CN108137592B - Deuterated compounds and compositions and methods thereof for treating leukemia - Google Patents

Abstract

The present invention provides novel compounds useful for the treatment of various blood cancers or their associated diseases or conditions, and pharmaceutical compositions and methods of preparation and use thereof.

Description

Deuterated compounds and compositions and methods thereof for treating leukemia

Priority claims and related patent applications

This application claims priority to U.S. provisional application serial No. 62/249,991 filed on 3/11/2015, the entire contents of which are incorporated herein by reference in their entirety.

Technical Field

The present invention relates generally to methods of treatment and management for certain diseases and conditions. More specifically, the present invention provides novel compounds comprising 5-fluoro-3-phenyl-2- [ (lS) -l- (9H-purin-6-ylamino) propyl ] quinazolin-4 (3H) -one with deuterium substitution at key positions, which are useful for the treatment of various leukemia and related diseases and conditions, as well as pharmaceutical compositions and methods of preparation and use thereof.

Background

Blood cancer (also known as hematological malignancy or liquid tumor) is a form of cancer that begins in cells of blood-forming tissues, such as bone marrow or cells of the immune system. Examples of hematologic cancers include acute and chronic leukemias, lymphomas, multiple myelomas, and myelodysplastic syndromes.

Leukemia is a group of cancers that usually begin in the bone marrow and result in large numbers of abnormal white blood cells. There are two types of leukemia: lymphocytic leukemia involves lymphocytes. Myeloid leukemia involves granulocytes. These leukocytes are important in combating infection. Lymphoma is a cancer that develops in the lymphatic system.

Leukemia can be a type known as acute leukemia, which is characterized by a rapid increase in the number of immature blood cells, rendering the bone marrow incapable of producing healthy blood cells. Immediate treatment is needed to slow down the rapid progression and accumulation of malignant cells that can spread to other organs of the body via the bloodstream.

Leukemia can be a type known as chronic leukemia, e.g., Chronic Lymphocytic Leukemia (CLL), characterized by an excessive accumulation of relatively mature, but still abnormal, white blood cells. It usually takes months or years to progress and cells are produced at a much higher rate than normal, resulting in many abnormal white blood cells. The chronic form is monitored from time to time prior to treatment for a period of time to ensure maximum efficacy of the treatment.

There are two general types of lymphoma depending on how the cancer spreads. In hodgkin lymphoma, the cancer spreads from one group of lymph nodes to another in a certain order. In non-hodgkin lymphoma, the cancer spreads from one group of lymph nodes to another in random order. Examples of non-hodgkin lymphomas include follicular B-cell non-hodgkin lymphoma (FL) and Small Lymphocytic Lymphoma (SLL). Myeloma is a cancer that causes plasma cells to form tumors in the bone marrow. Myeloma is commonly found in multiple sites in the body and is therefore commonly referred to as multiple myeloma.

Blood cancers such as CLL, FL, and SLL place an increasing burden on society, impairing the health and life of the affected people. Although drugs have been developed for the treatment of some of these diseases and conditions, the available treatments are often limited in clinical effectiveness and, at the same time, have undesirable side effects.

There is an urgent and growing need to provide innovative treatments and treatment methods with improved clinical effectiveness and reduced side effects.

Disclosure of Invention

The present invention provides novel, orally available phosphoinositide 3-kinase (PI3K) delta inhibitors. The compounds and pharmaceutical compositions disclosed herein are biochemically effective and physiologically active, as well as having improved pharmacokinetic and toxicological properties over idelalisib. The compounds disclosed herein are deuterium substituted and modified forms of idelalisis, wherein the hydrogen is substituted with deuterium at a key position in the molecule. The substitution positions are chosen according to the specific purpose to influence the pharmacokinetic and toxicological properties of the molecule. The resulting compounds have up to 7 deuterium substitutions and exhibit more desirable properties in terms of safety, efficacy and tolerability in the treatment of blood cancers and related diseases and conditions such as CLL, FL and SLL.

In one aspect, the present invention is generally directed to a compound having the structural formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each of which is D, and each of which is D,

in another aspect, the present invention relates generally to a pharmaceutical composition comprising a compound having the following structural formula or a pharmaceutically acceptable salt or ester thereof, effective for treating, preventing or ameliorating one or more blood cancers or associated diseases or conditions thereof in a mammal, including a human, and a pharmaceutically acceptable excipient, carrier or diluent:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each is D.

In yet another aspect, the present invention relates generally to a unit dosage form comprising a pharmaceutical composition disclosed herein. The unit dosage form is suitable for administration to a subject having one or more blood cancers, including advanced hematologic malignancies, or a disease or disorder associated therewith.

In yet another aspect, the present invention relates generally to methods for treating, alleviating or preventing a disease or disorder. The method comprises the following steps: administering to a subject in need thereof a pharmaceutical composition comprising a compound having the formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each is D.

In certain embodiments, the one or more hematologic cancers include diseases and conditions that can benefit from treatment with the compounds, pharmaceutical compositions, unit dosage forms, and methods of treatment disclosed herein, including any disease and disorder that can be addressed by inhibition of the delta isoform of PI3K, such as ALL, FL, and SLL.

In certain preferred embodiments, the method of treatment comprises administering to a subject in need thereof a pharmaceutical composition comprising a compound having the formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each is D.

Drawings

FIG. 1 shows certain exemplary compounds disclosed herein¹HNMR(CDCl₃)。

FIG. 2 shows certain exemplary compounds disclosed herein¹HNMR(CDCl₃)。

Fig. 3 shows specific exemplary LC-MS data for the compounds disclosed herein.

FIG. 4 shows certain exemplary compounds disclosed herein¹HNMR(CDCl₃)。

FIG. 5 shows certain exemplary compounds disclosed herein¹HNMR(CDCl₃)。

FIG. 6 shows certain exemplary compounds disclosed herein¹HNMR(CDCl₃)。

Fig. 7 shows specific exemplary HPLC data for compounds disclosed herein.

Figure 8 shows specific exemplary data for percent remaining compound versus incubation time. After 4 hours, D₇-the concentration of esdallas is approximately equal to 220% of esdallas.

FIG. 9 shows Eldallas and D in the formation of oxidative metabolites formed by purine epoxidation at carbon-8₇Specific exemplary comparisons of idelalisis.

Detailed Description

Definition of

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The general principles of Organic Chemistry, as well as specific functional moieties and reactivities are described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausaltito: 2006.

As used herein, "administering" of a disclosed compound encompasses delivering a compound as described herein, or a prodrug or other pharmaceutically acceptable derivative thereof, to a subject using any suitable formulation or route of administration, as discussed herein.

