Use of small molecule inhibitors of lactate dehydrogenase to treat acute and chronic kidney injury

Abstract

Methods and compositions are directed to treating and / or preventing AKI includes the regulation of lactate levels or the modulation of Mg2+ trafficking. In certain aspects treating and / or preventing AKI includes administering a lactate dehydrogenase (LDH) inhibitor or a mitochondrial RNA Splicing 2 protein (MRS2) inhibitor to a subject at risk of developing, suspected of having, or having AKI.

Description

OF SMALL MOLECULE INHIBITORS OF LACTATE DEHYDROGENASE TO TREAT ACUTE AND CHRONIC KIDNEY INJURYRELATED APPLICATION

[0001] This Application claims priority to US Provisional Patent Application 63 / 535,569 filed August 30, 2023 which is incorporated herein by reference in its entirety.STATEMENT REGARDING FEDERALLY FUNDED RESEARCH

[0002] None.FIELD

[0003] Aspects of the invention are directed generally to the field of medicine and particularly the field of nephrology.BACKGROUND

[0004] Acute kidney injury (AKI) is the abrupt loss of kidney function, resulting in the retention of urea and other nitrogenous waste products and in the dysregulation of extracellular volume and electrolytes. AKI can be fatal and requires immediate and intensive treatment. If treated appropriately and in time, AKI is often reversible. Causes of AKI are frequently classified as prerenal, intrinsic, or post-renal. Prerenal causes of AKI are those that decrease effective blood flow to the kidney. Sources of damage to the kidney itself are dubbed intrinsic causes. Post-renal causes of AKI are those resulting in urinary tract obstruction.

[0005] AKI is a common event associated with high morbidity and mortality. Ischemia is a major cause of AKI. During ischemia, a decrease in oxidative phosphorylation results in the accumulation of lactate. Levels of lactate are known to have diagnostic and prognostic value in ischemia and sepsis, but the notion that lactate may contribute to organ injury has received little attention.

[0006] The human Mitochondrial RNA Splicing 2 protein (MRS2) has been implicated in Mg2+ transport across mitochondrial inner membranes, thus playing an important role in Mg2+ homeostasis critical for mitochondrial integrity and function. Magnesium is the most abundant divalent cation in the body and regulates many metabolic pathways. In contrast to Ca2+, theother major divalent cation, virtually nothing is known about the trafficking of intracellular Mg2+ or its role in pathophysiology, such as tissue ischemia. Perturbations in intracellular Mg2+ concentrations can cause serious cellular dysfunction. For example, decreases in intracellular free Mg2+ lead to defective immune responses, mutations in the Na+ / Mg2+ exchanger causing chronic intracellular Mg2+ deficiency trigger neuronal damage, and overexpression of Mg2+ channels is a hallmark of several types of cancer, to name a few.

[0007] There remains a need for additional compositions and methods for treating and / or preventing ischemic acute kidney injury.SUMMARY

[0008] A solution to treating and / or preventing AKI includes the regulation of lactate levels or the modulation of Mg2+ trafficking. In certain aspects treating and / or preventing AKI includes administering a lactate dehydrogenase (LDH) inhibitor or a mitochondrial RNA Splicing 2 protein (MRS2) inhibitor to a subject at risk of developing, suspected of having, or having AKI. In certain aspects, a small molecule inhibitor of lactate dehydrogenase is sodium oxamate or (2-(5-(Cyclopropylmethyl)-4-(3-fluoro-4-sulfamoylbenzyl)-3-(3-((5-methylthi ophen- 2-yl)ethynyl)phenyl)-lH-pyrazol-l-yl)thiazole-4-carboxylic Acid, also known as NCATS- Sml441) or (6 / ?)-3-(2-Chlorophenoxy)-6-(4-morpholinophenyl) -6-(thiophen-3-yl)piperidine- 2,4- dione, also known as GNE-140.

[0009] Certain embodiments are directed to methods of treating a patient or organ (e.g., kidney) having or at risk of developing acute kidney injury or ischemic injury in general. The treatment can include administering an effective amount of an LDH inhibitor or a MRS2 inhibitor to the patient or organ (e.g., kidney). In certain aspects the patient is a mammal, e.g., human. In certain circumstances, the patient is elderly, pregnant, a surgical patient, or has been exposed to a nephrotoxic agent. In certain instances, a patient or subject is a surgical patient or will undergo a surgical procedure (e.g., a surgical procedure, including transplant surgery, is scheduled within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more days or weeks. The method of claim 20, wherein the lactate dehydrogenase inhibitor or the Mrs2 inhibitor is administered to the surgical patient prior to surgery. The LDH inhibitor or the MRS2 inhibitor can be administered to the patient or subject before; after; during; before and after; before and during; after and during; or before, after and during surgery. In certain aspects the LDH inhibitor or the MRS2 inhibitor canbe administered to a donor or a donor organ prior to harvesting of the organ (e.g., kidney(s)) for transplantation. In certain aspects the LDH inhibitor and / or the MRS2 inhibitor can be added to a preservation solution during storage or perfusion of kidneys while awaiting transplantation.

[0010] The nephrotoxic agent can be a drug, chemical, environment capable of causing acute kidney injury. The LDH inhibitor, the MRS2 inhibitor, or the LDH inhibitor and MRS2 inhibitor can be administered before; after; during; before and after; before and during; after and during; or before, after and during development of acute kidney injury or under conditions that put a patient or subject at risk of developing AKI.

[0011] In certain aspects acute kidney injury is characterized by a serum creatinine level of at least 1.5 times baseline, wherein baseline refers to the patient's serum creatinine level no more than 7 days prior, or an increase in serum creatinine of at least 0.3 mg / dL over the previous 48 hours. In other aspects the acute kidney injury is characterized by having a urine output of less than 0.5 mL / Kg over 6 hours or less than 0.3 mL / Kg over 12 hours. In other aspects the acute kidney injury is characterized by the patient having anuria for 12 or more hours. The lactate dehydrogenase inhibitor can be oxamate, NCATS-SM1441 or GNE-140. The MRS2 inhibitor can be CP ACC.

[0012] In certain aspects the nephrotoxic agent is a drug or chemical capable of causing acute kidney injury. The drug or chemical (e.g., chemotherapeutics and other pharmaceuticals) can be one or more of the compounds selected from the group consisting of: cisplatin; gentamicin; cephaloridine; cyclosporine; amphotericin; carbon tetrachloride; trichloroethylene; and dichloroacetylene.

