Methods for treating systemic lupus erythematosus and the use of biomarkers as predictors of clinical sensitivity to treatment.

Abstract

A method for determining the dosage of a therapeutic compound for treating a patient with systemic lupus erythematosus (SLE). [Solution] A method comprising (a) measuring the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a target sample, and (b) determining the dose of the therapeutic compound to be 0.45 mg or more or 0.15 mg or less per day if the score calculated based on the gene expression levels is higher than the reference level, wherein the therapeutic compound is a compound of formula I or a pharmaceutically acceptable salt thereof, the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, and the reference level is -1.38. JPEG2026062871000055.jpg46161

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the benefits of U.S. Provisional Patent Application No. 63 / 087,008, filed on 2 October 2020, which is incorporated herein by reference in its entirety.

[0002] In some embodiments, methods are provided herein for predicting and monitoring the clinical sensitivity and therapeutic response to compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, comprising (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, or for using specific biomarkers when determining the dose, in patients with various diseases and disorders, such as systemic lupus erythematosus (SLE). Kits for carrying out these methods are further provided. In certain embodiments, methods for determining the efficacy of a compound in the treatment of a disease are also provided herein. [Background technology]

[0003] Systemic lupus erythematosus (SLE) is a multi-organ autoimmune disease of unknown etiology with many clinical manifestations. While most organs can be involved, the most common manifestations are the skin, musculoskeletal system, and kidneys. SLE typically develops in young women of childbearing age, between 15 and 44 years of age. The prevalence of SLE is 300,000 in the United States, 4 million worldwide, and an annual incidence of 15,000 in the United States alone.

[0004] The etiology of SLE is likely to involve a series of components related to both genetic and environmental factors. Disease susceptibility is influenced by genes related to the immune response and major histocompatibility complex class I and II genes. Further susceptibility results from the interaction between the hormonal environment and the hypothalamic-pituitary-adrenal axis. Additionally, the onset of SLE is associated with an imperfect immune response that affects apoptotic cell clearance and immune complexes. Loss of immune tolerance, excessive T cell help, defective B cell suppression, and a shift in the immune response from T helper 1 (Th1) to Th2 and Th17 result in B cell hyperactivity and the production of pathogenic antibodies. External factors such as chemicals, drugs, ultraviolet light, diet, and viruses also contribute to the onset of the disease.

[0005] Since SLE is a waxing and waning disease, it is often controlled with NSAIDs or low-potency immunosuppressive drugs (antimalarials and low-dose corticosteroids) for milder symptoms (musculoskeletal symptoms, skin symptoms, and serositis). More prolonged and intensive use of corticosteroids and nonbiologic disease-modifying antirheumatic drugs (DMARDs) is the standard treatment available for treating patients with major organ involvement. In combination with standard treatment, there is biologic DMARD treatment to enhance the treatment of patients with more extensive disease. Belimumab, a monoclonal antibody and specific inhibitor of B lymphocyte stimulator, has recently been approved for use in combination with corticosteroids and other standard treatments for autoantibody-positive SLE. Additionally, rituximab, a B cell-depleting agent, is often used off-label as a rescue drug for patients who do not respond to standard treatment.

[0006] However, there is still a need for prophylactic or therapeutic agents that can be used to treat or prevent SLE. Numerous studies have been conducted for the purpose of providing compounds that can be safely and effectively used to treat SLE. Since the clinical efficacy of these compounds can usually only be measured with respect to the response of patients who require treatment for a minimum of several months, it cannot be easily and accurately predicted. Considering the drawbacks of conventional methods, there is a need to develop an efficient, sensitive, and accurate method for detecting, quantifying, and characterizing the pharmacodynamic activity of specific compounds. The present invention meets these and other needs.

Summary of the Invention

Means for Solving the Problems

[0007] In one aspect, a method for determining the dosage of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising: (a) obtaining a sample from the subject; (b) measuring the gene expression level of IKZF3 in the sample; and (c) when the score calculated based on the gene expression level of IKZF3 in the sample is higher than a reference level, determining that the dosage of the therapeutic compound is 0.45 mg or more per day, wherein the therapeutic compound is a compound of formula I:

Chemical formula

[0008] In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the score is the Log2 of the gene expression level of IKZF3 relative to the average gene expression level of two or more reference genes in the sample.

[0009] In some embodiments, the method comprises determining that the dosage of the therapeutic compound is 0.45 mg or more per day when the score is higher than -0.49.

[0010] In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0011] In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the baseline level.

[0012] In some embodiments, the method further comprises administering a dose of 0.45 mg or more of the therapeutic compound per day to the target.

[0013] In another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering a therapeutic compound to the subject at a dose of 0.45 mg or more per day, wherein the score calculated based on the gene expression level of IKZF3 in a sample from the subject is higher than the reference level, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0014] In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes. In some embodiments, the baseline level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0015] In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.7 mg / day.

[0016] In another embodiment, a method for determining the dose of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising: (a) obtaining a sample from the subject; (b) measuring the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample; and (c) determining the dose of the therapeutic compound to be 0.45 mg or more or 0.15 mg or less per day if the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0017] In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample.

[0018] In some embodiments, the method includes determining the dose of the therapeutic compound to be 0.45 mg or more per day, or 0.15 mg or less per day, if the score is higher than -1.38.

[0019] In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0020] In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.15 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.1 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.75 mg per day if the score is higher than the baseline level. In some embodiments, the above daily dose of the therapeutic compound is administered once daily. In some embodiments, the above daily dose of the therapeutic compound is administered every other day. In some embodiments, the above daily dose of the therapeutic compound is administered every three days. In some embodiments, the above daily dose of the therapeutic compound is administered once a week. For example, in some embodiments, the dose of the therapeutic compound is about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week.

[0021] In some embodiments, the method involves administering a therapeutic compound to a target at a dose of 0.45 mg or more or 0.15 mg or less per day.

[0022] In yet another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.15 mg or less per day, wherein the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L and RSAD2 in a sample from the subject is higher than the reference level, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0023] In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the score is greater than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0024] In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.7 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.1 mg / day. In yet another embodiment, the dose of the therapeutic compound is about 0.075 mg / day. In some embodiments, the above daily dose of the therapeutic compound is administered once daily. In some embodiments, the above daily dose of the therapeutic compound is administered every other day. In some embodiments, the above daily dose of the therapeutic compound is administered every three days. In some embodiments, the above daily dose of the therapeutic compound is administered once a week. For example, in some embodiments, the dose of the therapeutic compound is about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week.

[0025] In yet another embodiment, a method is provided herein for identifying subjects with systemic lupus erythematosus (SLE) that are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, comprising: (a) obtaining a sample from a subject; (b) determining (i) the gene expression level of IKZF3 or (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample; and (c) diagnosing that the subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression level of IKZF3 or IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0026] In some embodiments, the method includes the steps of determining the gene expression level of IKZF3 and diagnosing that the subject is likely to be responsive to a therapeutic compound if the score calculated based on the IKZF3 gene expression level is higher than a reference level. In some embodiments, the score is the Log2 of the IKZF3 gene expression level relative to a reference gene in the sample, or the score is the Log2 of the IKZF3 gene expression level relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the reference level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0027] In some embodiments, the method includes the steps of determining the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2, and diagnosing that a subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a baseline level. In some embodiments, the score is the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the baseline level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0028] In some embodiments, the method further includes administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound.

[0029] In yet another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering an effective amount of a therapeutic compound to the subject, wherein the subject has been determined to be likely to respond to the therapeutic compound by the method provided herein, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0030] In yet another embodiment, a method for determining the dose of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising: (a) obtaining a sample from the subject; (b) measuring (i) the gene expression level of IKZF3 and (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample; (c) determining a first score based on the gene expression level of IKZF3 and comparing the first score to a first reference level; (d) determining a second score based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 and comparing the second score to a second reference level; and (e) determining the dose of a therapeutic compound based on the first and second scores, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0031] In some embodiments, the first score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the first score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the first reference level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0032] In some embodiments, the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the second reference level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0033] In some embodiments, if the first score is higher than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day or 0.15 mg or less per day.

[0034] In other embodiments, if the first score is higher than the first reference level and the second score is lower than the second reference level, the dose of the therapeutic compound is determined to be 0.15 mg or less per day.

[0035] In yet another embodiment, if the first score is lower than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day.

[0036] In yet another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering a dose of a therapeutic compound to the subject, the dose of which is determined according to a method provided herein, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0037] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0038] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0039] In yet another embodiment, a method is provided herein for identifying subjects having systemic lupus erythematosus (SLE) that are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects having SLE to a therapeutic compound, comprising: (a) obtaining a sample from a subject; (b) determining the presence of the IKZF1 single nucleotide polymorphism (SNP) rs4917014 in the sample; and (c) diagnosing the subject as likely to be responsive to a therapeutic compound if at least one copy of the SNP rs4917014 is detected, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0040] In some embodiments, the method further includes administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound.

[0041] In yet another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering an effective amount of a therapeutic compound to the subject, wherein the subject has been determined to be likely to respond to the therapeutic compound by the method provided herein, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0042] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0043] In some embodiments, methods for treating subjects having systemic lupus erythematosus (SLE) are provided herein, comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.15 mg or less per day, wherein the subject has high type 1 IFN expression and / or gene signature, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0044] In some embodiments, if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from a subject is higher than a baseline level, the subject is determined to have high type 1 IFN expression and / or a gene signature. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the score is higher than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments, the dose of the therapeutic compound is approximately 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is approximately 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is approximately 0.6 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.7 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.1 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.075 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg every other day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg every three days. In some embodiments, the dose of the therapeutic compound is about 0.15 mg once a week. [Brief explanation of the drawing]

[0045] [Figure 1] This figure shows the distribution and overlap of the patient subsets with high IFN and high Aiolos levels in the study described in Example 2. [Figure 2]This figure shows the SRI-4 clinical response rates at week 24 in a subset of patients defined by their Aiolos and type 1 IFN gene signature status at baseline (Aiolos=IKZF3; type 1 IFN gene signatures=IFI27, IFI44, IFI44L, RSAD2; Δ=treatment effect (stratified difference vs placebo)). [Figure 3] This figure shows the SRI-4 clinical response rates to iverdide in SLE subsets, based on both type 1 IFN and Aiolos gene expression status. [Figure 4] This study shows the relationship between the Aiolos gene expression cutoff value, the SRI-4 clinical response rate to 0.45 mg iverdide, and the prevalence rate in the SLE research population. [Modes for carrying out the invention]

[0046] The methods provided herein are based in part on the finding that it is possible to predict the response of a subject having or suspected of having systemic lupus erythematosus (SLE) to a therapeutic compound (e.g., (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione), compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, using the level of a specific molecule (e.g., mRNA, cDNA, or protein) in a biological sample, or to determine the dose of the therapeutic compound for the subject.

[0047] 5.1 Definition As used herein, unless otherwise specified, the terms “to treat,” “to treat,” and “to treat” refer to an effect that occurs while a patient is suffering from a disease or disorder such as SLE, which reduces the severity of the disease or disorder or prevents or delays its progression.

[0048] The terms "susceptibility" or "being susceptible," when used in relation to treatment with a compound, are relative terms referring to the degree of the compound's effectiveness in reducing or mitigating the progression of the treated tumor or disease. For example, the term "increased susceptibility," when used in relation to the treatment of SLE in connection with a compound, indicates an increase of at least about 5% or more in the effectiveness of the SLE treatment.

[0049] As used herein, the terms “compound” and “therapeutic compound” are interchangeable and include compounds of formula I. Non-limiting examples of compounds include those disclosed in Section 5.5 below.

[0050] Where used herein, unless otherwise specified, the term “therapeutic dose” of a compound means an amount sufficient to provide a therapeutic benefit in the treatment or management of a disease or disorder (e.g., SLE), or to delay or minimize one or more symptoms associated with the presence of a disease or disorder (e.g., SLE). The therapeutic dose of a compound means the amount of a therapeutic agent, either alone or in combination with other therapies, that provides a therapeutic benefit in the treatment or management of a disease or disorder (e.g., SLE). The term “therapeutic dose” may also encompass an amount that improves overall treatment, reduces or avoids the symptoms or causes of a disease or disorder (e.g., SLE), or enhances the therapeutic efficacy of another therapeutic agent. The term also refers to an amount of a compound sufficient to induce a biological or medical response in a biological molecule (e.g., protein, enzyme, RNA, or DNA), cell, tissue, system, animal, or human, as sought by researchers, veterinarians, physicians, or clinicians.

[0051] The terms “responsiveness” or “responsive” when used in relation to treatment refer to the degree of effectiveness of the treatment in reducing or alleviating the symptoms of the disease being treated, for example, SLE. For example, when used in relation to the treatment of cells or subjects, the term “increased responsiveness” refers to an increase in effectiveness in reducing or alleviating the symptoms of the disease compared to a reference treatment (e.g., the same cells or subjects, or different cells or subjects), as measured using any method known in the art. In certain embodiments, the increase in effectiveness is at least about 5%, at least about 10%, at least about 20%, at least about 30%, at least about 40%, or at least about 50%.

[0052] As used herein, the terms “effective target response,” “effective patient response,” and “effective patient tumor response” refer to any increase in therapeutic benefit to the patient.

[0053] The term "likelihood" generally refers to an increase in the probability of an event occurring. When used in relation to the effectiveness of a treatment in a subject, the term "likelihood" generally refers to an increase in the probability of a disease progressing or its severity decreasing.

[0054] The term “predict” generally means to determine or notify in advance. When used to “predict” the effectiveness of a treatment, for example, the term “predict” may mean that the likelihood of the treatment outcome can be determined at the start of treatment, before the start of treatment, or before the treatment period has substantially progressed.

[0055] As used herein, the term “monitoring” generally refers to the control, supervision, regulation, observation, tracking, or surveillance of an activity. For example, the term “monitoring the effectiveness of a compound” refers to tracking its effectiveness in treating a patient’s disease or disorder (e.g., SLE). Similarly, when used individually or in a clinical trial in relation to patient compliance, the term “monitoring” refers to tracking or confirming that a patient is actually taking the drug being tested as prescribed. Monitoring can be carried out, for example, by tracking the expression of mRNA or protein biomarkers.

[0056] A "biological marker" or "biomarker" is a substance whose detection indicates a specific biological state. In some embodiments, biomarkers can be determined individually.

[0057] As used herein, the terms “expressed” or “expression” refer to transcription from a gene that yields an RNA nucleic acid molecule that is at least partially complementary to one region of the gene’s two nucleic acid strands. As used herein, the terms “expressed” or “expression” also refer to translation from an RNA molecule that yields a protein, polypeptide, or a portion thereof.

[0058] The term "level" refers to the quantity, accumulation, or rate of biomarker molecules. Levels can be expressed, for example, by the quantity or synthesis rate of messenger RNA (mRNA) encoded by a gene, the quantity or synthesis rate of polypeptides or proteins encoded by a gene, or the quantity or synthesis rate of biomolecules accumulated in cells or biological fluids. The term "level" also refers to the absolute or relative quantity of molecules in a sample, determined under steady-state or transient conditions.

[0059] As used herein, the terms “determine,” “measure,” “evaluate,” “assess,” and “assay” generally refer to any form of measurement, including determining whether or not an element is present. These terms include quantitative and / or qualitative decisions. Assessments may be relative or absolute. “Assessing the presence” may include determining the quantity of something that is present, and determining whether or not it is present.

[0060] As used herein, the term “sample” typically refers to a substance or mixture of substances in fluid form containing one or more components of the subject, but not necessarily.

[0061] As used herein, “biological sample” refers to a sample obtained from a living subject, including samples of living tissue or bodily fluid origin, and obtained, achieved, or collected in vivo or in situ. Such samples may be organs, tissues, and cells isolated from mammals, but are not limited to these. Exemplary biological samples include, but are not limited to, cell lysates, cell cultures, cell lines, tissues, oral tissues, gastrointestinal tissues, organs, organelles, bodily fluids, blood samples, urine samples, skin samples, etc.

[0062] As used herein, "tautomer" refers to isomeric forms of a compound that are in equilibrium with each other. The concentration of each isomeric form depends on the environment in which the compound is found, and may differ depending on whether the compound is in a solid state or in an organic solution or aqueous solution. For example, in aqueous solution, pyrazole may exhibit the following isomeric forms, called tautomers, of each other. [ka]

[0063] As used herein, unless otherwise specified, the term “pharmaceutically acceptable salt” includes non-toxic acid and base addition salts of the compound referred to by this term. Examples of acceptable non-toxic acid addition salts include those derived from organic and inorganic acids known in the art, such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, methanesulfonic acid, acetic acid, tartaric acid, lactic acid, succinic acid, citric acid, malic acid, maleic acid, sorbic acid, aconitic acid, salicylic acid, phthalic acid, embolic acid, and enanthic acid. Compounds having acidic properties can form salts with a variety of pharmaceutically acceptable bases. Bases that can be used to prepare pharmaceutically acceptable base addition salts of such acidic compounds are those that form non-toxic base addition salts, i.e., salts containing pharmaceutically acceptable cations such as alkali metal salts or alkaline earth metal salts (especially calcium, magnesium, sodium, or potassium salts). Suitable organic bases include, but are not limited to, N,N-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumaine (N-methylglucamine), lysine, and procaine.

[0064] As used herein, unless otherwise specified, the term “solvate” means a compound or salt thereof provided herein, further comprising a stoichiometric or non-stoichiometric amount of solvent bonded by non-covalent intermolecular forces. If the solvent is water, the solvate is a hydrate.

[0065] As used herein, unless otherwise specified, the term “stereoisomer” encompasses all enantiomerically / stereoisomerically pure and enantiomerically / stereoisomerically concentrated compounds of the present invention.

[0066] As used herein, unless otherwise specified, the term “stereoisomerically pure” means a composition containing one stereoisomer of a compound and substantially no other stereoisomers of that compound. For example, a stereoisomerically pure composition of a compound having one chiral center substantially does not contain the opposite enantiomer of the compound. A stereoisomerically pure composition of a compound having two chiral centers substantially does not contain the other diastereomer of the compound. A typical stereoisomerically pure compound contains more than about 80% by weight of one stereoisomer of the compound and less than about 20% by weight of the other stereoisomer of the compound; more preferably more than about 90% by weight of one stereoisomer of the compound and less than about 10% by weight of the other stereoisomer of the compound; even more preferably more than about 95% by weight of one stereoisomer of the compound and less than about 5% by weight of the other stereoisomer of the compound; and most preferably more than about 97% by weight of one stereoisomer of the compound and less than about 3% by weight of the other stereoisomer of the compound.

