours prior to receiving a partial hepatectomy.

38. The use of any one of claims 22-37, wherein the androgen receptor inhibitor is administered at least about 14 days prior to receiving a partial hepatectomy.

39. The use of any one of claims 22-38, wherein the androgen receptor inhibitor is continuously administered to the subject for at least several days after receiving a partial hepatectomy.

40. The use of any one of claims 23-39, wherein the androgen receptor inhibitor does not affect tumor growth kinetics in a subject comprising a tumor.41 . The use of any one of claims 22-40, wherein liver function of the subject increases after administering the androgen receptor inhibitor to the subject.

42. The use of any one of claims 22-41 , wherein the androgen receptor inhibitor is administered intravascularly, intravenously, subcutaneously, parenterally, pulmonarily, orally, topically, intramuscularly, buccally, sublingually, intranasally, transdermally, vaginally, rectally, or via subcutaneous implant.

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