[0013] Other embodiments of the invention are discussed throughout this application. Any embodiment discussed with respect to one aspect of the invention applies to other aspects of the invention as well and vice versa. Each embodiment described herein is understood to be embodiments of the invention that are applicable to all aspects of the invention. It is contemplated that any embodiment discussed herein can be implemented with respect to any method or composition of the invention, and vice versa. Furthermore, compositions and kits of the invention can be used to achieve methods of the invention.

[0014] The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and / or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.”

[0015] Throughout this application, the term “about” is used to indicate that a value includes the standard deviation of error for the device or method being employed to determine the value.

[0016] The use of the term “or” in the claims is used to mean “and / or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or.”

[0017] As used in this specification and claim(s), the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[0018] As used herein, the terms “comprises,” “comprising,” “includes,” “including,” “has,” “having,” “contains”, “containing,” “characterized by” or any other variation thereof, are intended to encompass a non-exclusive inclusion, subject to any limitation explicitly indicated otherwise, of the recited components. For example, a chemical composition and / or method that “comprises” a list of elements (e.g., components or features or steps) is not necessarily limited to only those elements (or components or features or steps), but may include other elements (or components or features or steps) not expressly listed or inherent to the chemical composition and / or method.

[0019] As used herein, the transitional phrases “consists of’ and “consisting of’ exclude any element, step, or component not specified. For example, “consists of’ or “consisting of’ used in a claim would limit the claim to the components, materials or steps specifically recited in the claim except for impurities ordinarily associated therewith (i.e., impurities within a given component). When the phrase “consists of’ or “consisting of’ appears in a clause of the body of a claim, rather than immediately following the preamble, the phrase “consists of’ or “consisting of’ limits only the elements (or components or steps) set forth in that clause; other elements (or components) are not excluded from the claim as a whole.

[0020] As used herein, the transitional phrases “consists essentially of’ and “consisting essentially of’ are used to define a chemical composition and / or method that includes materials, steps, features, components, or elements, in addition to those literally disclosed, provided that these additional materials, steps, features, components, or elements do not materially affect thebasic and novel characteristic(s) of the claimed invention. The term “consisting essentially of’ occupies a middle ground between “comprising” and “consisting of’.

[0021] Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.DESCRIPTION OF THE DRAWINGS

[0022] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of the specification embodiments presented herein.

[0023] FIG. 1A-1C. L-Lactate mediates AKI. (A) WT mice were treated with 2g / kg sodium acetate, sodium D-lactate or sodium L-lactate and then subjected to 22 min bilateral Ischemia Reperfusion Injury (IRI). Blood urea nitrogen (BUN) was measured 24h later as an indicator of kidney injury (n=4). Mice developed only mild kidney injury after this short duration of ischemia, but the degree of injury was markedly enhanced by the administration of L-lactate. In contrast, D-lactate, which is not produced by mammalian cells, had no effect on kidney injury. (B). Mice were treated with a moderate dose of cisplatin (15 mg / kg) and with 2g / kg sodium acetate, sodium D-lactate or sodium L-lactate. The acetate and lactate treatments were repeated 24 hours after cisplatin injection. BUN was measured 72 hours after cisplatin injection. Mice developed only mild kidney injury with this dose of cisplatin. However, treatment with L-lactate, but not D-lactate, markedly increased AKI severity. (C). Histologic examination of kidney tissue harvested 24 hours after IRI or 72 hours after cisplatin revealed minimal injury in the D-lactate treated mice but severe epithelial injury, inflammation and cast formation in the mice which received L-lactate.

[0024] FIG 2A-2C. LDH inhibition prevents ischemic acute kidney injury. (A). Mice were subjected to 26 minutes of bilateral kidney ischemia and kidneys were harvested 2 hours later and the level of L-lactate in kidney tissue was measured. Ischemia resulted in a 2-3 fold increasein lactate levels. Importantly, pretreatment of mice with sodium oxamate (300mg / kg), an inhibitor of LDH, 30 minutes prior to the ischemia prevented the increase in tissue lactate. (B). Mice were pretreated with vehicle or sodium oxamate and subjected to 26 minutes of bilateral kidney ischemia. BUN and creatinine levels were measured 24 hours after IRI as an index of AKI. Ischemia resulted in a marked increase in both BUN and creatinine in vehicle-treated mice whereas oxamate treated mice had no significant change in BUN or creatinine. (C) Histologic examination of kidney tissue harvested 24 hours after IRI showed extensive epithelial injury and neutrophil (Uy6G) infiltration in the vehicle treated mice but minimal injury and inflammation in oxamate-treated mice.

[0025] FIG 3A-3D. A high affinity inhibitor of LDH (NCATS-SM1441) prevents ischemic AKI. Mice were pretreated with vehicle or lOmg / kg NCATS-SM1441 and subjected to 26 minutes of bilateral kidney ischemia. (A) BUN and (B) creatinine were measured 24 hours after ischemia. Vehicle treated mice developed severe AKI whereas NCATS-SM1441-treated mice had no significant change in BUN or creatinine. (C) Histologic examination of kidney tissue harvested 24 hours after IRI showed extensive epithelial injury and neutrophil (Ly6G) infiltration in the vehicle treated mice but minimal injury and inflammation in NCATS-SM441 -treated mice. (D). Primary cultures of renal epithelial cells were exposed to normoxia or hypoxia for 24 hours in the presence or absence of lOmM oxamate or 5 M NCATS-SM1441. Cell viability was determined by Calcein retention. Cell viability was reduced by exposure to hypoxia but this was prevented by treatment with the either LDH inhibitor.

[0026] FIG. 4A-4C. A high affinity LDH inhibitor (GNE-140) prevents ischemic AKI. Mice were pretreated with vehicle or 5mg / kg GNE-140 and subjected to 26 minutes of bilateral kidney ischemia. (A) BUN and (B) creatinine were measured 24 hours after ischemia. Vehicle treated mice developed severe AKI whereas GNE-140-treated mice had no significant change in BUN or creatinine. (C) Primary cultures of renal epithelial cells were exposed to normoxia or hypoxia for 24 hours in the presence or absence of IpM or 5pM NCATS-SM1441. Cell viability was determined by Calcein retention. Cell viability was reduced by exposure to hypoxia but this was prevented by treatment with GNE-140.

[0027] FIG 5A-5C. LDH inhibition prevents cisplatin-induced AKI. (A). Mice were treated with 20mg / kg cisplatin to induce severe AKI in the presence or absence of pre-treatment wit lOmg / kg NCATS-SM1441. Creatinine and BUN were measured 72 hours later. Cisplatinresulted in severe AKI but this was completely prevented by NCATS-SM1441 . (B). Histologic examination of kidney tissue harvested 72 hours after cisplatin revealed extensive epithelial inury and cast formation. These changes were completely prevented by NCATS-Sml441. (C). Primary cultures of renal epithelial cells were exposed to saline or 50pM cisplatin for 24 hours in the presence of increasing concentrations of NCATS-SM1441. Cell viability was determined by Calcein retention. Cell viability was reduced by exposure to cisplatin but this was prevented by treatment with NCATS-SM1441 at sub-micromolar concentrations.