[0067] As used herein, unless otherwise specified, the term “stereoisomerically concentrated” means a composition comprising more than about 60% by weight of one stereoisomer of the compound, preferably more than about 70% by weight, and more preferably more than about 80% by weight of one stereoisomer of the compound. As used herein, unless otherwise specified, the term “enantiomerically pure” means a stereoisomerically pure composition of a compound having one chiral center. Similarly, the term “stereoisomerically concentrated” means a stereoisomerically concentrated composition of a compound having one chiral center.

[0068] It should also be noted that compounds can contain unnatural proportions of atomic isotopes in one or more atoms. For example, a compound may contain tritium ( 3 H), Iodine-125( 125 I), Sulfur-35( 35 S), or carbon-14 ( 14 It may be radioactively labeled with radioactive isotopes such as C, or deuterium ( 2 H), carbon-13 ( 13 C), or nitrogen-15( 15The compounds may be isotope-enriched with N, etc. As used herein, “isotopologs” are isotope-enriched compounds. The term “isotopically enriched” refers to an atom having an isotope composition other than the natural isotope composition of that atom. “Isotope-enriched” may also refer to a compound containing at least one atom having an isotope composition other than the natural isotope composition of that atom. The term “isotopologs” refers to the amount of each isotope present in a given atom. Radiolabeled and isotope-enriched compounds are useful as therapeutic agents, e.g., cancer and inflammation treatments, research reagents, e.g., binding assay reagents, and diagnostic agents, e.g., in vivo imaging agents. All isotope variants of compounds described herein, whether radioactive or not, are intended to be included within the scope of the embodiments provided herein. In some embodiments, isotopologs of compounds are provided, for example, isotopologs are deuterium, carbon-13, or nitrogen-15 enriched compounds. In some embodiments, the isotopologs provided herein are deuterium enriched compounds. In some embodiments, the isotopologs provided herein are deuterium-enriched compounds in which deuteration occurs at a chiral center. In some embodiments, isotopologs of compounds of formula I in which deuteration occurs on a chiral center are provided herein. In some embodiments, isotopologs of compounds C in which deuteration occurs at a chiral center are provided herein.

[0069] As used interchangeably herein, the terms “polypeptide” and “protein” refer to polymers of three or more amino acids linked in series via peptide bonds. The term “polypeptide” includes proteins, protein fragments, protein analogs, oligopeptides, etc. The term “polypeptide” as used herein may also refer to peptides. The amino acids constituting a polypeptide may be of natural origin or synthetic. Polypeptides can be purified from biological samples. Polypeptides, proteins, or peptides also include modified polypeptides, proteins, and peptides, such as glycopolypeptides, glycoproteins, or glycopeptides, or lipopeptides, lipopolypeptides, lipoproteins, or lipopeptides.

[0070] The terms “antibody,” “immunoglobulin,” or “Ig,” as used interchangeably herein, encompass fully assembled antibodies and antibody fragments that retain the ability to specifically bind to an antigen. Antibodies provided herein include, but are not limited to, synthetic antibodies, monoclonal antibodies, polyclonal antibodies, recombinant production antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, humanized antibodies, chimeric antibodies, intrabodies, single-chain Fv(scFv) (e.g., monospecific, bispecific, etc.), camelized antibodies, Fab fragments, F(ab') fragments, disulfide-linked Fv(sdFv), anti-idiotype (anti-Id) antibodies, and any of the epitope-binding fragments described above. In particular, antibodies provided herein include immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e., antigen-binding domains or molecules containing antigen-binding sites that immunospecifically bind to the CRBN antigen (e.g., one or more complementarity-determining regions (CDRs) of an anti-CRBN antibody). The antibodies provided herein may be of any class of immunoglobulin molecules (e.g., IgG, IgE, IgM, IgD, and IgA) or any subclass (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2). In some embodiments, the anti-CRBN antibody is a fully human antibody, e.g., a fully human monoclonal CRBN antibody. In certain embodiments, the antibody provided herein is an IgG antibody or a subclass thereof (e.g., human IgG1 or IgG4).

[0071] The terms “nucleic acid” and “polynucleotide” are used interchangeably herein to describe polymers of any length composed of nucleotides, such as deoxyribonucleotides or ribonucleotides, or synthetically produced compounds, that can hybridize with naturally occurring nucleic acids in a sequence-specific manner similar to the sequences of two naturally occurring nucleic acids, for example, and can participate in Watson-Crick base pairing interactions. Where used herein in the context of polynucleotide sequences, the term “bases” (or “base”) is synonymous with “nucleotides” (or “nucleotide”), i.e., the monomeric subunits of a polynucleotide. The terms “nucleoside” and “nucleotide” are intended to include moieties that include not only known purine and pyrimidine bases, but also other modified heterocyclic bases. Such modifications include methylated purines or pyrimidines, acylated purines or pyrimidines, alkylated ribose, or other heterocyclic bases. Furthermore, the terms “nucleoside” and “nucleotide” include moieties that include not only conventional ribose and deoxyribose sugars, but also other sugars. Modified nucleosides or nucleotides also include modifications on the sugar moiety, for example, by replacing one or more hydroxyl groups with halogen atoms or aliphatic groups, or by functionalization as ethers, amines, etc. "Analogs" refer to molecules having structural features recognized in the literature as mimics, derivatives, similar structures, or other similar terms, such as polynucleotides incorporating unnatural nucleotides, nucleotide mimics such as 2'-modified nucleosides, peptide nucleic acids, oligomeric nucleoside phosphonates, and any polynucleotides to which substituents such as protecting groups or linking moieties have been added.

[0072] The term "complementary" refers to specific binding between polynucleotides based on their sequences. As used herein, a first polynucleotide and a second polynucleotide are complementary if they bind to each other in a hybridization assay under stringent conditions, for example, if they produce a given or detectable level of signal in the hybridization assay. Polynucleotide segments are complementary if they follow conventional base pairing rules, e.g., A pairs with T (or U) and G pairs with C, but small regions (e.g., less than about 3 bases) of mismatches, insertions, or deletions may exist.

[0073] As used herein, the term “bonded” refers to direct or indirect adhesion. In the context of chemical structures, “bonded” (or “bonded”) may refer to the presence of a chemical bond that directly bonds two parts or indirectly bonds two parts (e.g., via a linking group or any other intervening part of the molecule). The chemical bond may be a covalent bond, an ionic bond, a coordination complex, a hydrogen bond, a van der Waals interaction, or a hydrophobic stacking, or may exhibit the properties of several types of chemical bonding. In certain examples, “bonded” includes embodiments in which the adhesion is direct and embodiments in which the adhesion is indirect.

[0074] As used herein, the term “scavenger” refers to a drug that binds to mRNA or protein and binds to mRNA or protein through interactions sufficient to concentrate mRNA or protein from a heterogeneous mixture.

[0075] As used herein, the term "probe" refers to a capture agent targeting a specific target mRNA biomarker sequence. Therefore, each probe in a probe set has its own target mRNA biomarker. A probe / target mRNA double helix is ​​a structure formed by hybridizing a probe to its target mRNA biomarker.

[0076] The terms “nucleic acid probe” or “oligonucleotide probe” refer to nucleic acids that can bind to target nucleic acids of complementary sequences, such as mRNA biomarkers provided herein, usually via complementary base pairing by forming hydrogen bonds. As used herein, probes may contain natural (e.g., A, G, C, or T) or modified bases (e.g., 7-deazaguanosine, inosine). Furthermore, the bases in the probe may be linked by bonds other than phosphodiester bonds, as long as they do not interfere with hybridization. Those skilled in the art will understand that, depending on the stringency of the hybridization conditions, probes may bind to target sequences that lack complete complementarity with the probe sequence. Probes are preferably directly labeled with a tag, such as a chromophore, luminophore, or chromogen, or indirectly labeled with biotin to which a streptavidin complex can later be bound. The presence or absence of a target mRNA biomarker can be detected by assaying for the presence or absence of a probe.

[0077] The term "stringent assay conditions" refers to conditions that are suitable for generating binding pairs of sufficiently complementary nucleic acids, such as probe and target mRNA, to provide the desired level of specificity in an assay, but are generally unsuitable for forming binding pairs between complementary binding members that are insufficient to provide the desired specificity. The term "stringent assay conditions" generally refers to a combination of hybridization and washing conditions.

[0078] As used herein, the term "label" or "detectable moiety" with respect to a nucleic acid refers to a composition which renders the nucleic acid detectable when linked thereto, e.g., by spectroscopic, photochemical, biochemical, immunochemical, or chemical means. Exemplary labels include, but are not limited to, radioisotopes, magnetic beads, metal beads, colloidal particles, fluorescent dyes, enzymes, biotin, digoxigenin, haptens, etc. A "labeled nucleic acid or oligonucleotide probe" is generally one that is covalently bound to the label via a linker or chemical bond, or non-covalently bound to the label via ionic bonds, van der Waals forces, electrostatic attraction, hydrophobic interactions or hydrogen bonds, and the presence of the nucleic acid or probe can be detected by detecting the presence of the label bound to the nucleic acid or probe.

[0079] As used herein, the term "polymerase chain reaction" or "PCR" generally refers to a procedure for amplifying small amounts of nucleic acids, RNA and / or DNA, as described, for example, in U.S. Patent No. 4,683,195. Generally, sequence information at or beyond the ends of the region of interest needs to be available so that oligonucleotide primers can be designed, and these primers will have the same or similar sequences as the opposite strands of the template to be amplified. The 5' terminal nucleotides of the two primers may coincide with the ends of the amplified material. PCR can be used to amplify specific RNA sequences, specific DNA sequences from total genomic DNA, and cDNA transcribed from total cellular RNA, bacteriophage, or plasmid sequences, etc. See generally Mullis et al., Cold Spring Harbor Symp. Quant. Biol. 1987, 51:263-273; PCR Technology (Stockton Press, NY, Erlich, ed., 1989).

[0080] The term "number of cycles" or "C T " as used herein in the context of the PCR method refers to the number of PCR cycles at which the fluorescence level exceeds a given set threshold level. C TThe measurement can be used, for example, to estimate the mRNA level in the original sample. T The measurement is of the C of a certain nucleic acid. T to another nucleic acid C T When subtracting from, "dC T " or "C T The term "difference" is often used in relation to scores.

[0081] The terms “about” or “approximately” mean an acceptable error of a particular value as determined by those skilled in the art, and depend in part on how the value is measured or determined. In certain embodiments, the terms “about” or “approximately” mean within 1, 2, 3, or 4 standard deviations. In certain embodiments, the terms “about” or “approximately” mean within 50%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.05% of a given value or range.

[0082] It should be noted that in the event of a discrepancy between the illustrated structure and the name assigned to it, the illustrated structure will be given greater weight. Furthermore, if the stereochemistry of a structure or part of a structure is not indicated, for example, by a thick or dashed line, the structure or part of a structure should be interpreted as encompassing all of its stereoisomers.

[0083] The embodiments provided herein utilize the prior art of molecular biology, microbiology, and immunology, which is within the scope of the skill of those skilled in the art, unless otherwise specified. Such art is well described in the literature. Examples of texts particularly suitable for advice include: Sambrook et al.,Molecular Cloning: A Laboratory Manual(2d ed.1989);Glover,ed.,DNA Cloning,Volumes I and II(1985);Gait,ed.,Oligonucleotide Synthesis(1984);Hames&Higgins,eds.Nucleic Acid Hybridization(1984);Hames&Higgins,eds.Transcription and Translation (1984); Freshney, ed., Animal Cell Culture: Immobilized Cells and Enzymes (IRL Press, 1986); Immunochemical Methods in Cell and Molecular Biology (Academic Press, London); Scopes, Protein Purification: Principles and Practice (Springer Verlag, NY, 2d ed. 1987); and Weir & Blackwell, eds. Handbook of Experimental Immunology, Volumes I-IV (1986).

[0084] 5.2 Biomarkers and their use In one embodiment, a method is provided herein for determining the dose of a therapeutic compound provided herein for treating a subject having systemic lupus erythematosus (SLE) based on the expression level of IKZF3 in a sample obtained from the subject.

[0085] More specifically, in some embodiments, the method provided herein includes (a) obtaining a sample from a subject, (b) measuring the gene expression level of IKZF3 in the sample, and (c) determining the dose of a therapeutic compound based on the measurement of the gene expression level of IKZF3, wherein the therapeutic compound is of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0086] In some embodiments, the method includes determining that the dose of the therapeutic compound is 0.45 mg or more per day if a score calculated based on the gene expression level of IKZF3 in the sample is higher than a reference level.

[0087] In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the method includes determining that the dose of the therapeutic compound is 0.45 mg or more per day if the score is higher than -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments of the various methods and kits provided herein (including those described in the above and below sections), the reference gene is a housekeeping gene.

[0088] In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the baseline level.

[0089] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0090] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0091] In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of 0.45 mg or more per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.45 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.5 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.55 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.6 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.65 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.7 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.75 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.8 mg per day.

[0092] Accordingly, in some embodiments, methods for treating subjects having systemic lupus erythematosus (SLE) are provided herein, comprising administering a therapeutic compound to the subject at a dose of 0.45 mg or more per day, wherein the score calculated based on the gene expression level of IKZF3 in a sample from the subject is higher than the reference level, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0093] In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes. In some embodiments, the baseline level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0094] In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.7 mg / day.

[0095] In some embodiments, methods for treating subjects having systemic lupus erythematosus (SLE) are provided herein, comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.15 mg or less per day, wherein the subject has high type 1 IFN expression and / or gene signature, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0096] In another embodiment, a method is provided herein for determining the dose of a therapeutic compound provided herein for treating a subject having systemic lupus erythematosus (SLE) based on high type 1 IFN expression and / or gene signature. In some embodiments, type 1 IFN expression and / or gene signature is determined based on the expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from the subject.

[0097] More specifically, in some embodiments, the method includes (a) obtaining a sample from a subject, (b) measuring the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample, and (c) determining the dose of a therapeutic compound based on the measurement in step (b), wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0098] In some embodiments, the method includes determining that the dose of the therapeutic compound is 0.45 mg or more or 0.15 mg or less per day if the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level.

[0099] Type 1 INF expression / signature can be determined based on any known method in the art. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the method includes determining the dose of the therapeutic compound to be 0.45 mg or more per day, or 0.15 mg or less per day, if the score is higher than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0100] In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the baseline level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.15 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.1 mg per day if the score is higher than the baseline level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.75 mg per day if the score is higher than the baseline level. The frequency of treatment can be adjusted, for example, once a day, every other day, every three days, or once a week. For example, in some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.15 mg per day, about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week if the score is higher than a baseline level.

[0101] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0102] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0103] In some embodiments, the method includes administering a dose of the therapeutic compound to a target of 0.45 mg or more per day or 0.15 mg or less per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.45 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.5 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.55 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.6 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.65 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.7 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.75 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.8 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to a target of about 0.15 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to the target at a dose of about 0.11 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to the target at a dose of about 0.10 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to the target at a dose of about 0.08 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to the target at a dose of about 0.075 mg per day. In some embodiments, the method further includes administering a dose of the therapeutic compound to the target at a dose of about 0.06 mg per day. The frequency of treatment can be adjusted, for example, once daily, every other day, every three days, or once a week. For example, in some embodiments, the method includes administering about 0.15 mg of the therapeutic compound daily, about 0.15 mg of the therapeutic compound every other day, about 0.15 mg of the therapeutic compound every three days, or about 0.15 mg of the therapeutic compound once a week.

[0104] Accordingly, in some embodiments, methods for treating subjects having systemic lupus erythematosus (SLE) are provided herein, comprising administering a therapeutic compound to the subject at a dose of 0.45 mg or more or 0.15 mg or less per day, wherein the subject is determined to have high type 1 IFN expression and / or genetic signature, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0105] In some embodiments, a subject is determined to have high type 1 IFN expression and / or a genetic signature if the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from the subject is higher than a baseline level. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the score is higher than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0106] In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.7 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg / day. In other embodiments, the dose of the therapeutic compound is about 0.1 mg / day. In yet another embodiment, the dose of the therapeutic compound is about 0.075 mg / day. The frequency of treatment can be adjusted, for example, once daily, every other day, every three days, or once a week. For example, in some embodiments, the method includes administering about 0.15 mg of the therapeutic compound daily, about 0.15 mg of the therapeutic compound every other day, about 0.15 mg of the therapeutic compound every three days, or about 0.15 mg of the therapeutic compound once a week.

[0107] In yet another embodiment, a method is provided herein for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, comprising: (a) obtaining a sample from the subject; (b) determining the gene expression level of IKZF3; and (c) diagnosing that the subject is likely to be responsive to the therapeutic compound if the score calculated based on the IKZF3 gene expression level is higher than a reference level, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0108] In some embodiments, the method includes the steps of determining the gene expression level of IKZF3 and diagnosing that the subject is likely to be responsive to a therapeutic compound if the score calculated based on the IKZF3 gene expression level is higher than a reference level. In some embodiments, the score is the Log2 of the IKZF3 gene expression level relative to a reference gene in the sample, or the score is the Log2 of the IKZF3 gene expression level relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the reference level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0109] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0110] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0111] In some embodiments, the method further includes administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound.

[0112] In yet another embodiment, a method is provided for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, comprising: (a) obtaining a sample from a subject; (b) determining the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample; and (c) diagnosing the subject as likely to be responsive to the therapeutic compound if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level, wherein the therapeutic compound is of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0113] In some embodiments, the method includes the steps of determining the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2, and diagnosing that a subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a baseline level. In some embodiments, the score is the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the baseline level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0114] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0115] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0116] In some embodiments, the method further comprises administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound. Thus, in some embodiments, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound by the method provided herein, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0117] In yet another embodiment, a method is provided herein for determining the dose of a therapeutic compound provided herein for treating a subject having systemic lupus erythematosus (SLE) based on the expression levels of IKZF3, IFI27, IFI44, IFI44L, and RSAD2.

[0118] More specifically, in some embodiments, the method includes (a) obtaining a sample from a subject; (b) measuring (i) the gene expression level of IKZF3 and (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample; (c) determining a first score based on the gene expression level of IKZF3 and comparing the first score to a first reference level; (d) determining a second score based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 and comparing the second score to a second reference level; and (e) determining the dose of a therapeutic compound based on the first and second scores, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0119] In some embodiments, the first score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the first score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the first reference level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0120] In some embodiments, the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the second reference level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0121] In some embodiments, if a first score is higher than a first reference level and a second score is higher than a second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day or 0.15 mg or less per day. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the reference level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the reference level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.15 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.1 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to about 0.75 mg per day if the score is higher than the baseline level. In yet another embodiment, the method includes determining the dose of the therapeutic compound to about 0.15 mg every other day if the score is higher than the baseline level. In yet another embodiment, the method includes determining the dose of the therapeutic compound to about 0.15 mg every three days if the score is higher than the baseline level. In yet another embodiment, the method includes determining the dose of the therapeutic compound to about 0.15 mg once a week if the score is higher than the baseline level.