As used herein, the term "effective amount" or "therapeutically effective amount" refers to an amount of a compound or pharmaceutical composition described herein sufficient to effect the intended use, including but not limited to disease treatment, as shown below. In some embodiments, the amount is effective to detectably kill or inhibit growth or spread of cancer cells; the size or number of tumors; or other measure of the level, stage, progression or severity of the cancer. The therapeutically effective amount may vary depending on the intended application or subject and disease condition being treated, e.g., the desired biological endpoint, the pharmacokinetics of the compound, the disease being treated, the mode of administration, and the weight and age of the patient, which can be readily determined by one of ordinary skill in the art. The term also applies to doses in the target cells that will induce a particular response, e.g., a decrease in cell migration. The specific dose will vary depending on, for example, the particular compound selected, the species of the subject and their age/existing health or risk of health, the dosing regimen followed, the severity of the disease, whether administered in combination with other agents, the time of administration, the tissue of administration and the physical delivery system carrying it.

As used herein, the term "treatment" or "treating" a disease or disorder refers to a method of reducing, delaying or ameliorating such conditions before or after onset. Treatment may be directed to more than one effect or symptom of the disease and/or underlying pathology. The purpose of treatment is to obtain beneficial or desired results, including but not limited to therapeutic benefits and/or prophylactic benefits. Therapeutic benefit refers to eradication or amelioration of the underlying disorder being treated. In addition, therapeutic benefit is achieved by eradicating or ameliorating more than one physiological symptom associated with the underlying disorder, such that an improvement is observed in the patient, although the patient may still be afflicted with the underlying disorder. For prophylactic benefit, the pharmaceutical compounds and/or compositions may be administered to patients at risk of developing a particular disease, or to patients reporting more than one physiological symptom of a disease, even though a diagnosis of the disease may not have been made. The treatment may be any reduction, and may be, but is not limited to, complete ablation of the disease or symptoms of the disease. The extent of such reduction or prevention is at least 5%, 10%, 20%, 40%, 50%, 60%, 80%, 90%, 95%, or 100% as measured by any standard technique, as compared to an equivalent untreated control.

The term "therapeutic effect" as used herein refers to a therapeutic benefit and/or a prophylactic benefit as described herein. A prophylactic effect includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, stopping, or reversing the progression of a disease or condition, or any combination thereof.

As used herein, the term "pharmaceutically acceptable ester" refers to an ester that hydrolyzes in vivo and includes esters that readily decompose within the human body to leave the parent compound or a salt thereof. Such esters may act as prodrugs as defined herein. Pharmaceutically acceptable esters include, but are not limited to, alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkyl esters of acidic groups including, but not limited to, carboxylic acids, phosphoric acids, phosphinic acids, sulfinic acids, sulfonic acids, and boronic acids. Examples of esters include formates, acetates, propionates, butyrates, acrylates and ethylsuccinates. Esters may be formed with hydroxy or carboxylic acid groups of the parent compound.

As used herein, the term "pharmaceutically acceptable enol ether" includes, but is not limited to, derivatives of the formula-C ═ C (or), where R may be selected from alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkyl. Pharmaceutically acceptable enol esters include, but are not limited to, derivatives of the formula-C ═ C (oc (o) R), where R may be selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkyl.

As used herein, "pharmaceutically acceptable forms" of the disclosed compounds include, but are not limited to, pharmaceutically acceptable salts, hydrates, solvates, isomers, prodrugs, and isotopically labeled derivatives of the disclosed compounds. In one embodiment, "pharmaceutically acceptable forms" include, but are not limited to, pharmaceutically acceptable salts, isomers, prodrugs, and isotopically labeled derivatives of the disclosed compounds. In some embodiments, "pharmaceutically acceptable forms" include, but are not limited to, pharmaceutically acceptable salts, stereoisomers, prodrugs, and isotopically labeled derivatives of the disclosed compounds.

In certain embodiments, the pharmaceutically acceptable form is a pharmaceutically acceptable salt. As used herein, the term "pharmaceutically acceptable salts" refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of subjects without excessive toxicity, irritation, allergic response, and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. Pharmaceutically acceptable salts are described in detail, for example, by Berge et al in J.pharmaceutical sciences (1977)66: 1-19. Pharmaceutically acceptable salts of the compounds provided herein include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are the salts of amino groups formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipates, alginates, ascorbates, aspartates, benzenesulfonates (bezenesulfonates), benzenesulfonates (besylate), benzoates, bisulfates, borates, butyrates, camphorates, camphorsulfonates, citrates, cyclopentanepropionates, digluconates, dodecylsulfates, ethanesulfonates, formates, fumarates, glucoheptonates, glycerophosphates, gluconates, hemisulfates, heptanoates, hexanoates, hydroiodides, 2-hydroxy-ethanesulfonates, lactobionates, lactates, laurates, malates, maleates, malonates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, oxalates, palmitates, pamoate, pectinates, persulfates, 3-phenylpropionates, Phosphates, picrates, pivalates, propionates, stearates, succinates, sulfates, tartrates, thiocyanates, p-toluenesulfonates, undecanoates, and valerates, and the like. In some embodiments, organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, lactic acid, trifluoroacetic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.

The salts can be prepared in situ during the isolation and purification of the disclosed compounds or prepared separately, e.g., by reacting the free base or free acid of the parent compound with a suitable base or acid, respectively. Pharmaceutically acceptable salts derived from suitable bases include alkali metals, alkaline earth metals, ammonium and N⁺(C_1-4Alkyl radical)⁴And (3) salt. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Other pharmaceutically acceptable salts include, when appropriate, non-toxic ammonium, quaternary ammonium and amine cations formed using counterions such as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, lower alkyl sulfonates and aryl sulfonates. Organic bases from which salts can be derived include, for example, primary, secondary and tertiary amines, substituted amines, including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the pharmaceutically acceptable base addition salt may be selected from the group consisting of ammonium, potassium, sodium, calcium and magnesium salts.

In certain embodiments, the pharmaceutically acceptable form is a "solvate" (e.g., hydrate). As used herein, the term "solvate" refers to a compound that further includes a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent intermolecular forces. The solvate can be the disclosed compound or a pharmaceutically acceptable salt thereof. When the solvent is water, the solvate is a "hydrate". Pharmaceutically acceptable solvates and hydrates are complexes that may include, for example, from 1 to about 100, or from 1 to about 10, or from 1 to about 2, about 3, or about 4 solvent or water molecules. It is to be understood that the term "compound" as used herein encompasses compounds and solvates of compounds, as well as mixtures thereof.

