Radiolabeled ligands of receptor interacting protein kinase 1 for positron emission tomography imaging

Abstract

Disclosed herein are compounds, and salts thereof, which inhibit Receptor Interacting Protein Kinase 1 (RIPK1). Also disclosed are compounds which may act as positron emission tomography (PET) ligands, pharmaceutical formulations, and methods of detection and evaluation of treatment for diseases associated with RIPK1.

Description

Attorney Docket No. MDA0091-401 -PCRADIOLABELED LIGANDS OF RECEPTOR INTERACTING PROTEIN KINASE 1 FOR POSITRON EMISSION TOMOGRAPHY IMAGING

[0001] This application claims the benefit of priority of United States Provisional Application No. 63 / 693,404, filed September 11, 2024, the entirety of which is incorporated herein by reference.

[0002] Receptor Interacting Protein Kinase 1 (RIPK1 ) has been reported to play a key role in the regulation of apoptotic or necroptotic cell death pathways, as well as coordinating the response to pro-inflammatory signaling in a number of cell types. RIPK1 can also, in different contexts, regulate apoptosis and inflammation. Given its role in inflammation, RIPK1 has been implicated in many diseases featuring chronic and acute inflammatory signaling, including viral infections, sepsis, retinal degeneration, traumatic brain injury, ischemic stroke, intracerebral hemorrhage, amyotrophic lateral sclerosis, acute kidney injury, myocardial reperfusion injury, Alzheimer's disease, ulcerative colitis, osteoarthritis, and others. In animal models of these diseases RIPK1 inhibitors such as necrostatin-1 have been shown to be effective, leading to the development of such molecules for clinical trials in a number of indications.

[0003] Despite this attention, there exists a need for compounds targeting RIPK1 that can serve as positron emission tomography (PET) ligands and methods for their use. These compounds could be useful in the detection, evaluation, and treatment of neurodegenerative disorders, particularly those disorders with an inflammatory component of cellular stress, along with neuropathies, retinal diseases, injuries to the CNS, autoimmune disorders, and inflammatory diseases, and cancer.

[0004] The present disclosure fulfills these and other needs, as evident in reference to the following disclosure.SUMMARY

[0005] Provided herein is a compound of structural Formula Ior a pharmaceutically acceptable salt thereof, whereinRlais chosen from halo, haloalkoxy, and R*;Attorney Docket No. MDA0091-401 -PCRlbis chosen from hydrogen, halo, alkyl, and R*;Rlcis chosen from hydrogen, halo, and R*;R2is chosen from halo and R*;R3is chosen from alkyl, halo, pinacol borane, and R*;R4is chosen from halo, pinacol borane, and R*; andR* is a radioactive isotope or a moiety comprising a radioactive isotope, wherein at least one of Rla, Rlb, Rlc, R2, R3, and R4is R*.

[0006] Also provided is a compound as disclosed herein, or a pharmaceutically acceptable salt thereof.

[0007] Also provided is a pharmaceutical formulation comprising a compound as disclosed herein, or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier.

[0008] Also provided is a method for positron emission tomography imaging in a subject, comprising the administration of a therapeutically effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, to a subject in need thereof, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound.

[0009] Also provided is a method for detecting a RIPK1 -mediated disease, comprising the administration of a therapeutically effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, to a subject in need thereof, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound.

[0010] Also provided is a method of evaluating a change in RIPK1 expression in a subject in response to a treatment for a RIPK1 -mediated disease in the subject, comprising a) administering to the subject a compound described herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation described herein, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b) administering to the subject a treatment for the RIPK1 -mediated disease; c) administering to the subject one or more subsequent doses of the compound; d) measuring subsequent amounts of RIPK1 expression in the subject; and e) comparing the first and subsequent amounts of RIPK1 expression.[Oi l] Also provided is a method for detecting a RIPK1 -expressing tumor, comprising the administration of a therapeutically effective amount of a compound described herein, or aAttorney Docket No. MDA0091-401 -PC pharmaceutically acceptable salt thereof, to a subject in need thereof, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound, wherein an increased amount of the tracer as compared to a control indicates a RIPK1- expressing tumor.

[0012] Also provided is a method of evaluating a change in RIPK1 expression in a subject in response to an anti-cancer treatment, comprising a) administering to the subject a compound described herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation described herein, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b) administering to the subject an anti-cancer treatment; c) administering to the subject one or more subsequent doses of the compound; d) measuring subsequent amounts of RIPK1 expression in the subject; and e) comparing the first and subsequent amounts of RIPK1 expression.|013 ] These and other aspects of the invention will be apparent upon reference to the following detailed description.BRIEF DESCRIPTION OF THE DRAWINGS

[0014] FIG. 1 depicts representative PET images of mice from the front I coronal view (top panel), top-down I dorsal view (middle panel), and side / saggital view (bottom panel), taken immediately after intravenous administration of the compound from Example 4. In the original color image, red areas indicate the highest concentration of radioligand, while blue areas represent the lowest concentration. In the converted greyscale image, the lower to medium radioligand concentrations are shown as progressively lighter areas and the highest- intensity concentrations (shown as red in the original color image) are shown as grey areas surrounded by a white border located in the abdomen of the subject mice. The greyscale representation of FIG. 2 and FIG. 4 were prepared based on the technique used in FIG. 1.

[0015] The compound appeared to accumulate prominently in the liver and gastrointestinal tract, suggesting rapid uptake and distribution through the circulatory system. This initial biodistribution supports the compound’s ability to penetrate systemic circulation and reach peripheral organs.Attorney Docket No. MDA0091-401 -PC

[0016] FIG. 2 depicts representative static images captured during a 30 minute dynamic scan. The images reflect radioligand distribution over time. The outlined regions in the image highlight regions of interest (ROIs) used for quantitative analysis, particularly in the brain. In the original color image, red areas indicate the highest concentration of radioligand, while blue areas represent the lowest concentration.

[0017] The compound demonstrated preferential accumulation in brain regions, including the cortex and hippocampus, which are known to express RIPK1. This image confirms target engagement in the brain, a critical requirement for PET imaging agents intended to assess neuroinflammation or neurodegeneration.

[0018] FIG. 3 depicts time-activity curves (TACs) for both control mice and 5xFAD transgenic mice, which model Alzheimer’ s disease. The curves plot standardized uptake value (SUV) over time (seconds), normalized to body weight.

[0019] The 5xFAD mice show elevated and sustained uptake in specific brain regions compared to controls, indicating higher RIPK1 expression or altered blood-brain barrier permeability. This data supports the compound’s selectivity and sensitivity for detecting R1PK1 -associated pathology.

[0020] FIG. 4 depicts PET images of mice taken from the front / coronal view (top panel), top-down / dorsal view (middle panel), and side / saggital view (bottom panel), 2 hours post-administration.

[0021] Compared to FIG. 1, these images show reduced signal intensity, indicating compound washout from non-target tissues. However, residual signal remains in brain regions, suggesting specific retention at RIPK1 -expressing sites.DETAILED DESCRIPTIONDefinitions

[0022] As used in the present specification, the following words and phrases are generally intended to have the meanings as set forth below, except to the extent that the context in which they are used indicates otherwise.

[0023] In the following description, certain specific details are set forth in order to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the invention may be practiced without these details. In other instances, well-known structures have not been shown or described in detail to avoid unnecessarily obscuring descriptions of the embodiments. Unless the context requires otherwise, throughoutAttorney Docket No. MDA0091-401 -PC the specification and claims which follow, the word “comprise” and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense, that is, as “including, but not limited to.” Further, headings provided herein are for convenience only and do not interpret the scope or meaning of the claimed invention.

[0024] Reference throughout this specification to “one embodiment” or “an embodiment” or “some embodiments” or “a certain embodiment” means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment. Thus, the appearances of the phrases “in one embodiment” or “in an embodiment” or “in some embodiments” or “in a certain embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

[0025] Also, as used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the content clearly dictates otherwise.

[0026] When ranges of values are disclosed, and the notation “from m ... to rii” or “between ni . . . and m” is used, where ni and n2 are the numbers, then unless otherwise specified, this notation is intended to include the numbers themselves and the range between them. This range may be integral or continuous between and including the end values. By way of example, the range “from 2 to 6 carbons” is intended to include two, three, four, five, and six carbons, since carbons come in integer units. Compare, by way of example, the range “from 1 to 3 pM (micromolar),” which is intended to include 1 pM, 3 pM, and everything in between to any number of significant figures (e.g., 1.255 pM, 2.1 pM, 2.9999 pM, etc.).

[0027] The term “alkoxy”, and, interchangeably, “(alkyl)oxy”, as used herein, refers to an alkyl radical attached to a molecule by oxygen.

[0028] The term “alkyl,” as used herein, refers to a straight-chain or branched-chain saturated, hydrocarbon radical containing from 1 to 20 carbon atoms. In some embodiments, alkyl will comprise from 1 to 10 carbon atoms. In some embodiments, alkyl will comprise from 1 to 8 carbon atoms. The term “alkylene” refers to a bivalent alkyl group.

[0029] The term “cryptand”, as used herein, refers to a bicyclic or polycyclic multidentate ligand capable of forming lipophilic complexes with a host ion, wherein the ligand completely surrounds said ion. In some embodiments, the cryptand is [2.2.2]cryptand, or 4,7,13,16,21,24-Hexaoxa-l,10-diazabicyclo[8.8.8]hexacosane, also known as Kryptofix.

[0030] The term “halo,” or “halogen,” as used herein, refers to fluorine, chlorine, bromine, or iodine.Attorney Docket No. MDA0091-401 -PC

[0031] The term “haloalkoxy,” as used herein, refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom.

[0032] The terms “radioactive isotope” or “radioisotope”, as used herein, refer to an unstable element which possess excess nuclear energy as a consequence of an excess of either neutrons or protons. These isotopes include, but are not limited to, hydrogen-3, carbon-11, carbon-14, nitrogen-13, oxygen-15, fluorine-18, phosphorus-32, phosphorus-33, sulfur-35, copper-64, gallium-67, gallium-68, bromine-78, rubidium-82, yttrium-86, zirconium-89, technetium-99, sodium-22, aluminium-26, potassium-40, strontium-83, iodine- 123, iodine- 124, iodine-125, and iodine-129. In some embodiments, the radioactive isotope is a positronemitting radioisotope.

[0033] The term “positron-emitting radioisotope”, as used herein, refers to a radioactive isotope that undergoes positron emission, or P+decay, in which a proton inside its nucleus is converted into a neutron while releasing a positron and an electron neutrino. Isotopes which undergo this decay include, but are not limited to, carbon-11, nitrogen- 13, oxygen- 15, fluorine-18, copper-64, gallium-68, bromine-78, rubidium-82, yttrium-86, zirconium-89, sodium-22, aluminium-26, potassium-40, strontium-83, and iodine- 124. The short-lived positron emitting isotopesnC (T1 / 2 = 20.4 min),13N (T = 10 min),15O (Tn = 2 min), and18F (T1 2= 110 min) used for positron emission tomography are typically produced by proton irradiation of natural or enriched targets.