[0122] In other embodiments, if the first score is higher than the first reference level and the second score is lower than the second reference level, the dose of the therapeutic compound is determined to be 0.15 mg or less per day. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.15 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.1 mg per day if the score is higher than the reference level. In yet another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.75 mg per day if the score is higher than the reference level.

[0123] In yet another embodiment, if the first score is lower than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.45 mg per day if the score is higher than the reference level. In some embodiments, the method includes determining the dose of the therapeutic compound to be about 0.5 mg per day if the score is higher than the reference level. In another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.6 mg per day if the score is higher than the reference level. In yet another embodiment, the method includes determining the dose of the therapeutic compound to be about 0.7 mg per day if the score is higher than the reference level.

[0124] In yet another embodiment, a method for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising administering a dose of a therapeutic compound to the subject, the dose of which is determined according to a method provided herein, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0125] In some embodiments, gene expression levels are measured by determining protein levels. In other embodiments, gene expression levels are measured by determining mRNA levels. In yet another embodiment, gene expression levels are measured by determining cDNA levels.

[0126] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0127] In another context, methods are provided herein for identifying subjects having systemic lupus erythematosus (SLE) who are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects having SLE to a therapeutic compound, based on the presence of one or two copies of a single nucleotide polymorphism (SNP) of IKZF1, namely rs4917014. As shown in Section 6 below, SLE patients are more likely to respond to treatment with the compounds of the present invention if at least one copy of this SNP (rs4917014) is detected.

[0128] More specifically, in some embodiments, the method includes (a) obtaining a sample from a subject, (b) determining the presence of the IKZF1 single nucleotide polymorphism (SNP) rs4917014 in the sample, and (c) diagnosing the subject as likely to be responsive to a therapeutic compound if at least one copy of the SNP rs4917014 is detected, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. In some embodiments, the method further comprises administering an effective amount of the therapeutic compound to a subject determined to be likely to be responsive to the therapeutic compound.

[0129] In some embodiments, methods for treating subjects having systemic lupus erythematosus (SLE) are provided herein, comprising administering an effective amount of a therapeutic compound to the subject, wherein the subject is determined to be likely to respond to the therapeutic compound by the method provided herein, and the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0130] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0131] In another embodiment, the use of the therapeutic compound provided herein in a method provided herein is provided herein. In yet another embodiment, the therapeutic compound provided herein for use in a method provided herein is provided herein. In yet another embodiment, the use of the therapeutic compound for manufacturing a pharmaceutical product for therapeutic and / or prophylactic treatment provided herein is provided herein. In some embodiments, the therapeutic compound is a compound of formula I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer or racemic mixture thereof.

[0132] In some embodiments, therapeutic compounds for use in treating subjects having systemic lupus erythematosus (SLE) are provided herein, and the dose of the therapeutic compound is determined according to a method provided herein.

[0133] For example, in some embodiments, therapeutic compounds for use in treating subjects having systemic lupus erythematosus (SLE) are provided herein, the compounds being used in doses of 0.45 mg or more per day, the score calculated based on the gene expression level of IKZF3 in a sample from the subject being higher than the baseline level, and the therapeutic compound being the compound of formula I or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer or racemic mixture thereof.

[0134] In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes. In some embodiments, the baseline level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.7 mg / day.

[0135] In some embodiments, therapeutic compounds are provided herein for use in treating subjects having systemic lupus erythematosus (SLE), the therapeutic compounds being administered in doses of 0.45 mg or more per day or 0.15 mg or less per day, and the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from the subject being higher than the reference level, the therapeutic compound being a compound of formula I or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the score is higher than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.7 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg per day, and in some cases, the dose of the therapeutic compound is about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week. In some embodiments, the dose of the therapeutic compound is about 0.1 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.075 mg / day.

[0136] In some embodiments, therapeutic compounds for use in treating subjects having systemic lupus erythematosus (SLE) are provided herein, the therapeutic compounds being used in doses of 0.45 mg or more or 0.15 mg or less per day, the subjects having high type 1 IFN expression and / or genetic signature, and the therapeutic compounds being compounds of formula I or their pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers or racemic mixtures. In some embodiments, a subject is determined to have high type 1 IFN expression and / or genetic signature if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from the subject is higher than a reference level. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the score is higher than -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments, the dose of the therapeutic compound is about 0.45 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.5 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.6 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.7 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.15 mg per day, and in some cases, the dose of the therapeutic compound is about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week. In some embodiments, the dose of the therapeutic compound is about 0.1 mg / day. In some embodiments, the dose of the therapeutic compound is about 0.075 mg / day.

[0137] In some embodiments of the various methods provided herein (for example, as described above), the method includes treating, preventing and / or managing systemic lupus erythematosus (SLE) or its symptoms by administering a therapeutically or prophylactically effective amount of compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, to a patient having SLE.

[0138] In one embodiment, a method for treating, preventing and / or managing SLE or its symptoms is provided herein, comprising administering a therapeutically effective amount of (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, or a pharmaceutically acceptable salt or solvate thereof, to a patient having SLE.

[0139] In one embodiment, a method for preventing SLE or its symptoms is provided herein, comprising administering an effective amount of compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, to a patient at risk of having SLE. In one embodiment, a method for preventing SLE or its symptoms is provided herein, comprising administering an effective amount of (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, or a pharmaceutically acceptable salt or solvate thereof, to a patient at risk of having SLE.

[0140] The term "systemic lupus erythematosus" is used herein interchangeably with SLE and lupus and refers to all signs (including remission and relapse) of the disease known in the art. In SLE, abnormal hyperactivity of B lymphocytes and massive abnormal production of immunoglobulin gamma (IgG) autoantibodies play a significant role. This pathological process leads to the isolation and destruction of Ig-coated cells, fixation and cleavage of complement proteins, and the release of chemokines, vasoactive peptides, and destructive enzymes into the tissues (Hahn BH. Systemic Lupus Erythematosus. In: Kasper DL, Braunwald E, Fauci AS, Hauser SL, Longo DL, Jameson, JL, editors. In: Harrison's Principles of Internal Medicine (16th edition). New York (US): McGraw-Hill; 2005. pp. 1960-1967).

[0141] Symptoms of SLE vary from person to person and can appear and disappear. In most patients, symptoms include joint pain and swelling. The joints most frequently affected are the fingers, hands, wrists, and knees. Some patients develop arthritis. Other common symptoms include chest pain on deep breaths, fatigue, fever without other cause, general discomfort, anxiety, or a feeling of malaise, hair loss, mouth pain, swollen lymph nodes, sensitivity to sunlight, and skin rashes. A "butterfly" rash covering the cheeks and bridge of the nose affects about half of SLE patients, and in some patients, the rash worsens in sunlight, and the rash can also be widespread.

[0142] Other symptoms may depend on which part of the body is affected and may include the following: Brain and nervous system: headaches, numbness, stabbing pain, seizures, vision problems, personality changes, Gastrointestinal tract: abdominal pain, nausea, and vomiting. Heart: Abnormal heart rhythm (arrhythmia), Lungs: Hemoptysis and dyspnea, and Skin: Uneven skin color, fingers that change color when cold (Raynaud's phenomenon).

[0143] In one embodiment, only skin symptoms appear in SLE, i.e., discoid lupus. In one embodiment, SLE is cutaneously dominant.

[0144] In one embodiment, a method for treating moderate, severe, or very severe SLE is provided herein. As used herein, the term “severe SLE” refers to an SLE condition in which the patient has one or more severe or life-threatening symptoms (e.g., hemolytic anemia, extensive cardiac or pulmonary involvement, renal disease, or central nervous system involvement).

[0145] Methods for achieving one or more clinical endpoints related to SLE are further provided herein, comprising administering an effective amount of compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, to a patient in need thereof.

[0146] The following methods are further provided herein for increasing overall survival, objective response rate, progression-free survival, progression-free survival and / or treatment success in patients with SLE, comprising administering to a patient an effective amount of compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0147] The dose of Compound I or its pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures administered to a patient may be variable and subject to the judgment of the healthcare professional. The dose of Compound I or its pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures will vary depending on factors such as the specific indication or symptom to be treated, prevented, or managed; the patient's age and condition; and the amount of any second active agent used. Generally, Compound I or its pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures may be administered 1 to 4 times a day or more at doses ranging from approximately 0.005 mg / kg to approximately 10 mg / kg of the patient's body weight, but the above dose may be appropriately modified depending on the patient's age, weight and medical condition, and the type of administration. In one embodiment, the dose is approximately 0.01 mg / kg to approximately 5 mg / kg of the patient's body weight, approximately 0.05 mg / kg to approximately 1 mg / kg of the patient's body weight, approximately 0.1 mg / kg to approximately 0.75 mg / kg of the patient's body weight, or approximately 0.25 mg / kg to approximately 0.5 mg / kg of the patient's body weight.

[0148] In one embodiment, a dose is given once per day. In any given case, the amount of compound I or its pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture administered will depend on factors such as the solubility of the active ingredient, the formulation used, and the route of administration. In one embodiment, the application of a topical concentration provides intracellular exposure or concentration of about 0.01 to 10 μM.

[0149] In certain embodiments, compound I or its pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures may be used in amounts ranging from about 0.1 mg to about 1000 mg per day, and may be adjusted in a conventional manner (e.g., the same amount administered daily during a treatment, prophylaxis, or control period), in cycles (e.g., a week of administration followed by a week of rest), or in amounts that increase or decrease over the course of treatment, prophylaxis, or control. In other embodiments, doses may range from about 1 mg to about 300 mg, about 0.1 mg to about 150 mg, about 1 mg to about 200 mg, about 10 mg to about 100 mg, about 0.1 mg to about 50 mg, about 1 mg to about 50 mg, about 10 mg to about 50 mg, about 20 mg to about 30 mg, or about 1 mg to about 20 mg. In other embodiments, the dose may be approximately 0.1 mg to approximately 100 mg, approximately 0.1 mg to approximately 50 mg, approximately 0.1 mg to approximately 25 mg, approximately 0.1 mg to approximately 20 mg, approximately 0.1 mg to approximately 15 mg, approximately 0.1 mg to approximately 10 mg, approximately 0.1 mg to approximately 7.5 mg, approximately 0.1 mg to approximately 5 mg, approximately 0.1 mg to approximately 4 mg, approximately 0.1 mg to approximately 3 mg, approximately 0.1 mg to approximately 2 mg, or approximately 1 mg to approximately 1 mg.

[0150] In some embodiments, compound 1A, or a pharmaceutically acceptable salt or solvate thereof, is administered. In one embodiment, the dose of compound 1A, or a pharmaceutically acceptable salt or solvate thereof, is 0.3 mg administered every other day. In one embodiment, the dose of compound 1A, or a pharmaceutically acceptable salt or solvate thereof, is 0.3 mg administered daily. In one embodiment, the dose of compound 1A, or a pharmaceutically acceptable salt or solvate thereof, is 0.6 mg and 0.3 mg administered on alternating days. In one embodiment, the dose of compound 1A, or a pharmaceutically acceptable salt or solvate thereof, is 0.6 mg administered daily.

[0151] In some embodiments, the patient may start treatment with a high dose, and if significant adverse effects persist, the dose may be adjusted, i.e., reduced accordingly. For example, the patient may start with a daily dose of 0.6 mg of compound 1A, or a pharmaceutically acceptable salt or solvate thereof, and if significant adverse effects persist, the dose may then be adjusted stepwise to 0.6 mg and 0.3 mg on alternating days, then 0.3 mg daily, and then 0.3 mg every other day.

[0152] Compound I, or any pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, may be combined with other pharmacologically active compounds ("second activators") in the methods and compositions provided herein. Certain combinations may act synergistically in the treatment of SLE and conditions and symptoms associated with SLE. Compound I, or any pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, may also act to mitigate adverse effects associated with certain second activators, and vice versa.

[0153] The methods and compositions provided herein may use one or more second active ingredients or agents. The second activator may be a large molecule (e.g., a protein) or a small molecule (e.g., a synthetic inorganic molecule, an organometallic molecule, or an organic molecule).

[0154] In another embodiment, the therapeutic method provided herein comprises the administration of a second therapeutic agent, the second therapeutic agent being an anti-inflammatory agent, such as a steroidal anti-inflammatory drug or a non-steroidal anti-inflammatory drug (NSAID), such as acetaminophen, naproxen, ibuprofen, or acetylsalicylic acid. In a more specific embodiment in which an NSAID is administered, a proton pump inhibitor (PPI), such as omeprazole, may also be administered. In one embodiment, the anti-inflammatory agent is a corticosteroid. In another embodiment, the anti-inflammatory agent is colchicine.

[0155] In another embodiment, the second therapeutic agent is an immunomodulatory or immunosuppressive compound such as azathioprine (Imuran®, Azasan®), methotrexate (Rheumatrex®, Trexall®), penicillamine (Depen®, Cuprimine®), cyclophosphamide (Cytoxan®), mycophenalate (CellCept®, Myfortic®), bosentan (Tracleer®), prednisone (Deltasone®, Liquid Pred®), and a PDE5 inhibitor. In another embodiment, if the affected individual has finger ulcers and pulmonary hypertension, a vasodilator such as prostacyclin (iloprost) may be administered.

[0156] In another embodiment, the second therapeutic agent is an HDAC inhibitor, such as romidepsin, vorinostat, panobinostat, valproic acid, or belinostat; or a biological agent, such as interleukin, an immunomodulatory monoclonal antibody, or Bacillus calmette-Guélain (BCG).

[0157] In another embodiment, the second therapeutic agent is an ActRII receptor inhibitor or an activin-ActRII inhibitor. ActRII receptor inhibitors include ActRIIA inhibitors and ActRIIB inhibitors. An ActRII receptor inhibitor may be a polypeptide containing the activin-binding domain of ActRII. In a particular embodiment, the activin-binding domain containing the polypeptide is ligated to the Fc portion of an antibody (i.e., a conjugate is generated containing the activin-binding domain containing the ActRII receptor polypeptide and the Fc portion of the antibody). In a particular embodiment, the activin-binding domain is ligated to the Fc portion of the antibody via a linker, such as a peptide linker.

[0158] Examples of non-antibody proteins selected for activin or ActRIIA binding, and methods for their design and selection, can be found in International Publication No. 2002 / 088171, International Publication No. 2006 / 055689, International Publication No. 2002 / 032925, International Publication No. 2005 / 037989, U.S. Patent Application Publication No. 2003 / 0133939, and U.S. Patent Application Publication No. 2005 / 0238646, each of which is incorporated herein by reference in whole.

[0159] In one embodiment, the inhibitor of the ActRII receptor is ACE-11. In another embodiment, the inhibitor of the ActRII receptor is ACE-536.

[0160] In another embodiment, the second therapeutic agent is a drug that has been conventionally used to treat SLE. Examples of such drugs include, but are not limited to, NSAIDs, corticosteroids, nonbiological disease-modifying antirheumatic drugs (DMARDs), and biological DMARD therapies (e.g., belimumab and rituximab).

[0161] Any combination of the above-mentioned therapeutic agents suitable for the treatment of SLE or its symptoms may be administered. Such therapeutic agents may be administered simultaneously or as separate therapeutic processes in any combination with compound I, or its pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures.

[0162] In certain embodiments, compound I, or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof, is administered to the patient periodically. Cycling therapy involves administering the active agent for a set period, followed by a set period of drug-free administration (i.e., discontinuation of administration), and repeating this cycle. Cycling therapy can reduce the development of tolerance to one or more treatments, avoid or reduce side effects of one treatment, and / or improve the effectiveness of the treatment.

[0163] Therefore, in one embodiment, the compounds provided herein are administered daily in single or divided doses in cycles of 4 to 6 weeks with rest periods of about 1 or 2 weeks. Cycling therapy further allows for increasing the frequency, number, and length of administration cycles. Thus, another embodiment involves administering the compounds provided herein over more cycles than is typical when administered alone. In yet another embodiment, the compounds provided herein are administered for a number of cycles greater than would typically cause dose-limiting toxicity in patients who have not also received a second active ingredient.

[0164] In one embodiment, the compound provided herein is administered daily for three or four weeks at a dose of about 0.03 mg to about 10 mg / day, followed by the remainder for one or two weeks. In other embodiments, the dose may be about 0.1 mg to about 8 mg, about 0.3 mg to about 6 mg, about 1 mg to about 4 mg, or about 2 mg, after which the drug may be discontinued.

[0165] In one embodiment, the compound and the second active ingredient provided herein are administered orally, with the compound administered 30 to 60 minutes prior to the second active ingredient during a 4 to 6-week cycle. In another embodiment, the combination of the compound and the second active ingredient provided herein is administered by intravenous infusion over approximately 90 minutes per cycle.

[0166] Typically, the number of cycles a patient receives combination therapy is approximately 1 to 24 cycles, 2 to 16 cycles, or 4 to 3 cycles.

[0167] 5.3. Methods for the detection and quantification of biomarkers In certain embodiments, a method for detecting and quantifying the protein level of a biomarker from a biological sample is provided herein, comprising contacting the protein in the sample with a first antibody that immunospecifically binds to the biomarker protein. In some embodiments, the method provided herein further comprises: (i) contacting the biomarker protein bound to the first antibody with a second antibody having a detectable label, wherein the second antibody immunospecifically binds to the biomarker protein and immunospecifically binds to an epitope on the biomarker protein different from that of the first antibody; (ii) detecting the presence of the second antibody bound to the biomarker protein; and (iii) determining the amount of the biomarker protein based on the amount of detectable label in the second antibody. In other embodiments, the method provided herein further includes (i) contacting a biomarker protein bound to a first antibody with a second antibody having a detectable label, wherein the second antibody is immunospecifically bound to the first antibody; (ii) detecting the presence of the second antibody bound to the first antibody; and (iii) determining the amount of biomarker protein based on the amount of detectable label in the second antibody.

[0168] In certain embodiments, a method for detecting and quantifying the RNA (e.g., mRNA) level of a biomarker from a biological sample is provided herein, comprising: (a) obtaining RNA from the sample; (b) contacting the RNA with a primer that specifically binds to a sequence in the RNA to generate a first DNA molecule having a sequence complementary to the RNA; (c) amplifying the DNA corresponding to a segment of a gene encoding the biomarker; and (d) determining the RNA level of the biomarker based on the amount of amplified DNA.

[0169] In certain embodiments of the various methods provided herein, two or more steps are performed sequentially. In other embodiments of the methods provided herein, two or more steps are performed in parallel (for example, simultaneously).