[0028] FIG 6A-6B. Treatment with an LDH inhibitor can rescue mice subjected to ischemic inury. (A). Mice were subjected to 26 minutes of bilateral IRI and then received either vehicle or lOmg / kg NCATS-SM1441 either 2 or 6 hours after the ischemia. Vehicle treated mice developed severe AKI as reflected by increases in creatinine and BUN. Mice which received NCATS- SM1441 2 hours after injury had significantly lower BUN and creatinine levels. Even when delayed for 6 hours after ischemia, NCATS-SM1441 reduced the severity of the AKI. (B). NGAL and KIMI are markers of kidney injury. Levels of NGAL aND KIMI mRNA were measured in kidney tissue harvested 24 hours after IRI. IRI resulted in large (400-600-fold) increase in both NGAL and KIMI expression. The expression of these markers was dramatically reduced when NCATS-SM1441 was administered 2 hours after ischemia and, to a lesser extent, when administered 6 hours after ischemia.

[0029] FIG. 7A-7D. Deletion of the Mrs2 mitochondrial Mg transporter prevents ischemic AKI. Wild type (WT) and Mrs2 deficient (Mrs2KO) mice were subjected to 26 minutes of bilateral renal ischemia. BUN (A) and creatinine (B) levels were markedly higher in WT as compared to Mrs2KO mice (n=9). (C) Tubular injury (right) and neutrophil infiltration (left) were significantly lower in Mrs2 KO mice than WT mice 72 h after IRI. (D) Primary cultures of renal epithelial cells from WT or Mrs2KO kidneys were subjected to 24 h of either normoxia or hypoxia. Cell viability was determined by Calcein retention (n=10 wells from 2 independent experiments). Exposure to hypoxia reduced cell viability in WT cells but Mrs2 KO cells were resistant to hypoxic cell death.

[0030] FIG. 8A-8D. Deletion of the Mrs2 mitochondrial Mg transporter prevents cisplatin- induced AKI. Wild type (WT) and Mrs2 deficient (Mrs2KO) mice were treated with 20mg / kg cisplatin to induce AKI. BUN (A) and creatinine (B) levels measured 72 hours after cisplatin injection were markedly higher in WT as compared to Mrs2KO mice. (C) Histologic analysis ofkidneys harvested 72 hours aftyer cisplatin showed extensive epithelial necrosis and cast formation in WT mice but well-preserved histology in the Mrs2 KO mice. (D) Primary cultures of renal epithelial cells from WT or Mrs2K0 kidneys were exposed to vehicle or 50 pM cisplatin for 24 hours. Cell viability was determined by Calcein retention. Exposure to cisplatin reduced cell viability in WT cells but Mrs2 KO cells were resistant to cisplatin cell death.

[0031] FIG. 9A-9C. Mrs2 Inhibition reduces ischemic and toxic AKI and hypoxic cell death.(A). WT mice were subjected to 26 min of bilateral IRI in presence or absence of pretreatment with 20mg / kg CP ACC, an inhibitor of the Mrs2 protein. Ischemia produced severe AKI in the untreated mice but this was prevented by CP ACC. (B). Primary renal epithelial cells were exposed to normoxia or hypoxia for 24 h in the presence of vehicle or 5 pM CPACC. Hypoxia reduced cell viability in the vehicle-treated cells but cell viability was better preserved by CPACC. (C). Pretreatment of mice with 20mg / kg CPACC reduced BUN and creatinine levels in response to 20 mg / kg cisplatin (CP).

[0032] FIG 10A-10D. LDH inhibition prevents the development of fibrosis after ischemic AKI. Mice were subjected to unilateral ischemia of 35 minutes duration, leaving the contralateral kidney intact, a model for progressive chronic kidney disease in the ischemic kidney. From day 3 after the ischemia through day 21, mice received either sodium oxamate 400mg / kg or sodium acetate 300mg / kg (control). Mice were sacrificed on day 28. (A). Oxamate treatment prevented renal atrophy. In the control mice, the ischemic kidney was smaller than the contralateral nonischemic kidney. In oxamate-treated mice, the weight of the ischemic kidney remained normal.(B) Oxamate prevented kidney fibrosis after ischemia. Kidney sections of the ischemic kidneys were stained for fibrosis with Picrosirius Red stain. The ischemic kidneys in the control mice had significantly greater fibrosis than kidneys from oxamate-treated mice. (C). Representative sections of the ischemic kidney from a control and an oxamate-treated mouse showing the abundance of fibrosis (dark) in the control kidney and no fibrosis in the oxamate kidney. (D). Western blot analysis showed dramatic accumulation of collagen 1A1 and SMA, markers of fibrosis, in the ischemic kidneys of control mice. Oxamate treatment prevented this accumulation. The non-ischemic kidneys from both groups of mice had no fibrosis.

[0033] FIG 11A-11C. Inhibition of Mrs2 prevents the development of CKD after ischemic AKI. Mice were subjected to unilateral ischemia of 35 minutes duration, leaving the contralateral kidney intact, a model for progressive chronic kidney disease in the ischemic kidney. From day 3after the ischemia through day 21, mice received either CP ACC 20mg / kg or saline every 3 days. On Day 27, the non-ischemic kidney was removed, leaving only the ischemic kidney. On Day 28, mice were sacrificed. (A). CP ACC preserved kidney function in the ischemic kidney. Upon removal of the non-ischemic kidney on Day 27, the BUN rose in the saline-treated mice while CP ACC-treated mice had continued low BUN levels. (B). CP ACC prevented kidney fibrosis after ischemia. Kidney sections of the ischemic kidneys were stained for fibrosis with Pircosirius Red stain. The ischemic kidneys in the saline-treated mice had significant fibrosis whereas the kidneys from CP ACC-treated mice were similar to sham (no ischemia) kidneys. (C). CP ACC treatment prevented the accumulation of Collagen 1 and smooth muscle actin (SMA), markers of fibrosis. Western blot analysis showed dramatic accumulation of collagen 1A1 and SMA in the ischemic kidneys of saline-treated mice. CP ACC treatment prevented this accumulation. Kidneys from non-ischemic (sham) mice also showed no Collagen or SMA.