[0034] The term “moiety comprising a radioactive isotope,” as used herein, refers to a covalently bonded structural group which comprises at least one radioactive isotope as defined herein. In one embodiment, the radioactive isotope is chosen from hydrogen-3, carbon-11, carbon-14, nitrogen-13, oxygen-15, fluorine-18, phosphorus-32, phosphorus-33, sulfur-35, copper-64, gallium-67, gallium-68, bromine-78, rubidium-82, yttrium-86, zirconium-89, technetium-99, sodium-22, aluminium-26, potassium-40, strontium-83, iodine- 123, iodine- 124, iodine-125, and iodine-129. In another embodiment, carbon-11, nitrogen- 13, oxygen-15, fluorine-18, copper-64, gallium-68, bromine-78, rubidium-82, yttrium-86, zirconium-89, sodium-22, aluminium-26, potassium-40, strontium-83, and iodine- 124. In a further embodiment the radioactive isotope is chosen from carbon- 11, nitrogen- 13, oxygen- 15, and fluorine-18. Examples of a moiety comprising a radioactive isotope includenCH3, -13NH2, -13NHCH3, -NH1 ]CH3, -N(nCH3)(CH3),13N(CH3)2, -N(] 1CH3)2, and -15OH, as well as other alkyl, amine, and alcohol groups comprising one or more radioactive isotopes.Attorney Docket No. MDA0091-401 -PC

[0035] The term “pinacol borane”, as used herein, refers to a functional group with the formulaPinacol borane may also be abbreviated as “Bpin”.

[0036] Asymmetric centers exist in the compounds and pharmaceutically acceptable salts thereof, disclosed herein. These centers are designated by the symbols “R” or “S,” depending on the configuration of substituents around the chiral carbon atom. It should be understood that the disclosure encompasses all stereochemical isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and 1 -isomers, and mixtures thereof. Individual stereoisomers of compounds, and pharmaceutically acceptable salts thereof, can be prepared synthetically from commercially available starting materials which contain chiral centers or by preparation of mixtures of enantiomeric products followed by separation such as conversion to a mixture of diastereomers followed by separation or recrystallization, chromatographic techniques, direct separation of enantiomers on chiral chromatographic columns, or any other appropriate method known in the art. Starting compounds, and pharmaceutically acceptable salts thereof, of particular stereochemistry are either commercially available or can be made and resolved by techniques known in the art. Additionally, the compounds, and pharmaceutically acceptable salts thereof, disclosed herein may exist as geometric isomers. The present disclosure includes all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers as well as the appropriate mixtures thereof.

[0037] Additionally, the compounds disclosed herein can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. In general, the solvated forms are considered equivalent to the unsolvated forms.

[0038] As used herein, the term “positron emission tomography imaging” refers to a type of functional imaging technique that utilizes compounds that comprise one of more positronemitting radioisotopes to visualize and measure, e.g., changes in physiological activities such as absorption, metabolism, distribution, chemical composition, protein expression, and blood flow. The act of “positron emission tomography imaging” can refer to the taking of detecting positron emission or the act of constructing an image based on positron emissions, typically achieved with the aid of software.

[0039] As used herein, “administering to a patient” refers to the process of introducing a composition or dosage form into the patient via an art-recognized means of introduction.Attorney Docket No. MDA0091-401 -PC

[0040] As used herein the term “cancer” is intended to be synonymous with “tumor” or “malignancy.” The cancer may be a hematologic malignancy or solid tumor.

[0041] As used herein, “anti-cancer treatment” can include any method that can be used to treat, ameliorate, or lessen the symptoms of cancer or a tumor or that can reduce the amount or cancerous cells or the amount or size of tumors.

[0042] The term “disease” as used herein is intended to be generally synonymous, and is used interchangeably with, the terms “disorder,” “syndrome,” and “condition” (as in medical condition), in that all reflect an abnormal condition of the human or animal body or of one of its parts that impairs normal functioning, is typically manifested by distinguishing signs and symptoms, and causes the human or animal to have a reduced duration or quality of life.

[0043] The term “combination therapy” means the administration of two or more therapeutic agents to treat a therapeutic condition or disorder described in the present disclosure. Such administration encompasses co- administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients or in multiple, separate capsules for each active ingredient. In addition, such administration also encompasses use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide beneficial effects of the drug combination in treating the conditions or disorders described herein.

[0044] The phrase “therapeutically effective” is intended to qualify the amount of active ingredients used in the treatment of a disease or disorder or on the effecting of a clinical endpoint. The precise therapeutically effective amount for a subject may depend upon, e.g., the subject’s size and health, the nature and extent of the condition, the therapeutics or combination of therapeutics selected for administration, and other variables known to those of skill in the art. The effective amount for a given situation is determined by routine experimentation and is within the judgment of the clinician.

[0045] As used herein, the term “treat,” “treating”, or “treatment” means the administration of therapy to an individual who already manifests at least one symptom of a disease or condition or who has previously manifested at least one symptom of a disease or condition. For example, “treating” can include alleviating, abating, or ameliorating a disease or condition symptoms, preventing additional symptoms, ameliorating the underlying metabolic causes of symptoms, inhibiting the disease or condition, e.g., arresting the development of the disease or condition, relieving the disease or condition, causing regression of the disease or condition, relieving a condition caused by the disease or condition, or stopping the symptoms of the disease or condition. For example, the termAttorney Docket No. MDA0091-401 -PC“treating” in reference to a disorder means a reduction in severity of one or more symptoms associated with that particular disorder. Therefore, treating a disorder does not necessarily mean a reduction in severity of all symptoms associated with a disorder and does not necessarily mean a complete reduction in the severity of one or more symptoms associated with a disorder.

[0046] The term “patient” is generally synonymous with the term “subject” and includes all mammals including humans. Examples of patients include humans, livestock such as cows, goats, sheep, pigs, and rabbits, and companion animals such as dogs, cats, rabbits, and horses. Preferably, the patient is a human.

[0047] Those skilled in the art will appreciate that the invention(s) described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the invention(s) includes all such variations and modifications. The invention(s) also includes all the steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations or any two or more of steps or features unless specifically stated otherwise.|048 ] The present invention(s) is not to be limited in scope by the specific embodiments described herein, which are intended for the purpose of exemplification only. Functionally equivalent products, compositions, and methods are clearly within the scope of the invention(s), as described herein.

[0049] It is appreciated that certain features of the invention(s), which are, for clarity, described in the context of separate embodiments, can also be provided in combination in a single embodiment. Conversely, various features of the invention(s), which are, for brevity, described in the context of a single embodiment, can also be provided separately or in any suitable subcombination.

[0050] Provided is a compound of structural Formula Ior a pharmaceutically acceptable salt thereof, whereinRlais chosen from halo, haloalkoxy, and R*;Rlbis chosen from hydrogen, halo, alkyl, and R*;Rlcis chosen from hydrogen, halo, and R*;Attorney Docket No. MDA0091-401 -PCR2is chosen from halo and R*;R3is chosen from alkyl, halo, pinacol borane, and R* ;R4is chosen from halo, pinacol borane, and R*; andR* is a radioactive isotope or a moiety comprising a radioactive isotope, wherein at least one of Rla, Rlb, Rlc, R2, R3, and R4is R*.

[0051] In some embodiments, the compound has structural Formula IAor a pharmaceutically acceptable salt thereof, whereinRlais chosen from halo and haloalkoxy;Rlbis chosen from hydrogen, halo, and alkyl;Rlcis chosen from hydrogen and halo;R3is -CH3and R4is18F or R3is -nCH3and R4is F.

[0052] In some embodiments, Rlais chosen from fluoro and trifluoromethoxy.

[0053] In some embodiments, Rlais fluoro.

[0054] In some embodiments, Rlbis chosen from hydrogen, fluoro, and methyl.

[0055] In some embodiments, Rlcis chosen from hydrogen and fluoro.

[0056] In some embodiments, Rlband Rlcare hydrogen.

[0057] In some embodiments, R2is fluoro.

[0058] In some embodiments, R3is R3is methyl and R4is R*.

[0059] In some embodiments, R3is R* and R4is chosen from fluoro, bromo, and pinacol borane .

[0060] In some embodiments, R4is fluoro.

[0061] In some embodiments, R* is a moiety comprising a radioactive isotope, wherein the moiety is chosen from alkyl, amino, and hydroxyl.

[0062] In some embodiments, R* is a moiety comprising a radioactive isotope, wherein the moiety is chosen from methyl, -NH2, -NHCH3, -N(CH3)2, and hydroxyl.

[0063] In some embodiments, R* is a moiety comprising a radioactive isotope chosen from -nCH3, -13NH2, -13NHCH3, -NHnCH3, -N(1 ]CH3)(CH3),13N(CH3)2, -N(1 ]CH3)2, and -15OH.Attorney Docket No. MDA0091-401 -PC

[0064] In some embodiments, R* is a moiety comprising a positron-emitting radioisotope or a positron-emitting radioisotope.

[0065] In some embodiments, R* is chosen from “CHr and18F.

[0066] In some embodiments, R* is18F.

[0067] In some embodiments, the structure of the compound

[0068] In some embodiments, Rlais fluoro and Rlband Rlcare hydrogen.

[0069] In some embodiments, Rlaand Rlbare fluoro and Rlcis hydrogen.

[0070] In some embodiments, Rlais fluoro, Rlbis methyl, and Rlcis hydrogen.

[0071] In some embodiments, Rlais fluoro, Rlbis methyl, and Rlcis fluoro.

[0072] In some embodiments, Rlais trifluoromethoxy and Rlband Rlcare hydrogen.

[0073] In some embodiments, R3isnCH3.

[0074] In some embodiments, R4is fluoro or bromo.

[0075] In some embodiments, R4is18F.

[0076] In some embodiments, R4is pinacol borane.

[0077] The compounds disclosed herein can exist as pharmaceutically acceptable salts. The present disclosure includes compounds listed herein in the form of salts, including acid addition salts. Suitable salts include those formed with both organic and inorganic acids.Such acid addition salts will normally be pharmaceutically acceptable. However, salts of non- pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question. Basic addition salts may also be formed and be pharmaceuticallyAttorney Docket No. MDA0091-401 -PC acceptable. For a more complete discussion of the preparation and selection of salts, refer to Pharmaceutical Salts: Properties, Selection, and Use (Stahl, P. Heinrich. Wiley-VCHA, Zurich, Switzerland, 2002).