[0170] 5.3.1 Method for determining mRNA levels in a sample Several methods for detecting or quantifying mRNA levels are known in the art. Examples include, but are not limited to, Northern blotting, ribonuclease-protected assays, and PCR-based methods. The mRNA sequences of biomarkers can be used to prepare probes that are at least partially complementary to the mRNA sequences. These probes can then be used to detect mRNA in a sample using any suitable assay, such as PCR-based methods, Northern blotting, or dipstick assays.

[0171] In other embodiments, nucleic acid assays can be prepared to test the activity of a compound in a biological sample. The assay typically comprises a solid support and at least one nucleic acid in contact with the support, the nucleic acid corresponding to at least a portion of mRNA whose expression has been altered during compound treatment in a patient, such as the mRNA of a biomarker. The assay may also have means for detecting the altered expression of mRNA in the sample.

[0172] The assay method can be varied depending on the type of mRNA information desired. Exemplary methods include, but are not limited to, Northern blotting and PCR-based methods (e.g., qRT-PCR). Methods such as qRT-PCR can also accurately quantify the amount of mRNA in a sample.

[0173] The presence of mRNA in a sample can be determined using any suitable assay platform. For example, the assay may take the form of a dipstick, membrane, tip, disk, test strip, filter, microsphere, slide, multiwell plate, or optical fiber. The assay system may have a solid support to which nucleic acid corresponding to mRNA is attached. The solid support may include, for example, plastic, silicon, metal, resin, glass, membrane, particle, precipitate, gel, polymer, sheet, sphere, polysaccharide, capillary, film, plate, or slide. The assay components can be prepared and packaged together as a kit for detecting mRNA.

[0174] Nucleic acids may be labeled to produce a population of labeled mRNA, if necessary. Generally, samples can be labeled using methods well known in the art (e.g., by using DNA ligases, terminal transferases, or by labeling the RNA backbone). See, for example, Ausubel et al., Short Protocols in Molecular Biology (Wiley & Sons, 3rd ed. 1995); Sambrook et al., Molecular Cloning: A Laboratory Manual (Cold Spring Harbor, NY, 3rd ed. 2001). In some embodiments, samples are labeled with fluorescent labels. Examples of fluorescent dyes include xanthene dyes, fluorescein dyes (e.g., fluorescein isothiocyanate (FITC), 6-carboxyfluorescein (FAM), 6-carboxy-2',4',7',4,7-hexachlorofluorescein (HEX), 6-carboxy-4',5'-dichloro-2',7'-dimethoxyfluorescein (JOE)), rhodamine dyes (e.g., rhodamine 110 (R110), N,N,N',N'-tetramethyl-6-carboxyrhodamine (TAMRA), 6-carboxy-X-rhodamine (ROX), 5-carboxyrhodamine 6G (R6G5 or G5), 6-carboxyrhodamine 6G) Examples of dyes include, but are not limited to, (R6G6 or G6), cyanine dyes (e.g., Cy3, Cy5 and Cy7), Alexa dyes (e.g., Alexa-fluor-555), coumarin, diethylaminocoumarin, umbelliferone, benzimide dyes (e.g., Hoechst33258), phenanthridine dyes (e.g., Texas Red), ethidium dyes, acridine dyes, carbazole dyes, phenoxazine dyes, porphyrin dyes, polymethine dyes, BODIPY dyes, quinoline dyes, pyrene, fluorescein chlorotriazinyl, eosin dyes, tetramethylrhodamine, lisamin, naphthofluorescein, etc.

[0175] Nucleic acids may be located at specific addressable positions on a solid support, each corresponding to at least a portion of the mRNA sequence that is differentially expressed in cells or patients during treatment with the compound.

[0176] A typical mRNA assay may include the steps of: (1) obtaining a target probe bound to a surface; (2) hybridizing an mRNA population to the surface-bound probe under conditions sufficient to provide specific binding; (3) washing after hybridization to remove nucleic acids that are not specifically bound to the surface-bound probe; and (4) detecting the hybridized mRNA. The reagents used in each of these steps and their usage conditions may vary depending on the specific application.

[0177] Hybridization can be carried out under appropriate hybridization conditions, where stringency may vary as desired. Typical conditions are sufficient to generate a probe / target complex on a solid surface between complementary binding members, i.e., between the surface-bound target probe and complementary mRNA in the sample. In certain embodiments, stringent hybridization conditions may be used.

[0178] Hybridization is typically performed under stringent hybridization conditions. Standard hybridization techniques (e.g., conditions sufficient to result in specific binding of target mRNA in the sample to the probe) are described in Kallioniemi et al., Science 1992, 258:818-821 and International Publication No. 93 / 18186. Several guides to general techniques are available, e.g., Tijssen, Hybridization with Nucleic Acid Probes, Parts I and II (Elsevier Amsterdam 1993). For a description of techniques suitable for in situ hybridization, see Gall et al., Meth. Enzymol. 1981, 21:470-480; Angerer et al., Genetic Engineering: Principles and Methods, Vol. 7, pgs 43-65 (Plenum Press, New York, Setlow and Hollaender, eds. 1985). The selection of appropriate conditions, including temperature, salt concentration, polynucleotide concentration, hybridization time, and stringency of washing conditions, depends on the experimental design, including the source of the sample, the identity of the capture agent, and the expected degree of complementarity, and can be determined as a routine experimental issue for those skilled in the art.

[0179] Those skilled in the art will readily recognize that similar stringency conditions can be provided by utilizing alternative but equivalent hybridization and washing conditions.

[0180] Following the mRNA hybridization procedure, surface-bound polynucleotides are typically washed to remove unbound nucleic acids. Washing can be performed using any convenient washing protocol, typically with stringent washing conditions, as described above. Hybridization of target mRNA with respect to the probe is then detected using standard techniques.

[0181] Other methods, such as PCR-based methods, can also be used to detect the expression of CRBN or proteins directly or indirectly affected by CRBN. Examples of PCR methods can be found in U.S. Patent No. 6,927,024, which is incorporated herein by reference in its entirety. Examples of RT-PCR methods can be found in U.S. Patent No. 7,122,799, which is incorporated herein by reference in its entirety. A method of fluorescence in situ PCR is described in U.S. Patent No. 7,186,507, which is incorporated herein by reference in its entirety.

[0182] In some embodiments, quantitative reverse transcription PCR (qRT-PCR) can be used for both the detection and quantification of RNA targets (Bustin et al., Clin. Sci. 2005, 109:365-379). Quantitative results obtained by qRT-PCR are generally more useful than qualitative data. Therefore, in some embodiments, qRT-PCR-based assays may be useful for measuring mRNA levels during cell-based assays. The qRT-PCR method is also useful for monitoring patient treatment. Examples of qRT-PCR-based methods can be found, for example, in U.S. Patent No. 7,101,663, which is incorporated in its entirety herein by reference.

[0183] In contrast to conventional reverse transcriptase-PCR and agarose gel analysis, qRT-PCR provides quantitative results. A further advantage of qRT-PCR is its relatively easy and convenient use. Instruments for qRT-PCR, such as the Applied Biosystems 7500, are commercially available, as are reagents such as TaqMan® Sequence Detection Chemistry. For example, TaqMan® gene expression assays can be used according to the manufacturer's instructions. These kits are pre-formulated gene expression assays for rapid and reliable detection and quantification of human, mouse, and rat mRNA transcripts. An exemplary qRT-PCR program is, for example, 2 minutes at 50°C, 10 minutes at 95°C, 15 seconds at 95°C for 40 cycles, followed by 1 minute at 60°C.

[0184] The number of cycles (C) over which the fluorescence signal associated with a specific amplicon accumulation crosses the threshold. T To determine (called), for example, use 7500 Real-Time PCR System Sequence Detection software or compare C T The data can be analyzed using a relative quantification calculation method. Using this method, the output is expressed as a multiplier change in expression levels. In some embodiments, the threshold level can be selected to be automatically determined by the software. In some embodiments, the threshold level is set to be above the baseline but low enough to remain within the exponential growth region of the amplification curve.

[0185] 5.3.2 Method for determining polypeptide or protein levels in a sample Several protein detection and quantization methods can be used to measure the level of biomarkers. Any suitable protein quantization method can be used. In some embodiments, antibody-based methods are used. Exemplary methods that can be used include, but are not limited to, immunoblotting (Western blotting), ELISA, immunohistochemistry, flow cytometry, cytometry bead arrays, and mass spectrometry. Several types of ELISA are commonly used, including direct ELISA, indirect ELISA, and sandwich ELISA.

[0186] 5.4. Subjects, samples, and cell types In certain embodiments, the various methods provided herein utilize samples (e.g., biological samples) from subjects or individuals (e.g., patients). Subjects may be patients, such as those with SLE. Subjects may be mammals, such as humans. Subjects may be male or female, and may be adults, children, or infants. Samples can be analyzed during the active phase of SLE or when SLE is inactive. In certain embodiments, two or more samples can be obtained from a subject.

[0187] In certain embodiments, the sample used in the methods provided herein includes bodily fluids from the subject. Non-limiting examples of bodily fluids include blood (e.g., whole blood), plasma, amniotic fluid, aqueous humor, bile, earwax, Cowper's fluid, bulbourethral gland fluid, chyle, oatmeal, female vaginal fluid, interstitial fluid, lymph, menstrual fluid, breast milk, mucus, pleural fluid, pus, saliva, sebum, semen, serum, sweat, tears, urine, vaginal lubrication, emetic fluid, water, feces, bodily fluids (including cerebrospinal fluid surrounding the brain and spinal cord), synovial fluid, intracellular fluid (fluid inside cells), and vitreous fluid (fluid in the eyeball). In some embodiments, the sample is a blood sample. Blood samples can be obtained using conventional techniques, such as those described, for example, in Innis et al, eds., PCR Protocols (Academic Press, 1990). Leukocytes can be isolated from blood samples using conventional techniques or commercially available kits, such as the RosetteSep kit (Stein Cell Technologies, Vancouver, Canada). Leukocytes, such as mononuclear cells, B cells, T cells, monocytes, granulocytes, or lymphocyte subpopulations, can be further isolated using conventional techniques, such as magnetically activated cell sorting (MACS) (Miltenyi Biotec, Auburn, California) or fluorescence-activated cell sorting (FACS) (Becton Dickinson, San Jose, California).

[0188] In one embodiment, the blood sample is approximately 0.1 mL to 10.0 mL, approximately 0.2 mL to 7 mL, approximately 0.3 mL to 5 mL, approximately 0.4 mL to 3.5 mL, or approximately 0.5 mL to 3 mL. In another embodiment, the blood sample is approximately 0.3, approximately 0.4, approximately 0.5, approximately 0.6, approximately 0.7, approximately 0.8, approximately 0.9, approximately 1.0, approximately 1.5, approximately 2.0, approximately 2.5, approximately 3.0, approximately 3.5, approximately 4.0, approximately 4.5, approximately 5.0, approximately 6.0, approximately 7.0, approximately 8.0, approximately 9.0, or approximately 10.0 mL.

[0189] In some embodiments, the specimen used in this method includes a biopsy (e.g., a tumor biopsy). The biopsy may be from any organ or tissue, such as skin, liver, lung, heart, colon, kidney, bone marrow, teeth, lymph nodes, hair, spleen, brain, breast, or other organ. Any biopsy technique known to those skilled in the art can be used to isolate the specimen from the subject, such as open biopsy, closed biopsy, core biopsy, incision biopsy, excision biopsy, or fine-needle aspiration biopsy.

[0190] In one embodiment, the sample used in the method provided herein is obtained from the subject before the subject receives treatment for a disease or disorder. In another embodiment, the sample is obtained from the subject while the subject is receiving treatment for a disease or disorder. In yet another embodiment, the sample is obtained from the subject after treatment for a disease or disorder. In various embodiments, the treatment includes administering a compound (for example, a compound provided in Section 5.5 below) to the subject.

[0191] In certain embodiments, the sample used in the method provided herein includes multiple cells. In certain embodiments, the number of cells used in the method provided herein ranges from a single cell to about 10 9 The range may be around 1 × 10⁶ cells. In some embodiments, the number of cells used in the methods provided herein is about 1 × 10⁶ cells. 4 pieces, about 5×10 4 pieces, about 1×10 5 pieces, about 5×10 5 pieces, about 1×10 6 pieces, about 5×10 6 pieces, about 1×10 7 pieces, about 5×10 7 pieces, about 1×10 8 pieces, about 5×10 8 individual or approximately 1 x 10 9 It is an individual.

[0192] The number and types of cells collected from the subject can be monitored, for example, by measuring changes in cell surface markers using standard cell detection techniques such as flow cytometry, cell sorting, immunocytochemistry (e.g., staining with tissue-specific or cell marker-specific antibodies), fluorescence-activated cell sorting (FACS), and magnetically activated cell sorting (MACS); by examining cell morphology using light or confocal microscopy; and / or by measuring changes in gene expression using techniques well known in the art, such as PCR and gene expression profiling. These techniques can also be used to identify cells that are positive for one or more specific markers.

[0193] In certain embodiments, a subset of cells is used in the methods provided herein. Methods for sorting and isolating specific populations of cells are well known in the Art and can be based on cell size, morphology, or intracellular or extracellular markers. Such methods include, but are not limited to, bead-based separation such as flow cytometry, flow sorting, FACS, and magnetic cell sorting; size-based separation (e.g., sieves, arrays of obstacles, or filters); sorting with microfluidics devices; antibody-based separation; sedimentation; affinity adsorption; affinity extraction; density gradient centrifugation; and laser capture microdissection. Fluorescence-activated cell sorting (FACS) is a well known method for separating cells from particles based on the fluorescence properties of the particles (Kamarch, Methods Enzymol. 1987, 151:150-165). Laser excitation of the fluorescent portion in individual particles results in a small charge that allows for the electromagnetic separation of positive and negative particles from a mixture. In one embodiment, cell surface marker-specific antibodies or ligands are labeled with different fluorescent labels. Cells are processed via a cell sorter, allowing for cell separation based on their ability to bind to the antibody used. FACS sorted particles can be deposited directly into individual wells of 96-well or 384-well plates to facilitate separation and cloning.

[0194] In one embodiment, RNA (e.g., mRNA) or protein is purified from the tumor, and the presence or absence of biomarkers is measured by gene or protein expression analysis. In a specific embodiment, the presence or absence of biomarkers is measured by quantitative real-time PCR (qRT-PCR), microarray, flow cytometry, or immunofluorescence. In another embodiment, the presence or absence of biomarkers is measured by ELISA or other similar methods known in the art.

[0195] 5.5 Compounds In certain embodiments, compound I for use in the methods and compositions provided herein, including combination therapies, is of the formula: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0196] In one embodiment, the compound has the following structure: (S)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, or tautomer thereof.

[0197] In one embodiment, the compound is (S)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione

[0198] In one embodiment, the compound is a pharmaceutically acceptable salt of (S)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione.

[0199] In one embodiment, the compound is (S)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione hydrochloride.

[0200] In one embodiment, the compound has the following structure: (R)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, or tautomer thereof.

[0201] In one embodiment, the compound is (R)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione.

[0202] In one embodiment, the compound is a pharmaceutically acceptable salt of (R)-3-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-piperidine-2,6-dione.

[0203] In one embodiment, the compound is 3-[4-(4-morpholine-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, 3-[4-(4-morphyoline-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride, (R)-3-[4-(4-morphyoline-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, ( Selected from (R)-3-[4-(4-morphhilin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride, (S)-3-[4-(4-morphhilin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione, and (S)-3-[4-(4-morphhilin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0204] Compound I, or any pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, tautomers, or racemic mixtures thereof, can be prepared by methods known to those skilled in the art, for example, by following the procedures described in U.S. Patent Application Publication No. 2011 / 0196150, which is incorporated herein by reference in whole.

[0205] An exemplary method for preparation is described in Example 1.

[0206] The various compounds provided herein contain one or more chiral centers and may exist as a mixture of enantiomers (e.g., a racemic mixture) or a mixture of diastereomers. The methods provided herein encompass the use of stereoisomerically pure forms of such compounds and mixtures of those forms. For example, a mixture containing equal or unequal amounts of enantiomers of a particular compound can be used in the methods provided herein. These isomers can be synthesized or resolved asymmetrically using standard techniques such as chiral columns or chiral resolving agents. See Jacques et al., Enantiomers, Racemates and Resolutions (Wiley-Interscience, New York, 1981); Wilen et al., Tetrahedron 1977, 33:2725-2736; Eliel, Stereochemistry of Carbon Compounds (McGraw-Hill, NY, 1962); and Wilen, Tables of Resolving Agents and Optical Resolutions, p.268 (Eliel, ed., Univ. of Notre Dame Press, Notre Dame, IN, 1972).

[0207] Isotope-enriched analogs of the compounds provided herein are also provided herein. Isotope enrichment (e.g., deuteration) of pharmaceuticals to improve pharmacokinetic ("PK"), pharmacodynamic ("PD"), and toxicity profiles has been previously demonstrated in several classes of drugs. For example, see Lijinsky et al., Food Cosmet. Toxicol. 20:393 (1982); Lijinsky et al., J. Nat. Cancer Inst., 69:1127 (1982); Mangold et al., Mutation Res. 308:33 (1994); Gordon et al., Drug Metab. Dispos. 15:589 (1987); Zello et al., Metabolism, 43:487 (1994); Gately et al., J., Nucl. Med. 27:388 (1986); Wade D, Chem. Biol. Interact. 117:191 (1999).

[0208] Without being limited to any particular theory, isotopic enrichment of drugs can be used, for example, to (1) reduce or eliminate undesirable metabolites, (2) increase the half-life of the parent drug, (3) reduce the number of doses required to achieve the desired effect, (4) reduce the amount of dose required to achieve the desired effect, (5) increase the formation of active metabolites, if any, and / or (6) reduce the production of harmful metabolites in specific tissues, and / or to create more effective and / or safer drugs for combination therapy, whether or not the combination therapy is intentional.

[0209] Replacing an atom with one of its isotopes often alters the rate of a chemical reaction. This phenomenon is known as the rate isotope effect ("KIE"). For example, if a CH bond is cleaved during the rate-determining step of a chemical reaction (i.e., the step with the highest transition state energy), using deuterium instead of hydrogen will decrease the reaction rate and slow down the process. This phenomenon is known as the deuterium rate isotope effect ("DKIE"). (See, for example, Foster et al., Adv. Drug Res. vol.14, pp.1-36 (1985); Kushner et al., Can. J. Physiol. Pharmacol. vol.77, pp.79-88 (1999)).