In certain embodiments, the pharmaceutically acceptable form is a prodrug. As used herein, the term "prodrug" (or "prodrug") refers to a compound that is converted in vivo to produce the disclosed compound or a pharmaceutically acceptable form of the compound. A prodrug may be inactive when administered to a subject, but converted in vivo to an active compound, for example, by hydrolysis (e.g., hydrolysis in blood). In certain instances, the prodrugs have improved physical and/or delivery properties over the parent compound. The prodrug may increase the bioavailability of the compound when administered to a subject (e.g., by allowing for enhanced absorption into the blood following oral administration), or it enhances delivery to a biological compartment of interest (e.g., the brain or lymphatic system) relative to the parent compound. Exemplary prodrugs include derivatives of the disclosed compounds having enhanced aqueous solubility or active transport through the intestinal membrane relative to the parent compound.

Prodrug compounds generally provide the advantages of solubility, histocompatibility, or delayed release in mammalian organisms (see, e.g., Bundgard, h., Design of produgs (1985), pp.7-9,21-24(Elsevier, Amsterdam)). A discussion of prodrugs is provided in Higuchi, T.T., et al, "Pro-drugs as Novel Delivery Systems", A.C.S.Symposium series, Vol.14 and Bioreversible Carriers in Drug Design, ed.Edward B.Roche, American Pharmaceutical Association and Pergamon Press,1987, both of which are incorporated herein by reference in their entirety. Exemplary advantages of the prodrugs may include, but are not limited to, their physical properties, such as increased aqueous solubility compared to parenteral administration of the parent compound at physiological pH, or they may enhance absorption from the digestive tract, or they may enhance drug stability for long term storage.

As used herein, the term "pharmaceutically acceptable" excipient, carrier or diluent refers to a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in the delivery or transport of an agent of interest from one organ or part of the body to another organ or part of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials that can be used as pharmaceutically acceptable carriers include: sugars such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered gum tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols, such as propylene glycol; polyols such as glycerol, sorbitol, mannitol and polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; ringer's solution; ethanol; a phosphate buffer solution; and other non-toxic compatible materials employed in pharmaceutical formulations. Wetting agents, emulsifiers and lubricants, such as sodium lauryl sulfate, magnesium stearate and polyethylene oxide-polypropylene oxide copolymers, as well as coloring, mold release, coating, sweetening, flavoring and perfuming agents, preservatives and antioxidants, may also be present in the composition.

As used herein, the term "subject" refers to any animal (e.g., a mammal), including but not limited to humans, non-human primates, rodents, and the like, which will be the recipient of a particular treatment. Generally, the terms "subject" and "patient" are used interchangeably herein when referring to a human subject.

As used herein, "low dose" refers to a dose that is at least 5% (e.g., at least 10%, 20%, 50%, 80%, 90%, or even 95%) lower than the minimum standard recommended dose of a particular compound formulated for a given route of administration for the treatment of any human disease or condition. For example, a low dose of an agent that lowers glucose levels and is formulated for administration by inhalation will be different than a low dose of the same agent formulated for oral administration.

As used herein, "high dose" means a dose that is at least 5% (e.g., at least 10%, 20%, 50%, 100%, 200%, or even 300%) higher than the highest standard recommended dose of a particular compound for treating any human disease or condition.

The compounds of the present invention are preferably isolated and purified after their preparation to obtain a composition containing an amount equal to or greater than 95% by weight ("substantially pure"), which is then used or formulated as described herein. In certain embodiments, the compounds of the present invention are greater than 99% pure.

Solvates and polymorphs of the compounds of the invention are also encompassed herein. Solvates of the compounds of the invention include, for example, hydrates.

Detailed description of the invention

The present invention provides novel chemical entities that are biochemically efficient and physiologically active, as well as having improved pharmacokinetic and toxicological properties over 5-fluoro-3-phenyl-2- [ (lS) -l- (9H-purin-6-ylamino) propyl ] quinazolin-4 (3H) -one (idelaris) shown below.

Ai dallas si

Is an inhibitor of phosphatidylinositol 3-kinase (PI3K) approved for use in combination with rituximab in the treatment of patients with relapsed Chronic Lymphocytic Leukemia (CLL) for which rituximab alone would be considered a suitable treatment due to other complications. Also indicated is the use of idelalisis for the treatment of patients with recurrent follicular B-cell non-hodgkin's lymphoma (FL) or recurrent Small Lymphocytic Lymphoma (SLL) who have received at least two previous systemic therapies (system therapies).

The US FDA's prescription information for idelalisis contains a black box warning for fatal and/or severe adverse reactions. Recently, six clinical trials investigating combinations of idelalisin with other treatments were discontinued for patients with hematological malignancies due to reports of increased rates of adverse events including death.

(http://www.fda.gov/Drugs/DrugSafety/ucm490618.htm)

Clinical studies have revealed severe toxicity associated with idelalisis, including hepatotoxicity, severe diarrhea, colitis, pneumonia and intestinal perforation. Fatal and/or severe hepatotoxicity occurs in 14% of patients treated with idelalisis. Fatal and/or severe diarrhea or colitis occurs in 14% of patients treated with idelalisia. In addition, fatal and severe pneumonia can occur in idelalis-treated patients.

Severe adverse reactions were also reported when used with rituximab. For example, severe adverse reactions were shown in 49% of patients treated with idelalisib and rituximab. The most frequent serious adverse reactions were pneumonia (17%), fever (9%), sepsis (8%), febrile neutropenia (5%) and diarrhea (5%). Adverse reactions leading to the discontinuation of rituximab occurred in 10% of patients. The most common adverse reactions leading to treatment discontinuation are hepatotoxicity and diarrhea/colitis. 35% of patients have dose interruption (dose interruption) and 15% have dose reduction (dose reduction) due to adverse reactions or laboratory abnormalities. The recommended maximum starting dose for idelalisis is 2 times daily for oral administration of 150 mg.

The compounds disclosed herein are deuterium substituted forms of idelalisis, wherein the hydrogen is substituted with deuterium at a critical position in the molecule. The substitution positions are chosen according to the specific purpose to influence the pharmacokinetic and toxicological properties of the molecule.

The preferred compounds shown above have 7 deuterium substitutions and exhibit excellent safety, efficacy and tolerability characteristics. The compounds provide new and improved options in the treatment of blood cancers such as CLL and related diseases and conditions. The compounds may also be used to treat FL and SLL (e.g., recurrent FL or SLL, particularly after a previous systemic treatment).

The main metabolite GS-5637 of Idelalis is less than or equal to 10 μm (clinical free C of the metabolite)_minAbout 17-fold) does not significantly inhibit class I PI3K kinase activity in an in vitro enzyme assay. Therefore, it is expected that the metabolite does not significantly contribute to the therapeutic effect of the drug.