[0078] The term “pharmaceutically acceptable salt,” as used herein, represents salts or zwitterionic forms of the compounds or disclosed herein. The salts can be prepared during the final isolation and purification of the compounds or separately by reacting the appropriate compound in the form of the free base with a suitable acid. Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (besylate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethansulfonate (isethionate), lactate, maleate, malonate, DL-mandelate, mesitylenesulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylproprionate, phosphonate, picrate, pivalate, propionate, pyroglutamate, succinate, sulfonate, tartrate, L-tartrate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, para-toluenesulfonate (p-tosylate), and undecanoate. Also, basic groups in the compounds disclosed herein can be quatemized with methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dimethyl, diethyl, dibutyl, and diamyl sulfates; decyl, lauryl, myristyl, and steryl chlorides, bromides, and iodides; and benzyl and phenethyl bromides. Examples of acids which can be employed to form pharmaceutically acceptable addition salts include inorganic acids such as hydrochloric, hydrobromic, sulfuric, and phosphoric, and organic acids such as oxalic, maleic, succinic, and citric. Salts can also be formed by coordination of the compounds with an alkali metal or alkaline earth ion. Hence, the present disclosure contemplates sodium, potassium, magnesium, and calcium salts of the compounds disclosed herein, and the like.

[0079] Basic addition salts can be prepared during the final isolation and purification of the compounds by reacting a carboxy group with a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation or with ammonia or an organic primary, secondary, or tertiary amine. The cations of pharmaceutically acceptable salts include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as nontoxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, A,A-dimethylaniline, A-methylpiperidine, A-methy 1 morpholine, dicyclohexylamine, procaine, dibenzylamine, A,A-dibenzylphenethylamine, 1-ephenamine,Attorney Docket No. MDA0091-401 -PC and Ar-dibenzylethylenediamine. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine.

[0080] While it may be possible for the compounds, or pharmaceutically acceptable salts thereof, of the subject disclosure to be administered as the raw chemical, it is also possible to present them as a pharmaceutical formulation.

[0081] Also provided is a pharmaceutical formulation comprising a compound as disclosed herein, or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. Typically, these methods include the step of bringing into association a compound, or a pharmaceutically acceptable salt thereof, of the subject disclosure (“active ingredient”) with the carrier which constitutes one or more accessory ingredients. In general, formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both and then, if necessary, shaping the product into the desired formulation.

[0082] Preferred unit dosage formulations are those containing an effective dose, or an appropriate fraction thereof, of the active ingredient.

[0083] Compounds, or pharmaceutically acceptable salts thereof, may be administered at a dose of between 0. 1 to 50 mg. In some embodiments, the compounds are administered at a dose of between 5 to 8 mg. In some embodiments, the compounds are administered in a sufficient dose to deliver between 100 and 300 microcuries (pCi) of radiation to the subject. In some embodiments, the compounds are administered in a sufficient dose to deliver approximately 250 pCi of radiation to the subject. The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.

[0084] The compounds, or pharmaceutically acceptable salts thereof, can be administered in various modes. In some embodiments, the pharmaceutical formulation is formulated for intravenous administration.

[0085] A PET scan can be used to determine the amount of positron emitting radiotracer (e.g., a compound as disclosed herein comprising a radioactive isotope or a moiety comprising a radioactive isotope) taken up by cells. Standardized uptake value, SUV, (also referred to as the dose uptake ratio, DUR) is a widely used, robust PET quantifier, calculated as a ratio of tissue radioactivity concentration (for example in units [kBq / mL]) at time T,Attorney Docket No. MDA0091-401 -PCCPET(T), and administered dose (for example in units [MBq]) at the time of injection divided by body weight (BW, usually in units [kg]). SUVBW =C PET(T) / (Dose / Weight). The step of determining an amount of the positron-emitting compound (radiotracer) may also be measured by detection of gamma radiation (e.g., by use of e.g. scintillation detectors, semiconductors, and the like).

[0086] Also provided is a method for positron emission tomography imaging in a subject, comprising the administration of a therapeutically effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, to a subject in need thereof and subjecting the subject to a positron emission tomography (PET) scan.

[0087] In a further embodiment, the method for positron emission tomography imaging in a subject further comprises one or more steps chosen from: a) measuring the amount of gamma radiation emitted from a tissue; b) measuring the concentration of the radiotracer in a tissue; and c) determining an amount of the radiotracer in a tissue.

[0088] Also provided is a method for detecting a RIPK1 -mediated disease, comprising the administration of a therapeutically effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, to a subject in need thereof, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound.

[0089] Also provided is a method of evaluating a change in RIPK1 expression in a subject in response to a treatment for a RIPK1 -mediated disease in the subject, comprising a) administering to the subject a compound described herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation described herein, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b) administering to the subject a treatment for the RIPK1 -mediated disease; c) administering to the subject one or more subsequent doses of the compound; d) measuring subsequent amounts of RIPK1 expression in the subject; and e) comparing the first and subsequent amounts of RIPK1 expression.

[0090] In some embodiments, the amount of the compound is determined within an inflammatory, apoptotic, or necroptotic process of the RIPK1 -mediated disease.

[0091] In some embodiments, the inflammatory process is RIPK1 -dependent cytokine release.

[0092] In some embodiments, the necroptotic process is RIPK1 -dependent necroptosis.Attorney Docket No. MDA0091-401 -PC

[0093] In some embodiments, the RIPK1 -mediated disease is chosen from a neurological disease, a neuropathy, an autoimmune disorder, an inflammatory disease, or cancer.

[0094] In some embodiments, the RIPK1 -mediated disease is a neurological disease.

[0095] In some embodiments, the neurological disease is accompanied by an inflammatory component of cellular stress.

[0096] In some embodiments, the neurological disease is chosen from Multiple Sclerosis, Neimann-Pick disease, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, Lewy body dementia, frontotemporal dementia, glutamine expansion disease, Huntington’s disease, Kennedy’s disease, and spinocerebellar ataxia.

[0097] In some embodiments, the RIPK1 -mediated disease is a neuropathy.

[0098] In some embodiments, the neuropathy is chosen from diabetic neuropathy and chemotherapy-induced neuropathy.

[0099] In some embodiments, the RIPK1 -mediated disease is a retinal disease.

[0100] In some embodiments, the retinal disease is chosen from macular degeneration and retinitis.101011 In some embodiments, the RIPK1 -mediated disease is an autoimmune disorder.

[0102] In some embodiments, the autoimmune disorder is chosen from ulcerative colitis, rheumatoid arthritis, psoriasis, lupus, and inflammatory bowel disease.

[0103] In some embodiments, the RIPK1 -mediated disease is an inflammatory disease.

[0104] In some embodiments, the inflammatory disease is found in one or more organs chosen from the lung, heart, kidney, and liver.

[0105] In some embodiments, the RIPK1 -mediated disease is cancer.

[0106] In some embodiments, the cancer can be treated by promoting an appropriate immune response to the tumor.

[0107] In some embodiments, the appropriate immune response to the tumor comprises, or results in, one or more of the following: a. an increase in the number or activity, or degree of tumor infiltration, of cytotoxic T- lymphocytes and / or natural killer cells; b. an increase in the number or activity of M 1 macrophages in the tumor microenvironment and / or a decrease in the in the number or activity of M2 macrophages in the tumor microenvironment; c. a decrease in the number or activity of regulatory T cells; and d. a decrease in the number or activity of myeloid-derived suppressor cells.

[0108] In some embodiments, the cancer is a myelodysplastic syndrome (MDS).Attorney Docket No. MDA0091-401 -PC

[0109] In some embodiments, the cancer is acute myeloid leukemia (AML).

[0110] In some embodiments, the amount of the compound is determined with positron emission tomography imaging.

[0111] In some embodiments, the positron emission tomography imaging confirms uptake and distribution into the brain.

[0112] Also provided is a method for detecting a RIPK1 -expressing tumor, comprising the administration of a therapeutically effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, to a subject in need thereof, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound, wherein an increased amount of the tracer as compared to a control indicates a RIPK1- expressing tumor.

[0113] Also provided is a method of evaluating a change in RIPK1 expression in a subject in response to an anti-cancer treatment, comprising a) administering to the subject a compound described herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation described herein, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b) administering to the subject an anti-cancer treatment; c) administering to the subject one or more subsequent doses of the compound; d) measuring subsequent amounts of RIPK1 expression in the subject; and e) comparing the first and subsequent amounts of RIPK1 expression.

[0114] As used herein, “anti-cancer treatment” can include any method that can be used to treat, ameliorate, or lessen the symptoms of cancer or a tumor or that can reduce the amount or cancerous cells or the amount or size of tumors. Treatments can include surgery, radiotherapy, chemotherapy, targeted therapy, immunotherapy, or any combination thereof. In some aspects, administration can be in combination with one or more additional therapeutic agents. The phrases “combination therapy”, “combined with” and the like refer to the use of more than one treatment simultaneously to increase the response. Such therapies can be administered prior to, simultaneously with, or following administration of one another.