[0034] FIG. 12. Lactate levels are elevated in human kidneys with AKI. Kidney biopsies obtained from subjects with ischemic AKI and healthy subjects (HRT) were analyzed for lactate content using mass spectroscopic imaging. Left: Representative mass spec image of lactate in a healthy (HRT) and AKI kidney biopsy showing areas (bright dots) of lactate accumulation in the AKI sample. Right: Summary data of 6 healthy and 6 AKI samples demonstrating a significantly higher tissue lactate content in the AKI samples.DESCRIPTION

[0035] The following discussion is directed to various embodiments of the invention. The term “invention” is not intended to refer to any particular embodiment or otherwise limit the scope of the disclosure. Although one or more of these embodiments may be preferred, the embodiments disclosed should not be interpreted, or otherwise used, as limiting the scope of the disclosure, including the claims. In addition, one skilled in the art will understand that the following description has broad application, and the discussion of any embodiment is meant only to be an example of that embodiment, and not intended to imply that the scope of the disclosure, including the claims, is limited to that embodiment.I. Acute Kidney Injury (AKI)

[0036] Acute kidney injury carries a high mortality and incidence appears to be increasing (Brown et al., Biomed Res Int. 2016, 4278579; Sawhney and Fraser, Adv Chronic Kidney Dis. 2017, 24(4): 194-204; Hsu et al., J Am Soc Nephrol. 2013, 24(l):37-42). In addition to its shortterm consequences, AKI also increases the risk and rate of progression for chronic kidney disease (CKD) and perhaps cardiovascular events (Chawla et al., New England Journal of Medicine. 2014, 371 (1): 58-66; Ikizler et al., Kidney Int. 2021, 99(2):456-65). It is unlikely that the short-term and long-term morbidity and mortality and associated cost will be reduced until there is a better understanding of the cellular and molecular mechanisms of renal injury and recovery.

[0037] The proximal tubule cell is metabolically active and requires high levels of ATP production to support transcellular solute transport. Mitochondrial dysfunction is a hallmark of several forms of experimental AKI (Funk and Schnellmann, American Journal of Physiology- Renal Physiology. 2012, 302(7):F853-F64; Brooks et al., The Journal of Clinical Investigation. 2009, 119(5): 1275-85; Qi et al., Toxicology and applied pharmacology. 2007, 222(1): 105-10; Liu et al., Hum Exp Toxicol. 2020, 39(5): 673 -82; Li et al., Am J Physiol Renal Physiol. 2020, 319(2):F229-F44). In the presence of decreased oxidative phosphorylation, glycolysis is increased with the production of lactate from pyruvate mediated by the enzyme lactate dehydrogenase (LDH) (Glancy et al., J Physiol. 2021, 599(3):863-88). The cellular release of LDH is widely used as a marker of cell death in vitro (Moran and Schnellmann, Journal of Pharmacological and Toxicological Methods. 1996, 36(l):41-44) and clinically in such conditions as hemolysis or tumor lysis (Gupta, Inflammation. 2022, 45(6):2091 - 123 ; Kato et al., Blood. 2006, 107(6):2279-85; Khan et al., Endocr Metab Immune Disord Drug Targets. 2020, 20(6):855-68). However, the possibility that LDH could be a mediator of cell damage has not been addressed. The studies described herein used both low affinity and high affinity inhibitors of LDH to establish that the inhibition of LDH is highly effective at preventing ischemic and drug-induced kidney injury in vivo and reducing hypoxic and drug-induced cell death in vitro.II. Methods of Treating AKI

[0038] Provided herein are methods for treating acute kidney injury in a patient or subject (e.g., human) having or at risk of developing acute kidney injury. Such methods compriseadministering an effective amount of an LDH inhibitor or a MRS2 inhibitor to the patient. In one embodiment, the LDH inhibitor and / or the MRS2 inhibitor is administered prior to the patient or subject developing AKI. Alternatively, the LDH inhibitor or the MRS2 inhibitor can be administered after the patient or subject develops acute kidney injury.

[0039] As used herein, the term “acute kidney injury” or “AKI”, previously referred to as “acute renal failure” or “ARF”, refers to an acute clinical syndrome characterized by rapid decline in renal functions, which is caused by a number of factors such as a reduction in renal blood flow, glomerulonephritis, use of nephrotoxic antibiotics and anticancer agents.

[0040] Acute kidney injury is defined by an abrupt decrease in kidney function, and can be diagnosed based on when a patient or subject exhibits changes in one or more of serum creatinine level, glomerular filtration rate, or urine output. For example, acute kidney injury can be characterized by a serum creatinine level of at least 1.5 times baseline, wherein baseline refers to the patient's serum creatinine level no more than 7 days prior. For example, a patient having acute kidney injury can have a serum creatinine level is 1.5 to 1.9 times baseline (Stage 1 AKI), 2.0 to 2.9 times baseline (Stage 2 AKI), or 3.0 or more times baseline (Stage 3 AKI). Alternatively, acute kidney injury can be characterized by an increase in serum creatinine of at least 0.3 mg / dL or at least 0.4 mg / dL.

[0041] Alternatively, acute kidney injury can be characterized by the patient or subject having a urine output of less than 0.5 mL / Kg over 6 hours (Stage 1), less than 0.5 mL / Kg over 12 hours (Stage 2), less than 0.3 mL / Kg over 12 hours, or anuria for 12 or more hours (Stage 3).

[0042] Acute kidney injury can occur with specific kidney diseases (e.g., acute interstitial nephritis, acute glomerular and vasculitic renal diseases); non-specific conditions (e.g., ischemia, toxic injury); as well as extrarenal pathology (e.g., prerenal azotemia or acute postrenal obstructive nephropathy). More than one of these conditions may coexist in the same patient or subject and, more importantly, epidemiological evidence supports the notion that even mild, reversible AKI has important clinical consequences, including increased risk of death. Furthermore, because the manifestations and clinical consequences of AKI can be quite similar (even indistinguishable) regardless of whether the etiology is predominantly within the kidney or predominantly from outside stresses on the kidney, the syndrome of AKI encompasses both direct injury to the kidney as well as acute impairment of function.

[0043] In some embodiments, the patient or subject being treated for AKI has, is suspected of having, or at risk of developing diabetes, underlying renal insufficiency, nephritic syndrome, atherosclerotic disease, sepsis, hypotension, hypoxia, myoglobinuria-hematuria, or liver disease. In other embodiments, the patient is elderly, pregnant, a surgical patient, or has been exposed to a nephrotoxic agent. In a specific embodiment, the patient being treated for AKI is a surgical patient. Accordingly, in certain embodiments, the compound(s) are administered to a surgical patient prior to, during, and / or after surgery.