[0210] The magnitude of the DKIE can be expressed as the ratio of the rate of a given reaction in which a CH bond is broken to the rate of the same reaction in which deuterium is replaced by hydrogen. The DKIE can range from about 1 (without isotope effects) to very large numbers, such as over 50, meaning that the reaction can be slowed down by more than 50 times when deuterium is replaced by hydrogen. Without being limited by any particular theory, high DKIE values ​​can be partly attributable to a phenomenon known as tunneling, which is a result of the uncertainty principle. Tunneling occurs because the hydrogen atom has a small mass, and a proton-accompanied transition state can form in the absence of the necessary activation energy. Since deuterium has a larger mass than hydrogen, the probability of it undergoing this phenomenon is statistically much lower.

[0211] Tritium ("T") is a radioactive isotope of hydrogen used in research, fusion reactors, neutron generators, and radiopharmaceuticals. Tritium is a hydrogen atom with two neutrons in its nucleus and an atomic weight close to 3. It occurs naturally in the environment at very low concentrations and is most commonly found as T2O. Tritium decays slowly (half-life = 12.3 years), releasing low-energy beta particles that cannot penetrate the outer layers of human skin. Internal exposure is the primary hazard associated with this isotope, but it must be ingested in large quantities to pose a significant health risk. Compared to deuterium, less tritium must be consumed before it reaches dangerous levels. The substitution of hydrogen with tritium ("T") results in an even stronger bond than with deuterium, giving a numerically greater isotopic effect.

[0212] Similarly, carbon 13 C or 14 C, sulfur 33 S, 34 S or 36 S, nitrogen 15 N and oxygen 17 O or 18 Substitutions of isotopes of other elements, including but not limited to oxygen, result in similar kinetic isotopic effects.

[0213] Isotope enrichment at specific locations of the compounds provided herein may produce detectable KIEs that affect the pharmacokinetic, pharmacological, and / or toxicological profiles of the compounds provided herein compared to similar compounds with natural isotopic compositions. In one embodiment, deuterium enrichment occurs at the CH bond cleavage site during metabolism.

[0214] 5.6 Pharmaceutical Compositions Pharmaceutical compositions can be used to prepare individual single-unit dosage forms. The pharmaceutical compositions and dosage forms provided herein comprise the compounds provided herein, or their pharmaceutically acceptable salts, solvates, hydrates, stereoisomers, racemates, clathrates, or prodrugs. The pharmaceutical compositions and dosage forms may further comprise one or more excipients.

[0215] The pharmaceutical compositions and dosage forms provided herein may also contain one or more additional active ingredients. Examples of any second or additional active ingredients are disclosed above.

[0216] The single-unit dosage forms provided herein are suitable for oral, mucosal (e.g., nasal, sublingual, vaginal, buccal, or rectal), parenteral (e.g., subcutaneous, intravenous, bolus injection, intramuscular, or intra-arterial), topical (e.g., eye drops or other ophthalmic preparations), transdermal, or transdermal administration to patients. Examples of dosage forms include, but are not limited to, tablets; caplets; capsules such as soft-elastic gelatin capsules; cachets; lozenges; drops; dispersions; suppositories; powders; aerosols (e.g., nasal sprays or inhalers); gels; suspensions (e.g., aqueous or non-aqueous liquid suspensions, oil-in-water emulsions, or water-in-oil liquid emulsions), solutions, and elixirs; liquid dosage forms suitable for oral or mucosal administration to patients; liquid dosage forms suitable for parenteral administration to patients; other ophthalmic preparations suitable for eye drops or topical administration; and sterile solids (e.g., crystalline or amorphous solids) that can be reconstituted to provide liquid dosage forms suitable for parenteral administration to patients.

[0217] The composition, form, and type of dosage forms typically vary depending on their use. For example, a dosage form used for the acute treatment of a disease may contain one or more of the active ingredients in greater quantities than a dosage form used for the chronic treatment of the same disease. Similarly, a parenteral dosage form may contain one or more of the active ingredients in smaller quantities than an oral dosage form used to treat the same disease. It will be readily apparent to those skilled in the art that these and other methods of using a particular dosage form are different from one another. For example, Remington's Pharmaceutical Sciences, 20 th See ed., Mack Publishing, Easton PA (2000).

[0218] In one embodiment, the pharmaceutical composition and dosage form comprises one or more excipients. Suitable excipients are well known to those skilled in the art of pharmaceuticals, and non-limiting examples of suitable excipients are provided herein. Whether a particular excipient is suitable for incorporation into a pharmaceutical composition or dosage form depends on a variety of factors well known in the art, including, but not limited to, how the dosage form is administered to a patient. For example, an oral dosage form such as a tablet may contain an excipient that is not suitable for use in a parenteral dosage form. The suitability of a particular excipient may also depend on the particular active ingredient in the dosage form. For example, the degradation of some active ingredients may be accelerated by certain excipients, such as lactose, or by exposure to water. Active ingredients containing primary or secondary amines are particularly susceptible to such accelerated degradation. Therefore, pharmaceutical compositions and dosage forms are provided that contain, if any, little to no, other monosaccharides or disaccharides of lactose. As used herein, the term “lactose-free” means that the amount of lactose present, if present, is insufficient to substantially increase the rate of degradation of the active ingredient.

[0219] Lactose-free compositions are well known in the art and may include, for example, excipients described in the United States Pharmacopeia (USP) 25-NF20 (2002). Generally, lactose-free compositions contain pharmaceutically compatible and pharmaceutically acceptable amounts of active ingredients, binders / fillers, and lubricants. In one embodiment, a lactose-free dosage form contains an active ingredient, microcrystalline cellulose, pregelatinized starch, and magnesium stearate.

[0220] Since water can promote the decomposition of some compounds, anhydrous pharmaceutical compositions and dosage forms containing an active ingredient are also provided. For example, the addition of water (e.g., 5%) is widely accepted in the pharmaceutical field as a means to simulate long-term storage in order to determine properties such as shelf life or the stability of the formulation over time. See, for example, Jens T. Carstensen, Drug Stability: Principles & Practice, 2d. Ed., Marcel Dekker, NY, NY, 1995, pp. 379-80. In fact, water and heat can promote the decomposition of some compounds. Thus, since moisture and / or humidity are commonly encountered during the manufacture, handling, packaging, storage, transportation, and use of formulations, the effect of water on formulations can be very important.

[0221] Anhydrous pharmaceutical compositions and dosage forms can be prepared using anhydrous or low-moisture-containing ingredients and low-moisture or low-humidity conditions. Pharmaceutical compositions and dosage forms containing at least one active ingredient including lactose and a primary or secondary amine are anhydrous when substantial contact with moisture and / or humidity during manufacture, packaging, and / or storage is anticipated.

[0222] Anhydrous pharmaceutical compositions should be prepared and stored such that their anhydrous nature is maintained. Thus, in one embodiment, anhydrous compositions are packaged using materials known to prevent exposure to water so that they can be included in a suitable prescription kit. Examples of suitable packaging include, but are not limited to, sealed foil, plastic, unit dose containers (e.g., vials), blister packs, and strip packs.

[0223] Pharmaceutical compositions and dosage forms containing one or more compounds that reduce the rate of decomposition of the active ingredient are also provided. Such compounds, referred to herein as "stabilizers," include, but are not limited to, antioxidants such as ascorbic acid, pH buffers, or salt buffers.

[0224] Similar to the amount and type of excipients, the amount and specific type of active ingredient in a dosage form may vary depending on factors such as the route of administration to the patient, although this is not limited to these factors. In one embodiment, the dosage form contains an amount of about 0.10 to about 500 mg of the compound provided herein. In other embodiments, the dosage form contains an amount of about 0.1, 1, 2, 5, 7.5, 10, 12.5, 15, 17.5, 20, 25, 50, 100, 150, 200, 250, 300, 350, 400, 450, or 500 mg of the compound provided herein.

[0225] In other embodiments, the dosage form contains the second active ingredient in amounts of 1 to about 1000 mg, about 5 to about 500 mg, about 10 to about 350 mg, or about 50 to about 200 mg. Of course, the specific amount of the second active agent depends on the specific drug used, the disease or disorder being treated or managed, the amount of the compound provided herein, and any additional active agents administered concurrently to the patient.

[0226] Oral dosage form Pharmaceutical compositions suitable for oral administration can be provided in various dosage forms, including, but are not limited to, tablets (e.g., chewable tablets), caplets, capsules, and liquids (e.g., flavored syrups). Such dosage forms contain a predetermined amount of the active ingredient and can be prepared by pharmaceutical methods well known to those skilled in the art. For general information, see Remington's Pharmaceutical Sciences, 20th ed., Mack Publishing, Easton PA (2000).

[0227] The oral dosage forms provided herein are prepared by closely mixing and combining the active ingredient with at least one excipient, in accordance with conventional pharmaceutical formulation techniques. Excipients can take a wide variety of forms depending on the desired form of the preparation for administration. For example, excipients suitable for use in oral liquid or aerosol dosage forms include, but are not limited to, water, glycols, oils, alcohols, flavoring agents, preservatives, and colorants. Examples of excipients suitable for use in solid oral dosage forms (e.g., powders, tablets, capsules, and caplets) include, but are not limited to, starch, sugars, microcrystalline cellulose, diluents, granulators, lubricants, binders, and disintegrants.

[0228] In one embodiment, the oral dosage form is a tablet or capsule, in which case a solid excipient is used. In another embodiment, the tablet may be coated by standard aqueous or non-aqueous techniques. Such dosage forms can be prepared by any pharmaceutical method. Generally, pharmaceutical compositions and dosage forms are prepared by uniformly and closely mixing the active ingredient with a liquid carrier, a finely divided solid carrier, or both, and then, if necessary, shaping the product into the desired form.

[0229] For example, tablets can be prepared by compression or molding. Compressed tablets can be prepared by compressing a free-flowing active ingredient, such as a powder or granules mixed with an excipient, using appropriate machinery. Molded tablets can be prepared by molding a mixture of powder compounds moistened with an inert liquid diluent using appropriate machinery.

[0230] Examples of excipients that may be used in the oral dosage forms provided herein include, but are not limited to, binders, fillers, disintegrants, and lubricants. Suitable binders for use in pharmaceutical compositions and dosage forms include, but are not limited to, corn starch, potato starch or other starches, gelatin, natural and synthetic rubber such as acacia, sodium alginate, alginic acid, other alginates, powdered tragacanth, guar gum, cellulose and its derivatives (e.g., ethylcellulose, cellulose acetate, calcium carboxymethylcellulose, sodium carboxymethylcellulose), polyvinylpyrrolidone, methylcellulose, pregelatinized starch, hydroxypropyl methylcellulose (e.g., numbers 2208, 2906, 2910), microcrystalline cellulose, and mixtures thereof.

[0231] Suitable forms of microcrystalline cellulose include, but are not limited to, materials marketed as AVCEL-PH-101, AVCEL-PH-103, AVCEL RC-581, and AVCEL-PH-105 (available from FMC Corporation, American Viscose Division, Avicel Sales, Marcus Hook, PA) and mixtures thereof. A specific binder is a mixture of microcrystalline cellulose and sodium carboxymethylcellulose, marketed as AVCEL RC-581. Suitable anhydrous or low-moisture excipients or additives include AVCEL-PH-103 (trademark) and 1500 LM starch.

[0232] Examples of fillers suitable for use in the pharmaceutical compositions and dosage forms provided herein include, but are not limited to, talc, calcium carbonate (e.g., granules or powder), microcrystalline cellulose, powdered cellulose, dextrate, kaolin, mannitol, silicic acid, sorbitol, starch, pregelatinized starch, and mixtures thereof. In one embodiment, the binder or filler in the pharmaceutical composition is present in about 50 to about 99% by weight of the pharmaceutical composition or dosage form.

[0233] Disintegrants may be used in a composition to provide tablets that disintegrate when exposed to an aqueous environment. Tablets with too much disintegrant may disintegrate during storage, while tablets with too little may not disintegrate at the desired rate or under the desired conditions. Therefore, a solid oral dosage form can be formed using a sufficient amount of disintegrant, neither too much nor too little, that would adversely alter the release of the active ingredient. The amount of disintegrant used varies depending on the type of formulation and is readily apparent to those skilled in the art. In one embodiment, the pharmaceutical composition contains about 0.5 to about 15% by weight of disintegrant, or about 1 to about 5% by weight of disintegrant.

[0234] Disintegrants that can be used in pharmaceutical compositions and dosage forms include, but are not limited to, agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, polaritrin potassium, sodium starch glycolate, potato or tapioca starch, other starches, pregelatinized starch, other starches, clay, other algins, other celluloses, gums, and mixtures thereof.

[0235] Lubricants that can be used in pharmaceutical compositions and dosage forms include, but are not limited to, calcium stearate, magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate, talc, hydrogenated vegetable oils (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar, and mixtures thereof. Additional lubricants include, for example, siloid silica gel (AEROSIL200, manufactured by WRGrace Co. of Baltimore, MD), synthetic silica solidification aerosol (commercially available from Degussa Co. of Plano, TX), CAB-O-SIL (a pyrogenic silicon dioxide product sold by Cabot Co. of Boston, MA), and mixtures thereof. When used, lubricants may be used in amounts less than about 1 weight percent of the pharmaceutical composition or dosage form into which they are incorporated.

[0236] In one embodiment, the solid oral dosage form comprises the compounds provided herein, anhydrous lactose, microcrystalline cellulose, polyvinylpyrrolidone, stearic acid, colloidal anhydrous silica, and gelatin.

[0237] Controlled-release dosage form The active ingredients, such as compounds, provided herein can be administered by controlled release means or delivery devices well known to those skilled in the art. Examples include U.S. Patent Nos. 3,845,770, 3,916,899, 3,536,809, 3,598,123, and 4,008,719, 5,674,533, 5,059,595, 5,591,767, 5,120,548, 5,073,543, 5,639,476, 5,354,556, 5,639,480, 5,733,566, 5,739,108, and 5,891,474, each of which are incorporated herein by reference. Examples include, but are not limited to, those described in the following specifications: the Book, Specification No. 5,922,356, Specification No. 5,972,891, Specification No. 5,980,945, Specification No. 5,993,855, Specification No. 6,045,830, Specification No. 6,087,324, Specification No. 6,113,943, Specification No. 6,197,350, Specification No. 6,248,363, Specification No. 6,264,970, Specification No. 6,267,981, Specification No. 6,376,461, Specification No. 6,419,961, Specification No. 6,589,548, Specification No. 6,613,358, and Specification No. 6,699,500. Such dosage forms can be used, for example, to provide sustained or controlled release of one or more active ingredients, using hydroxypropyl methylcellulose, other polymer matrices, gels, permeable membranes, osmotic systems, multilayer coatings, microparticles, liposomes, microspheres, or combinations thereof, to provide desired release profiles in various proportions. Suitable controlled-release formulations known to those skilled in the art, including those described herein, can be readily selected for use with the active ingredients provided herein. Accordingly, the compositions provided include single-unit dosage forms suitable for oral administration, such as, but not limited to, tablets, capsules, gel caps, and caplets adapted for controlled release.

[0238] All controlled-release drugs share the common goal of improving the pharmacotherapy achieved by their uncontrolled counterparts. Ideally, the use of optimally designed controlled-release formulations in medical procedures is characterized by a minimal amount of active pharmaceutical ingredient used to cure or control a condition in the shortest possible time. Advantages of controlled-release formulations include extended drug activity, reduced administration frequency, and increased patient compliance. Furthermore, controlled-release formulations can be used to influence other properties such as the time of action onset or drug blood levels, and therefore the occurrence of side effects (e.g., adverse events).

[0239] Most controlled-release formulations are designed to initially release a rapid amount of the drug (active ingredient) to produce the desired therapeutic effect, and then gradually and continuously release other amounts of the drug to maintain this level of therapeutic or preventive effect over a longer period. To maintain this constant level of drug in the body, the drug must be released from the dosage form at a rate that replaces the amount of drug metabolized and excreted from the body. The controlled release of the active ingredient can be stimulated by a variety of conditions, including but not limited to pH, temperature, enzymes, water, or other physiological conditions or compounds.

[0240] In certain embodiments, the drug may be administered by intravenous infusion, implantable osmotic pump, transdermal patch, liposome, or other mode of administration. In one embodiment, a pump may be used (see Sefton, CRC Crit.Ref.Biomed.Eng.14:201(1987); Buchwald et al., Surgery 88:507(1980); Saudek et al., N.Engl.J.Med.321:574(1989)). In another embodiment, a polymer material may be used. In yet another embodiment, the controlled-release system may be placed on the target at an appropriate site determined by those skilled in the art, i.e., requiring only a portion of the systemic dose (see, for example, Goodson, Medical Applications of Controlled Release, vol.2, pp.115-138(1984)). Other controlled-release systems are described in a review by Langer (Science 249:1527-1533(1990)).The active ingredients can be dispersed in a solid internal matrix, such as polymethyl methacrylate, polybutyl methacrylate, plasticized or unplasticized polyvinyl chloride, plasticized nylon, plasticized polyethylene terephthalate, natural rubber, polyisoprene, polyisobutylene, polybutadiene, polyethylene, ethylene-vinyl acetate copolymer, silicone rubber, polydimethylsiloxane, silicone carbonate copolymer, hydrogels of acrylic and methacrylic acid esters, collagen, crosslinked polyvinyl alcohol, and crosslinked partially hydrolyzed polyvinyl acetate, which form an outer polymer film, for example. It is surrounded by polyethylene, polypropylene, ethylene / propylene copolymer, ethylene / ethyl acrylate copolymer, ethylene / vinyl acetate copolymer, silicone rubber, polydimethylsiloxane, neoprene rubber, chlorinated polyethylene, polyvinyl chloride, vinyl chloride copolymer with vinyl acetate, vinylidene chloride, ethylene and propylene, polyethylene terephthalate which is an ionomer, butyl rubber epichlorohydrin rubber, ethylene / vinyl alcohol copolymer, ethylene / vinyl acetate / vinyl alcohol terpolymer, and ethylene / vinyl oxyethanol copolymer, and is insoluble in body fluids. The active ingredient is then diffused through the outer polymer membrane in a release rate control step. The proportion of the active ingredient in such a parenteral composition depends largely on its specific properties and the needs of the target.

[0241] Parenteral dosage form Parenteral dosage forms can be administered to patients by a variety of routes, including but not limited to subcutaneous, intravenous (including bolus injection), intramuscular, and intra-arterial. In some embodiments, administration of parenteral dosage forms bypasses the patient's natural defenses against contaminants, and therefore, in these embodiments, parenteral dosage forms can be sterile or sterilized before administration to the patient. Examples of parenteral dosage forms include, but are not limited to, solutions ready for injection, dried products ready to be dissolved or suspended in a pharmaceutically acceptable vehicle for injection, suspensions ready for injection, and emulsions.