[0115] The term “chemotherapy” or “chemotherapeutic agent” as used herein refers to any therapeutic agent used to treat cancer. Examples of chemotherapeutic agents include, but are not limited to, (i) anti-microtubules agents comprising vinca alkaloids (vinblastine, vincristine, vinflunine, vindesine, and vinorelbine), taxanes (cabazitaxel, docetaxel, larotaxel,Attorney Docket No. MDA0091-401 -PC ortataxel, paclitaxel, and tesetaxel), epothilones (ixabepilone), and podophyllotoxin (etoposide and teniposide); (ii) antimetabolite agents comprising anti-folates (aminopterin, methotrexate, pemetrexed, pralatrexate, and raltitrexed), and deoxynucleoside analogues (azacitidine, capecitabine, carmofur, cladribine, clofarabine, cytarabine, decitabine, doxifluridine, floxuridine, fludarabine, fluorouracil, gemcitabine, hydroxycarbamide, mercaptopurine, nelarabine, pentostatin, tegafur, and thioguanine); (iii) topoisomerase inhibitors comprising Topoisomerase I inhibitors (belotecan, camptothecin, cositecan, gimatecan, exatecan, irinotecan, lurtotecan, silatecan, topotecan, and rubitecan) and Topoisomerase II inhibitors (aclarubicin, amrubicin, daunorubicin, doxorubicin, epirubicin, etoposide, idarubicin, merbarone, mitoxantrone, novobiocin, pirarubicin, teniposide, valrubicin, and zorubicin); (iv) alkylating agents comprising nitrogen mustards (bendamustine, busulfan, chlorambucil, cyclophosphamide, estramustine phosphate, ifosamide, mechlorethamine, melphalan, prednimustine, trofosfamide, and uramustine), nitrosoureas (carmustine (BCNU), fotemustine, lomustine (CCNU), N-Nitroso-N-methylurea (MNU), nimustine, ranimustine semustine (MeCCNU), and streptozotocin), platinum-based (cisplatin, carboplatin, dicycloplatin, nedaplatin, oxaliplatin and satraplatin), aziridines (carboquone, thiotepa, mytomycin, diaziquone (AZQ), triaziquone and triethylenemelamine), alkyl sulfonates (busulfan, mannosulfan, and treosulfan), non-classical alkylating agents (hydrazines, procarbazine, triazenes, hexamethylmelamine, altretamine, mitobronitol, and pipobroman), tetrazines (dacarbazine, mitozolomide and temozolomide); (v) anthracycline / anthracenedione agents comprising doxorubicin and daunorubicin and derivatives of these compounds including epirubicin and idarubicin, pirarubicin, aclarubicin, and mitoxantrone; (vi) enzyme inhibitors agents comprising FI inhibitor (Tipifamib), CDK inhibitors (Abemaciclib, Alvocidib, Palbociclib, Ribociclib, and Seliciclib), Prl inhibitor (Bortezomib, Carfilzomib, and Ixazomib), Phi inhibitor (Anagrelide), IMPDI inhibitor (Tiazofurin), LI inhibitor (Masoprocol), PARP inhibitor (Niraparib, Olaparib, Rucaparib), HDAC inhibitor (Belinostat, Panobinostat, Romidepsin, Vorinostat), and PIKI inhibitor (Idelalisib); (vii) receptor antagonist agent comprising ERA receptor antagonist (Atrasentan), Retinoid X receptor antagonist (Bexarotene), Sex steroid receptor antagonist (Testolactone); (viii) anticancer antibiotics such as bleomycins, mitomycin C, and actinomycin; and (ix) other ungrouped anticancer agents comprising Amsacrine, Trabectedin, Retinoids (Alitretinoin Tretinoin) Arsenic trioxide, Asparagine depleters (Asparaginase / Pegaspargase), Celecoxib, Demecolcine Elesclomol, Elsamitrucin, Etoglucid, Lonidamine, Lucanthone, Mitoguazone, Mitotane, Oblimersen, Omacetaxine mepesuccinate, and Eribulin.Attorney Docket No. MDA0091-401 -PC

[0116] The term “immunotherapy” refers to any type of therapy that ameliorates, treats, or prevents a malignancy in a subject by assisting or boosting the subject’s immune system in eradicating cancerous cells. Modulating the immune system includes inducing, stimulating, or enhancing the immune system as well as reducing, suppressing, or inhibiting the immune system. Immunotherapy can be active or passive. Passive immunotherapy relies on the administration of drugs, such as monoclonal antibodies directed against the target to eliminate it. For example, tumor-targeted monoclonal antibodies have demonstrated clinical efficacy to treat cancer. Active immunotherapy aims to induce cellular immunity and establish immunological memory against the target agent. Active immunotherapy includes, but is not limited to, vaccination, and immune modulators.

[0117] Types of immunotherapy include, for example, immune checkpoint inhibitors, T- cell transfer therapy (i.e., adoptive cell therapy, adoptive immunotherapy, or immune cell therapy), monoclonal antibodies (e.g., monoclonal antibodies that can mark cancer cells so that they will be better identified and destroyed by the immune system), treatment vaccines (e.g., Sipuleucel-T, T-VEC), and immune system modulators [e.g., cytokines, interferons, interleukins (e.g., IL-2; IL- 11), granulocyte-macrophage colony-stimulating factor (GM- CSF) and granulocyte colony- stimulating factor (G-CSF), BCG], and immunomodulatory drugs such as thalidomide, lenalidomide, pomalidomide, imiquimod.

[0118] Also provided are methods of inhibiting at least one RIPK1 function comprising the step of contacting RIPK1 with a compound as described herein, or a pharmaceutically acceptable salt thereof. The cell phenotype, cell proliferation, activity of RIPK1 , change in biochemical output produced by active RIPK1 , expression of RIPK1 , or binding of RIPK1 with a natural binding partner may be monitored. Such methods may be modes of treatment of disease, biological assays, cellular assays, biochemical assays, or the like.

[0119] Also provided are methods of treatment of a RIPK1 -mediated disease comprising the administration of a therapeutically effective amount of a compound as disclosed herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

[0120] A “treatment for a RIPK1 -mediated disease,” as used herein, can be a treatment for a neurological disease, a neuropathy, an autoimmune disorder, an inflammatory disease, or cancer. Such treatments can be chosen from pharmaceutical interventions (e.g., drugs, biologies, including RIPK1 inhibitors), immunotherapies (e.g., checkpoint inhibitors, mono- and bispecific monoclonal antibodies, cytokines, CAR-T and other targeted immune cell therapies, and antibody-drug conjugates), targeted therapies, chemotherapies, radiotherapies, and the like.Attorney Docket No. MDA0091-401 -PC

[0121] Also provided is a method of inhibition of RIPK1 comprising contacting RIPK1 with a compound as disclosed herein, or a pharmaceutically acceptable salt thereof.

[0122] Also provided is a method of modulation of a RIPK1 -mediated function in a subject comprising the administration of a therapeutically effective amount of a compound as disclosed herein, or a pharmaceutically acceptable salt thereof.

[0123] Further embodiments include the embodiments disclosed in the following Schemes and Examples, which are not to be construed as limiting in any way.

[0124] In the Examples below and throughout the disclosure, the following abbreviations may be used: RT = Room Temperature; SM = Starting Material; MeCN or ACN = acetonitrile; CDI = I , I '-carbonyldii midazole; DCE = dichloroethane; DCM = dichloromethane; DIEA or DIPEA = N,N-Diisopropylethylamine; DMF = dimethylformamide; DMSO = dimethylsulfoxide; EtrN or TEA = triethylamine; EtOAc = ethyl acetate; EtOH = ethanol; H2O = water; MeCN = acetonitrile; MeOH = methanol; n- BuLi = zi-butyllithium; NMP = A-methyl-2-pyrrolidone; PE = petroleum ether; TFA = trifluoroacetic acid; TFA A = trifluoroacetic anhydride; THF = tetrahydrofuran; HATU = 1- |bis(dimethylamino)methylene |- 1H- 1 ,2,3-triazolo| 4,5-b Ipyridinium 3-oxide hexafluorophosphate; NCS = N-chlorosuccinimide;1H-NMR = Proton Nuclear magnetic Resonance; LCMS = Liquid chromatography-mass spectrometry; TLC = thin layer chromatography; and HPLC = High Performance Liquid Chromatography. Other abbreviations may be used and will be familiar in context to those of skill in the art.Schemes

[0125] The following schemes may be used to practice the invention.Scheme I101 Formula I

[0126] Referring to Scheme I, Step 1, to a solution of a compound of Formula 101 in an organic solvent, such as dichloromethane, is added a compound of formula 102 and a base, such as triethylamine, and a coupling agent, such as hexafluorophosphate azabenzotriazoleAttorney Docket No. MDA0091-401 -PC tetramethyl uranium (HATU). The mixture is stirred, optionally at ambient temperature. In some embodiments, the mixture is stirred from 1-3 h. The product, a compound of Formula I, is isolated and purified using methods known in the art. Individual enantiomers can be separated by using methods known in the art, such as chiral chromatography.

[0127] Additionally, compounds of Formula I may be transformed to other compounds of Formula I via additional steps. As a non-limiting example, to a solution of a compound of Formula I containing a bromine functional group in an organic solvent, such as 1 ,4-dioxane, is added a base, such as potassium phosphate, an oxidizing agent, such as silver oxide, a palladium catalyst, such as Pd(dppf)Ch, and bis(pinacolato)diboron. The mixture is stirred, optionally at ambient temperature. In some embodiments, the mixture is stirred from 16-24 h. The product, a compound of Formula I containing a pinacol borane functional group, is isolated and purified using methods known in the art.

[0128] As a further non-limiting example, to a reactor vessel containing a mixture of [18F]Huoride, a cryptand, such as Kryptofix, and a base, such as a potassium carbonate, in an organic solvent, such as acetonitrile, is added a compound of Formula I containing a pinacol borane functional group. The mixture is stirred, optionally at elevated temperatures. In some embodiments, the mixture is stirred for 20-40 minutes. The product, a compound of Formula I containing a radiolabeled18F fluorine, is isolated and purified using methods known in the art.Examples

[0129] The invention is further illustrated by the following non-limiting Examples.EXAMPLES 1 A and IB(5-(5-Bromo-2-fluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(fluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanoneMethyl 3-(hydroxymethyI)bicyclo[l.l.l]pentane-l-carboxylate (Compound 2)Attorney Docket No. MDA0091-401 -PC90.0% yield 1 2

[0130] To a solution of compound 1 (105 g, 617 mmol, 1.00 eq) in THF (1.10 L) was dropwise added BHs^THF (1.00 M, 740 mL, 1.20 eq) at N2 atmosphere at 0 °C for 1 hr. The mixture was stirred at 0 °C for 2 hrs. TLC (Petroleum ether / Ethyl acetate = 2 / 1) indicated compound 1 was consumed completely. The mixture was cooled to 0 °C, quenched with MeOH (400 mL), and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiC , Petroleum ether / Ethyl acetate = 1 / 0 to 0 / 1) to give compound 2 (175 g, 1.12 mol, 90.0% yield) as a yellow oil. *H NMR: (400 MHz, CDCh) 3 3.685 (s, 3H), 3.638 (s, 2H), 1.98 (s, 6H).Methyl 3-(((methylsulfonyl)oxy)methyl)bicyclo[l.l.l]pentane-l-carboxylate (Compound74.3% yield3)2 3

[0131] To a solution of compound 2 (175 g, 1.12 mol, 1.00 eq) and TEA (340 g, 3.36 mol, 468 mL, 3.00 eq) in DCM (1.75 L) was added MS2O (293 g, 1.68 mol, 1.50 eq) at 0 °C. The mixture was stirred at 0 °C for 4 hrs. Analysis of the reaction by TLC (Petroleum ether / Ethyl acetate = 1 / 1) indicated compound 2 was consumed completely. The reaction mixture was washed with water (1.00 L x 2) and brine (1.00 L x 2), dried over Na2SO4, filtered and concentrated under reduced pressure to give compound 3 (195 g, 832 mmol, 74.3% yield) as a yellow solid.1H NMR: (400 MHz, CDCh) S 4.26 (s, 2H), 3.69 (s, 3H), 3.02 (s, 3H), 2.10 (s, 6H).Methyl 3-(fluoromethyl)bicyclo[l.l.l]pentane-l-carboxylate (Compound 4)Used directly3 4

[0132] To a solution of compound 3 (195 g, 832 mmol, 1.00 eq) in DMSO (2.00 L) was added CsF (379 g, 2.50 mol, 3.00 eq). The mixture was stirred at 100 °C for 4 hrs. Analysis of the reaction by TLC (Petroleum ether / Ethyl acetate = 5 / 1) showed Reactant 1 (Rf = 0.28)Attorney Docket No. MDA0091-401 -PC was consumed completely. The reaction mixture was filtered and the filtrate was without further work-up. Compound 4 (131 g, crude) was obtained as a yellow liquid.3-(Fluoromethyl)bicyclo[l.l.l]pentane-l-carboxylic acid (Compound 5)

