WO2016079527A1 - Combination therapy - Google Patents

Abstract

A combination therapy comprising simultaneous or sequential administration of a Smac mimetic and an NSAID.

Description

Title

Combination Therapy

Field of the Invention

This invention is in the field of compositions and methods to treat proliferative disorders including cancers.

Background of the Invention

Inhibitors of Apoptosis Proteins (lAPs) are naturally occurring intra-cellular proteins that suppress caspase-dependent apoptosis. Second mitochondria- derived activator of caspases (Smac), also known as DIABLO, is an intracellular protein that functions to antagonize, i.e., inhibit, the activity of lAPs. In normal healthy cells, Smac and lAPs function together to maintain the viability of healthy cells. However, in certain disease states, e.g., cancers and other proliferative disorders, lAPs are not adequately antagonized and therefore prevent apoptosis and cause or exacerbate abnormal proliferation and survival.

Smac mimetics are synthetic small molecules that mimic the structure and IAP antagonist activity of the four N-terminal amino acids of Smac. When

administered to animals suffering proliferative disorders, the Smac mimetics antagonize lAPs, causing an increase in apoptosis among abnormally proliferating cells. Various Smac mimetics are in development for use in the treatment of proliferative disorders. Smac mimetics have also been shown to promote apoptosis in chronically infected cells while sparing uninfected cells and are in development for treatment of viral and other infections.

Non-steroidal anti-inflammatory drugs (NSAIDs) are a class of drugs that has analgesic, antipyretic, and anti-inflammatory properties. Examples include, without limitation celecoxib, rofecoxib, etodolac, diclofenac, meloxicam, indomethacin, naproxen, ibuprofen and aspirin. NSAIDs have been shown to induce apoptosis in colorectal cells. NSAIDs appear to induce apoptosis via the release of Smac from the mitochondria (PNAS, November 30, 2004, vol.

101 : 16897-16902). TNFa-related apoptosis-inducing ligand (TRAIL) is a cytokine that is produced and secreted by most normal tissue cells. TRAIL causes apoptosis in abnormally proliferating cells by binding to certain death receptors including DR4 and DR5. TRAIL receptor agonists, such as anti-DR4 antibodies, anti-DR5 antibodies, and anti-DR4/DR5 antibodies are under investigation as possible cancer

therapeutics.

IAP antagonists have been shown to enhance the cytotoxicity of TRAIL and combination therapies comprising co-administration of an IAP antagonist and a TRAIL receptor agonist are under investigation.

Summary of the Invention

This invention, in one aspect, is a method of treating a disorder that is amenable to treatment with an IAP antagonist in a mammalian subject, e.g., a human patient, by internally administering to the subject an effective amount of (1 ) an IAP antagonist, e.g., a Smac mimetic, (2) a TRAIL receptor agonist, e.g., an anti- DR4 antibody or an anti-DR5 antibody, and (3) an anti-inflammatory agent other than a TNFa inhibitor, e.g., a non-steroidal antiinflammatory drug (NSAID).

In a more specific illustrative embodiment, this invention is a method of treating a disorder that is amenable to treatment with an IAP antagonist in a mammalian subject, e.g., a human patient, by internally administering to the subject an effective amount of (1 ) an IAP antagonist, e.g., a Smac mimetic such as birinapant, and (2) an NSAID.

In other aspects, the invention comprises a method of treating a cancer, the symptoms of which disorder or disease can be ameliorated by pro-apoptotic therapy, in a mammalian subject in need thereof, e.g., a human, or a companion animal, a food animal, or a sporting animal, that comprises internally

administering to the subject an effective amount of a Smac mimetic such as birinapant and an effective amount of a TRAIL receptor agonist and also internally administering to the subject an effective amount of an anti-inflammatory agent that is not a TNFa inhibitor, e.g., an NSAID. The skilled person will understand that the invention in this or other embodiments does not require (but does not exclude) co-administration of a TRAIL receptor agonist.

In additional illustrative embodiments, the invention comprises such method that further comprises administering one or more additional cancer therapies, e.g., radiation, chemotherapy, immunotherapy, photodynamic therapy, and

combinations thereof in addition to the Smac mimetic such as birinapant and the NSAID.

In a further illustrative embodiment, the invention comprises a method of treating an autoimmune disease in a mammal in need thereof, including, for example, rheumatoid arthritis, systemic lupus erythematosus, psoriasis, and idiopathic thrombocytopenic purpura (Morbus Werlhof) that comprises internally

administering to the animal an effective amount of a Smac mimetic and of a TRAIL receptor agonist and also internally administering to the subject an effective amount of an anti-inflammatory agent that is not a TNFa inhibitor. In a more specific illustrative embodiment, this invention is a method of treating an autoimmune disease in a mammal in need thereof by internally administering to the subject an effective amount of (1 ) an IAP antagonist, e.g., a Smac mimetic, and (2) an NSAID.