[0242] Suitable vehicles that can be used to provide parenteral dosage forms are well known to those skilled in the art. Examples include Water for Injection USP; aqueous vehicles such as, but not limited to, sodium chloride injection, Ringer's injection, dextrose injection, dextrose and sodium chloride injection, and lactated Ringer's injection; water-miscible vehicles such as, but not limited to, ethyl alcohol, polyethylene glycol, and polypropylene glycol; and non-aqueous vehicles such as, but not limited to, corn oil, cottonseed oil, peanut oil, sesame oil, ethyl oleate, isopropyl myristate, and benzyl benzoate, but are not limited thereto.

[0243] Compounds that increase the solubility of one or more of the active ingredients disclosed herein can also be incorporated into parenteral dosage forms. For example, cyclodextrins and their derivatives can be used to increase the solubility of the compounds provided herein. See, for example, U.S. Patent No. 5,134,127, which is incorporated herein by reference.

[0244] Topical and mucosal dosage forms The topical and mucosal dosage forms provided herein include, but are not limited to, sprays, aerosols, solutions, emulsions, suspensions, eye drops or other ophthalmic formulations, or other forms known to those skilled in the art. See, for example, Remington’s Pharmaceutical Sciences, 16 th , 18 th and 20 th eds., Mack Publishing, Easton PA (1980, 1990 and 2000); and Introduction to Pharmaceutical Dosage Forms, 4th ed. Lea & Febiger, Philadelphia (1985). Dosage forms suitable for treating mucosal tissue in the oral cavity can be formulated as mouth washes or oral gels.

[0245] Appropriate excipients (e.g., carriers and diluents) and other materials that can be used to provide the topical and mucosal dosage forms incorporated herein are well known to those skilled in the pharmaceutical art and depend on the specific tissue to which a given pharmaceutical composition or dosage form is applied. In one embodiment, excipients include, but are not limited to, water, acetone, ethanol, ethylene glycol, propylene glycol, butane-1,3-diol, isopropyl myristate, isopropyl palmitate, mineral oil, and mixtures thereof, which are non-toxic and form pharmaceutically acceptable solutions, emulsions, or gels. Humectants or water-retaining agents may also be added to pharmaceutical compositions and dosage forms. Examples of additional components are well known in the art. For example, Remington's Pharmaceutical Sciences, 16 th ,18 th and 20 th See eds., Mack Publishing, Easton PA (1980, 1990, and 2000).

[0246] The pH of a pharmaceutical composition or dosage form may also be adjusted to improve the delivery of one or more active ingredients. Delivery can also be improved by adjusting the polarity, ionic strength, or tonicity of the solvent carrier. Compounds such as stearates may also be added to the pharmaceutical composition or dosage form to alter the hydrophilicity or lipophilicity of one or more active ingredients and improve delivery. In other embodiments, stearates can act as a lipid vehicle for the formulation, as an emulsifier or surfactant, or as a delivery enhancer or penetration enhancer. In other embodiments, the properties of the resulting composition can be further adjusted using salts, solvates, hydrates, prodrugs, clathrates, or stereoisomers of the active ingredients.

[0247] 5.7 Kit In one embodiment, a kit for the method provided herein is provided herein.

[0248] In some embodiments, a kit for determining the dose of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, the kit comprising a drug for obtaining a sample from the subject and measuring the gene expression level of IKZF3 in the sample, and the therapeutic compound being a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0249] In some embodiments, the kit further includes instructions for determining a dose of the therapeutic compound of 0.45 mg or more per day if a score calculated based on the gene expression level of IKZF3 in the sample is higher than a reference level. In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0250] In another embodiment, a kit for determining the dose of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising a drug for obtaining a sample from the subject and measuring the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0251] In some embodiments, the kit further includes instructions for determining the dose of the therapeutic compound to be 0.45 mg or more per day or 0.15 mg or less per day if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level. In some embodiments, the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0252] In further embodiments, kits are provided herein for identifying subjects with systemic lupus erythematosus (SLE) that are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, comprising a drug for obtaining a sample from a subject and determining (i) the gene expression level of IKZF3 or (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0253] In some embodiments, the kit further includes instructions for diagnosing that a subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression level of IKZF3 or the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a baseline level. In some embodiments, the instructions include the steps of determining the gene expression level of IKZF3 and diagnosing that a subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression level of IKZF3 is higher than a baseline level. In some embodiments, the score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the score is the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the baseline level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0254] In some embodiments, the description includes the steps of determining the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2, and diagnosing that a subject is likely to be responsive to a therapeutic compound if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than a reference level. In some embodiments, the score is the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the Log2 of the mean of the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the reference level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0255] In yet another embodiment, a kit for determining the dose of a therapeutic compound for treating a subject having systemic lupus erythematosus (SLE) is provided herein, comprising (a) obtaining a sample from the subject, and (b) a drug for measuring (i) the gene expression level of IKZF3 and (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0256] In some embodiments, the kit further includes instructions for the steps of: determining a first score based on the gene expression level of IKZF3 and comparing the first score to a first reference level; determining a second score based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 and comparing the second score to a second reference level; and determining the dose of a therapeutic compound based on the first and second scores. In some embodiments, the first score is the Log2 of the gene expression level of IKZF3 relative to a reference gene in the sample, or the Log2 of the gene expression level of IKZF3 relative to the mean gene expression level of two or more reference genes in the sample. In some embodiments, the first reference level is -0.49. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof. In some embodiments, the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to a reference gene in the sample, or the second score is the Log2 of the mean gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the mean of two or more reference genes in the sample. In some embodiments, the second reference level is -1.38. In some embodiments, the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

[0257] In some embodiments, the description includes determining that if the first score is higher than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is 0.45 mg or more per day or 0.15 mg or less per day.

[0258] In other embodiments, the description includes determining that the dose of the therapeutic compound is 0.15 mg or less per day if the first score is higher than the first reference level and the second score is lower than the second reference level.

[0259] In yet another embodiment, the description includes determining that the dose of the therapeutic compound is 0.45 mg or more per day if the first score is lower than the first reference level and the second score is higher than the second reference level.

[0260] In yet another embodiment, a kit is provided herein for identifying subjects with systemic lupus erythematosus (SLE) that are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, comprising (a) obtaining a sample from a subject, and (b) a drug for determining the presence of the IKZF1 single nucleotide polymorphism (SNP) rs4917014 in the sample, wherein the therapeutic compound is a compound of formula I: [ka] or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof.

[0261] In some embodiments, the kit further includes instructions for diagnosing that if at least one copy of SNP rs4917014 is detected, the subject is likely to be responsive to the therapeutic compound.

[0262] In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate. In some embodiments, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione. In yet another embodiment, the compound is (S)-3-[4-(4-morphhirin-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride.

[0263] In some embodiments, the kit includes a drug for determining protein levels. In other embodiments, the kit includes a drug for determining mRNA levels. In yet another embodiment, the kit includes a drug for determining cDNA levels.

[0264] In certain embodiments, kits for detecting mRNA levels of one or more biomarkers are provided herein. In certain embodiments, the kit comprises one or more probes that specifically bind to the mRNA of one or more biomarkers. In certain embodiments, the kit further comprises a washing solution. In certain embodiments, the kit further comprises reagents for performing a hybridization assay, means for mRNA isolation or purification, means for detection, and positive and negative controls. In certain embodiments, the kit further comprises instructions for using the kit. The kits can be adapted for home use, clinical use, or research use.

[0265] In certain embodiments, kits for detecting protein levels of one or more biomarkers are provided herein. In certain embodiments, the kit includes a dipstick coated with an antibody that recognizes a protein biomarker, a washing solution, reagents for performing the assay, means for protein isolation or purification, means for detection, and positive and negative controls. In certain embodiments, the kit further includes instructions for using the kit. The kits can be adapted for home use, clinical use, or research use.

[0266] Such kits may include, for example, dipsticks, membranes, tips, disks, test strips, filters, microspheres, slides, multiwell plates, or optical fibers. The solid support of the kit may be, for example, plastic, silicon, metal, resin, glass, membrane, particles, precipitate, gel, polymer, sheet, sphere, polysaccharide, capillary, film, plate, or slide. Biological samples may be, for example, cell cultures, cell lines, tissues, organs, organelles, body fluids, blood samples, urine samples, or skin samples.

[0267] In another embodiment, the kit includes a solid support, a nucleic acid bound to the support that is complementary to at least 20, 50, 100, 200, 350, or more bases of mRNA, and means for detecting mRNA expression in a biological sample.

[0268] In certain embodiments, the pharmaceutical kit or assay kit comprises a compound or a pharmaceutical composition thereof in a container, and further comprises components for isolating RNA in one or more containers. In another specific embodiment, the pharmaceutical kit or assay kit comprises a compound or pharmaceutical composition in a container, and further comprises components for performing RT-PCR, qRT-PCR, deep sequencing, or microarray in one or more containers.

[0269] In certain embodiments, the kits provided herein utilize means for detecting biomarker expression by quantitative real-time PCR (qRT-PCR), microarrays, flow cytometry, or immunofluorescence. In other embodiments, biomarker expression is measured by ELISA-based methodologies or other similar methods known in the art.

[0270] In another specific embodiment, the pharmaceutical kit or assay kit comprises a compound or a pharmaceutical composition thereof in a container, and further comprises components for isolating proteins in one or more containers. In yet another specific embodiment, the pharmaceutical kit or assay kit comprises a compound or a pharmaceutical composition in a container, and further comprises components for performing flow cytometry or ELISA in one or more containers.

[0271] In another embodiment, a kit for measuring biomarkers is provided herein that supplies the materials necessary to measure the abundance of one or more gene products of a biomarker or a subset of biomarkers (e.g., one, two, three, four, five, or more biomarkers) provided herein. Such a kit may include materials and reagents necessary to measure RNA or protein. In some embodiments, such a kit includes a microarray, which consists of oligonucleotides and / or DNA and / or RNA fragments that hybridize to one or more gene products of a biomarker or a subset of biomarkers provided herein, or any combination thereof. In some embodiments, such a kit may include primers for PCR of either or both the RNA product or a cDNA copy of the RNA product of the biomarker or a subset of biomarkers. In some embodiments, such a kit may include primers for PCR and probes for qPCR. In some embodiments, such a kit may include multiple primers and multiple probes, some of which have different fluorophores to enable simultaneous measurement of multiple gene products of a biomarker or a subset of biomarkers provided herein. In some embodiments, such a kit may further include materials and reagents for producing cDNA from RNA. In some embodiments, such a kit may include antibodies specific to the protein products of the biomarkers or subsets of biomarkers provided herein. Such a kit may further include materials and reagents for isolating RNA and / or proteins from a biological sample. Furthermore, such a kit may include materials and reagents for synthesizing cDNA from RNA isolated from a biological sample. In some embodiments, such a kit may include a computer program product embedded in a computer-readable medium for predicting whether a patient is clinically susceptible to a compound. In some embodiments, the kit may include a computer program product embedded in a computer-readable medium along with instructions.

[0272] In some embodiments, such kits measure the expression of one or more nucleic acid products of a biomarker or a subset of biomarkers provided herein. According to this embodiment, the kit may include materials and reagents necessary to measure the expression of a specific nucleic acid product of a biomarker or a subset of biomarkers provided herein. For example, a microarray or RT-PCR kit may be constructed for a specific condition and may contain only the reagents and materials necessary to measure the level of a specific RNA transcript of a biomarker or a subset of biomarkers provided herein in order to predict whether a patient is clinically sensitive to a compound. Alternatively, in some embodiments, the kit may include materials and reagents necessary to measure the expression of a specific nucleic acid product of a gene other than a biomarker provided herein. For example, in a particular embodiment, the kit includes reagents and materials necessary to measure the expression levels of at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50 or more genes of the biomarkers provided herein, in addition to the reagents and materials necessary to measure the expression levels of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50 or more genes of the biomarkers provided herein.In other embodiments, the kit includes reagents and materials necessary to measure the expression levels of at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50 or more biomarkers provided herein, as well as 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 300, 350, 400, 450 or more genes that are not biomarkers provided herein. In a particular embodiment, the kit includes at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, or more of the genes of the biomarkers provided herein, as well as 1-10, 1-10 This invention includes reagents and materials necessary for measuring the expression levels of non-biomarker genes provided herein, in the ranges of 0, 1-150, 1-200, 1-300, 1-400, 1-500, 1-1000, 25-100, 25-200, 25-300, 25-400, 25-500, 25-1000, 100-150, 100-200, 100-300, 100-400, 100-500, 100-1000, or 500-1000.

[0273] In the case of nucleic acid microarray kits, the kit generally includes probes mounted on the surface of a solid support. In one such embodiment, the probes may be either oligonucleotides or longer probes, including probes in the range of 150 to 800 nucleotides in length. The probes may be labeled with detectable labels. In certain embodiments, the probes are specific to one or more gene products of biomarkers provided herein. A microarray kit may include instructions for performing an assay and methods for interpreting and analyzing data resulting from performing the assay. In certain embodiments, the kit may include instructions for predicting whether a patient is clinically sensitive to a compound. The kit may also include hybridization reagents and / or reagents necessary to detect the signal produced when the probe hybridizes to a target nucleic acid sequence. Generally, the materials and reagents for a microarray kit are contained in one or more containers. Each component of the kit is generally contained in its own appropriate container.

[0274] In certain embodiments, the nucleic acid microarray kit includes, in addition to reagents and materials necessary to measure the expression levels of at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50 or more biomarkers provided herein, materials and reagents necessary to measure the expression levels of genes of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50 or more biomarkers provided herein, or combinations thereof. In other embodiments, the nucleic acid microarray kit includes reagents and materials necessary to measure the expression levels of at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50 or more genes of the biomarkers provided herein, or any combination thereof, and 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 300, 350, 400, 450 or more genes that are not of the biomarkers provided herein.In another embodiment, the nucleic acid microarray kit includes at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50 or more genes of biomarkers provided herein, or any combination thereof, and The set includes reagents and materials necessary to measure the expression levels of 1-10, 1-100, 1-150, 1-200, 1-300, 1-400, 1-500, 1-1000, 25-100, 25-200, 25-300, 25-400, 25-500, 25-1000, 100-150, 100-200, 100-300, 100-400, 100-500, 100-1000, or 500-1000 genes that are not biomarkers provided herein.

[0275] For quantitative PCR, kits generally include pre-selected primers specific to a particular nucleic acid sequence. Quantitative PCR kits may also include enzymes suitable for amplifying nucleic acids (e.g., polymerases such as Taq polymerase), deoxynucleotides, and buffers necessary for the amplification reaction. Quantitative PCR kits may also include probes specific to nucleic acid sequences that are state-related or state-indicating. Probes may or may not be labeled with fluorophores. Probes may or may not be labeled with quencher molecules. In some embodiments, quantitative PCR kits also include components suitable for reverse transcription RNA, including enzymes for reverse transcription (e.g., reverse transcriptases such as AMV, MMLV), and primers, along with the deoxynucleotides and buffers necessary for the reverse transcription reaction. Each component of a quantitative PCR kit generally comes in its own suitable container. Therefore, these kits generally include separate containers suitable for each of the individual reagents, enzymes, primers, and probes. Furthermore, quantitative PCR kits may include instructions for carrying out the reaction, as well as methods for interpreting and analyzing the data resulting from the carrying out of the reaction. In certain embodiments, the kit includes instructions for predicting whether a patient is clinically sensitive to the compound.

[0276] In the case of antibody-based kits, the kit may include, for example, (1) a first antibody (which may or may not be conjugated to a solid support) that binds to the peptide, polypeptide, or protein of interest, and optionally (2) a second different antibody that binds to either the first antibody or the peptide, polypeptide, or protein and is conjugated to a detectable label (e.g., fluorescent label, radioisotope, or enzyme). In certain embodiments, the peptide, polypeptide, or protein of interest is related to or represents a condition (e.g., disease). Antibody-based kits may also include beads for performing immunoprecipitation. Each component of an antibody-based kit is generally contained in its own appropriate container. Thus, these kits generally include separate containers suitable for each antibody and reagent. Furthermore, antibody-based kits may include instructions for performing the assay, as well as methods for interpreting and analyzing data resulting from the performance of the assay. In certain embodiments, the kit includes instructions for predicting whether a patient is clinically susceptible to the compound.

[0277] In one embodiment, the kit provided herein comprises a compound provided herein, or a pharmaceutically acceptable salt, solvate, stereoisomer, isotopolog, prodrug, hydrate, cocrystal, clathrate, or polymorph thereof. The kit may further include, but is not limited to, additional activators, including those disclosed herein.

[0278] The kits provided herein may further include devices used for administering the active ingredient. Examples of such devices include, but are not limited to, syringes, infusion bags, patches, and inhalers.

[0279] The kit may further include a pharmaceutically acceptable vehicle that can be used to administer cells or blood for transplantation, as well as one or more active ingredients. For example, if the active ingredient is provided in a solid form that must be reconstituted for parenteral administration, the kit may include a sealed container of a suitable vehicle from which the active ingredient can be dissolved to form a particulate-free sterile solution suitable for parenteral administration. Examples of pharmaceutically acceptable vehicles include, but are not limited to, water for injection (USP); aqueous vehicles (such as sodium chloride injection, Ringer's solution injection, dextrose injection, dextrose and sodium chloride injection, and lactated Ringer's solution injection); water-miscible vehicles (such as ethyl alcohol, polyethylene glycol, and polypropylene glycol); and non-aqueous vehicles (such as corn oil, cottonseed oil, peanut oil, sesame oil, ethyl oleate, isopropyl myristate, and benzyl benzoate).

[0280] In certain embodiments of the methods and kits provided herein, a solid phase support is used for protein purification, sample labeling, or the performance of a solid phase assay. Examples of solid phases suitable for performing the methods disclosed herein include beads, particles, colloids, single surfaces, tubes, multi-well plates, microtiter plates, slides, membranes, gels, and electrodes. When the solid phase is a granular material (e.g., beads), it is distributed into the wells of a multi-well plate to allow parallel processing of the solid phase support in one embodiment.

[0281] For example, but not limited to, any combination of the embodiments listed above with respect to one or more reagents such as nucleic acid primers and solid supports should be noted as being intended for any of the various methods and / or kits provided herein.

[0282] Specific embodiments of the present invention are illustrated by the following non-limiting examples. [Examples]

[0283] The following embodiments are carried out using standard techniques that are well known to those skilled in the art and are commonplace unless otherwise specified. The embodiments are intended to be illustrative only.