Although GS-56363 is an important metabolite in humans (exposure [ AUC ] 3.3 times that of Idelalisib), it is only a minor metabolite in animal species (exposure [ AUC ] to GS-56363 is 0.7-3.6%, 4.4-8.9% and 16-66% of Idelalisib in rats, rabbits and dogs, respectively). Studies to date have shown that this oxidation product (formed by the epoxidation of a purine on carbon-8) can be responsible for the skin, reproductive and phototoxicity of idelalisia.

GS-56363 has significant inhibitory activity against Ste 20-like kinase (SLK) and lymphocyte directed kinase (LOK) at clinically relevant concentrations. Pharmacologically active oxidation products at carbon-8 of the purine ring antagonize LOK and SLK kinases that can activate lymphocyte responses and lead to skin disorders.

Adverse fetal effects were associated with the inhibition of SLK by the eltoralis metabolite GS-56363. SLK is expressed in the muscle and neuronal lineages in developing embryos and inactivation of this kinase has led to embryonic fetal death. Findings in rat experiments indicate that if idellaris is taken during pregnancy, there is a risk to the developing fetus.

It is clear that GS-5656336 has phototoxicity on cultured cells in vitro experiments (IC 5016-23. mu.g/mL; ERC_max5-7). In tissue distribution studies, it appears that there is some retention of drug-related substances in the pigmented tissue. Due to ai DynastyLaris and GS-5656363 absorb light, distribute and remain in the pigmented skin, and show that the metabolite GS-56363 is phototoxic in vitro, and some phototoxic skin reactions can be seen in vivo.

In vitro phototoxicity studies in embryonic mouse fibroblast BALB/c 3T3 cell lines using Neutral Red (Neutral Red) uptake as a marker of cell viability in the presence or absence of ultraviolet A (UVA) light exposure were reported. This study showed that the major human metabolite GS-56363 induces phototoxicity in the presence of UVA exposure. This metabolite has a very high exposure in humans (3.3 times higher in esdallas) and is responsible for several serious toxicities.

In addition, GS-5656363 inhibits the catalytic activity of cytochrome P450 enzymes CYP2C9, CYP3A, CYP2C8 and CYP2C 19. CYP2C8, CYP2C9 or UGT1 Al substrates were administered simultaneously to some patients, particularly to about 30% of patients participating in NHL and CLL trials with sensitive CYP2C19 substrates including lansoprazole and omeprazole. The incidence of diarrhea and skin rash is higher in patients taking Proton Pump Inhibitors (PPIs). These adverse events are associated with both PPI and idelalisis. Exposure to idelalisis is similar in patients taking acid-reducing agents (ARA) compared to patients not taking these agents. Overlapping adverse events or increased exposure to PPI (CYP2C19 substrate) may explain increased incidence of adverse events. Higher exposure (5-12 fold) of PPIs has been observed in weak CYP2C19 metabolisers and dose-response to diarrhea and infection has been observed. Thus, GS-56311 may cause toxicity through drug-drug interactions (DDI) in patients taking other drugs, since the plasma concentration of GS-56311 is much higher in humans than that of Idelalisib.

The present invention aims to reduce toxic-causing metabolites while increasing parent drug exposure. The present invention provides compounds that are orally available phosphoinositide 3-kinase (PI3K) delta inhibitors. In particular, the compound targets the delta isoform of P110 delta, PI3K, which is one of the proteins responsible for leukemia and other cell growth.

In one aspect, the present invention is generally directed to a compound having the structural formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each of which is D, and each of which is D,

in another aspect, the present invention relates generally to a pharmaceutical composition comprising a compound having the following structural formula or a pharmaceutically acceptable salt or ester thereof, effective for treating, preventing or ameliorating one or more blood cancers or associated diseases or conditions thereof in a mammal, including a human, and a pharmaceutically acceptable excipient, carrier or diluent:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each of which is D, and each of which is D,

in yet another aspect, the present invention relates generally to a unit dosage form comprising a pharmaceutical composition disclosed herein. The unit dosage form is suitable for administration to a subject having one or more blood cancers, including advanced hematologic malignancies, or a disease or disorder associated therewith.

In yet another aspect, the present invention relates generally to methods for treating, alleviating or preventing a disease or disorder. The method comprises the following steps: administering to a subject in need thereof a pharmaceutical composition comprising a compound having the formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each is D.

In certain embodiments, the one or more hematologic cancers include diseases and conditions that can benefit from treatment with the compounds, pharmaceutical compositions, unit dosage forms, and methods of treatment disclosed herein, including any disease and disorder that can be addressed by inhibition of the delta isoform of PI3K, such as ALL, FL, and SLL.

In certain preferred embodiments, the method of treatment comprises administering to a subject in need thereof a pharmaceutical composition comprising a compound having the formula:

wherein R is₁、R₂、R₃、R₄、R₅、R₆And R₇Each is D.

The one or more additional anti-cancer agents may be small molecules, chemotherapeutic agents, peptides, polypeptides or proteins, antibodies, antibody-drug conjugates, aptamers, or nucleic acid molecules.

In certain embodiments, the one or more additional anti-cancer agents are chemotherapeutic agents, which are compounds useful in the treatment of cancer. Examples of chemotherapeutic agents include erlotinib (b: (a))

Genentech/OSI Pharm.), bortezomib (

Millennium Pharm, fulvestrant (

AstraZeneca), sotriptan (SU11248, Pfizer), letrozole (I), (II)

Novartis), imatinib mesylate (

Novartis), PTK787/ZK222584(Novartis), oxaliplatin (A) ((B)

Sanofi), 5-FU (5-fluorouracil), leucovorin, rapamycin (Sirolimus,

wyeth), lapatinib (

GSK572016, Glaxo Smith Kline), lonafarnib (SCH 66336), sorafenib (BAY43-9006, Bayer Labs), and gefitinib (R: (R) ((R))

AstraZeneca), AG1478, AG1571(SU 5271; sugen), alkylating agents such as thiotepa and