In a further illustrative embodiment, the invention comprises a method of treating an intracellular infection, in particular, a chronic infection, in a mammal in need thereof, including, for example,

a virus selected from the group consisting of Human papillomaviruses, Herpes viruses including herpes simplex 1/2, varicella zoster, Epstein-Barr virus (EBV), cytomegalovirus (CMV), HHV-6/7, HTLV, Human

papovaviruses, including JC virus and BK virus, adeno and parvoviruses, HIV, HBV and HCV.

bacteria selected from the group consisting of Salmonella spp., Ehrlichia spp., Mycobacteria spp., Spirochetes, Legionella spp., Listeria spp., Rickettsia spp., Chlamydia spp., Mycoplasma spp., Coxiella spp., Yersinia spp., Francisella spp., Brucella spp., Neisseria spp, and Nocardia spp., fungus or yeast selected from the group consisting of Histoplasma spp., Aspergillus spp., Cryptococcus spp., and Pneunocystis jirovecii, protozoa selected from the group consisting of Trypanosomatids (e.g., Leishmania spp.), Apicomplexans, including liver forms of Plasmodium spp., Toxoplasma spp., and Cryptosporidium spp.,

among others, that comprises internally administering to the animal an effective amount of a Smac mimetic and of a TRAIL receptor agonist and also internally administering to the subject an effective amount of an anti-inflammatory agent that is not a TNFa inhibitor. The skilled person will understand that the invention does not require co-administration of a TRAIL receptor agonist.

In related aspects, the invention is directed to a method of preventing the occurrence or reducing the severity of adverse events associated with treatment of a subject with an IAP antagonist, e.g., a Smac mimetic, e.g., birinapant, or with co-treatment of a subject with an IAP antagonist and a TRAIL agonist, e.g., symptoms of a cranial nerve palsy such as VI cranial nerve palsy or Bell's palsy, that comprises co-treating the patient with an anti-inflammatory drug regimen (excluding TNFa inhibitors), e.g., an NSAID.

In related aspects, the invention comprises co-administration of an IAP antagonist, e.g., a Smac mimetic, and an anti-inflammatory agent other than a TNFa inhibitor, e.g., a NSAID. Such aspects comprise a method of reducing the incidence and/or severity of adverse events such as Bell's palsy or sixth nerve palsy (also known as abducens nerve palsy) associated with IAP antagonist therapy, e.g., Smac mimetic therapy, in a patient, said method comprising administering to the patient an effective amount of an anti-inflammatory agent (but not a TNFa inhibitor), e.g., an NSAID, while the patient is undergoing IAP antagonist therapy, e.g., Smac mimetic therapy.

In certain illustrative embodiments, the IAP antagonist is birinapant, which has the chemical name: N-{1 S-[2R-(6,6'-Difluoro-3'-{4S-hydroxy-1 -[2S-(2S- methylamino-propionylamino)-butyryl]-pyrrolidin-2R-ylmethyl}-1 H, 1 Ή- [2,2 biindolyl-3-ylmethyl)-4S-hydroxy-pyrrolidine-1 -carbonyl]-propyl}-2S- meth lamino-propionamide and which has the chemical formula:

wherein R5 is -CH2CH3, or a pharmaceutically acceptable salt thereof.

Birinapant is described as Compound 15 in US8603816. Structurally similar Smac mimetics are disclosed, e.g., in US7517906 and US8022230.

Detailed Description of the Invention

In accordance with this invention, a Smac mimetic alone or in combination with a TRAIL agonist, is used in the treatment of proliferative disorders, e.g.: various benign tumors or malignant tumors (cancer), benign proliferative diseases (e.g., psoriasis, benign prostatic hypertrophy, and restenosis), autoimmune diseases (e.g., autoimmune proliferative glomerulonephritis, lymphoproliferative

autoimmune responses), or infectious disease.

Some embodiments of the invention include inducing apoptosis of cells, particularly pathologically proliferating cells. The methods can be carried out in vitro or in vivo. The term, "coadministration," is not limited to simultaneous coadministration but more generally refers to a treatment regimen that comprises administration of an IAP antagonist (or an IAP antagonist and a TRAIL agonist) and an anti-inflammatory. Thus, e.g., the NSAID therapy may be initiated prior to (e.g., one day to one week prior to, or even longer), concurrently with (e.g., within moments before or after to within 24 hours before or after), or after (e.g. , one day to one week after) initiation of treatment with the IAP antagonist. Preferably, treatment with the anti-inflammatory is initiated prior to appearance of symptoms of a cranial nerve palsy but in some cases the NSAID therapy may not be initiated until after appearance of such symptoms, which could be several weeks or months after initiation of IAP antagonist therapy.

Without limiting the scope of this invention to a particular mechanism of action, it is believed that NSAIDs unexpectedly counteract the on-target mechanism of birinapant and thereby reduce the frequency and/or severity of cranial nerve palsies. Thus, an aspect of this invention is co-administration of an NSAID in order to counteract the IAP antagonist activity of a Smac mimetic (e.g. , the clAP degradative and XIAP inhibitory activities of a Smac mimetic discussed herein) while sufficiently preserving the pro-apoptotic effects of the Smac mimetic such that the Smac mimetic is useful in promoting apoptosis of abnormally proliferating (or infected) cells without occurrence of, or with reduced frequency and/or severity of, cranial nerve palsies.