[0044] In another embodiment, the patient being treated for AKI has been exposed to a nephrotoxic agent (a drug or chemical capable of causing AKI). Drugs or chemicals capable of causing acute kidney injury include, but are not limited to chemotherapies, antimicrobials, or other pharmaceuticals or biologies, such as cisplatin; gentamicin; cephaloridine; cyclosporine; amphotericin; carbon tetrachloride; trichloroethylene; dichloroacetylene or the like.

[0045] As such, a patient or subject with any of the conditions mentioned above is at risk for developing AKI and can be treated with one or more LDH and / or MRS2 inhibitor in accordance with the disclosed methods to reduce the likelihood of developing AKI or ameliorate AKI.

[0046] In another embodiment, the patient or subject has sustained a recent episode of AKI. Such a patient or subject is at increased risk of developing chronic kidney disease, an irreversible condition. Such a patient can be treated with a LDH or MRS2 inhibitor early after the episode of AKI to reduce the risk of developing chronic kidney disease.

[0047] In another embodiment, a kidney harvested from a donor for kidney transplantation can develop kidney injury during the organ procurement or preservation process. Treatment of the donor or donor organ with a LDH inhibitor or MRS2 inhibitor can reduce kidney injury and improve the function of the transplanted kidney. Likewise, inclusion of LDH or MRS2 inhibitor in the organ preservation solutions can prolong organ viability and improve transplant outcomes.

[0048] Kidney transplantation is one process for treating end-stage renal disease. This procedure involves transplanting a healthy kidney from a donor into a patient whose kidneys have irreversibly lost their ability to function d. The transplant not only aims to restore kidney function but also to prevent the life-threatening complications associated with prolonged kidney failure, such as severe electrolyte imbalances, intractable fluid overload, and uremia.

[0049] The kidney transplant process, from harvest to transplant, involves several steps. Initially, the kidney is either harvested from a living donor, who undergoes a surgical procedureto remove one kidney while remaining healthy with the other, or from a deceased donor shortly after death. Once harvested, the kidney(s) is / are immediately preserved in a cold, sterile solution to slow cellular metabolism and prevent damage. This organ is then transported, often in specialized containers that maintain low temperatures, to the recipient's location. Upon arrival, the transplant surgery begins. Surgeons connect the renal artery and vein of the donor kidney to the recipient's iliac artery and vein, respectively, ensuring immediate blood flow. The ureter of the new kidney is then attached to the bladder, allowing urine production to commence shortly after blood flow is established. The entire surgical process aims to minimize ischemia time, the period when the kidney is without blood flow, to ensure the best possible outcome for the transplanted kidney's function. Certain methods described herein can be used to ameliorate ischemic damage by exposing the donor organ to LDH and / or MRS2 inhibitors. In certain aspects an organ preservation fluid or solution can contain an effective amount of an LDH inhibitor and / or a MRS2 inhibitor that is added to preservation solutions during organ storage or perfusion of a organ while awaiting transplant surgery.III. Inhibitors for Treating AKI

[0050] Lactate dehydrogenase is an enzyme found in nearly all living cells. LDH catalyzes the conversion of pyruvate to lactate and back, as it converts NAD+to NADH and back. The term dehydrogenase refers to an enzyme that transfers a hydride from one molecule to another. LDH inhibitors include but are not limited to fluoropyruvic acid, fluoropyruvate, levulinic acid, levulinate, oxamic acid, N-substituted oxamic acids, oxamate, oxalic acid, oxalate, betabromopropionate, beta-chloropropionate, malonate, sodium formaldehyde bisufite, chloroacetic acid, alpha-chloropropionate, alpha-bromopropionate, beta-iodopropionate, acrylate, acetoin, malic acid, glycolate, diglycolate, acetamide, acetaldehyde, acetylmercaptoacetic acid, alpha ketobutyrate, thioglycolic acid, nicotinic acid, alpha-ketoglutarate, butanedione, hydroxypyruvic, chloropyruvic, bromopyruvic, 2,3-dihydroxy-6-methyl-4-(l-methylethyl)-l-naphthoic acid, diethyl pyrocarbonate, hexyl N,N-diethyloxamate, 3 -acetylpyridine adenine dinucleotide, 7-p- Trifluoromethylbenzyl-8-deoxyhemigossylic acid, dihydroxynaphthoic acids, N-substituted oxamic acids, gossypol, gossylic iminolactone, derivatives of gossypol, dihydroxynaphthoic acid, 2,3-dihydroxy-6-methyl-4-(l-methylethyl)-l-naphthoic acid, blue dye, reactive blue dye #2 (Cibacron Blue 3G-A) urea, methylurea and hydantoic acid, glyoxylate, hydroxybutyrate, 4-hydroxyquinoline-2-3 carboxylic acids, sodium bisulfite, dieldrin, L-(+) beta monofluorolactic acid, fluoro-lactic acid, tartronic acid, mesotartarate, sesquiterpene 8-deoxyhemigossylic acid (2,3-dihydroxy-6-methyl-4-(l-methylethyl)-l-naphthoic acid), or analogues of these chemicals. In certain embodiments the LDH inhibitor is Oxamate, NCATS-SM1441, GNE-140, or small molecules with similar activity.

[0051] MRS2 is a magnesium transporter that mediates the influx of magnesium into the mitochondrial matrix. An example of a MRS2 inhibitor is CP ACC or analogs thereof.IV. Pharmaceutical Compositions and Administration ThereofA. Additional Therapeutic Agents

[0052] Pharmaceutical compositions are disclosed that include one or more LDH inhibitor and / or MRS2 inhibitor and typically at least one additional substance, such as an excipient, a known therapeutic other than those of the present disclosure, and combinations thereof. In some embodiments, an LDH inhibitor or a MRS2 inhibitor can be used in combination with other agents known to have beneficial, additive or synergistic activity with the LDH inhibitor or the MRS2 inhibitor.B. Excipients and Dosage Forms

[0053] “Pharmaceutically acceptable excipient” and “pharmaceutically acceptable carrier” refer to a substance that aids in the formulation and / or administration of an active agent to and / or absorption by a subject and can be included in the compositions of the present disclosure without causing a significant adverse toxicological effect on the subject. Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, lactated Ringer's, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer's solution), alcohols, oils, gelatins, carbohydrates such as lactose, amylose or starch, fatty acid esters, hydroxymethycellulose, polyvinyl pyrrolidine, and colors, and the like. Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and / or aromatic substances and the like that do not deleteriously react with or interfere with the activity of the compounds provided herein. One of ordinary skill in the art will recognize that other pharmaceutical excipients are suitable for use with disclosed compounds.