[0133] To a solution of compound 4 (131 g, 832 mmol, 1.00 eq) in DMSO (1.50 L) and H2O (150 mL) was added NaOH (133 g, 3.33 mol, 4.00 eq). The mixture was stirred at 25 °C for 4 hrs. Analysis of the reaction by TLC (Petroleum ether / Ethyl acetate = 5 / 1) indicated compound 4 (Rf = 0.69) was consumed completely and one new spot (Rf = 0.20) formed.The residue was diluted with H2O (2.00 L). The aqueous phase was extracted with Petroleum ether (1.00 L x 3). The combined aqueous phase acidified to pH ~ 2 with aqueous 2 M hydrochloric. The aqueous phase was extracted with Ethyl acetate (1.00 L x 3), washed with aqueous 2 M hydrochloric (1.00 L x 2), dried over Na2SO4, concentrated under reduced pressure to give compound 5 (59.5 g, 413 mmol) as a yellow solid. ’H NMR: (400 MHz, CDCI3) 8 10.27 (s, 1H), 4.39 (d, J = 47.6 Hz, 2H), 2.11 (s, 6H).19F NMR: (400 MHz, CDCh) <5 -223.98. l-Bromo-4-fluoro-2-methylbenzene (Compound B-l)B-1a

[0134] To a solution of compound B-la (188 g, 994 mmol, 1.00 eq) in dry THF (2.00 L) was added dropwise LDA (2.00 M, 497 mL, 1.00 eq) under argon atmosphere at -78 °C.After stirring for 2 hrs, a solution of DMF (366 g, 5.01 mol, 385 mL, 5.04 eq) in THF (500 mL) was added dropwise. The mixture was returned to 20 °C and stirred for 1 hr. Analysis of the reaction by LCMS showed 61.6% of desired compound was detected. Then, the mixture was poured into water (2.00 L) and the mixture was neutralized with IN hydrochloric acid. After extraction with ethyl acetate (2.00 L), the extract was washed with saturated brine (1.00 L) and dried with magnesium sulfate. The residue was used directly in the next step without further purification. Compound B-l (216 g, crude) was obtained as a yellow liquid. LCMS: (ES+) C8H6BrFO requires: 216, found: 217 [M+H]+.Attorney Docket No. MDA0091-401 -PC(E)-3-(5-Bromo-2-fluoro-4-methylphenyl)acrylaldehyde (Compound B-2)17.3% yield over two steps BrB-1 B-2

[0135] To a solution of compound B-l (108 g, 497 mmol, 1.00 eq) in toluene (1.50 L) was added 2-(triphenyl-phosphanylidene)acetaldehyde (151 g, 497 mmol, 1.00 eq) separately in two batches. The mixture was stirred at 80 °C for 16 hrs. Analysis of the reaction by TLC (Petroleum ether / Ethyl acetate = 10 / 1) showed several new spots (Rf = 0.24, 0.38 (Pl) and 0.43) were detected in batch 1 and batch 2. The two batches were combined together to concentrated in vacuum. The residue was purified by column chromatography (SiCb, Petroleum ether / Ethyl acetate = 1 / 0 to 100 / 1) to give a compound B-2 (52.0 g, 189 mmol, 17.3 yield over two steps, 88.4% purity) as a yellow solid. LCMS: (ES+) CioHgBrFO requires: 242 / 4, found: 245 [M+H]+.5-(5-Bromo-2-fluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazole (Compound B)

[0136] To a solution of NHiNFL’HiO (18.4 g, 312 mmol, 17.8 mL, 85.0% purity, 1.65 eq) in EtOH (500 mL) was added AcOH (19.3 g, 321 mmol, 18.4 mL, 1.70 eq) and compound B-2 (52.0 g, 189 mmol, 1 .00 eq) in THF (100 mL) at 0 °C. The mixture was stirred at 90 °C for 48 hrs. Analysis of the reaction by LCMS showed 75.7% of the desired product. The reaction was diluted ethyl acetate (500 mL), washed with NaHCCL (200 mL x 3), the organic layer was dried with NazSCL and concentrated in vacuum. TLC (Petroleum ether / Ethyl acetate = 2 / 1) showed several new spots (Rf = 0.15, 0.39 (Pl) and 0.61). The residue was purified by column chromatography CSiCL, Petroleum ether / Ethyl acetate = 30 / 1 to 10 / 1) to give compound B (53.0 g, 149 mmol, 79.1% yield, 72.6% purity) as a yellow solid. LCMS: (ES+) CioHioBrFN2requires: 256 / 8, found: 259 [M+H]+.Attorney Docket No. MDA0091-401 -PC(5-(5-Bromo-2-fluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(fluoromethyl)bicydo[l.l.l]pentan-l-yl)methanone (Compound 6b)

[0137] To a solution of compound 5 (15.5 g, 107 mmol, 1.00 eq) and compound B (38.1 g, 107 mmol, 1.00 eq) in DCM (100 mL) was added HATU (61.3 g, 161 mmol, 1.50 eq) and DIEA (27.8 g, 215 mmol, 37.5 mL, 2.00 eq). The mixture was stirred at 20 °C for 1 hr. Analysis of the reaction by LCMS showed 41.4% of desired product. The mixture was concentrated in vacuum. TLC (Petroleum ether / Ethyl acetate = 2 / 1) showed several new spots (Rf = 0.29, 0.42 (Pl) and 0.57). The residue was purified by column chromatography (SiCh, Petroleum ether / Ethyl acetate) to give compound 6b (28.0 g, 67.1 mmol, 62.4 % yield, 91.8% purity) as a yellow solid. LCMS: (ES+) Ci?Hi7BrF2N2O requires: 382 / 4, found: 385 [M+H]+. 'H NMR: (400 Hz, CDCh) 87.19 (d, J = 7.2 Hz, 1H), 6.95-6.92 (m, 2H), 5.46 (dd, Ji = 6.0 Hz, J2= 12.0 Hz, 1H), 4.40 (d, J = 48 Hz, 2H), 3.39-3.31 (m, 1H), 2.77-2.71 (m, 1H), 2.33 (s, 3H), 2.18 (s, 6H).19F NMR: (400 Hz, CDCh) S -120.82, -223.35.

[0138] The racemic compound 6b was further separated by SFC (column: REGIS (s,s) WHELK-01 (250mm*50mm,10 um);mobile phase: [CO2-MeOH(0.1%NH3H2O)];B%:65%, isocratic elution mode) to give both enantiomers as peak 1 and peak 2.

[0139] Example 1A (R enantiomer, 7.00 g, 17.7 mmol, 26.2% yield, 97.0% purity) was obtained as a yellow solid.: LCMS: (ES+) Ci?Hi7BrF2N2O requires: 382 / 4, found: 383 [M+H]+. 'H NMR: (400 Hz, CDCh) 87.19 (d, J = 7.2 Hz, 1H), 6.95-6.92 (m, 2H), 5.46 (dd, Ji = 6.0 Hz, J2= 12.0 Hz, 1H), 4.40 (d, J = 48 Hz, 2H), 3.39-3.31 (m, 1H), 2.77-2.71 (m,Attorney Docket No. MDA0091-401 -PC1H), 2.33 (s, 3H), 2.18 (s, 6H).19F NMR: (400 Hz, CDCh) 3 -120.82, -223.35. SFC (Rt = 1.310min, ee = 100%).

[0140] Example IB (S enantiomer, 8.80 g, 22.2 mmol, 30.2% yield, 96.8% purity) was obtained as a yellow solid. LCMS: (ES+) CnHnBrF ^O requires: 382 / 4, found: 385 [M+H]+. ’H NMR: (400 Hz, CDCh) 37.19 (d, J = 7.2 Hz, 1H), 6.95-6.92 (m, 2H), 5.46 (dd, 7 / = 5.2 Hz, 72= 12.0 Hz, 1H), 4.40 (d, 7 = 48 Hz, 2H), 3.39-3.31 (m, 1H), 2.77-2.71 (m, 1H), 2.33 (s, 3H), 2.18 (s, 6H).19F NMR: (400 Hz, CDCh) 3 -120.82, -223.35. SFC (Rt = 2.419 min, ee = 100%).EXAMPLE 2(R)-(5-(2-FIuoro-4-methyl-5-(4,4,5,5-tetramethyI-l,3,2-dioxaborolan-2-yl)phenyl)-4,5- dihydro- lH-pyrazol-l-yl)(3-(fluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanone

[0141] To a solution of the compound of Example 1A (7.00 g, 17.7 mmol, 1.00 eq) in dioxane (100 mL) was added B2Pin2 (9.00 g, 35.4 mmol, 2.00 eq), K3PO4 (11.3 g, 53.2 mmol, 3.00 eq), Pd(dppf)Ch.CH2Ch (2.89 g, 3.54 mmol, 0.20 eq) and Ag2O (6.16 g, 26.6 mmol, 1.50 eq). The mixture was stirred at 100 °C for 16 hrs. Analysis of the reaction by LCMS showed 61 .7% of desired product. The mixture was concentrated in vacuum and analysis by TLC (Petroleum ether / Ethyl acetate = 2 / 1) showed several new spots (Rf = 0.24, 0.52 (Pl) and 0.78). The residue was purified by column chromatography (S1O2, Petroleum ether / Ethyl acetate = 30 / 1 to 8 / 1) to give the title compound (6.60 g, 14.9 mmol, 83.9% yield, 97.0% purity) as an off-white solid. LCMS: (ES+) C23H29BF2N2O3 requires: 430, found: 431 [M+H]+. SFC: (Rt = 0.785 min, ee = 100%). ’H NMR: (400 Hz, CDCh) <57.50 (d, 7 = 8.8 Hz , 1H), 6.94-6.81 (m, 2H), 5.49 (dd, 7 / = 5.2 Hz, J2= 12.0 Hz, 1H), 4.40 (d, 7 = 48 Hz, 2H), 3.34-3.27 (m, 1H), 2.78-2.72 (m, 1H), 2.48 (s, 3H), 2.18 (s, 6H), 1.31 (d, 7 = 3.6 Hz , 12H).19F NMR: (400 Hz, CDCh 3 -115.57, -223.27.EXAMPLE 3Attorney Docket No. MDA0091-401 -PC(S)-(5-(2-Fluoro-4-methyl-5-(4,4,5,5-tetramethyI-l,3,2-dioxaborolan-2-yl)phenyl)-4,5- dihydro- lH-pyrazol-l-yl)(3-(fluoromethyl)bicydo[l.l.l]pentan-l-yl)methanone