The methods of the invention can include administration of an IAP antagonist, or coadministration of an IAP antagonist and a TRAIL agonist, with or without one or more additional IAP antagonists and with or without one or more additional active pharmaceutical ingredients. Administration of multiple agents can be simultaneous or sequential. Useful additional chemotherapeutic agents include, but are not limited to, alkylating agents (e.g., cyclophosphamide,

mechlorethamine, chlorambucil, melphalan), anthracyclines (e.g. , daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, valrubicin), cytoskeletal disruptors (e.g. , paclitaxel, docetaxel), epothilones (e.g., epothilone A, epothilone B, epothilone D), inhibitors of topoisomerase I and I I (e.g., irinotecan, topotecan, etoposide, teniposide, tafluposide), nucleotide analogs precursor analogs (e.g. , azacitidine, azathioprine, capecitabine, cytarabine, doxifluridine, fluorouracil, gemcitabine, mercaptopurine, methotrexate, tioguanine), peptide antibiotics (e.g. , bleomycin), platinum-based agents (e.g., carboplatin, cisplatin, oxaliplatin), retinoids (e.g. , all-trans retinoic acid), and vinca alkaloids and derivatives (e.g., vinblastine, vincristine, vindesine, vinorelbine). In some embodiments, the chemotherapeutic agents include fludarabine, doxorubicin, paclitaxel, docetaxel, camptothecin, etoposide, topotecan, irinotecan, cisplatin, carboplatin, oxaliplatin, amsacrine, mitoxantrone, 5-fluorouracil, or gemcitabine. Combination therapies can also employ such biological agents as a Type I or a Type III interferon, e.g., Interferon-a, lnterferon-β and/or lnterferon-λ.

In some embodiments, patients in need of Smac mimetic therapy are selected for co-administration of an NSAID as provided by this invention on the basis of risk of a cranial nerve palsy. Such patients include, e.g., patients with a history of cranial nerve palsy or symptoms thereof or of other cranial nerve disorders such as trigeminal neuralgia or symptoms thereof. Such patient may have exhibited such symptoms as a consequence of IAP antagonist therapy or otherwise or may have a family history or a genetic marker indicating predisposition or possible predisposition to a cranial nerve palsy (or trigeminal neuralgia), or symptoms of either or both.

In related aspects of the invention, patients receiving Smac mimetic therapy are monitored specifically for symptoms of a cranial nerve palsy. In some aspects, patients exhibiting such symptoms are then co-treated with an NSAID and/or have their dose of IAP antagonist reduced including possibly having the IAP antagonist therapy terminated or at least suspended for a period of time, e.g., until the symptoms resolve.

In all embodiments of this invention the cranial nerve palsy can be VI cranial nerve palsy or Bell's palsy. Adverse events associated with other cranial nerves, e.g., trigeminal neuralgia, can also be impacted by the practice of this invention.

IAP antagonists include, without limitation, the Smac mimetics disclosed in US 7,244,851 ; US 7,517,906; US 7,419,975; US 7,589, 1 18; US 7,932,382; US 7,345,081 ; US 7,244,851 ; US 7,674,787; US 7,772, 177; US 7,989,441 ; US 8, 163,792; US 8,278,293; US8445440; US8445473; US8552003; US85751 13; US 8,815,927; US8551955; US 8,716,236; US8835393 ; US 8,859,541 ; US 8,889,712; US 8,883,771 ; US8907092; US8993523; WO201 1098904; WO2012080260; WO2014009495; WO201401 1712; WO2014023708;

WO2014025759; WO2014026882; WO2014031487; WO2014044622;

WO2014047024; WO2014055461 ; WO2014056755; WO2014056867;

WO2014056871 ; WO2014060767; WO2014060768; WO2014060770;

WO2014074658; WO2014074665; WO2014085489; WO2014090709;

WO2015006524; WO2015071393; WO2015073072; WO2015092420;

WO2015109391.

Many IAP antagonists, but not all, are commonly within the genus of monovalent or bivalent Smac mimetics that have the general structure:

[P1 -P2-P3-P4] (Formula I)

or

[Ρ1 -Ρ2-Ρ3-Ρ4]-Ι_-[Ρ1 '-Ρ2'-Ρ3'-Ρ4'] (Formula II) wherein P1 -P2-P3- and P1 '-P2'-P3'- correspond to peptide replacements, i.e., peptidomimetics, of the N-terminal Ala-Val-Pro- tripeptide of mature Smac and P4 and P4' correspond to amino acid replacements of the fourth N-terminal amino acid, Phe, Tyr, lie, or Val, and L is a linking group or bond covalently linking [P1 -P2-P3-P4] to [Ρ1 '-Ρ2'-Ρ3'-Ρ4'].

For example, without limitation, a Smac mimetic may reside in the following genus of compounds of Formula I or of Formula II:

P1 and P1 ' are NHR¹-CHR²-C(0)-;

P2 and P2' are -NH-CHR³-C(0)-;

P3 and P3' are pyrrolidine, pyrrolidine fused to a cycloalkyi, or pyrrolidine fused to a heterocycloalkyl having a -N- heteroatom, optionally substituted in each case, and wherein the pyrrolidine of P3/P3' may be bound to P2/P2' by an amide bond;

P4 and P4' are -M-Q_p-R⁷.