[0054] The particular mode of administration and the dosage regimen will be selected by the attending clinician, taking into account the particulars of the case (e.g., the subject, the disease, the disease state involved, the particular treatment, and whether the treatment is prophylactic). Treatment can involve daily or multi-daily or less than daily (such as weekly or monthly etc.) doses over a period of a few days to months, or even years. However, a person of ordinary skill in the art would immediately recognize appropriate and / or equivalent doses looking at dosages of approved compositions for guidance.C. Methods of Administration

[0055] The precise amount of compound administered to provide a “therapeutically effective amount” to the subject will depend on the mode of administration, the type, and severity of the AKI, and on the characteristics of the subject, such as general health, age, sex, body weight, and tolerance to drugs. The skilled artisan will be able to determine appropriate dosages depending on these and other factors. When administered in combination with other therapeutic agents, e.g., when administered in combination with an anti-cancer agent, an “effective amount” of any additional therapeutic agent(s) will depend on the type of drug used. Suitable dosages are known for approved therapeutic agents and can be adjusted by the skilled artisan according to the condition of the subject, the type of condition(s) being treated and the amount of a compound of the invention being used by following, for example, dosages reported in the literature and recommended in the current Physician's Desk Reference .

[0056] The term “therapeutically effective amount” means an amount when administered to the subject which results in beneficial or desired results, including clinical results, e.g., inhibits, suppresses or reduces the symptoms of the condition being treated in the subject as compared to a control. For example, a therapeutically effective amount can be given in unit dosage form (e.g., 0.1 mg to about 50 g per day, alternatively from 1 mg to about 5 grams per day; and in another alternatively from 10 mg to 1 gram per day).

[0057] The terms “administer”, “administering”, “administration”, and the like, as used herein, refer to methods that may be used to enable delivery of compositions to the desired site of biological action. These methods include, but are not limited to, intravenous, intraarterial, intraarticular (in the joints), intramuscular, intratumoral, intradermal, intraperitoneal, subcutaneous, orally, topically, intrathecally, inhalationally, transdermally, rectally, and the like. Administration techniques that can be employed with the agents and methods described herein are found in e g., Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon; and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa.

[0058] As used herein, the terms “co-administration”, “administered in combination with”, and their grammatical equivalents, are meant to encompass administration of two or more therapeutic agents to a single subject, and are intended to include treatment regimens in which the agents are administered by the same or different route of administration or at the same ordifferent times. In some embodiments the one or more compounds described herein will be coadministered with other agents. These terms encompass administration of two or more agents to the subject so that both agents and / or their metabolites are present in the subject at the same time. They include simultaneous administration in separate compositions, administration at different times in separate compositions, and / or administration in a composition in which both agents are present, co-formulation. Thus, in some embodiments, the therapeutic compositions described herein and the other agent(s) are administered in a single composition. In some embodiments, the compounds described herein and the other agent(s) are admixed in the composition.

[0059] The particular mode of administration and the dosage regimen will be selected by the attending clinician, taking into account the particulars of the case (e.g., the subject, the disease, the disease state involved, the particular treatment, and whether the treatment is prophylactic). Treatment can involve daily or multi-daily doses over a period of a few days. However, a person of ordinary skill in the art would immediately recognize appropriate and / or equivalent doses looking at dosages of approved compositions for treating disease using the disclosed compounds for guidance.

[0060] Pharmaceutical compositions are disclosed that include an LDH inhibitor and / or a MRS2 inhibitor, and typically at least one additional substance, such as an excipient, a known therapeutic other than those of the present disclosure, and combinations thereof.

[0061] The pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration. In an embodiment, the composition is formulated in accordance with routine procedures as a pharmaceutical composition adapted for intravenous, intraarterial, subcutaneous, intramuscular, oral, intranasal, or topical administration to human beings. In preferred embodiments, the pharmaceutical composition is formulated for intravenous or intraarterial administration.V. Examples

[0062] The following examples as well as the figures are included to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples or figures represent techniques discovered by the inventors to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the presentdisclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.EXAMPLE 1

[0063] Acute kidney injury (AKI) is a common event associated with high morbidity and mortality. Ischemia is a major cause of AKI. During ischemia, a decrease in oxidative phosphorylation results in the accumulation of lactate. Levels of lactate are known to have diagnostic and prognostic value in ischemia and sepsis, but the notion that lactate may contribute to organ injury has received little attention. Magnesium is the most abundant divalent cation in the body and regulates many metabolic pathways. In contrast to Ca2+, the other major divalent cation, virtually nothing is known about the trafficking of intracellular Mg2+or its role in pathophysiology, such as tissue ischemia.

[0064] Recent work has shed light on these important issues. LDH, the enzyme responsible for the conversion of pyruvate to lactate, is expressed at high levels in the kidney. The release of LDH has long been used as a measure of cell injury. It was determined that LDH and lactate are not only markers of ischemia but are critical mediators of ischemic kidney injury. Specifically, inhibition of LDH reduces (FIG.s 2-5), while exogenous lactate increases, ischemic and cisplatin-induced acute kidney injury (FIG. 1). In addition, it was recently reported that intracellular lactate triggers the release of Mg2+from the endoplasmic reticulum with subsequent uptake into mitochondria and that the mitochondrial uptake is mediated by the MRS2 transporter. Genetic (FIG. 7 and 8) or pharmacologic inhibition (FIG. 9) of MRS2 was found to dramatically reduce ischemic and cisplatin-induced kidney injury. Thus, important and novel roles for lactate, LDH and MRS2 have been identified in ischemic and cisplatin-induced kidney injury. It is contemplated that these pathways can be targeted to treat, prevent, or ameliorate acute kidney injury.

[0065] It is contemplated that during kidney injury, a shift to glycolysis results in lactate production via LDH which, in turn, triggers the translocation of Mg2+from the ER to mitochondria and initiation of mitochondrial cell death pathways. This theory is based on extensive data which clearly indicate that inhibition of LDH or Mrs2 decrease ischemic AKI andAKI induced by the nephrotoxic drug, cisplatin, in vivo and decrease hypoxic and cisplatin- induced cell death in vitro.