[0284] 6.1 Example 1: Preparation of (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione hydrochloride [ka] (1) 3-hydroxy-2-methylbenzoate methyl ester [ka] 3-hydroxy-2-methylbenzoic acid (105 g, 690 mmol) was added to 800 mL of MeOH in a 2 L three-necked round-bottom flask equipped with a condenser, thermometer, and stirring rod, followed by the addition of 250 mL of MeOH. H2SO4 (10 mL, 180 mmol) was added to the solution. The reaction mixture was stirred at 62 °C for 17 hours. The solvent was removed under vacuum. The residue (200 mL) was slowly added to 600 mL of water at room temperature, forming a white solid. The suspension was stirred in an ice bath for 30 minutes and filtered. The solid was washed with 5 × 250 mL of water and dried to obtain methyl 3-hydroxy-2-methylbenzoate as a white solid (100 g, yield 87%). The compound was used in the next step without further purification: LCMS MH=167; 1 H NMR(DMSO-d6)δ 2.28(s,3H,CH3),3.80(s,3H,CH3),6.96-7.03(m,1H,Ar),7.09(t,J=7.8Hz,1H,Ar),7.14-7.24(m,1H,Ar),9.71(s,1H,OH).

[0285] (2) 3-(tert-butyl-dimethyl-silanyloxy)-2-methyl-benzoate methyl ester [ka] In a 1 L three-necked RB flask equipped with a stirring rod and thermometer, DMF (300 mL), methyl 3-hydroxy-2-methylbenzoate (90 g, 542 mmol), and imidazole (92 g, 1,354 mmol) were added. TBDMS-Cl (90 g, 596 mmol) was added to the above solution in fractions, and the internal temperature was controlled to 15-19°C for 20 minutes, after which the internal temperature dropped to less than 1°C. The ice bath was removed, and the reaction mixture was stirred at room temperature for 16 hours. The reaction mixture was added to ice water (500 mL), and the resulting solution was divided into two parts (700 mL x 2). Each part was extracted with Depositphotos (700 mL). Each organic layer was washed with cold water (350 mL) and brine (350 mL). The organic layers were combined and dried over MgSO4. The combined organic layers were concentrated to obtain 3-(tert-butyl-dimethyl-silanyloxy)-2-methyl-benzoate methyl ester as a light brown oil (160 g, crude yield 100%). The compound was used in the next step without further purification: LCMS MH=281; 1 H NMR(DMSO-d6)δ -0.21(s,6H,CH3,CH3),0.73-0.84(m,9H,CH3,CH3,CH3),2.10(s,3H,CH3),3.60(s,3 H,CH3),6.82(dd,1H,Ar),6.97(t,J=7.9Hz,1H,Ar),7.13(dd,J=1.1,7.7Hz,1H,Ar).

[0286] (3) 2-bromomethyl-3-(tert-butyl-dimethyl-silanyloxy)methyl benzoate [ka] NBS (49.8 g, 280 mmol) was added to methyl 3-(tert-butyldimethylsilyloxy)-2-methylbenzoate (78.4 g, 280 mmol) in methyl acetate (500 mL) at room temperature to obtain an orange suspension. The resulting reaction mixture was heated in an oil bath at 40°C and illuminated under reflux with a 300 wt solar bulb for 4 hours. The reaction mixture was cooled and washed with Na2SO3 solution (2 × 600 mL, 50% saturated), water (500 mL), and brine (600 mL). The organic layer was dried over MgSO4 and decolorized with charcoal. The organic layer was concentrated to obtain 2-bromomethyl-3-(tert-butyl-dimethyl-silanyloxy)-methyl benzoate as a light brown oil (96 g, crude yield 91%). The compound was used in the next step without further purification: LCMS M-Br=279; 1 H NMR(DMSO-d6)δ 0.05-0.11(m,6H,CH3,CH3),0.82(s,9H,CH3,CH3,CH3),3.65(s,3H,CH3),4.74(s,2H ,CH2),6.94(dd,J=1.3,8.1Hz,1H,Ar),7.10-7.20(m,1H,Ar),7.21-7.29(m,1H,Ar).

[0287] (4) 4-Carbamoyl methyl butyrate [ka] Methyl 4,5-diamino-5-oxopentanoate hydrochloride (70.4 g, 358 mmol) was added to a stirred solution of methyl 2-(bromomethyl)-3-(tert-butyldimethylsilyloxy)benzoate (137.5 g, 325 mmol) in acetonitrile (1100 mL) in a 2 L round-bottom flask. DIPEA (119 ml, 683 mmol) was added to the suspension over 10 minutes using an addition funnel. The suspension was stirred at room temperature for 1 hour, and the mixture was heated in an oil bath at 40°C for 23 hours. The reaction mixture was concentrated under vacuum. The residue was stirred in ether (600 mL), and a white solid precipitated. The mixture was filtered, and the solid was washed with ether (400 mL). The filtrate was washed with HCl (1 N, 200 mL), NaHCO3 (saturated, 200 mL), and brine (250 mL). The aqueous acid layer and the basic layer were kept separately. Next, the solid was further washed with ether (250 mL), and the liquid was washed with the above-mentioned acidic and basic solutions. The two organic layers were combined and concentrated under vacuum to obtain 4-[4-(tert-butyl-dimethyl-silanyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-4-carbamoyl-butyrate methyl ester as a brown oil (152 g, crude yield 115%, purity 77% by 1H NMR). The compound was used in the next step without further purification: LCMS MH=407.

[0288] (5) 4-Carbamoyl-4-(4-hydroxy-1-oxo-1,3-dihydro-isoindole-2-yl)-methyl butyrate [ka] To a stirred, cold solution of methyl 5-amino-4-(4-(tert-butyldimethylsilyloxy)-1-oxoisoindorin-2-yl)-5-oxopentanoate (152 g, 288 mmol) in DMF (500 mL) and water (55 mL), K2CO3 (19.89 g, 144 mmol) was gradually added over 5 minutes. The resulting reaction mixture was stirred at room temperature for 40 minutes. The reaction mixture was cooled in an ice bath. HCl (12 M, 23.99 ml, 288 mmol) was slowly added to the mixture. After the addition, acetonitrile (280 mL) was added to the mixture, and a solid precipitated. The mixture was stirred at room temperature for 10 minutes and filtered. The solid was washed with acetonitrile (50 mL x 4). The filtrate was concentrated under high vacuum to obtain a yellow oily substance (168 g). The oil was dissolved in acetonitrile (600 mL) and stirred at room temperature for 10 minutes. The mixture was filtered, and the solid was washed with acetonitrile (25 mL x 2). The filtrate was concentrated under high vacuum to obtain a yellow oily substance (169 g), which was added to a mixture of water (1200 mL) and ether (1000 mL). The mixture was stirred for 3 minutes, and the layers were separated. The aqueous solution was concentrated under high vacuum, and the residue was stirred in acetonitrile (160 mL). After stirring overnight, a white solid was formed. The mixture was filtered to obtain 4-carbamoyl-4-(4-hydroxy-1-oxo-1,3-dihydro-isoindole-2-yl)-methyl butyrate as a white solid (46 g, yield 54%). The filtrate was concentrated, and the residue was further crystallized in acetonitrile (60 mL) to obtain more 4-carbamoyl-4-(4-hydroxy-1-oxo-1,3-dihydro-isoindole-2-yl)-methyl butyrate as a white solid (11.7 g, yield 14%). The filtrate was concentrated, and the residue was purified by ISCO chromatography to obtain a larger amount of methyl 4-carbamoyl-4-(4-hydroxy-1-oxo-1,3-dihydro-isoindole-2-yl)-butyrate as a white solid (13.2 g, 15% yield). The total product obtained was 70.9 g in 83% yield: LCMS MH=293; 1H NMR(DMSO-d6)δ 1.95-2.34(m,4H,CH2,CH2),3.51(s,3H,CH3),4.32(d,J=17.6Hz,1H,CHH),4.49(d,J=17.4Hz,1H,CHH),4.73(dd,J=4.7,10.2Hz ,1H,CHH),6.99(dd,J=0.8,7.9Hz,1H,Ar),7.10-7.23(m,2H,Ar,NHH),7.25-7.38(m,1H,Ar),7.58(s,1H,NHH),10.04(s,1H,OH).

[0289] (6)3-(4-((4-(morpholinomethyl)benzyl)oxy)-1-oxoisoindorin-2-yl)piperidine-2,6-dione [ka] Step 1: Triphenylphosphine (polymer-supported 1.6 mmol / g, 12 g, 18.8 mmol) was added to a solution of 3-(4-hydroxy-1-oxo-1,3-dihydro-isoindole-2-yl)-piperidine-2,6-dione (2.5 g, 8.56 mmol) in THF (60 mL). The mixture was stirred at room temperature for 15 minutes. Diisopropyl azodicarboxylate (3.96 mL, 18.8 mmol) was added at 0°C, and the mixture was stirred at 0°C for 30 minutes. (4-morpholine-4-ylmethylphenyl)-methanol (2.62 g, 12.4 mmol) was added at 0°C, and the mixture was warmed to room temperature and stirred overnight at room temperature. The reaction mixture was filtered, and the filtrate was concentrated. The resulting oily substance was purified by eluting with methylene chloride and methanol (gradient; the product appeared at 6% methanol) using a silica gel column to obtain 4-carbamoyl-4-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-methyl butyrate (2.2 g, yield 54%). The product was used in the next step without further purification.

[0290] Step 2: Potassium tert-butoxide (0.51 g, 4.57 mmol) was added at 0°C to a THF solution (50 mL) of 4-carbamoyl-4-[4-(4-morpholine-4-ylmethyl-benzyloxy)-1-oxo-1,3-dihydro-isoindole-2-yl]-methyl butyrate (2.2 g, 4.57 mmol). The mixture was stirred at 0°C for 10 minutes and quenched with 1N HCl (5 mL, 5 mmol), followed by saturated NaHCO3 (25 mL). The mixture was extracted with ethyl acetate (2 × 50 mL). The organic layer was washed with water (30 mL) and brine (30 mL), dried over MgSO4, and concentrated. To the obtained solid, ethyl acetate (10 mL), followed by hexane (10 mL), was added under stirring. The suspension was filtered to obtain 3-(4-((4-(morpholinomethyl)benzyl)oxy)-1-oxoisoindorin-2-yl)piperidine-2,6-dione as a white solid (1.5 g, 73% yield). HPLC: Waters Symmetry C 18 , 5μm, 3.9×150mm, 1mL / min, 240nm, gradient to 95 / 5 acetonitrile / 0.1%H3PO4 in 5 minutes: t R =4.78min(97.5%);mp:210~212℃; 1 1H NMR (DMSO-d6)δ 1.86-2.09(m,1H,CHH),2.29-2.38(m,4H,CH2,CH2),2.44(dd,J=4.3,13.0Hz,1H ,CHH),2.53-2.64(m,1H,CHH),2.82-2.99(m,1H,CHH),3.46(s,2H,CH2),3.52-3 .61(m,4H,CH2,CH2),4.18-4.51(m,2H,CH2),5.11(dd,J=5.0,13.3Hz,1H,NCH), 5.22(s,2H,CH2),7.27-7.38(m,5H,Ar),7.40-7.53(m,3H,Ar),10.98(s,1H,NH) 13C NMR(DMSO-d6)δ 22.36,31.21,45.09,51.58,53.14,62.10,66.17,69.41,114.97,115.23,127.64,128.99 ,129.81,129.95,133.31,135.29,137.68,153.50,168.01,170.98,172.83;LCMS:465;Analysis value C 25 H 27 Calculated values ​​for N3O5 + 0.86H2O: C, 64.58; H, 6.23; N, 9.04; Measured values: C, 64.77; H, 6.24; N, 8.88.

[0291] (S)-3-(4-((4-(morpholinomethyl)benzyl)oxy)-1-oxoisoindorin-2-yl)piperidine-2,6-dione and (R)-3-(4-((4-(morpholinomethyl)benzyl)oxy)-1-oxoisoindorin-2-yl)piperidine-2,6-dione were prepared from 3-(4-((4-(morpholinomethyl)benzyl)oxy)-1-oxoisoindorin-2-yl)piperidine-2,6-dione by chiral separation.

[0292] 6.2 Example 2: Patient subsets enriched with biomarkers The pharmacokinetics, pharmacodynamics (PD), efficacy, and safety of oral iverdimide ((S)-3-[4-({4-[(morpholine-4-yl)methyl]phenyl}methoxy)-1-oxo-1,3-dihydro-2H-isoindole-2-yl]piperidine-2,6-dione-hydrogen chloride (1 / 1)) were evaluated in a Phase 2b trial in subjects with active autoantibody-positive SLE.

[0293] Adult SLE patients (N=288) with a history of 6 months or longer and a SLE disease activity index (SLEDAI 2K) ≥6 were randomized to placebo (n=83), iverdimide 0.15 mg QD (n=42), 0.3 mg QD (n=82), or 0.45 mg QD (n=81). Clinical response was determined for all patients at week 24 by the SLE Responder Index 4 (SRI-4) within a pre-specified biomarker enrichment subset, based on the expression of Ikaros, Aiolos, type 1 IFN signatures (IFI27, IFI44, IFI44L, RSAD2), and other gene modules (Modular Immune Profile Test, DxTerity Diagnostics, Inc.) measured from fingertip blood samples taken at baseline. The primary objective of this study was to evaluate the clinical efficacy of three doses of iverdide (0.45 mg once daily [QD], 0.30 mg once daily [QD], or 0.15 mg once daily [QD]) compared to placebo for the treatment of active SLE, using the SLE Responder Index at week 24.

[0294] Among all subjects with baseline gene expression data (N=287), 179 subjects (62.4%) had type 1 IFN-high levels, and 110 subjects (38.3%) had Aiolos-high levels. These patient subsets partially overlapped. Type 1 IFN-high levels comprised 39.1% of the Aiolos-high subset, and the Aiolos-high subset comprised 63.6% of the IFN-high subset. 高 Aileolos 低 The group included 109 individuals (39.9%), and IFN 低 Aileolos 高 Forty individuals (13.9%) were included in the group. IFN 高 Aileolos 高 Seventy subjects (24.4%) fit the "double positive" group. Conversely, the "double negative" group (IFN) 低 Aileolos 低 A subgroup of 68 subjects (23.7%) was excluded (Figure 1).

[0295] Therapeutic efficacy in subgroups based on baseline mRNA and DNA. Subgroup analyses of SRI responses based on baseline mRNA and DNA expression (4) showed larger effect sizes (with a suggested trend) compared to the overall group for high IKZF3 (0.45 mg QD iverdide), high type 1 IFN (0.15 mg and 0.45 mg QD iverdide), and high Ikaros IFN module (0.45 mg QD iverdide) (Table 1).

[0296] [Table 1]

[0297] [Table 2]

[0298] The best response rates were observed in the type 1 IFN-high subset and the Aiolos-high subset. The results for Ikaros in the type 1 IFN-high subset were very similar to those in the type 1 IFN-high subset, due to the essentially identical patient populations identified by these two gene modules.

[0299] As shown in Figure 2, high-dose iverdide (0.45 mg) had a significant SRI-4 therapeutic effect compared to placebo in patients within the Aiolos high subset at baseline (32.9%; P=0.011). Low-dose (0.15 mg) or medium-dose (0.3 mg) iverdide did not show significant efficacy in the Aiolos high subset.

[0300] In the baseline type 1 IFN-high subset, the low dose of 0.15 mg demonstrated a significant 25.6% SRI-4 therapeutic effect compared to placebo (P=0.032), while the high dose of 0.45 mg demonstrated a significant 26.8% SRI-4 therapeutic effect compared to placebo (P=0.006) (Figure 2).

[0301] In addition, as shown in Figure 3, the low dose of 0.15 mg and the high dose of 0.45 mg are IFN 高 Aileolos 高 It was effective in the double-positive population, with a similar SRI-4 treatment effect compared to placebo in 29.8% of cases. IFN 高 Aileolos 低 In the group, the low dose of 0.15 mg showed a 26.8% SRI-4 therapeutic effect compared to placebo, which was slightly better than the high dose of 0.45 mg. Conversely, IFN 低 Aileolos 高 Within the subset, only the high dose of 0.45 mg was clinically effective compared to placebo, with a 35.0% SRI-4 treatment efficacy. Finally, double-negative IFN 低 Aileolos 低 In the population, iverdome did not have clinical efficacy (Figure 3).

[0302] Due to the unimodal distribution of Aiolos gene expression, the optimal cutoff for Aiolos(IKZF3) gene expression was investigated using the prevalence vs. cutoff analysis shown in Figure 4. At a pre-specified cutoff value of -0.49, the estimated response rate was 60–65%, consistent with the observed results. Using a higher Aiolos cutoff of -0.1 could achieve a higher response rate of 80–85%, but the prevalence of such a subset was small, representing only 10% of the study population. Given the desired minimum patient subset size of 35% or more, the pre-specified Aiolos cutoff value of -0.49 is near optimal (Figure 4).

[0303] Improvement of 50% or more from baseline in the CLASI activity score. CLASI activity scores at week 24 were analyzed. Improvements of more than 50% from baseline were observed in CLASI activity scores at week 24. The results for the overall population at each time point, as well as for subgroups based on baseline characteristics and mRNA and DNA expression, are described below.

[0304] The distinction between the 0.45 mg QD iverdide treatment group and the placebo treatment group was evident at week 8 (stratified difference: 14.3%; 95% CI: -0.17, 28.07; p=0.055), week 12 (stratified difference: 12.2%; 95% CI: -2.97, 26.66; p=0.118), week 16 (stratified difference: 18.2%; 95% CI: 2.94, 32.43; p=0.020), and week 20 (stratified difference: 12.3%; 95% CI: -2.91, 26.80; p=0.117) in the ITT population using NRI. At these points in time, the stratified difference from placebo ranged from 0.4% to 6.9% in the 0.30 mg QD iverdide treatment group and from -0.1% to 4.0% in the 0.15 mg QD iverdide treatment group.

[0305] Among subjects with a baseline CLASI activity score ≥ 8 (81 subjects in total), the stratified difference from placebo in the percentage of subjects with a CLASI activity score improvement of 50% or more from baseline was in the 0.45 mg QD iverdide treatment group from 9.4% (week 16) to 32.0% (week 4; 95% CI: 2.05, 54.92; p=0.043), in the 0.30 mg QD iverdide treatment group from 3.1% (week 16) to 23.6% (week 8), and in the 0.15 mg QD iverdide treatment group from 10.6% (week 16) to 35.3% (week 20; 95% CI: 1.06, 61.54; p=0.056).