The variable substituents can be, for example:

R¹: -H or -CH3; R²: C1 -6 alkyi, C1 -6 alkoxy, optionally substituted, e.g., -CH3, -CH2CH3 or - CH20H;

R³: C1 -6 alkyi, C1 -6 alkoxy, C3-C7 cycloalkyl or heterocycloalkyl, or C6-C8 aryl or heteroaryl, optionally substituted in each case;

M: a covalent bond, C1 -6 alkylene, substituted C1 -C6 alkylene such as but not limited to -C(O)-, or C3-C7 cycloalkyl or heterocycloalkyl, optionally substituted in each case;

Q: a covalent bond, C1 -6 alkylene, substituted C1 -C6 alkylene, -0-, -NR⁸- , or C3-C7 cycloalkyl or heterocycloalkyl, optionally substituted in each case;

P: 0 or 1 ;

R⁷: cycloalkyl, heterocycloalkyl, cycloalkylaryl, alkylaryl, alkyi heteroaryl, aryl or heteroaryl, optionally substituted in each case;

R⁸: -H or C1 -6 alkyi.

In compounds of Formula II, L is a linking group or bond covalently linking [P1 - P2-P3-P4] to [Ρ1 '-Ρ2'-Ρ3'-Ρ4'].

"Alkyi" (monovalent) and "alkylene" (divalent) when alone or as part of another term (e.g., alkoxy) mean branched or unbranched, saturated aliphatic

hydrocarbon group, having up to 12 carbon atoms unless otherwise specified. Examples of particular alkyi groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n- butyl, iso-butyl, sec-butyl, tert-butyl, n-pentyl, 2- methylbutyl, 2,2-dimethylpropyl, n-hexyl, 2- methylpentyl, 2,2-dimethylbutyl, n- heptyl, 3-heptyl, 2-methylhexyl, and the like. The term, "lower," when used to modify alkyi, alkenyl, etc., means 1 to 4 carbon atoms, branched or linear so that, e.g., the terms "lower alkyi", "C_rC alkyi" and "alkyi of 1 to 4 carbon atoms" are synonymous and used interchangeably to mean methyl, ethyl, 1 -propyl, isopropyl, 1 -butyl, sec-butyl or t-butyl. Examples of alkylene groups include, but are not limited to, methylene, ethylene, n-propylene, n-butylene and 2-methyl- butylene.

The term substituted alkyi refers to alkyi moieties having substituents replacing one or more hydrogens on one or more (often no more than four) carbon atoms of the hydrocarbon backbone. Such substituents are independently selected from the group consisting of: a halogen (e.g., I, Br, CI, or F, particularly fluoro(F)), hydroxy, amino, cyano, mercapto, alkoxy (such as a C₁-C6 alkoxy, or a lower (Ci- C ) alkoxy, e.g., methoxy or ethoxy to yield an alkoxyalkyl), aryloxy (such as phenoxy to yield an aryloxyalkyl), nitro, oxo (e.g., to form a carbonyl), carboxyl (which is actually the combination of an oxo and hydroxy substituent on a single carbon atom), carbamoyl (an aminocarbonyl such as NR₂C(0)-, which is the substitution of an oxo and an amino on a single carbon atom), cycloalkyi (e.g., a cycloalkylalkyl), aryl (resulting for example in aralkyls such as benzyl or phenylethyl), heterocyclylalkyl (e.g., heterocycloalkylalkyl), heteroaryl (e.g., heteroarylalkyl), alkylsulfonyl (including lower alkylsulfonyl such as

methylsulfonyl), arylsulfonyl (such as phenylsulfonyl), and -OCF₃ (which is a halogen substituted alkoxy). The invention further contemplates that several of these alkyl substituents, including specifically alkoxy, cycloalkyi, aryl,

heterocyclylalkyl and heteroaryl, are optionally further substituted as defined in connection with each of their respective definitions provided below. In addition, certain alkyl substituent moieties result from a combination of such substitutions on a single carbon atom. For example, an ester moiety, e.g., an alkoxycarbonyl such as methoxycarbonyl, or tert-butoxycarbonyl (Boc) results from such substitution. In particular, methoxycarbonyl and Boc are substituted alkyls that result from the substitution on a methyl group (-CH₃) of both an oxo (=0) and an unsubstituted alkoxy, e.g., a methoxy (CH₃-0) or a tert-butoxy ((CH₃)₃C-0-), respectively replacing the three hydrogens. Similarly, an amide moiety, e.g., an alkylaminocarbonyl, such as dimethylaminocarbonyl or methylaminocarbonyl, is a substituted alkyl that results from the substitution on a methyl group (-CH₃) of both an oxo (=0) and a mono-unsubstitutedalkylamino or,

diunsubstitutedalkylamino, e.g., dimethylamino (-N-(CH₃)₂), or methylamino (-NH- (CH₃)) replacing the three hydrogens (similarly an arylaminocarbonyl such as diphenylaminocarbonyl is a substituted alkyl that results from the substitution on a methyl group (-CH₃) of both an oxo (=0) and a mono- unsubstitutedaryl(phenyl)amino). Exemplary substituted alkyl groups further include cyanomethyl, nitromethyl, hydroxyalkyls such as hydroxymethyl, trityloxymethyl, propionyloxymethyl, aminoalkyls such as aminomethyl, carboxylalkyls such as carboxymethyl, carboxyethyl, carboxypropyl, 2,3- dichloropentyl, 3-hydroxy-5-carboxyhexyl, acetyl (e.g., an alkanoyl, where in the case of acetyl the two hydrogen atoms on the -CH₂ portion of an ethyl group are replaced by an oxo (=0)), 2-aminopropyl, pentachlorobutyl, trifluoromethyl, methoxyethyl, 3-hydroxypentyl, 4-chlorobutyl, 1 ,2-dimethyl-propyl,