[0066] Lactate and AKI. Proximal tubule cells are highly metabolically active, relying on ATP generation mainly from fatty acid oxidation (van der Rijt et al., Nature Reviews Nephrology, 2022, 18(9):588-603). After kidney injury, oxidative phosphorylation is impaired and glycolysis is stimulated to produce ATP, albeit less efficiently than oxidative phosphorylation (Lan et al., J Am Soc Nephrol. 2016, 27(11):3356-67; Li et al. Am J Physiol Renal Physiol. 2020, 319(2):F229-F44). Since the mitochondrial metabolism of pyruvate is impaired, pyruvate is converted to lactate. Elevated tissue and blood lactate levels occur in experimental and human AKI and correlate with the severity of injury and mortality rates (Li et al. Am J Physiol Renal Physiol. 2020, 319(2):F229-F44; Legouis et al., Nat Metab. 2020;2(8):732-43; Zager et al., J Am Soc Nephrol. 2014, 25(5):998- 1012; Shen et al., American Journal of Physiology-Renal Physiology. 2020, 318(3):F689-F701; Li et al., Eur Rev Med Pharmacol Sci. 2018;22(16):5082-90). However, the role of lactate as a mediator of AKI is not clear. Our studies indicate that exogenous lactate increases the susceptibility to AKI in vivo. Specifically, exogenous L-, but not D-lactate, resulted in severe AKI in response to a short period of ischemia (22 minutes) or a moderate dose of cisplatin (15mg / kg) which alone produced only a mild increase in BUN and histologic damage (FIG. 1). This dose of lactate produced circulating lactate levels of 10-15 mM. As noted below, L-, but not D-lactate triggers Mg2release from the ER. These results support the hypothesis that lactate is a mediator of AKI.

[0067] Inhibition of LDH prevents AKI. The proximal tubule cell is metabolically active and requires high levels of ATP production to support transcellular solute transport. Mitochondrial dysfunction is a hallmark of several forms of experimental AKI (Li et al., Am J Physiol Renal Physiol. 2020, 319(2):F229-F44; Funk and Schnellmann, American Journal of Physiology-Renal Physiology. 2012, 302(7):F853-F64; Brooks et al., American Journal of Physiology-Renal Physiology. 2012, 302(7):F853-F64; Qi et al., Toxicology and applied pharmacology. 2007, 222(1): 105-10; Liu et al., Hum Exp Toxicol. 2020, 39(5):673-82). In the presence of decreased oxidative phosphorylation, glycolysis is increased with the production of lactate from pyruvate mediated by the enzyme lactate dehydrogenase (LDH) (Glancy et al., J Physiol. 2021, 599(3):863-88). LDH is encoded by two genes, LDHA, which converts pyruvate to lactate utilizing NADH, and LDHB, which converts lactate to pyruvate utilizing NAD. Osis et al. (Am JPhysiol Renal Physiol. 2021, 320(5):F706-Fl 8) found that both LDHA and LDHB are expressed in the normal kidney with LDHA having broader expression, particularly in the proximal tubules, and LDHB expressed more in distal segments. Hypoxia increased the expression of both LDHA and LDHB in cultured renal epithelial cells, but expression of both genes tended to decrease in vivo after ischemia (Osis et al., Am J Physiol Renal Physiol. 2021, 320(5):F706-F18). Likewise, Zager et al. (PloS one. 2013, 8(6):e66776) reported a decrease in kidney LDH content after ischemia which they attributed to leakage from damaged cells. Indeed, the cellular release of LDH is widely used as a marker of cell death in vitro (Moran and Schnellmann, Journal of Pharmacological and Toxicological Methods. 1996, 36(l):41-4) and clinically in such conditions as hemolysis or tumor lysis (Gupta, Inflammation. 2022, 45(6):2091-123; Kato et al., Blood. 2006, 107(6):2279-85; Khan et al., Endocr Metab Immune Disord Drug Targets. 2020, 20(6):855-68). However, the possibility that LDH could be a mediator of cell damage has not been recognized. Since the conversion of pyruvate to lactate regenerates NAD, LDH supports further glycolytic ATP production which could be protective in states of decreased oxidative phosphorylation. Likewise, the production of lactate and its subsequent release could provide metabolic substrate to other cells (the lactate shuttle (Brooks, Cell Metab. 2018, 27(4):757-85)), as has been shown for astrocytes and neurons (Park et al., frontiers in Physiology. 2021, 12). However, our studies found that inhibition of LDH with oxamate prevented the accumulation of kidney lactate during ischemia and dramatically reduced ischemic AKI as judged by creatinine and BUN levels, histologic damage and neutrophil infiltration (FIG. 2). The LDH inhibitor NCATS-SM1441 also reduced both ischemic (FIG. 3) and cisplatin kidney injury (FIG. 5). Likweise, the LDH inhibitor GNE-140 reduced ischemic AKI in vivo (FIG. 4). LDH inhibition also reduced the extent of cell death in renal epithelial cells exposed to hypoxia or cisplatin in vitro (FIG.s 3D, 4C and 5C). These results indicate that LDH is a mediator of AKI and that inhibition of LDH can prevent ischemic and toxic AKI.

[0068] Inhibition of LDH after ischemia reduces the severity of ischemic AKI. Since ischemia cannot always be predicted clinically, such as cardiac surgery, we examined the ‘window of opportunity’ by administering the LDH inhibitor 2 or 6 hr after ischemia (FIG. 6). IRI produced severe AKI in vehicle (saline) treated mice as judged by BUN, creatinine and expression of KIMI and NGAL at 24h after IR. Administration of NCATS-SM1441 2 hr afterreperfusion provided dramatic protection. Delaying the administration until 6 hours after reperfusion resulted in modest (but still significant) protection.

[0069] Inhibition of Intracellular Magnesium trafficking prevents AKI. It was discovered that the glycolytic end-product lactate is a ligand which induces ER Mg2+release and subsequent mito-Mg2+uptake. The release of ER Mg2+was specific for L-lactate rather than D-lactate and required cellular entry of lactate. Further, it was shown that lactate-induced mito-Mg2+entry is mediated by MRS2, a highly conserved mitochondrial protein related to the bacterial CorA Mg2+transport protein (Schindl et al., Biophysical Journal. 2007, 93(11):3872-83). Bilateral ischemia reperfusion in wild type (WT) and mice lacking MRS2 (Mrs2KO) was performed (FIG. 7). WT mice developed severe renal dysfunction associated with severe tubular injury and renal inflammation. These changes were dramatically reduced in Mrs2KO mice. Likewise, treatment of WT mice with cisplatin (20 mg / kg) produced severe AKI as reflected by increases in BUN, creatinine and histologic damage, but this effect was reduced in Mrs2 KO mice (FIG. 8). Likewise, primary cultures of renal epithelial cells from WT mice were more susceptible to hypoxic cell death and cisplatin-induced cell death than were cells from Mrs2KO kidneys (FIG. 7D and 8D). Cation hexamines and pentamines were shown to inhibit the CorA protein (Kucharski et al., J Biol Chem. 2000, 275(22): 16767-73), which is the bacterial homologue to MRS2. Several of the CorA inhibitors were screened and found that chloropentaammine cobalt(III)chloride (CP ACC) was a highly potent inhibitor of MRS2-mediated Mg2+transport in isolated mitochondria and intact hepatocytes (Madaris et al., Sci Reports. 2023). It was found that CPACC (20 mg / kg IP) prevented ischemic AKI when administered prior to ischemia (FIG. 9A) and also reduced hypoxic cell death in cultured renal epithelial cells (FIG. 9B). Likewise, pretreatment of mice with CPACC prevented cisplatin-induced AKI (FIG. 9C).