[0306] Subgroup analyses of CLASI improvement due to mRNA and DNA expression were performed at week 24 in subjects with a baseline CLASI activity score of ≥8. The results regarding the proportion of subjects with a CLASI activity score of 50% or more from baseline can be summarized as follows. IKZF3 High: Compared to 77.8% (7 / 9 subjects) for placebo, the rates for 0.45 mg, 0.30 mg, and 0.15 mg of QD iverdide were 77.8% (7 / 9 subjects), 33.3% (3 / 9 subjects), and 50.0% (2 / 4 subjects), respectively. • Type 1 IFN module performance: 61.9% (13 / 21 subjects), 52.4% (11 / 21 subjects), and 66.7% (6 / 9 subjects), compared to 46.7% (7 / 15 subjects) for placebo. • Ikaros IFN module performance: 63.6% (14 / 22 subjects), 52.4% (11 / 21 subjects), and 54.5% (6 / 11 subjects), compared to 46.7% (7 / 15 subjects) for placebo.

[0307] BILAG-based composite Lupus evaluation Among the overall population with a baseline BILAG2004 score of 1A or 2B, the proportion of subjects achieving a BICLA response at week 24, based on NRI analysis in the ITT population, was 37.3% in the 0.45 mg QD iverdide group (stratified difference from placebo: 1.0%), 33.3% in the 0.30 mg QD iverdide group (stratified difference from placebo: -3.6%), 37.1% in the 0.15 mg QD iverdide group (stratified difference from placebo: 0.3%), and 36.9% in the placebo group. The treatment comparison for BICLA with placebo (33.3%) was greater in the subgroup of subjects with higher baseline IKZF3, but only for 0.45 mg QD iverdide (52.0%) and 0.15 mg QD iverdide (41.7%).

[0308] Lupus low disease activity Of all subjects, the percentage of subjects achieving an LLDAS response at week 24 was 19.8% in the 0.45 mg QD iverdide group (stratified difference from placebo: 6.9%; 95% CI: -4.82, 18.56; p=0.221), 17.1% in the 0.30 mg QD iverdide group (stratified difference from placebo: -6.45, 16.49; p=0.357), and 19.0% in the 0.15 mg QD iverdide group (stratified difference from placebo: 6.7%; 95% CI: -6.51, 22.32; p=0.311). In the placebo group based on NRI analysis of the ITT population, the percentage was 13.3%. The percentages of LLDAS responders for key mRNA and DNA subgroups were as follows: IKZF1 elevated: 23.4%, 17.0%, and 14.3% for iverdide 0.45 mg, 0.30 mg, and 0.15 mg QDs, compared to 12.5% ​​for placebo. IKZF3 high: 27.8%, 9.4%, and 14.3%, respectively, compared to 11.1% in the placebo group. • Type 1 IFN module height: 17.5%, 14.3%, and 24.0%, respectively, compared to 8.3% for placebo. • Ikaros IFN module high: 16.1%, 14.3%, and 20.0%, respectively, compared to 8.2% for placebo.

[0309] Relationship between drug dose, drug concentration, and response PK evaluation was performed. The exposure-response analysis included subjects from trial SLE-002, and these subjects were evaluated using the final population PK model and AUC from efficacy data. ss It possessed both of the post-hoc estimated iverdomid PK metrics.

[0310] Logistic regression analysis of binomial response SRIs (4) (non-responder or responder) showed that iverdimide AUC in the subjects across the dose range of 0.15 mg to 0.45 mg QD. ss The relationship was not shown (p=0.927).

[0311] The effect of iverdide exposure on SRI(4) response was further evaluated by multivariate logistic modeling. Categorical variables of baseline oral OCS (criterion: yes), baseline antimalarial drug use (criterion: yes), type I IFN, and IKZF3(Aiolos) signature (criterion: low) were tested by regression analysis. Subjects with a high IFKZ3(Aiolos) genetic signature were associated with a better response (p=0.033) (Table 2). Other subject characteristics (baseline use of oral corticosteroids or antimalarial drugs and high IFN1 genetic signature) were not correlated with the response associated with increased exposure to iverdide.

[0312] [Table 3]

[0313] Conclusion on effectiveness This study met the primary endpoint, which is the proportion of subjects achieving an SRI(4) response at week 24. The percentage of subjects achieving an SRI(4) response at week 24 was 54.3% in the 0.45 mg QD iverdamide group compared to 34.9% in the placebo group, with a stratified difference of 19.4% from placebo (95% CI: 4.12, 33.42; p=0.011). The stratified differences from placebo for the 0.30 mg QD group and the 0.15 mg QD group were 5.0% and 11.4%, respectively. OC analysis supported a linear dose-response relationship, and other sensitivity analyses supported the findings in the primary analysis. Subgroup analyses showed that the placebo response rate for the primary efficacy endpoint was higher (48.6%) in the South American region, including Mexico, which accounted for 42.7% of enrollments (34.9%) than in the overall population. Analysis of biomarker subgroups showed a greater efficacy of iverdide treatment in subjects with elevated baseline IKZF3, type 1 IFN, and Ikaros IFN compared to the overall population (stratified differences from placebo in the 0.45 mg QD iverdide treatment group at week 24 for the ITT population were 32.9%, 26.8%, and 24.3%, respectively).

[0314] Pharmacogenetic evaluation Clinical efficacy (SRI[4] response) was evaluated between subjects according to single nucleotide polymorphisms (SNPs) at the Ikaros (IKZF1) locus rs4917014 (0 copies vs. 1 or 2 copies). The SNP rs4917014 is as described in Westra et al, Nat Genet. 45(10):1238-1243 (2013). Specifically, the SNP rs4917014 is located at chr7:50266267 (GRCh38.p12) (allele: T>C / T>G). The SNP rs4917014 is associated with SLE and affects IKZF1 expression. For example, the mutant rs4917014*T allele strongly increases the 3'-UTR expression level of IKZF1 and decreases C1QB expression, which are two features found in SLE.

[0315] Among subjects with zero copies of the IKZF1 rs4917014 protective minor allele (G), the SRI(4) stratification difference between the 0.45 mg QD iverdide treatment group and placebo was 6.1% (95% CI: -17.04 to 28.38, p=0.618). Among subjects with one or two copies of the IKZF1 rs4917014 protective minor allele (G), the SRI(4) stratification difference between the 0.45 mg QD iverdide treatment group and placebo was 21.3% (95% CI: -0.94 to 41.03, p=0.047). Therefore, the clinical efficacy of 0.45 mg QD iverdide was observed only among subjects with one or two copies of the IKZF1 rs4917014 protective minor allele (G).

[0316] Overall conclusion Findings from the 24-week placebo-controlled phase of this study suggest that iverdimide may be an effective treatment for active SLE, particularly in patients with high expression of IKZF3 or type 1 IFN.

[0317] From the foregoing, it will be understood that while certain embodiments are described herein for illustrative purposes, various modifications can be made without departing from the spirit and scope of what is provided herein. All references mentioned above are incorporated herein by reference in their entirety.

Claims

1. A method for determining the dose of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE), (a) A step of obtaining a sample from the subject, (b) A step of measuring the gene expression level of IKZF3 in the sample, (c) If the score calculated based on the gene expression level of IKZF3 in the sample is higher than the reference level, the step of determining the dose of the therapeutic compound to be 0.45 mg or more per day, Includes, The therapeutic compound is a compound of formula I: 【Chemistry 1】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for determining the dosage of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE).

2. The method according to claim 1, wherein the score is Log2 of the gene expression level of IKZF3 relative to the reference gene in the sample, or the score is Log2 of the gene expression level of IKZF3 relative to the average gene expression level of two or more reference genes in the sample.

3. The method according to claim 2, comprising determining that the dose of the therapeutic compound is 0.45 mg or more per day if the score is higher than -0.

49.

4. The method according to claim 2 or 3, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH, and combinations thereof.

5. The method according to any one of claims 1 to 4, comprising determining the dose of the therapeutic compound to about 0.45 mg per day if the score is higher than the reference level.

6. The method according to any one of claims 1 to 4, comprising determining the dose of the therapeutic compound to about 0.5 mg per day if the score is higher than the reference level.

7. The method according to any one of claims 1 to 4, comprising determining the dose of the therapeutic compound to about 0.6 mg per day if the score is higher than the reference level.

8. The method according to any one of claims 1 to 4, comprising determining the dose of the therapeutic compound to about 0.7 mg per day if the score is higher than the reference level.

9. The method according to any one of claims 1 to 8, further comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more per day.

10. A method for treating a subject having systemic lupus erythematosus (SLE), comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more per day, The score calculated based on the gene expression level of IKZF3 in the sample from the aforementioned subject was higher than the reference level. The therapeutic compound is a compound of formula I: 【Chemistry 2】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

11. The method according to claim 10, wherein the score is Log2 of the gene expression level of IKZF3 relative to a reference gene, or the score is Log2 of the gene expression level of IKZF3 relative to the average gene expression level of two or more reference genes.

12. The method according to claim 11, wherein the reference level is -0.

49.

13. The method according to claim 11 or 12, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

14. The method according to any one of claims 10 to 13, wherein the dose of the therapeutic compound is about 0.45 mg / day.

15. The method according to any one of claims 10 to 13, wherein the dose of the therapeutic compound is about 0.5 mg / day.

16. The method according to any one of claims 10 to 13, wherein the dose of the therapeutic compound is about 0.6 mg / day.

17. The method according to any one of claims 10 to 13, wherein the dose of the therapeutic compound is about 0.7 mg / day.

18. A method for determining the dose of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE), (a) A step of obtaining a sample from the subject, (b) A step of measuring the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the sample, (c) If the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than the reference level, the step of determining the dose of the therapeutic compound to be 0.45 mg or more per day or 0.15 mg or less per day, Includes, The therapeutic compound is a compound of formula I: 【Transformation 3】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for determining the dosage of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE).

19. The method according to claim 18, wherein the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the average of two or more reference genes in the sample.

20. The method according to claim 19, comprising determining the dose of the therapeutic compound to be 0.45 mg or more per day, or 0.15 mg or less per day, if the score is higher than -1.

38.

21. The method according to claim 19 or 20, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

22. The method according to any one of claims 18 to 21, comprising determining the dose of the therapeutic compound to about 0.45 mg per day if the score is higher than the reference level.

23. The method according to any one of claims 18 to 21, comprising determining the dose of the therapeutic compound to about 0.5, 0.6, or 0.7 mg per day if the score is higher than the reference level.

24. The method according to any one of claims 18 to 21, comprising determining the dose of the therapeutic compound to about 0.15 mg per day if the score is higher than the reference level, wherein the dose of the therapeutic compound is optionally about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week.

25. The method according to any one of claims 18 to 21, comprising determining the dose of the therapeutic compound to about 0.1 mg per day if the score is higher than the reference level.

26. The method according to any one of claims 18 to 25, further comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.75 mg or less per day.

27. A method for treating a subject having systemic lupus erythematosus (SLE), comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.15 mg or less per day, The scores calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in the samples from the aforementioned subjects were higher than the reference level. The therapeutic compound is a compound of formula I: 【Chemistry 4】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

28. The method according to claim 27, wherein the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the average of two or more reference genes in the sample.

29. The method according to claim 28, wherein the score is higher than -1.

38.

30. The method according to claim 28 or 29, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

31. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.45 mg / day.

32. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.5 mg / day.

33. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.6 mg / day.

34. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.7 mg / day.

35. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.15 mg per day, and optionally the dose of the therapeutic compound is about 0.15 mg every other day, about 0.15 mg every three days, or about 0.15 mg once a week.

36. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.1 mg / day.

37. The method according to any one of claims 27 to 30, wherein the dose of the therapeutic compound is about 0.075 mg / day.

38. A method for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, (a) A step of obtaining a sample from the subject, (b) A step of determining the gene expression level of (i) IKZF3 or (ii) IFI27, IFI44, IFI44L, and RSAD2 in the sample, (c) If the score calculated based on the gene expression level of IKZF3 or the gene expression levels of IFI27, IFI44, IFI44L and RSAD2 is higher than the reference level, the step of diagnosing that the subject is likely to be responsive to the therapeutic compound, Includes, The therapeutic compound is a compound of formula I: 【Transformation 5】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to therapeutic compounds, or for predicting the responsiveness of subjects with SLE to therapeutic compounds.

39. The method according to claim 38, comprising the steps of determining the gene expression level of IKZF3, and diagnosing that the subject is likely to be responsive to the therapeutic compound if the score calculated based on the gene expression level of IKZF3 is higher than the reference level.

40. The method according to claim 39, wherein the score is Log2 of the gene expression level of IKZF3 relative to the reference gene in the sample, or the score is Log2 of the gene expression level of IKZF3 relative to the average gene expression level of two or more reference genes in the sample.

41. The method according to claim 40, wherein the reference level is -0.

49.

42. The method according to claim 40 or 41, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

43. The method according to claim 38, comprising the steps of: determining the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2; and diagnosing that the subject is likely to be responsive to the therapeutic compound if the score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 is higher than the reference level.

44. The method according to claim 43, wherein the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the average of two or more reference genes in the sample.

45. The method according to claim 44, wherein the reference level is -1.

38.

46. The method according to claim 44 or 45, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

47. The method according to any one of claims 38 to 46, further comprising administering an effective amount of the therapeutic compound to a subject that has been determined to be highly likely to be responsive to the therapeutic compound.

48. A method for treating a subject having systemic lupus erythematosus (SLE), comprising administering an effective amount of a therapeutic compound to the subject, The subject is determined to be highly likely to be responsive to the therapeutic compound according to any one of claims 38 to 46. The therapeutic compound is a compound of formula I: 【Transformation 6】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

49. A method for determining the dose of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE), (a) A step of obtaining a sample from the subject, (b) (i) a step of measuring the gene expression level of IKZF3 in the sample and (ii) the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2, (c) A step of determining a first score based on the gene expression level of IKZF3 and comparing the first score with a first reference level, (d) A step of determining a second score based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2, and comparing the second score with a second reference level, (e) A step of determining the dose of the therapeutic compound based on the first score and the second score, Includes, The therapeutic compound is a compound of formula I: 【Transformation 7】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for determining the dosage of a therapeutic compound for treating a subject with systemic lupus erythematosus (SLE).

50. The method according to claim 49, wherein the first score is Log2 of the gene expression level of IKZF3 relative to the reference gene in the sample, or the first score is Log2 of the gene expression level of IKZF3 relative to the average gene expression level of two or more reference genes in the sample.

51. The method according to claim 50, wherein the first reference level is -0.

49.

52. The method according to claim 50 or 51, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

53. The method according to any one of claims 49 to 52, wherein the second score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the second score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the average of two or more reference genes in the sample.

54. The method according to claim 53, wherein the second reference level is -1.

38.

55. The method according to claim 53 or 54, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

56. The method according to any one of claims 49 to 55, wherein if the first score is higher than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day or 0.15 mg or less per day.

57. The method according to any one of claims 49 to 55, wherein if the first score is higher than the first reference level and the second score is lower than the second reference level, the dose of the therapeutic compound is determined to be 0.15 mg or less per day.

58. The method according to any one of claims 49 to 55, wherein if the first score is lower than the first reference level and the second score is higher than the second reference level, the dose of the therapeutic compound is determined to be 0.45 mg or more per day.

59. A method for treating a subject with systemic lupus erythematosus (SLE), This includes administering a dose of the therapeutic compound to the subject, The dose of the therapeutic compound is determined according to the method described in any one of claims 49 to 58. The therapeutic compound is a compound of formula I: 【Transformation 8】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

60. The method according to any one of claims 1 to 59, wherein the gene expression level is measured by determining the protein level.

61. The method according to any one of claims 1 to 59, wherein the gene expression level is measured by determining the mRNA level.

62. The method according to any one of claims 1 to 59, wherein the gene expression level is measured by determining the cDNA level.

63. The method according to any one of claims 1 to 62, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydroisoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate.

64. The method according to any one of claims 1 to 62, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione.

65. The method according to any one of claims 1 to 62, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydroisoindo-2-yl]piperidine-2,6-dione hydrochloride.

66. A method for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to a therapeutic compound, or for predicting the responsiveness of subjects with SLE to a therapeutic compound, (a) A step of obtaining a sample from the subject, (b) A step of determining the presence of the IKZF1 single nucleotide polymorphism (SNP) rs4917014 in the sample, (c) If at least one copy of SNP rs4917014 is detected, the step of diagnosing the subject as likely to be responsive to the therapeutic compound, Includes, The therapeutic compound is a compound of formula I: 【Chemistry 9】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for identifying subjects with systemic lupus erythematosus (SLE) who are likely to be responsive to therapeutic compounds, or for predicting the responsiveness of subjects with SLE to therapeutic compounds.

67. The method according to claim 66, further comprising administering an effective amount of the therapeutic compound to a subject that has been determined to be highly likely to be responsive to the therapeutic compound.

68. A method for treating a subject having systemic lupus erythematosus (SLE), comprising administering an effective amount of a therapeutic compound to the subject, It has been determined that the subject is highly likely to be responsive to the therapeutic compound according to the method of claim 66. The therapeutic compound is a compound of formula I: 【Chemistry 10】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

69. The method according to any one of claims 66 to 68, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydroisoindo-2-yl]piperidine-2,6-dione or a pharmaceutically acceptable salt thereof, solid form, solvate, hydrate, tautomer, stereoisomer or racemate.

70. The method according to any one of claims 66 to 68, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydro-isoindo-2-yl]piperidine-2,6-dione.

71. The method according to any one of claims 66 to 68, wherein the compound is (S)-3-[4-(4-morphyl-4-ylmethylbenzyloxy)-1-oxo-1,3-dihydroisoindo-2-yl]piperidine-2,6-dione hydrochloride.

72. A method for treating a subject having systemic lupus erythematosus (SLE), comprising administering a therapeutic compound to the subject in a dose of 0.45 mg or more or 0.15 mg or less per day, The subject has high type 1 IFN expression and / or gene signature, The therapeutic compound is a compound of formula I: 【Chemistry 11】 or a pharmaceutically acceptable salt, solvate, hydrate, stereoisomer, tautomer, or racemic mixture thereof. A method for treating patients with systemic lupus erythematosus (SLE).

73. The method according to claim 72, wherein if a score calculated based on the gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 in a sample from the subject is higher than a reference level, the subject is determined to have the high type 1 IFN expression and / or gene signature.

74. The method according to claim 73, wherein the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the reference gene in the sample, or the score is the Log2 of the average gene expression levels of IFI27, IFI44, IFI44L, and RSAD2 relative to the average of two or more reference genes in the sample.

75. The method according to claim 74, wherein the score is higher than -1.

38.

76. The method according to claim 74 or 75, wherein the reference gene is selected from the group consisting of TFRC, ACTB, GAPDH and combinations thereof.

77. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.45 mg / day.

78. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.5 mg / day.

79. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.6 mg / day.

80. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.7 mg / day.

81. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.15 mg / day.

82. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.1 mg / day.

83. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.075 mg / day.

84. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is approximately 0.15 mg every other day.

85. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is approximately 0.15 mg every three days.

86. The method according to any one of claims 74 to 76, wherein the dose of the therapeutic compound is about 0.15 mg once a week.

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