pentafluoroethyl, alkyloxycarbonylmethyl, allyloxycarbonylaminomethyl, carbamoyloxymethyl, methoxymethyl, ethoxymethyl, t- butoxymethyl,

acetoxymethyl, chloromethyl, bromomethyl, iodomethyl, trifluoromethyl, 6- hydroxyhexyl, 2,4-dichloro (n-butyl), 2-amino (iso-propyl), cycloalkylcarbonyl (e.g., cyclopropylcarbonyl) and 2-carbamoyloxyethyl. Particular substituted alkyls are substituted methyl groups. Examples of substituted methyl group include groups such as hydroxymethyl, protected hydroxymethyl (e.g.,

tetrahydropyranyl- oxymethyl), acetoxymethyl, carbamoyloxymethyl,

trifluoromethyl, chloromethyl, carboxymethyl, carboxyl (where the three hydrogen atoms on the methyl are replaced, two of the hydrogens are replaced by an oxo (=0) and the other hydrogen is replaced by a hydroxy (-OH)), tert-butoxycarbonyl (where the three hydrogen atoms on the methyl are replaced, two of the hydrogens are replaced by an oxo (=0) and the other hydrogen is replaced by a tert-butoxy (-0-C(CH₃)₃), bromomethyl and iodomethyl. When the specification and especially the claims refer to a particular substituent for an alkyl, that substituent can potentially occupy one or more of the substitutable positions on the alkyl. For example, reciting that an alkyl has a fluoro substituent, would embrace mono-, di-, and possibly a higher degree of substitution on the alkyl moiety.

The term substituted alkylene refers to alkylene moieties having substituents replacing one or more hydrogens on one or more (often no more than four) carbon atoms of the hydrocarbon backbone where the alkylene is similarly substituted with groups as set forth above for alkyl.

Alkoxy is -O-alkyl. A substituted alkoxy is -O-substituted alkyl, where the alkoxy is similarly substituted with groups as set forth above for alkyl. One substituted alkoxy is acetoxy where two of the hydrogens in ethoxy (e.g., -0-CH₂-CH₃) are replaced by an oxo, (=0) to yield -0-C(0)-CH₃; another is an aralkoxy where one of the hydrogens in the alkoxy is replaced by an aryl, such as benzyloxy, and another is a carbamate where two of the hydrogens on methoxy (e.g., -0-CH₃) are replaced by oxo (=0) and the other hydrogen is replaced by an amino (e.g., - NH₂, -NHR or -NRR) to yield, for example, -0-C(0)-NH₂. A lower alkoxy is -0- lower alkyl.

"Alkenyl" (monovalent) and "alkenylene" (divalent) when alone or as part of another term mean an unsaturated hydrocarbon group containing at least one carbon-carbon double bond, typically 1 or 2 carbon-carbon double bonds, which may be linear or branched and which have at least 2 and up to 12 carbon atoms unless otherwise specified. Representative alkenyl groups include, by way of example, vinyl, allyl, isopropenyl, but-2-enyl, n-pent-2-enyl, and n-hex-2-enyl.

The terms substituted alkenyl and substituted alkenylene refer to alkenyl and alkenylene moieties having substituents replacing one or more hydrogens on one or more (often no more than four) carbon atoms of the hydrocarbon backbone. Such substituents are independently selected from the group consisting of: halo (e.g., I, Br, CI, F), hydroxy, amino, cyano, alkoxy (such as C_rC₆ alkoxy), aryloxy (such as phenoxy), nitro, mercapto, carboxyl, oxo, carbamoyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, alkylsulfonyl, arylsulfonyl and -OCF₃.

"Alkynyl" means a monovalent unsaturated hydrocarbon group containing at least one carbon-carbon triple bond, typically 1 carbon-carbon triple bond, which may be linear or branched and which have at least 2 and up to 12 carbon atoms unless otherwise specified. Representative alkynyl groups include, by way of example, ethynyl, propargyl, and but-2-ynyl.

"Cycloalkyl" when alone or as part of another term means a saturated or partially unsaturated cyclic aliphatic hydrocarbon group (carbocycle group), having 3 to 8 carbon atoms unless otherwise specified, such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl, and further includes polycyclic, including fused cycloalkyls such as 1 ,2,3,4-tetrahydonaphthalenyls (1 ,2,3,4-tetrahydonaphthalen- 1 -yl, and 1 ,2,3,4-tetrahydonaphthalen-2-yl), indanyls (indan-1yl, and indan-2-yl), isoindenyls (isoinden-1 -yl, isoinden-2-yl, and isoinden-3-yl) and indenyls (inden- 1 -yl, inden-2-yl and inden-3-yl). A lower cycloalkyl has from 3 to 6 carbon atoms and includes cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

The term substituted cycloalkyl refers to cycloalkyl moieties having substituents replacing one or more hydrogens on one or more (often no more than four) carbon atoms of the hydrocarbon backbone. Such substituents are

independently selected from the group consisting of: halo (e.g., I, Br, CI, F), hydroxy, amino, cyano, alkoxy (such as C_rC₆ alkoxy), substituted alkoxy, aryloxy (such as phenoxy), nitro, mercapto, carboxyl, oxo, carbamoyl, alkyl, substituted alkyls such as trifluoromethyl, aryl, substituted aryls, heterocyclyl, heteroaryl, alkylsulfonyl, arylsulfonyl and -OCF₃. When the specification and especially the claims refer to a particular substituent for a cycloalkyl, that substituent can potentially occupy one or more of the substitutable positions on the cycloalkyl. For example, reciting that a cycloalkyl has a fluoro substituent, would embrace mono-, di-, and a higher degree of substitution on the cycloalkyl moiety.