[0070] Inhibition of LDH or MRS2 prevents fibrosis after kidney ischemia. AKI is linked to the development of chronic kidney disease (Amdur et al., Kidney Int. 2009, 76(10): 1089-97; Lo et al., Kidney Int. 2009, 76(8): 893 -9). Renal lactate may promote fibrosis (Shen et al., American Journal of Physiology-Renal Physiology. 2020, 318(3):F689-F701; Li et al., Eur Rev Med Pharmacol Sci. 2018, 22(16): 5082-90) and it has recently determined that CPACC reduces liver fibrosis in response to a high fat diet (Madaris et al., Sci Reports. 2023). Mice were treated with vehicle (sodium acetate) or sodium oxamate (300 mg / kg / d) or CPACC (20 mg / kg / d) beginning 72 hours after unilateral ischemia, when injury is well established, and continued daily for 18days. The unilateral ischemia model, leaving the contralateral kidney intact, was used to minimize the impact of azotemia, allow longer clamp times (35 minutes) and promote the development of fibrosis in the ischemic kidney (Yang et al., Nat Med. 2010, 16(5):535-43). The non-ischemic kidney was removed on day 27 and mice were sacrificed on day 28 after ischemia. Blood was analyzed for BUN and the kidneys analyzed for size, fibrosis by histochemistry (Picrosirius and Trichrome stain), tubular atrophy, tubular dilation and glomerulosclerosis, and abundance of ECM components (collagen 1 and ot-SMA). Oxamate-treated mice (FIG. 10) and CP ACC-treated mice (FIG. 11) had preserved renal size (FIG. 10A) and function (FIG. 11 A) in the ischemic kidney. Saline-treated ischemic kidneys developed fibrosis as shown by Western blot for collagen and SMA (FIG. 10D and 11C) and Picrosirius Red staining (FIG. 10B and C and FIG. 1 IB) while the CP ACC or oxamate-treated kidneys had no fibrosis.

[0071] Human kidneys with AKI have high lactate levels. Although elevated kidney lactate levels have been found in animal models of AKI (e g., FIG. 2A), this has never been documented in humans with AKI. We analyzed kidney biopsies from humans with ischemic AKI using imaging mass spectroscopy to measure lactate levels in the tissue (FIG. 12). Compared to biopsies from healthy individuals, the samples from subjects with AKI had significantly higher tissue lactate levels. This result suggests that our findings from mouse models of AKI related to lactate and LDH are relevant to human AKI.

Claims

CLAIMS1. A method of treating a subject having or at risk of developing acute kidney injury comprising administering an effective amount of a lactate dehydrogenase inhibitor or a Mrs2 inhibitor to the subject.

2. The method of claim 1, wherein the subject is elderly, pregnant, a surgical patient, or has been exposed to a nephrotoxic agent, wherein the nephrotoxic agent is a drug or chemical capable of causing acute kidney injury.

3. The method of claim 1, wherein the lactate dehydrogenase inhibitor or the Mrs2 inhibitor is administered after developing acute kidney injury.

4. The method of claim 3, wherein the acute kidney injury is characterized by a serum creatinine level of at least 1.5 times baseline, wherein baseline refers to serum creatinine level no more than 7 days prior.

5. The method of claim 4, wherein the serum creatinine level is 1.5 to 1.9 times baseline, and the acute kidney injury is characterized by an increase in serum creatinine of at least 0.3 mg / dL.

6. The method of claim 4, wherein the serum creatinine level is 3.0 or more times baseline.

7. The method of claim 3, wherein the acute kidney injury is characterized by an increase in serum creatinine of at least 0.3 mg / dL.

8. The method of claim 7, wherein the acute kidney injury is characterized by an increase in serum creatinine of at least 0.4 mg / dL, wherein the acute kidney injury is characterized by a glomerular filtration rate of 60-89 mL / min / 1.73 m2.

9. The method of claim 3, wherein the acute kidney injury is characterized by a glomerular filtration rate of less than 90 mL / min / 1.73 m2.

10. The method of claim 9, wherein the glomerular filtration rate is 60-89 mL / min / 1 .73 m2, wherein the acute kidney injury is characterized by the subject having a urine output of less than 0.5 mL / Kg over 6 hours.

11. The method of claim 9, wherein the glomerular filtration rate is less than 15 mL / min / 1.73 m2.

12. The method of claim 3, wherein the acute kidney injury is characterized by the subject having a urine output of less than 0.5 mL / Kg over 6 hours.

13. The method of claim 12, wherein the urine output is less than 0.3 mL / Kg over 12 hours.

14. The method of claim 3, wherein the acute kidney injury is characterized by the subject having anuria for 12 or more hours.

15. The method of claim 1, wherein the lactate dehydrogenase inhibitor is oxamate, NCATS- SM1441 or GNE-140.

16. The method of claim 1, wherein the MRS2 inhibitor is CP ACC.

17. The method of claim 1, wherein the subject is has or is suspected of having acute interstitial nephritis, acute glomerular renal disease, acute vasculitic renal disease, ischemia, toxic injury, prerenal axotemia, or acute postrenal destructive nephropathy.

18. The method of claim 1, wherein the subject has diabetes, underlying renal insufficiency, nephritic syndrome, atherosclerotic disease, sepsis, hypotension, hypoxia, myoglobinuriahematuria, or liver disease.

19. The method of claim 1, wherein the subject has been exposed to a nephrotoxic agent.

20. The method of claim 19, wherein the nephrotoxic agent is a drug or chemical capable of causing acute kidney injury, wherein the drug or chemical is one or more of compound selectedfrom cisplatin; gentamicin; cephaloridine; cyclosporine; amphotericin; carbon tetrachloride; trichloroethylene; and dichloroacetylene.

21. The method of claim 1, wherein the subject is a surgical patient.

22. The method of claim 21, wherein the lactate dehydrogenase inhibitor or the MRS2 inhibitor is administered to the surgical patient prior to, during, and / or after surgery or any combination thereof.

23. A method of preserving a harvested organ wherein a lactate dehydrogenase inhibitor or a MRS2 inhibitor is administered to an organ donor, an organ to be harvested, or a harvested organ prior to, during, and / or after harvesting kidney(s) for transplantation.

24. The method of claim 23, wherein the lactate dehydrogenase inhibitor or the MRS2 inhibitor is added to preservation solutions during storage or perfusion of harvested donor organ while awaiting organ transplant surgery.

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