[0142] To a solution of compound of Example IB (8.80 g, 22.2 mmol, 1.00 eq) in dioxane (100 mL) was added B2Pin2 (11.3 g, 44.5 mmol, 2.00 eq), K3PO4 (14.1 g, 66.6 mmol, 3.00 eq), Pd(dppf)C12.CH2Ch (3.63 g, 4.45 mmol, 0.20 eq) and Ag2O (7.73 g, 33.3 mmol, 1.50 eq). The mixture was stirred at 100 °C for 16 hrs. LCMS showed 53.2% of desired product. The mixture was concentrated in vacuum. Analysis of the reaction by TLC (Petroleum ether / Ethyl acetate = 2 / 1) showed several new spots (Rf = 0.24, 0.52 (Pl) and 0.78). The residue was purified by column chromatography (SiCL, Petroleum ether / Ethyl acetate = 30 / 1 to 8 / 1) to give the title compound (5.50 g, 12.3 mmol, 55.4% yield, 96.4% purity) as an off-white solid. LCMS: (ES+) C23H29BF2N2O3 requires: 430, found: 431 [M+H]+. SFC: (Rt = 0.876 min, ee = 100%).!H NMR: (400 Hz, CDCh) <57.50 (d, 7 = 9.2 Hz , 1H), 6.94-6.81 (m, 2H), 5.49 (dd, 7 / = 5.2 Hz, J2= 12.0 Hz, 1H), 4.40 (d, 7 = 48 Hz, 2H), 3.34-3.27 (m, 1H), 2.78-2.72 (m, 1H), 2.48 (s, 3H), 2.18 (s, 6H), 1.31 (d, 7 = 3.6 Hz , 12H).19F NMR: ( 400 Hz, CDCh) d -115.58, -223.28.EXAMPLE 4(S)-(5-(5-Fluoro

[0018] -2-fluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(fluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanone

[0143] The title compound was synthesized by the copper-mediated deboronation- fluorination reaction in full automation on an automated synthesizer from the corresponding precursor, the compound of Example 3. [18F]Fluoride (as HF) was produced by a GEAttorney Docket No. MDA0091-401 -PCPETtrace cyclotron (GE Healthcare) by 16 MeV irradiation of enriched |sO|I hO target. Approximately 40 GBq of [18F]Fluoride was trapped on a QMA light Sep-Pak cartridge and the18O water eluted into a collection vial. Kryptofix carbonate solution [Kryptofix (8.0 mg, 21.2 pmol), potassium carbonate (1.1 mg, 8.0 pmol), acetonitrile (0.650 mL), and water (0.200 mL)] was used to elute the QMA cartridge into the reaction vessel. The [18F]Fluoride / Kryptofix / carbonate mixture was azeotropically dried at 120 °C under a mixture of nitrogen pressure and vacuum using 1 mL of anhydrous acetonitrile. 8 mg of the pinacol boronate ester 7-2 precursor was introduced at once in the reactor vessel. Labeling occurred for 20 min at 110°C, then the reactor was cooled to 80 °C. The reaction mixture was cooled down to 40 °C and aqueous ascorbic acid solution (25%) was added. The mixture was then filtered through an alumina cartridge and glass fiber filter to remove unreacted fluoride and copper-derived solid residues. The crude was purified using a Cl 8 semi -preparative HPLC column with an isocratic mobile phase (50% acetonitrile, 0.085% orthophosphoric acid in water, flow rate 4.0 mL / min). The collected fraction was diluted in water (50 mL) and reformulated via solid phase extraction (SPE) on a tC18 cartridge. Pure compound was eluted off the cartridge with ethanol (1 mL) into a double neck vial containing 0.9% normal saline (10 mL).

[0144] The identity and radiochemical purity of the product were determined by radio- HPLC using the following conditions: HPLC Agilent 1200. Column: Agilent Zorbax Eclipse XDB-C8 4.6 x 150 mm, 5 pm. Mobile Phase A: 0.1% Formic acid in water. Mobile Phase B: 0.1% Formic acid in acetonitrile. Gradient: 0 minutes 50% A; 0 to 5 minutes 5% A; 5 to 10 minutes 50% A.Retention time of reference standard : 4.418 min at 250 nm; retention time of the radioactive product: 4.524 min.EXAMPLE 5(R)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(fluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanoneAttorney Docket No. MDA0091-401 -PC

[0145] Compound 5c was synthesized from 3-(((tert- butyldipheny Isi ly 1 )oxy )methy I )bicyclo[ 1 . 1 . 1 Ipentane- 1 -carboxy lie acid and 5-(2,5-difluoro- 4-methylphenyl)-4,5-dihydro-lH-pyrazole using HATU in DMF as the coupling reagent. Compound 5d was synthesized from compound 5c using HC1 in dioxane and DCM as the solvent for deprotection of the TBS group. Compound 5d-l was separated from the 5d mixture of enantiomers by SFC purification. The title compound was synthesized from 5d-l in DCM using DAST as the fluorinating reagent, similar to the procedure used in Example 7 MS (ES+) C17H17F3N2O requires: 322, found: 323 [M+H]+. ’H NMR (400 MHz, CDCI3) S 6.95 (s, 1H), 6.89-6.85 (m, 1H), 6.70-6.66 (m, 1H), 5.51-5.46 (m, 1H), 4.47 (s, 1H), 4.35 (s,1H), 3.40-3.32 (m, 1H), 2.77-2.71 (m, 1H), 2.22 (s, 3H), 2.19 (s, 6H).EXAMPLE 6(S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(fluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanone

[0146] Compound 5d-2 was separated from 5d by SFC purification similarly to what was described in Example 5. The title compound was synthesized in DCM using DAST as the fluorinating reagent, similar to the procedure described in Example 7, to give the titleAttorney Docket No. MDA0091-401 -PC compound. MS (ES+) C17H17F3N2O requires: 322, found: 323 [M+H]+.]H NMR (500 MHz, CDCI3) 5 6.94 (d, J = 1.7 Hz, 1H), 6.86 (dd, J = 10.2, 6.2 Hz, 1H), 6.67 (dd, J = 9.4, 6.2 Hz, 1H), 5.48 (dd, J = 12.0, 5.3 Hz, 1H), 4.45 (s, 1H), 4.35 (s, 1H), 3.40 - 3.30 (m, 1H), 2.78 - 2.69 (m, 1H), 2.21 (d, J = 2.0 Hz, 3H), 2.18 (s, 6H).EXAMPLE 7(S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3- (difluoromethyl)bicyclo[ 1.1.1 ]pentan- 1 -yl) methanoneStep 1: (S)-3-(5-(2,5-Difluoro-4-methyIphenyl)-4,5-dihydro-lH-pyrazole-l- carbonyl)bicyclo[l.l.l]pentane-l-carbaldehyde (7a)5d-2 7a

[0147] To a solution of oxalyl chloride (0.18 mL, 2.0 mmol) in DCM (19.5 mL) was added DMSO (0.29 mL, 4.1 mmol) and the resulting mixture was stirred at -78 °C for 10 min. A solution of 5d-2 (500 mg, 1.6 mmol) in DCM (3.9 mL) was added dropwise and stirred for 15 min, then TEA (1.3 mL, 9.4 mmol) was added dropwise. The resulting mixture was stirred for 1 hr at -78 °C then allowed to reach room temperature. The solution was poured into aqueous saturated NaHCCL, and the layers were separated. The aqueous phase was extracted with DCM (3 x), the combined organic layers were washed with aqueous saturated NaCl, dried over Na2SO4, filtered and concentrated under reduced pressure. The crude product was re-dissolved in EtOAc and washed with 1 M HC1, and brine, extracted aqueous layers once with EtOAc, combined organics, dried over MgSO4, filtered, concentrated, and purified by flash chromatography (0 - 60 % EtOAc:IPA 4: 1 in hexanes, 25 min) to give the title compound (350 mg, 1.1 mmol, 70 % yield) as a white solid. MS(ES+) C17H16F2N2O2 requires: 318, found: 319 / 337 [M+H]+ / [M+H30]+. 1H NMR (600 MHz,Attorney Docket No. MDA0091-401 -PCDMSO) 5 9.56 (s, 1H), 7.27 (s, 1H), 7.17 (dd, J = 10.7, 6.2 Hz, 1H), 6.78 (dd, J = 9.2, 6.7 Hz, 1H), 5.37 (dd, J = 12.2, 5.3 Hz, 1H), 3.44 (dd, J = 18.9, 12.1 Hz, 1H), 2.75 - 2.70 (m, 1H), 2.28 (s, 6H), 2.19 (s, 3H).Step 2: (S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(difluoromethyl)bicyclo[l.l.l]pentan-l-yl)methanone

[0148] To a solution of 7a (20 mg, 0.063 mmol) in DCM (314 pL) was added DAST (9.96 pL, 0.075 mmol) and the resulting mixture was stirred at 25°C for 16 h. The residue was purified by mass-triggered preparative HPLC (Mobile phase: A = 0.1% TFA / H2O, B = 0.1% TFA / MeCN; Gradient: B = 10 - 90%; 12 min; Column: C18) to give the title compound (12 mg, 0.035 mmol, 56.1 % yield) as a tan solid. LCMS: (ES+) C17H16F4N2O requires: 340, found: 341 [M+H]+. ’H NMR (500 MHz, DMSO) 8 7.28 - 7.24 (m, 1H), 7.20 - 7.13 (m, 1H), 6.81 - 6.74 (m, 1H), 6.07 (t, 7= 56.2 Hz, 1H), 5.40 - 5.33 (m, 1H), 3.48 - 3.39 (m, 1H), 2.76 - 2.67 (m, 1H), 2.21 - 2.17 (m, 3H), 2.13 (s, 6H).EXAMPLE 8(S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-((trifluoromethoxy)methyl)bicyclo[l.l.l]pentan-l-yl)methanone5d-2 Example 8

[0149] To a suspension of silver trifluoromethane sulfonate (64 mg, 0.25 mmol), KF (15 mg, 0.25 mmol), and Selectfluor (24 mg, 0.069 mmol) was added dropwise 5d-2 dissolved inAttorney Docket No. MDA0091-401 -PCEtOAc (310 pL) which was previously degassed with N2 for 1 minutes. The reaction was capped under nitrogen, 2-fluoropyridine (24 mg, 0.25 mmol) and trimethyl(trifluoromethyl)silane (23 pL, 0.16 mmol) were added and the reaction mixture was stirred to 25 °C for 16 h. The reaction mixture was filtered through a silica plug and concentrated under reduced pressure. The residue was purified by mass-triggered preparative HPLC (Mobile phase: A = 0.1% TFA / H2O, B = 0.1% TFA / MeCN; Gradient: B = 10 - 90%; 12 min; Column: XBridge C18, 5 pm, 19 mm x 150 mm) to give the title compound (7 mg, 0.018 mmol, 28.9 % yield) as a tan solid. LCMS: (ES+) C18H17F5N2O2 requires: 388, found: 389 [M+H]+. ’H NMR (600 MHz, DMSO) 5 7.26 - 7.22 (m, 1H), 7.19 - 7.13 (m, 1H), 6.78 - 6.72 (m, 1H), 5.39 - 5.33 (m, 1H), 4.13 (s, 2H), 3.47 - 3.39 (m, 1H), 2.74 - 2.66 (m, 1H), 2.22 - 2.18 (m, 3H), 2.05 (s, 6H).EXAMPLE 9((S)-5-(2,5-difhioro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(l- fluoroethyl)bicyclo[l.l.l]pentan-l-yl)methanoneStep 1((S)-5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(l- hydroxyethyl)bicydo[l.l.l]pentan-l-yl)methanone (9a)