Examples of cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, tetrahydronaphthyl and indanyl.

"Aryl" when used alone or as part of another term means an aromatic carbocyclic group whether or not fused having the number of carbon atoms designated, or if no number is designated, from 6 up to 14 carbon atoms. Particular aryl groups include phenyl, naphthyl, biphenyl, phenanthrenyl, naphthacenyl, indolyl, and the like (see e. g. Lang's Handbook of Chemistry (Dean, J. A., ed) 13^th ed. Table 7-2 [1985]).

The term substituted aryl refers to aryl moieties having substituents replacing one or more hydrogens on one or more (usually no more than six) carbon atoms of the aromatic hydrocarbon core. Such substituents are independently selected from the group consisting of: halo (e.g., I, Br, CI, F), hydroxy, amino, cyano, alkoxy (such as C_rC₆ alkoxy and particularly lower alkoxy), substituted alkoxy, aryloxy (such as phenoxy), nitro, mercapto, carboxyl, carbamoyl, alkyl, substituted alkyl (such as trifluoromethyl), aryl, -OCF₃, alkylsulfonyl (including lower alkylsulfonyl), arylsulfonyl, heterocyclyl and heteroaryl. Examples of such substituted phenyls include but are not limited to a mono-or di (halo) phenyl group such as 2-chlorophenyl, 2- bromophenyl, 4-chlorophenyl, 2,6- dichlorophenyl, 2,5-dichlorophenyl, 3,4-dichlorophenyl, 3-chlorophenyl, 3- bromophenyl, 4-bromophenyl, 3,4-dibromophenyl, 3-chloro-4-fluorophenyl, 2- fluorophenyl; 3-fluorophenyl, 4-fluorophenyl, a mono-or di (hydroxy) phenyl group such as 4-hydroxyphenyl, 3- hydroxyphenyl, 2,4-dihydroxyphenyl, the protected- hydroxy derivatives thereof; a nitrophenyl group such as 3-or 4-nitrophenyl; a cyanophenyl group, for example, 4-cyanophenyl; a mono-or di (lower alkyl) phenyl group such as 4-methylphenyl, 2,4-dimethylphenyl, 2- methylphenyl, 4- (iso-propyl) phenyl, 4-ethylphenyl, 3- (n-propyl) phenyl; a mono or di (alkoxy) phenyl group, for example, 3,4-dimethoxyphenyl, 3-methoxy-4-benzyloxyphenyl, 3- methoxy-4- (1 -chloromethyl) benzyloxy-phenyl, 3-ethoxy phenyl, 4- (isopropoxy) phenyl, 4- (t-butoxy) phenyl, 3-ethoxy-4-methoxy phenyl; 3-or 4- trifluoromethylphenyl; a mono- or dicarboxyphenyl or (protected carboxy) phenyl group such 4-carboxy phenyl,; a mono-or di (hydroxymethyl) phenyl or (protected hydroxymethyl) phenyl such as 3- (protected hydroxymethyl) phenyl or 3,4-di (hydroxymethyl) phenyl; a mono-or di (aminomethyl) phenyl or (protected aminomethyl) phenyl such as 2- (aminomethyl) phenyl or 2, 4- (protected aminomethyl) phenyl; or a mono-or di (N- (methylsulfonylamino)) phenyl such as

3- (N- methylsulfonylamino) phenyl. Also, the substituents, such as in a disubstituted phenyl group, can be the same or different, for example, 3-methyl-

4- hydroxyphenyl, 3- chloro-4-hydroxyphenyl, 2-methoxy-4-bromophenyl, 4-ethyl- 2-hydroxy phenyl, 3-hydroxy-4- nitrophenyl, 2-hydroxy-4-chlorophenyl, as well as for trisubstituted phenyl groups where the substituents are different, as for example 3-methoxy-4-benzyloxy-6-methyl sulfonylamino, 3- methoxy-4- benzyloxy-6-phenyl sulfonylamino, and tetrasubstituted phenyl groups where the substituents are different such as 3-methoxy-4-benzyloxy-5-methyl-6-phenyl sulfonylamino. Particular substituted phenyl groups are 2-chlorophenyl, 2- aminophenyl, 2-bromophenyl, 3- methoxyphenyl, 3-ethoxy-phenyl, 4- benzyloxyphenyl, 4-methoxyphenyl, 3-ethoxy-4- benzyloxyphenyl, 3,4- diethoxyphenyl, 3-methoxy-4-benzyloxyphenyl, 3-methoxy-4- (1 - chloromethyl) benzyloxy-phenyl, 3-methoxy-4- (1 -chloromethyl) benzyloxy-6-methyl sulfonyl aminophenyl groups. When the specification and especially the claims refer to a particular substituent for an aryl, that substituent can potentially occupy one or more of the substitutable positions on the aryl. For example, reciting that an aryl has a fluoro substituent, would embrace mono-, di-, tri, tetra and a higher degree of substitution on the aryl moiety. Fused aryl rings may also be substituted with the substituents specified herein, for example with 1 , 2 or 3 substituents, in the same manner as substituted alkyl groups. The terms aryl and substituted aryl do not include moieties in which an aromatic ring is fused to a saturated or partially unsaturated aliphatic ring.