cyclophosphamide; alkyl sulfates such as busulfan, improsulfan and piposulfan; aziridines such as benzodidopa (benzodipa), carboquone (carboquone), metridopa (meteredopa), and ulidopa (uredopa); vinyl imines and methyl melamines (melamines) including altretamine (altretamine), triethylenemelamine (triethyleneamine), triethylenephosphoramide (triethylenephosphoramide), triethylenemercaptophosphamide (triethylenethiophosphamide), and trimethylolmelamine (trimethylamine); annona squamosa lactones (acetogenins) (in particular)Are bullatacin (bullatacin) and bullatacin (bullatacinone)); camptothecin (including the synthetic analogue topotecan); bryostatin; a caristatin (callystatin); CC-1065 (including its adozelesin (adozelesin), carvelesin (carzelesin), and bizelesin (bizelesin) synthetic analogs); cryptophycins (especially cryptophycins 1 and 8); dolastatin (dolastatin); duocarmycins (duocarmycins) (including synthetic analogs, KW-2189 and CB1-TM 1); shogaol (eleutherobin); coprinus atrata base (pancratistatin); sarcandra glabra alcohol (sarcodictyin); spongistatin (spongistatin); nitrogen mustards such as chlorambucil, naphazel, chlorophosphamide, estramustine, ifosfamide, mechlorethamine hydrochloride, melphalan, neonebichin (novembichin), benzene mustard cholesterol, prednimustine, trofosfamide (trofosfamide), uracil mustard (uracil musard); nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine and ranimustine; antibiotics such as enediyne antibiotics (actinomycin), in particular, calicheamicin (gammall) and calicheamicin (omell) (Angew chem. int. Ed. Engl. (1994)33:183-186), daptomycin (dynemicin), including daptomycin A, bisphosphonates, such as clodronate, esperamicin (esperamicin), and neocarzinostain chromophores (neocarzinostain chromophoropterin) and related chromoprotein enediyne antibiotic chromophores, clarithromycin (acrinomycin), actinomycin, apramycin (aureomycin), azaserine (azaserine), bleomycin (bleomycin), actinomycin, carubicin (carvacrin), actinomycin (camycin), carcinomycin (carcinomycin), daunomycin (carbamycin), daunomycin (5-5), daunomycin (daunomycin D), daunomycin D-183-186), daunomycin (dynemycin-A), daunomycin A, daunomycin (gentin), daunomycin A, daunomycin (daunomycin A), daunomycin (daunomycin A-5, daunomycin A-D), daunomycin (daunomycin A-D), daunomycin A-D, a,

(doxorubicin), morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrroline-doxorubicin and deoxydoxorubicin), epirubicin, isoxabixin(esonicicin), idarubicin (idarubicin), marijumycin (marcelomycin), mitomycins such as mitomycin C, mycophenolic acid, nogalamycin (nogalamycin), olivomycin (olivomycin), pelomycin (polyplomycin), pofiomycin (porfiromycin), puromycin, triformycin (queramycin), rodobicin (rodorubicin), streptonigrin (streptonigrin), streptozotocin (streptozocin), tubercidin (tubicidin), ubenimex (ubenimex), stastatin (zinostatin), zorubicin (zorubicin); antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs such as denopterin, methotrexate, pteropterin (pteropterin), trimetrexate (trimetrexate); purine analogs such as fludarabine (fludarabine), 6-mercaptopurine, thiamine (thiamniprine), thioguanine; pyrimidine analogs such as cyclocytidine (ancitabine), azacitidine (azacitidine), 6-azauridine (6-azauridine), carmofur (carmofur), cytarabine, dideoxyuridine (dideoxyuridine), deoxyfluorouridine (doxifluridine), enocitabine (enocitabine), floxuridine; androgens such as carposterone (calusterone), drostandrosterone propionate (dromostanolone propionate), epitioandrostanol (epitiostanol), mepiquitane (mepiquitazone), testolactone (testolactone); anti-adrenals such as aminoglutethimide (aminoglutethimide), mitotane (mitotane), trilostane (trilostane); folic acid replenisher such as leucovorin; acetoglucurolactone (acegultone); (ii) an aldophosphamide glycoside; (ii) aminolevulinic acid; eniluracil (eniluracil); ambridine (amsacrine); bessburyl (beslabucil); bisantrene; edatrexate (edatraxate); desphosphamide (defofamine); colchicine (demecolcine); diazaquinone (diaziqutone); eflornithine (elformithine); ammonium etitanium acetate; epothilone (epothilone); ethydine (etoglucid); gallium nitrate; a hydroxyurea; lentinan; lonidamine (lonidainine); maytansinoids (maytansinoids) such as maytansine (maytansine) and ansamitocins (ansamitocins); mitoguazone (mitoguzone); mitoxantrone (mitoxantrone); mopidanol (mopidanmol); diamine nitracridine (nitrarine); pentostatin (pentostatin); egg ammoniaNitrogen mustard (phenamett); pirarubicin (pirarubicin); losoxantrone (losoxantrone); podophyllinic acid (podophyllic acid); 2-acethydrazide; procarbazine (procarbazine);

polysaccharide complex (JHS Natural Products, Eugene, Oreg.); razoxane (rizoxane); rhizomycin (rhizoxin); azofurans (sizofurans); germanium spiroamines (spirogyranium); tenuazonic acid (tenuazonic acid); triimine quinone (triaziquone); 2, 2' -trichlorotriethylamine; trichothecenes (trichothecenes), in particular the T-2 toxin, verrucin A (verrucin A), bacillocin A (roridin A) and serpentinine (anguidine)); uratan; vindesine (vindesine); dacarbazine (dacarbazine); mannitol mustard (mannomustine); dibromomannitol (mitobronitol); dibromodulcitol (mitolactol); pipobromane (pipobroman); gatifloxacin (gacytosine); arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxanes, e.g.

(paclitaxel; Bristol-Myers Squibb Oncology, Princeton, N.J.),

(hydrogenated castor oil free), albumin-engineered nanoparticle formulations of paclitaxel (American Pharmaceutical Partners, Schaumberg,111.), and

(docetaxel; Rhone-Poulenc Rorer, Antony, France); chlorambucil (chlorembucil);

(gemcitabine); 6-mercaptoguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine (vinblastine); etoposide (VP-16); ifosfamide; mitoxantrone; vincristine;

(vinorelbine); nuantro (novantrone); teniposide (teniposide); edatrexate (edatrexate); daunorubicin (daunomycin); aminopterin (aminopterin); capecitabine

Ibandronate (ibandronate); CPT-11; topoisomerase inhibitor RFS 2000; difluoromethyl ornithine (DMFO); retinoids (retinoids) such as retinoic acid (retinoic acid); and pharmaceutically acceptable salts, acids and derivatives of any of the above.

In certain preferred embodiments, the one or more additional anti-cancer agents are selected from monoclonal antibodies against CD20 protein, e.g., rituximab.

Rituximab is a chimeric monoclonal antibody against the CD20 protein, which is found mainly on the surface of B cells of the immune system. Rituximab is used to treat diseases characterized by excessive numbers of B cells, hyperactivated B cells, or dysfunctional B cells, including many lymphomas, leukemias, transplant rejection, and autoimmune disorders (autoimmune disorders).

The compounds disclosed herein may be used as second-line drugs against patients who have relapsed CLL. These compounds may be used in combination with rituximab in patients when CLL has relapsed after a previous cancer treatment. These compounds are particularly effective in patients with the p53 mutation, which otherwise tend to confer a poor prognosis in CLL patients.