[0150] To a cooled -78°C solution of compound 7a (100 mg, 0.314 mmol) in THF (3.14 mL) were added methylmagnesium bromide (157 pL, 0.471 mmol) dropwise. The resulting mixture was stirred at -78°C for 2 hr. At -78°C aqueous saturated NH4CI was added andAttorney Docket No. MDA0091-401 -PC allowed to reach room temp, and the layers were separated. The aqueous phase was extracted with EtOAc (3x), the combined organic layers were washed with aqueous saturated NaCl, dried over Na2SC>4, filtered and concentrated under reduced pressure. The residue was purified via silica gel chromatography (10 - 100 % EtOAc in hexanes to give compound 9a (115 mg, 0.344 mmol, 99 % yield) as a pale yellow liquid. MS (ES+) C18H20F2N2O2 requires: 334, found: 335 [M+H]+.Step 2((S)-5-(2,5-difluoro-4-methylphenyI)-4,5-dihydro-lH-pyrazol-l-yl)(3-(l- fluoroethyl)bicyclo[l.l.l]pentan-l-yl)methanone (Example 9)

[0151] To a solution of compound 9a (20 mg, 0.060 mmol) in toluene (150 pL) were added pyridine-2-sulfonyl fluoride (10.60 mg, 0.066 mmol) and DBU (18.03 pL, 0.120 mmol) and the resulting mixture was stirred at 20°C for 48hrs. TBAF (65.8 pL, 0.066 mmol) was added as a fluorine source and stirred overnight. H2O was added, and the layers were separated. The aqueous phase was extracted with EtOAc (3x), the combined organic layers were washed with aqueous saturated NaCl, dried over Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by mass-triggered preparative HPLC (Mobile phase: A = 0.1% TFA / H2O, B = 0.1% TFA / MeCN; Gradient: B = 30 - 70%; 20 min; Column: C18) to give the title compound (3.7 mg, 0.011 mmol, 18.39 % yield). MS (ES+) C18H19F3N2O requires: 336, found: 337 [M+H]+.]H NMR (600 MHz, DMSO) 5 7.25 - 7.22 (m, 1H), 7.16 (dd, J = 10.6, 6.2 Hz, 1H), 6.74 (dd, J = 9.7, 6.1 Hz, 1H), 5.37 (dd, J = 12.1, 5.2 Hz, 1H), 4.67 (dq, J = 48.9, 6.3 Hz, 1H), 3.43 (ddd, J = 18.8, 12.1, 1.6 Hz, 1H), 2.70 (ddd, J = 18.8, 5.3, 1.8 Hz, 1H), 2.19 (d, J = 1.8 Hz, 3H), 1.99 (dddd, J = 40.2, 9.4, 4.3, 1.5 Hz, 6H), 1.20 (dd, J = 23.9, 6.3 Hz, 3H).EXAMPLE 10Attorney Docket No. MDA0091-401 -PC(S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(l,l- difluoroethyl)bicydo[l.l.l]pentan-l-yl)methanoneStep 1(S)-l-(3-(5-(2,5-difluoro-4-methyIphenyl)-4,5-dihydro-lH-pyrazole-l- carbonyl)bicyclo[l.l.l]pentan-l-yl)ethan-l-one (10a)

[0152] To a solution of 9a (50 mg, 0.150 mmol) in DCM (1 mL) was added Dess-Martin Periodinane (95 mg, 0.224 mmol) and the resulting mixture was stirred at 20°C for 0.5hr. aqueous saturated NaHCCh was added and the layers were separated. The aqueous phase was extracted with DCM (3x), the combined organic layers were washed with aqueous saturated NaCl, dried over NaiSCL, filtered, and concentrated under reduced pressure. The residue was purified via silica gel chromatography (10 - 100 % EtOAc in hexanes) to give compound 10a (22 mg, 0.066 mmol, 44.3 % yield) as white amorphous material. MS (ES+) C18H18F2N2O2 requires: 332, found: 333 [M+H]+. ’H NMR (500 MHz, DMSO) 5 7.26 (s, 1H), 7.17 (dd, J = 10.6, 6.2 Hz, 1H), 6.76 (dd, J = 9.7, 6.2 Hz, 1H), 5.37 (dd, J = 12.1, 5.3 Hz, 1H), 3.49 - 3.38 (m, 1H), 2.76 - 2.67 (m, 1H), 2.26 (s, 6H), 2.19 (s, 3H), 2.10 (s, 3H).Step 2(S)-(5-(2,5-difluoro-4-methylphenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3-(l,l- difluoroethyl)bicyclo[l.l.l]pentan-l-yl)methanone

[0153] To a solution of compound 10a (20 mg, 0.060 mmol) in DCM (0.3 mL) at 0°C was added DAST (0.024 mL, 0.181 mmol) and the resulting mixture was stirred at 20°C overnight. Aqueous saturated Na2SO3 was added and the layers were separated. The aqueous phase was extracted with DCM (3x), the combined organic layers were washed with aqueousAttorney Docket No. MDA0091-401 -PC saturated NaCl, dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified via silica gel chromatography (10 - 100 % EtOAc in hexanes) to give the title compound (10.4 mg, 0.029 mmol, 48.8 % yield) as white amorphous material. MS (ES+) C18H18F4N2O requires: 354, found: 355 [M+H]+.]H NMR (500 MHz, DMSO) 5 7.26 (d, J = 1.7 Hz, 1H), 7.17 (dd, J = 10.6, 6.2 Hz, 1H), 6.77 (dd, J = 9.8, 6.1 Hz, 1H), 5.37 (dd, J = 12.1, 5.3 Hz, 1H), 3.49 - 3.39 (m, 1H), 2.77 - 2.66 (m, 1H), 2.24 - 2.16 (m, 3H), 2.12 (d, J = 4.3 Hz, 6H), 1.57 (t, J = 18.7 Hz, 3H).

[0154] Compounds containing anC radiolabel can be prepared following procedures known for the radiolabeling of pinacol borane esters with “CHsI. A non- limiting representative example is described below.EXAMPLE 11(S)-(5-(2,5-difluoro-4-(methyl-nC)phenyl)-4,5-dihydro-lH-pyrazol-l-yl)(3- (fhioromethyl)bicydo [1.1.1] pentan- 1 -yl)methanone

[0155] “CHsI can be prepared from "CCb, which can be produced by bombardment with protons in a nitrogen gas mixture (N2 + 1% O2) via the nuclear reaction14N(p,n)nC. ThenCO2can be processed through an automated synthesis module to producenCH3l .Typically,nCO2 can be converted tonCH4 in the presence of hydrogen gas and a Ni catalyst at 350°C for about 2 minutes.nCH4 can be captured in a trapping column at -75°C. ThenCH4 can be converted to1'CH d by reacting with sublimed I2, which can be prepared by heating I2 crystals at 100°C. ThenCH3l can be trapped by a Porapak Q at approximately room temperature and can be released at 200°C. The flow rate of He gas can be used for movement of materials within the synthesis and purification equipment within 10 - 120 mL / min. The1'CHsI can be transferred to a second module directly inside a reactor containing DMF, the boronic acid / ester precursor and a Pd catalyst. Once the transfer is completed, the mixture can be heated to 60°C for a specified time, typically 10 minutes. The reaction mixture can be cooled down to 40°C, diluted with a mixture 1 :1 acetonitrile / water and purified through semi-prep HPLC. The fraction collected during the HPLC purification can be purified via SPE and the final product eluted using ethanol, typically 1 mL. TheAttorney Docket No. MDA0091-401 -PC borane ester precursor in this Example can be prepared using similar procedures as those described for Examples 2 and 3 from the appropriate starting materials.

[0156] The following radiolabeled Examples may be prepared with procedures that were similar to the examples disclosed herein and can generally be made by methods disclosed herein from the appropriate starting materials.Attorney Docket No. MDA0091-401 -PC

[0157] The activity of the compounds described herein is illustrated in the following assays. The compounds described herein can be tested for efficacy in the treatment or prevention of symptoms or indications of RIPK1 -mediated diseases using techniques well known to those in the art.Biological Activity Assays

[0158] The activity of compounds disclosed herein is illustrated in the following assays.In Vivo Evaluation of Fluorescent Compounds

[0159] A dynamic 30-min micro-PET scan was performed on a SPECT / PET / CT Bruker Albira system (Bruker Corporation, Billerica, MA) immediately after intravenously injecting 5xFAD transgenic mice with the compound of Example 4. The compound was dosed at -200 pCi via intravenous (IV) administration. Mice were kept under anesthesia throughout the scan and data was collected from 0 to 30 mins after IV administration. PET time frames were: 12 x 10 seconds, 3 x 60 seconds, 5 x 300 seconds. A maximum likelihood expectation maximization (MLEM) method was used to reconstruct PET images with attenuation, randoms, scatter and decay corrections (volume size: 20 x 20 mm, voxel size 0.5 mm) (FIGS.1 and 2). For quantitative analyses, dynamic PET images were co-registered to a standardized mouse brain atlas available in the FUSION application within PMOD. The PMOD software 3.5 (PMOD Technologies, Zurich, Switzerland) was used to createAttorney Docket No. MDA0091-401 -PC predefined brain ROIs from the template and transformed to the co-registered PET images. This allowed the collection of time-activity curves (TACs) (FIG. 3). The uptake of radioactivity was standardized to body weight and the dose of radioactivity injected to yield standardized uptake value (SUV). A PET scan was performed 2 hours post-administration of Example 4 on a SPECT / PET / CT Bruker Albira system to confirm compound washout (FIG. 4).

[0160] Together, these results demonstrate that the compound:• Rapidly distributes systemically (FIG. 1).• Accumulates in RIPKl-rich brain regions (FIG. 2).• Shows differential uptake in disease models (FIG. 3).• Washes out from non-target tissues while retaining signal in target areas (FIG. 4).

[0161] These findings support the compound’s functionality as a PET ligand for imaging RIPK1 -mediated diseases, such as neurodegenerative disorders. The imaging data validate its potential for non-invasive diagnosis and monitoring of disease progression or therapeutic response.