"Heterocyclic group", "heterocyclic", "heterocycle", "heterocyclyl",

"heterocycloalkyl" or "heterocyclo" alone and when used as a moiety in a complex group, are used interchangeably and refer to any mono-, bi-, or tricyclic, saturated or unsaturated, non-aromatic hetero-atom-containing ring system having the number of atoms designated, or if no number is specifically

designated then from 5 to about 14 atoms, where the ring atoms are carbon and at least one heteroatom and usually not more than four heteroatoms (i.e., nitrogen, sulfur or oxygen). Included in the definition are any bicyclic groups where any of the above heterocyclic rings are fused to an aromatic ring (i.e., an aryl (e.g., benzene) or a heteroaryl ring). In a particular embodiment the group incorporates 1 to 4 heteroatoms. Typically, a 5- membered ring has 0 to 1 double bonds and a 6-or 7-membered ring has 0 to 2 double bonds and the nitrogen or sulfur heteroatoms may optionally be oxidized (e. g. SO, S0₂), and any nitrogen heteroatom may optionally be quaternized. Particular unsubstituted non-aromatic heterocycles include morpholinyl (morpholino), pyrrolidinyls, oxiranyl, indolinyls, 2,3-dihydoindolyl, isoindolinyls, 2,3-dihydoisoindolyl, tetrahydroquinolinyls, tetrahydroisoquinolinyls, oxetanyl, tetrahydrofuranyls, 2,3- dihydrofuranyl, 2H-pyranyls, tetrahydropyranyls, aziridinyls, azetidinyls, 1 -methyl- 2-pyrrolyl, piperazinyls and piperidinyls. The term substituted heterocyclo refers to heterocyclo moieties having substituents replacing one or more hydrogens on one or more (usually no more than six) atoms of the heterocyclo backbone. Such substituents are

independently selected from the group consisting of: halo (e.g., I, Br, CI, F), hydroxy, amino, cyano, alkoxy (such as C1-C6 alkoxy), substituted alkoxy, aryloxy (such as phenoxy), nitro, carboxyl, oxo, carbamoyl, alkyl, substituted alkyl (such as trifluoromethyl), -OCF₃, aryl, substituted aryl, alkylsulfonyl (including lower alkylsulfonyl), and arylsulfonyl. When the specification and especially the claims refer to a particular substituent for a heterocycloalkyl, that substituent can potentially occupy one or more of the substitutable positions on the

heterocycloalkyl. For example, reciting that a heterocycloalkyl has a fluoro substituent, would embrace mono-, di-, tri, tetra and a higher degree of substitution on the heterocycloalkyl moiety.

"Heteroaryl" alone and when used as a moiety in a complex group refers to any mono-, bi-, or tricyclic aromatic ring system having the number of atoms designated, or if no number is specifically designated then at least one ring is a 5-, 6-or 7-membered ring and the total number of atoms is from 5 to about 14 and containing from one to four heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur (Lang's Handbook of Chemistry, supra). Included in the definition are any bicyclic groups where any of the above heteroaryl rings are fused to a benzene ring. The following ring systems are examples of the heteroaryl groups denoted by the term "heteroaryl": thienyls (alternatively called thiophenyl), furyls, imidazolyls, pyrazolyls, thiazolyls, isothiazolyls, oxazolyls, isoxazolyls, triazolyls, thiadiazolyls, oxadiazolyls, tetrazolyls, thiatriazolyls, oxatriazolyls, pyridyls, pyrimidinyls (e.g., pyrimidin-2-yl), pyrazinyls, pyridazinyls, thiazinyls, oxazinyls, triazinyls, thiadiazinyls, oxadiazinyls, dithiazinyls, dioxazinyls, oxathiazinyls, tetrazinyls, thiatriazinyls, oxatriazinyls, dithiadiazinyls, imidazolinyls, dihydropyrimidyls, tetrahydropyrimidyls, tetrazolo [1 , 5-b] pyridazinyl and purinyls, as well as benzo-fused derivatives, for example benzoxazolyls, benzofuryls, benzothienyls, benzothiazolyls, benzothiadiazolyl, benzotriazolyls, benzoimidazolyls, isoindolyls, indazolyls, indolizinyls, indolyls, naphthyridines, pyridopyrimidines, phthalazinyls, quinolyls, isoquinolyls and quinazolinyls.