Any suitable route of administration may be employed, for example, parenteral, intravenous, subcutaneous, intramuscular, intraventricular, intracorporeal, intraperitoneal, rectal or oral administration. The most appropriate mode of administration for a particular patient will depend on the nature and severity of the disease or condition being treated, or the nature of the treatment being used and the nature of the active compound.

Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the compounds described herein or derivatives thereof are admixed with at least one inert conventional excipient (or carrier) as described below: sodium citrate or dicalcium phosphate or (i) fillers or extenders (extenders) such as starches, lactose, sucrose, glucose, mannitol and silicic acid, (ii) binders such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and acacia, (iii) wetting agents such as glycerol, (iv) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates and sodium carbonate, (v) solution retarders (solution reducers) such as paraffin, (vi) absorption accelerators such as quaternary ammonium compounds, (vii) wetting agents such as cetyl alcohol and glycerol monostearate, (viii) adsorbents such as kaolin and bentonite, and (ix) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate or mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. Solid compositions of a similar type may also be employed as fillers in soft-filled and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other materials known in the art.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1, 3-butylene glycol, dimethylformamide, oils, in particular, cottonseed oil, groundnut oil, corn germ oil, olive oil, castor oil, sesame oil, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, or mixtures of these substances and the like. In addition to such inert diluents, the compositions may include additional agents such as wetting agents, emulsifying agents, suspending agents, sweetening, flavoring, or perfuming agents.

The materials, compositions, and components disclosed herein can be used for, can be used in combination with, can be used in the preparation of, or are the product of, the disclosed methods and compositions. It is to be understood that when combinations, subsets, interactions, groups, etc. of these materials are disclosed that while specific reference of each various individual and collective combinations and permutation of these compounds may not be explicitly disclosed, each is specifically contemplated and described herein. For example, if a method is disclosed and discussed and a number of modifications that can be made to a number of molecules included in the method are discussed, each combination and permutation of the method and the modifications that are possible are specifically contemplated unless indicated to the contrary. Likewise, any subset or combination of these is also specifically contemplated and disclosed. This concept applies to all aspects of this disclosure including, but not limited to, steps in methods of using the disclosed compositions. Thus, if there are a variety of additional steps that can be performed it is understood that each of these additional steps can be performed with any specific method step or combination of method steps of the disclosed methods and that each such combination or subset of combinations is specifically contemplated and should be considered disclosed.

Certain compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, R-and S-enantiomers, diastereomers, (D) -isomers, (L) -isomers, racemic mixtures thereof, and other mixtures thereof, which fall within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers and mixtures thereof are intended to be included in the present invention.

Isomer mixtures comprising any of a variety of isomer ratios may be used in accordance with the present invention. For example, where only two isomers are combined, mixtures comprising ratios of 50:50, 60:40, 70:30, 80:20, 90:10, 95:5, 96:4, 97:3, 98:2, 99:1, or 100:0 isomers are contemplated by the present invention. One of ordinary skill in the art will readily appreciate that similar ratios are contemplated for more complex isomer mixtures.

For example, if a particular enantiomer of a compound of the invention is desired, it may be prepared by asymmetric synthesis or by derivatization with a chiral auxiliary, wherein the resulting mixture of diastereomers is separated and the ancillary groups are cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (e.g., amino) or an acidic functional group (e.g., carboxyl), diastereomeric salts are formed with a suitable optically active acid or base, followed by resolution of the diastereomers thus formed by fractional crystallization or chromatographic methods well known in the art, and subsequent recovery of the pure enantiomers.

Examples

₇Synthesis of D-Idelalis (Compound J-D7)

Synthesis of Compound D

1.9g L-2-aminobutyric acid d₆(17.4mmoL) was dissolved in 18mL of 1M sodium hydroxide solution and 13mL of MeOH, cooled in an ice bath, and added dropwise to 4.8mL of Boc₂O (20.9 mmoL). The reaction was stirred at room temperature overnight. MeOH was removed using rotary evaporation, then the pH was adjusted to-2 with 1M hydrochloric acid solution, extracted with ethyl acetate, the organic layers were combined, washed with brine, then dried over anhydrous magnesium sulfate, then filtered, the solvent was removed by rotary evaporation, and dried with an oil pump under vacuum. The product was collected as a colorless or pale yellow sticky wax (3.5g, 96% yield, including traces of ethyl acetate residue).¹HNMR(CDCl₃) Shown in fig. 1.

Synthesis of Compound E

4.45g (17.1mmoL) of Compound C are suspended in 10mL of thionyl chloride and 50. mu.L of DMF is added dropwise. The reaction mixture was refluxed for about 2 hours. Thionyl chloride was removed via rotary evaporation. Toluene was used to help remove the residue.

3.40g (16.2mmoL) of Compound D and 3.1mL Triethylamine (TEA) were dissolved in 20mL anhydrous DCM and cooled for 5-10min while on ice bath. Will come fromThe above intermediate of compound C was also dissolved in 20mL Dichloromethane (DCM). The intermediate solution was slowly added dropwise to the cold solution. The reaction mixture was stirred at room temperature over the weekend. The reaction mixture was extracted with water, sodium bicarbonate (saturated), 5% citric acid, water and brine, and over MgSO₄(anhydrous) drying. After filtration and concentration, the target compound was purified using flash column (flash column). Compound E was collected as a beige foamy solid (6.4g, yield 82%).¹HNMR(CDCl₃) Shown in fig. 2. LC-MS is shown in fig. 3.

Synthesis of Compound G

A solution of compound E (14mmoL) in acetic acid (60mL) was treated with zinc dust (5.13g, 6 equivalents) added in 3 portions. The reaction mixture was cooled to below 35 ℃ between additions. After stirring overnight at ambient temperature, the solid was filtered off and washed with acetic acid (5 mL). The filtrate was concentrated in vacuo, dissolved in EtOAc, and washed with water. The aqueous layer was extracted with EtOAc. The combined organic layers were washed with water, saturated sodium bicarbonate and saturated brine, dried over anhydrous magnesium sulfate and concentrated to a slurry. The slurry was purified by flash chromatography to give compound G as an off-white foamy solid (3.74G, 66% yield).¹HNMR(CDCl₃) Shown in fig. 4.

Synthesis of Compound H

A solution of compound G (9.17mmoL) in DCM (10mL) was treated with trifluoroacetic acid (TFA) (10 mL). The reaction mixture was stirred for 1 hour, concentrated in vacuo, and partitioned between DCM and 10% potassium carbonate (aqueous layer pH-10 after extraction). The aqueous layer was extracted with additional DCM and the combined organic layers were washed with water and brine and dried over anhydrous magnesium sulfate. The solution was concentrated to provide compound H (2.42g, 87% yield) as an off-white solid.¹HNMR(CDCl₃) Shown in fig. 5.