[0162] It is expected that other radiolabeled compounds disclosed herein will similarly be efficacious in methods of positron emission tomography (PET) imaging and related uses, such as detecting RIPK1 -mediated disease, evaluating levels or change in RIPK1 expression, and detecting or monitoring changes in a RIPK1 -expressing tumor.Human U937 Cellular Necroptosis Assay

[0163] The human monocytic cell line U937 (CRL-1593.2) was purchased from ATCC. The cells were routinely maintained in RPMI-1640 Medium (Gibco, Catalog #11875-093) supplemented with 10% heat inactivated fetal bovine serum (Gibco, Catalog #16140-071), 100 units / mL penicillin and 100 pg / mL streptomycin (Gibco, Catalog #15140-122), in a humidified incubator (37°C, 5% CO2). For the assay, cells were resuspended in RPMI-1640 phenol red free Media (Gibco, Catalog # 11835-030) supplemented with 10% fetal bovine serum (Sigma, Catalog #F2442), 100 units / mL penicillin and 100 pg / mL streptomycin. Cells were stimulated with 25 ng / mL human TNF alpha (Cell Sciences, Catalog #CSI15659B) and 25pM z-VAD-FMK (R&D Systems, Catalog #FMK001) followed by seeding 5000 cells per well in a volume of 40 pL to a white, CulturPlate-384 (Perkin Elmer, Catalog #6007680). Stock solutions of the test compounds were prepared in 100% DMSO (Sigma, Catalog #D2650) and serially diluted 1:3 using 100% DMSO. Compounds were additionally diluted 1:40 in assay medium, and 10 pL / well was transferred to the plate. Following the compoundAttorney Docket No. MDA0091-401 -PC addition, the plate was incubated at 37°C and 5% CO2 for 22 h. After 22 h, viability was assessed with the addition of 20 pL of Cell Titer-Gio 2.0 (Promega, Catalog #G9243). The tissue culture plate was shaken on an orbital shaker at 300 RPM for 15 minutes at rt in the dark. Luminescence was measured using a PerkinElmer Envision™ plate reader. IC50 values were calculated using a four-parameter logistic curve fit using Genedata Screener software. Results for the “cold” (i.e., representative compounds lacking a radiolabel) compounds are shown below.Table 1. hU937 activity of cold compounds101641 All references, patents or applications, U.S. or foreign, cited in the application are hereby incorporated by reference as if written herein in their entireties. Where any inconsistencies arise, material literally disclosed herein controls.

[0165] From the foregoing description, one skilled in the art can easily ascertain the essential characteristics of this disclosure, and without departing from the spirit and scope thereof, can make various changes and modifications of the disclosure to adapt it to various usages and conditions.

Claims

Attorney Docket No. MDA0091-401 -PCCLAIMSWhat is claimed is:

1. A compound of structural Formula Ior a pharmaceutically acceptable salt thereof, whereinRlais chosen from halo, haloalkoxy, and R*;Rlhis chosen from hydrogen, halo, alkyl, and R*;Rlcis chosen from hydrogen, halo, and R*;R2is chosen from halo and R*;R3is chosen from alkyl, halo, pinacol borane, and R*;R4is chosen from halo, pinacol borane, and R*; andR* is a radioactive isotope or a moiety comprising a radioactive isotope, wherein at least one of Rla, Rlb, Rlc, R2, R3, and R4is R*.

2. The compound of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R2is fluoro.

3. The compound of claim 1 or 2, or a pharmaceutically acceptable salt thereof, wherein Rlais chosen from fluoro and trifluoromethoxy.

4. The compound of claim 3, or a pharmaceutically acceptable salt thereof, wherein Rlais fluoro.

5. The compound of any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, wherein Rlbis chosen from hydrogen, fluoro, and methyl.

6. The compound of any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, wherein Rlcis chosen from hydrogen and fluoro.

7. The compound of any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, wherein Rlband Rlcare hydrogen.

8. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein Rlais fluoro and Rlband Rlcare hydrogen.

9. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein Rlaand Rlbare fluoro and Rlcis hydrogen.Attorney Docket No. MDA0091-401 -PC10. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein Rlais fluoro, Rlbis methyl, and Rlcis hydrogen.

11. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein Rlais fluoro, Rlbis methyl, and Rlcis fluoro.

12. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein Rlais trifluoromethoxy and Rlband Rlcare hydrogen.

13. The compound of any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, wherein R3is methyl and R4is R*.

14. The compound of any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, wherein R3is R* and R4is chosen from fluoro, bromo, and pinacol borane.

15. The compound of claim 14, or a pharmaceutically acceptable salt thereof, wherein R4is fluoro.

16. The compound of any one of the preceding claims, or a pharmaceutically acceptable salt thereof, wherein R* is a moiety comprising a positron-emitting radioisotope or a positronemitting radioisotope.

17. The compound of claim 16, or a pharmaceutically acceptable salt thereof, wherein R* is chosen fromnCH3and18F.

18. The compound of claim 17, or a pharmaceutically acceptable salt thereof, wherein R* is 1SF.

19. The compound of claim 1 , having structural Formula IAor a pharmaceutically acceptable salt thereof, whereinRlais chosen from halo and haloalkoxy;Rlbis chosen from hydrogen, halo, and alkyl;Rlcis chosen from hydrogen and halo; andR3is -CH3and R4is18F, or R3is -nCH3and R4is F.

20. The compound of claim 1 , or a pharmaceutically acceptable salt thereof, having a structural formula chosen fromAttorney Docket No. MDA0091-401 -PC21. A pharmaceutical formulation comprising a compound as recited in any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier.

22. The pharmaceutical formulation as recited in claim 21, formulated for intravenous administration.

23. A method for positron emission tomography (PET) imaging in a subject, comprising administering to the subject a radiotracer comprising a compound of any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation of claim 21 or 22, and subjecting the subject to a positron emission tomography (PET) scan.

24. The method of claim 23, further comprising one or more steps chosen from: a. measuring the amount of gamma radiation emitted from a tissue; b. measuring the concentration of the radiotracer in a tissue; and c. determining an amount of the radiotracer in a tissue.

25. A method of detecting a RIPK1 -mediated disease in a subject, comprising administering to the subject a compound of any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation of claim 21 or 22, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound.

26. A method of evaluating a change in RIPK1 expression in a subject in response to a treatment for a RIPK1 -mediated disease in the subject, comprising a. administering to the subject a compound of any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation ofAttorney Docket No. MDA0091-401 -PC claim 21 or 22, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b. administering to the subject a treatment for the RIPK1 -mediated disease; c. administering to the subject one or more subsequent doses of the compound; d. measuring subsequent amounts of RIPK1 expression in the subject; and e. comparing the first and subsequent amounts of RIPK1 expression.

27. The method as recited in claim 25 or 26, wherein the amount of the compound is determined within an inflammatory, apoptotic, or necroptotic process of the RIPK1- mediated disease.

28. The method as recited in claim 27, wherein the inflammatory process is RIPK1- dependent cytokine release.

29. The method as recited in claim 28, wherein the necroptotic process is RIPK1 -dependent necroptosis.

30. The method as recited in any one of claims 25 to 29, wherein the RIPK1 -mediated disease is chosen from a neurological disease, a neuropathy, an autoimmune disorder, an inflammatory disease, or cancer.

31. The method as recited in claim 30, wherein the RIPK1 -mediated disease is a neurological disease.

32. The method as recited in claim 31, wherein the neurological disease is accompanied by an inflammatory component of cellular stress.

33. The method as recited in claim 31, wherein the neurological disease is chosen from Multiple Sclerosis, Neimann-Pick disease, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, Lewy body dementia, frontotemporal dementia, glutamine expansion disease, Huntington’s disease, Kennedy’s disease, and spinocerebellar ataxia.

34. The method as recited in claim 30, wherein the RIPK1 -mediated disease is a neuropathy.

35. The method as recited in claim 34, wherein the neuropathy is chosen from diabetic neuropathy and chemotherapy -induced neuropathy.

36. The method as recited in claim 30, wherein the RIPK1 -mediated disease is a retinal disease.

37. The method as recited in claim 36, wherein the retinal disease is chosen from macular degeneration and retinitis.

38. The method as recited in claim 30, wherein the RIPK1 -mediated disease is an autoimmune disorder.Attorney Docket No. MDA0091-401 -PC39. The method as recited in claim 38, wherein the autoimmune disorder is chosen from ulcerative colitis, rheumatoid arthritis, psoriasis, lupus, and inflammatory bowel disease.

40. The method as recited in claim 30, wherein the RIPK1 -mediated disease is an inflammatory disease.

41. The method as recited in claim 40, wherein the inflammatory disease is found in one or more organs chosen from the lung, heart, kidney, and liver.

42. The method as recited in claim 30, wherein the RIPK1 -mediated disease is cancer.

43. The method as recited in claim 42, wherein the cancer can be treated by promoting an appropriate immune response to the tumor.

44. The method as recited in claim 43, wherein the appropriate immune response to the tumor comprises, or results in, one or more of the following: a) an increase in the number or activity, or degree of tumor infiltration, of cytotoxic T-lymphocytes and / or natural killer cells; b) an increase in the number or activity of Ml macrophages in the tumor microenvironment and / or a decrease in the in the number or activity of M2 macrophages in the tumor microenvironment; c) a decrease in the number or activity of regulatory T cells; and d) a decrease in the number or activity of myeloid-derived suppressor cells.

45. The method as recited in claim 42, wherein the cancer is a myelodysplastic syndrome (MDS).

46. The method as recited in claim 42, wherein the cancer is acute myeloid leukemia (AML).

47. The method as recited in any one of the preceding claims, wherein the amount of the compound is determined with positron emission tomography imaging.

48. The method as recited in claim 47, wherein the positron emission tomography imaging confirms uptake and distribution into the brain.

49. A method of detecting or monitoring changes in a RIPK1 -expressing tumor in a subject, comprising administering to the subject a compound of any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation of claim 21 or 22, subjecting the subject to a positron emission tomography (PET) scan, and determining an amount of the compound, wherein an increased amount of the tracer as compared to a control indicates a RIPK1 -expressing tumor.

50. A method of evaluating a change in RIPK1 expression in a subject in response to an anticancer treatment, comprising:Attorney Docket No. MDA0091-401 -PC a. administering to the subject a compound of any one of claims 1 to 20, or a pharmaceutically acceptable salt thereof, or a pharmaceutical formulation of claim 21 or 22, subjecting the subject to a positron emission tomography (PET) scan, and measuring a first amount of RIPK1 expression in the subject by determining an amount of the compound; b. administering to the subject an anti-cancer treatment; c. administering to the subject one or more subsequent doses of the compound; d. measuring subsequent amounts of RTPK1 expression in the subject; and e. comparing the first and subsequent amounts of RIPK1 expression.

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