Synthesis of Compound I

To a 1L flask was added 6-chloropurine (30g, 194.1 mmole, 1 eq.), 3, 4-dihydropyran (24.5g, 291.1mmoL, 1.5 eq.), and PTSA monohydrate (2.95g, 15.5mmoL, 8% eq.), followed by additionEtOAc (240mL) was added. The mixture was refluxed for 2 hours. After the mixture was cooled, it was washed with NaHCO₃Wash (250mL) to adjust pH 7-8 and wash 3 times with brine 150 mL. The EtOAc layer was washed with Na₂SO₄Dried and concentrated to dryness. The residue was purified by short silica gel column (silica gel plug) with hexanes: EtOAc (2:1, 1:1 and 1:2) to give 6-chloro-9- (tetrahydro-2H-pyran-2-yl) -9H-purine (31.6g, 67%) as an off-white solid.

At-40 ℃ under N₂To a 100mL round bottom flask, n-butyllithium (2.5M, 23.5mL, 58.7mmoL, 1.4 equivalents) was added dropwise to diisopropylamine (5.94g, 58.7mmoL, 1.4 equivalents) in Tetrahydrofuran (THF) (40 mL). The temperature of the mixture was raised to-10 ℃. The mixture was then cooled to-70 ℃. While maintaining the temperature below-68 ℃, a solution of 6-chloro-9- (tetrahydro-2H-pyran-2-yl) -9H-purine (10g, 41.9mmoL, 1 eq) in THF (20mL) was added dropwise. The mixture was stirred for 1 hour, then 5mL of D was added₂And O. The temperature of the mixture was raised to 10 ℃, 2N HCl was added to the mixture to adjust the pH to 8. The separated THF layer was concentrated. The aqueous phase was extracted with EtOAc (100 mL. times.2). The EtOAc layer was combined with concentrated THF, washed with brine (75 mL. times.2), and over Na₂SO₄And (5) drying. The solvent was evaporated to give a red oil. The above H-D exchange process was repeated twice. The crude product was purified by short silica gel column with hexanes: EtOAc (5:1, 4:1, 3:1, 2.5:1) to give compound I as a yellow oil (5.6g, 56% yield).

Synthesis of Compound K

Compound H (2.5g, 8.24mmoL, 1 eq) and compound I (1.97g, 8.24mmoL, 1 eq) were mixed with Ν, Ν -Diisopropylethylamine (DIPEA) (3.2g, 24.7mmoL, 3 eq) and tert-butyl-OD (13mL) in a 150mL sealed tube. The mixture was heated at 88 ℃ by means of an oil bath. After 40 hours, the mixture was concentrated to dryness. The residue was recrystallized from isopropanol (50mL) to give compound K as a yellow solid (3g, yield 55%).

Synthesis of Compound J-D7

To compound K (3.2g, 6.3mmoL) in EtOH (4mL) was added 6N HCl (3.5 mL). Mixing the raw materialsThe material was stirred at room temperature for 1 hour and diluted with water (20 mL). Adding NaHCO dropwise to the mixture₃Aqueous solution (3g in 100mL of water) and a white solid precipitated. The suspension was stirred for 30min and filtered. The solid was washed with water (25mL × 2) and dried under vacuum at 50 ℃ to give compound K-D7 as an off-white solid (2.3g, yield 86%).¹HNMR(CDCl₃) Shown in fig. 6.¹H-NMR (500MHz, DMSO-d 6): 12.93(s,1H),8.17(s,1H),7.74-7.81(m,2H),7.48-7.54(m,5H),7.41-7.43(d,1H),7.22-7.27(dd, 1H); HPLC 97.0% purity (method: 150mm C18 reverse phase column, gradient 5-95% ACN + 0.1% trifluoroacetic acid over 11 min; wavelength: 254 nm): retention time: 6.44 min. HPLC is shown in fig. 7.

₇Pharmacokinetic and pharmacokinetic studies of Idelalisib and D-Idelalisib using human hepatocytes

This study was conducted to evaluate the stability of test compounds when metabolized by cryopreserved human hepatocytes.

Cryopreserved hepatocytes represent a widely accepted experimental system for evaluation of drug properties including metabolic stability, metabolite identification, drug-drug interaction potential (potential), and hepatotoxicity potential. Cryopreserved human hepatocytes were obtained from IVAL LLC. Donor characteristics of the hepatocyte batches used for the study (HH1009) were as follows:

the test article is administered directly or in vitro via a solvent compatible with the test system. The study was performed in uncoated 24-well plates.

Cell-free (NC) negative controls consisted of the addition of test article but no hepatocytes. These samples represent possible chemical degradation and/or adsorption to the surface (adsorption) and "sticking" of specific compounds. Cell-free controls were also run at similar time points (i.e., T ═ 0min, 30min, 60min, 120min, 180min, and 240 min). Cell-free controls were performed in incubation medium as per the requirements of the sponsor and run as a single incubation.

Cryopreserved hepatocytes were thawed in a 37 ℃ water bath and placed on ice. Thawed hepatocytes were treated with Universal Cryopreserved Recovery Medium^TM(UCRM^TM) Resuscitated and centrifuged at 100 × g for 5 min to remove residual cryopreservative (cryopreservant). The hepatocyte pellets (pellet) were resuspended in William's E medium HIM (William's E based medium HIM). Viability and cell concentration were determined using a hemocytometer based on trypan blue exclusion. The cell suspension was adjusted to 1.11X 10 per mL⁶Cells were plated and placed on ice until use.

The final reaction mixture for hepatocyte metabolism was 1X 10 per mL of HIM⁶Individual liver cells and test article or negative control.

The study was designed so that each group of reference (Idelalis) and test article (D) was present₇-idelalisis). The final concentrations of the reference and test samples at the beginning of incubation were 2 μm. Each test article including the reference article was prepared as a 20,000-fold stock solution in DMSO at a concentration of 40 mM. Each test article was mixed with a reference article in equal volumes to prepare pooled 10,000-fold DMSO stocks of 20mM each. This DMSO stock solution was diluted 1000-fold in HIM to prepare a 10-fold dose stock solution (dousing stock) of 20 μm. This dose stock was diluted to 2 μ Μ when added to the medium containing hepatocytes or to the blank medium.

Incubation of hepatocytes with 2 μm test article + reference article at 37 ℃ maintained at a constant temperature and 5% CO₂And 95% equilibrium air in a humidified atmosphere incubator in triplicate for periods of 0, 30, 60, 120, 180 and 240 minutes and at a hepatocyte concentration of 1 × 10 per mL⁶And (4) cells. Negative controls included samples with 2 μ Μ of the test article + reference article at the same six time points only, in the absence of hepatocytes (incubation medium). These controls were performed under the same conditions. The total reaction volume was 500 μ L (0.450mL hepatocyte suspension +0.050mL incubation buffer media with 10 × test compound or positive control).All samples except the negative control were run in triplicate.

The reaction was initiated with the addition of 0.050mL of the appropriate test chemical in buffer (mix-up and down movement of multichannel pipettor to mix the test product well with the incubation medium mixture) and placed in a 37 ℃ incubator. At the indicated time points, 50 μ L of sample was collected from each treatment group, followed by the addition of 100 μ L of ice-cold acetonitrile containing 1 μ g/mL of amisulbutamide (internal standard). Internal standard was added to all samples.

The concentration of the test sample:one concentration (2.0 μm) of test + reference was used for all incubations.

Positive control:a reference substance at 2 μm in each group was used as a positive control.

And (4) terminating:after incubation, the reaction was terminated as described previously. The total mixture after termination was cryopreserved for LCMS analysis.

Figure 8 shows the percentage of remaining compound versus incubation time. After 4 hours, D₇-the concentration of esdallas is approximately equal to 220% of esdallas. The result shows D₇Idelalisib has a longer half-life and AUC. This substantial difference indicates D₇Excellent DMPK properties of idelalisis.

Toxicity of the oxidation products

As noted herein, studies have shown that oxidation products (formed by purine epoxidation on carbon-8) can be responsible for skin toxicity, reproductive toxicity, and phototoxicity of idelalisia. Selectively modifying the molecular structure to reduce metabolite formation can result in minimizing toxic side effects in human applications. Samples generated from the processes described herein (paragraphs [0086] - [0097 ]) were analyzed to compare metabolite formation.

FIG. 9 compares Eldallas for D in the formation of oxidative metabolites formed by purine epoxidation on carbon-8₇-idelalisis. Results support D₇Idelalisib slows down metabolism that can cause toxicity in the treatment of human diseases.

IC50 measurement of PI3K δ

The study of IC50 to measure lipid kinase PI3K δ was performed on a uniform time resolved fluorescence (HTRF) platform. The PIP3 product was detected by displacement of biotin-PIP 3 from an energy transfer complex consisting of an europium-labeled anti-GST monoclonal antibody, a GST-labeled Pleckstrin Homology (PH) domain, biotinylated PIP3, and Streptavidin-Allophycocyanin (Streptavidin-Allophycocyanin, (APC)). Excitation of europium in the complex results in energy transfer to the APC and fluorescence emission at 665 nM. The PIP3 product formed by PI 3-kinase (h) activity displaces biotin-PIP 3 from the complex, resulting in a loss of energy transfer and thus a decrease in signal. The method comprises the following 3 steps: first, the kinase reaction with PIP2 substrate was performed in the presence of ATP, then the reaction was quenched with stop solution and finally detected by addition of detection mixture followed by incubation. The control inhibitor was PI-103. The emission ratio was converted to μm PIP3 production based on PIP3 standard curve. Nonlinear regression to obtain standard curves and IC50 values was performed using Graphpad Prism software.

Idelalisi and D₇Idelalisib was received as a 2mg/mL stock in DMSO. Two compounds were tested against 1 PI3K isoform. These compounds were tested in a 10 dose IC50 mode starting from a concentration of 2 μ Μ. The control compound PI-103 was tested in 10 dose IC50 at 3-fold serial dilutions starting at 10 μ Μ. The reaction was carried out at 10 μ M ATP. HTRF assay format was used for PI 3K. Curve fitting was performed when the enzyme activity at the highest concentration of compound was below 65%.

Idelalisi and D₇Calculated IC50 values for Idelalisib were 3.5nM and 1.1 nM.

TABLE 1IC50 of PI3K delta

Compound ID	IC50*(M)-PI3K δ
		Ai dallas si	3.52E-09
D7-Idelalisib	1.05E-09
		PI-103	4.54E-09

Applicants' disclosure is described herein in preferred embodiments with reference to the drawings, wherein like reference numerals designate identical or similar elements. Reference throughout this specification to "one embodiment," "an embodiment," or similar language means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases "in one embodiment," "in an embodiment," and similar language throughout this specification may, but do not necessarily, all refer to the same embodiment.

The described features, structures, or characteristics of the applicants' disclosure may be combined in any suitable manner in one or more embodiments. In the following description, numerous specific details are set forth in order to provide a thorough understanding of embodiments of the invention. One skilled in the relevant art will recognize, however, that applicants' compositions and/or methods can be practiced without the specific details, or with other methods, components, materials, and so forth. In other instances, well-known structures, materials, or operations are not shown or described in detail to avoid obscuring aspects of the disclosure.

In this specification and the appended claims, the singular forms "a", "an" and "the" include plural references unless the context clearly dictates otherwise.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present disclosure, the preferred methods and materials are now described. The methods described herein may be performed in any order that is logically possible, except in the particular order disclosed.

INCORPORATION BY REFERENCE

References to other documents, such as patents, patent applications, patent publications, periodicals, books, articles, web page content, are mentioned in this disclosure. All such documents are hereby incorporated by reference in their entirety for all purposes. Any material, or portion thereof, that is said to be incorporated by reference herein, but which conflicts with existing definitions, statements, or other disclosure material set forth herein will only be incorporated to the extent that no conflict arises between that incorporated material and the present disclosure material. In the event of a conflict, the conflict is resolved in favor of the present disclosure as the preferred disclosure.

Identity of

The representative examples are intended to aid in the description of the invention and are not intended, nor should they be construed, to limit the scope of the invention. Indeed, various modifications of the invention and many further embodiments thereof, in addition to those shown and described herein, will become apparent to those skilled in the art from the entirety of this document, including the examples and referenced scientific and patent documents included herein. The examples contain important additional information, exemplification and guidance which can be adapted to the practice of this invention in its various embodiments and equivalents thereof.

Claims (8)

1. A compound, or a pharmaceutically acceptable salt thereof, having the structural formula:

2. a pharmaceutical composition comprising a compound having the formula:

3. a unit dosage form comprising the pharmaceutical composition of claim 2.

4. Use of a pharmaceutical composition comprising a compound having the formula:

5. the use of claim 4, wherein the one or more hematological malignancies is selected from Chronic Lymphocytic Leukemia (CLL), follicular B-cell non-Hodgkin's lymphoma (FL), and Small Lymphocytic Lymphoma (SLL).

6. The use according to any one of claims 4 or 5, wherein the compound is administered in combination with one or more other anti-cancer agents.

7. The use of claim 6, wherein the one or more other anti-cancer agents is selected from a monoclonal antibody against CD20 protein.

8. The use of claim 7, wherein the one or more additional anti-cancer agents is rituximab.

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