The term substituted heteroaryl refers to heteroaryl moieties (such as those identified above) having substituents replacing one or more hydrogens on one or more (usually no more than six) atoms of the heteroaryl backbone. Such substituents are independently selected from the group consisting of: halo (e.g., I, Br, CI, F), hydroxy, amino, cyano, alkoxy (such as C_rC₆ alkoxy), aryloxy (such as phenoxy), nitro, mercapto, carboxyl, carbamoyl, alkyl, substituted alkyl (such as trifluoromethyl), -OCF₃, aryl, substituted aryl, alkylsulfonyl (including lower alkylsulfonyl), and arylsulfonyl. When the specification and especially the claims refer to a particular substituent for a heteroaryl, that substituent can potentially occupy one or more of the substitutable positions on the heteroaryl. For example, reciting that a heteroaryl has a fluoro substituent, would embrace mono-, di-, tri, tetra and a higher degree of substitution on the heteroaryl moiety. Particular "heteroaryls" (including "substituted heteroaryls") include; 1 H- pyrrolo[2,3-£>]pyridine, 1 , 3-thiazol-2-yl, 4- (carboxymethyl)-5-methyl-1 , 3- thiazol- 2-yl, 1 ,2,4-thiadiazol-5-yl, 3- methyl-1 , 2,4-thiadiazol-5-yl, 1 ,3,4-triazol-5-yl, 2- methyl-1 ,3,4-triazol-5-yl, 2-hydroxy-1 ,3,4- triazol-5-yl, 2-carboxy-4-methyl-1 ,3,4- triazol-5-yl , 1 , 3-oxazol-2-yl, 1 , 3,4-oxadiazol-5-yl, 2-methyl-1 , 3,4-oxadiazol-5-yl, 2- (hydroxymethyl)- 1 , 3,4-oxadiazol-5-yl, 1 , 2,4-oxadiazol-5-yl, 1 , 3,4-thiadiazol- 5-yl, 2-thiol-1 , 3,4-thiadiazol-5-yl, 2- (methylthio)-l , 3,4-thiadiazol-5-yl, 2-amino-1 , 3,4-thiadiazol-5-yl, 1 H-tetrazol-5-yl, 1 -methyl-1 H- tetrazol-5-yl, 1 -(1 - (dimethylamino) eth-2-yl)-1 H-tetrazol-5-yl, 1 -(carboxymethyl)-1 H-tetrazol-5-yl, I- (methylsulfonic acid)-IH-tetrazol-5-yl, 2-methyl-IH-tetrazol-5-yl, 1 , 2,3-triazol-5-yl, 1 -methyl-1 , 2,3-triazol-5-yl, 2-methyl-1 , 2,3-triazol-5-yl, 4-methyl-1 , 2,3-triazol-5- yl, pyrid-2-yl N- oxide, 6-methoxy-2- (n-oxide)-pyridaz-3-yl, 6-hydroxypyridaz-3-yl, 1 -methylpyrid-2-yl, 1 - methylpyrid-4-yl, 2-hydroxypyrimid-4-yl, 1 ,4, 5,6-tetrahydro- 5, 6-dioxo-4-methyl-as-thazin-3-yl, 1 , 4,5, 6-tetrahydro-4- (formylmethyl)-5, 6- dioxo-as-triazin-3-yl, 2,5-dihydro-5-oxo-6-hydroxy- astriazin-3-yl, 2,5-dihydro-5- oxo-6-hydroxy-as-triazin-3-yl , 2,5-dihydro-5-oxo-6- hydroxy-2-methyl-astriazin-3- yl , 2,5-dihydro-5-oxo-6-hydroxy-2-methyl-as-triazin-3-yl, 2,5-dihydro-5-oxo-6- methoxy-2-methyl-as-triazin-3-yl, 2,5-dihydro-5-oxo-as-triazin-3-yl, 2,5- dihydro- 5-oxo-2-methyl-as-triazin-3-yl, 2,5-dihydro-5-oxo-2, 6-dimethyl-as-triazin-3-yl, tetrazolo [1 , 5-b] pyridazin-6-yl, 8-aminotetrazolo [1 , 5-b] -pyridazin-6-yl, quinol-2- yl, quinol-3-yl, quinol-4-yl, quinol-5-yl, quinol-6-yl, quinol-8-yl, 2-methyl-quinol-4- yl, 6-fluoro-quinol-4-yl, 2-methyl,8-fluoro-quinol-4-yl, isoquinol-5-yl, isoquinol-8-yl, isoquinol-1 -yl, and quinazolin-4-yl. An alternative group of "heteroaryl" includes: 5-methyl-2-phenyl-2H-pyrazol-3-yl, 4- (carboxymethyl)-5-methyl-1 , 3-thiazol-2-yl, 1 , 3,4-triazol-5-yl, 2-methyl-1 , 3,4-triazol-5-yl, 1 H-tetrazol-5-yl, 1 -methyl-1 H- tetrazol-5-yl, 1 -(1 -(dimethylamino) eth-2 -y I) -I H -tetrazo l-5-y I , l-(carboxymethyl)- 1 H-tetrazol-5-yl, 1 - (methylsulfonic acid)-IH- tetrazol-5-yl, 1 , 2,3-triazol-5-yl, 1 ,4, 5,6- tetrahydro-5,6-dioxo-4-methyl-as-triazin-3-yl, 1 , 4,5, 6-tetrahydro-4- (2- formylmethyl)-5, 6-dioxo- as-triazin-3-yl, 2, 5-dihydro-5-oxo-6-hydroxy-2-methyl- as-triazin-3-yl , 2,5-dihydro-5- oxo-6-hydroxy-2-methyl-as-triazin-3-yl, tetrazolo [1 , 5-b] pyridazin-6-yl, and 8-aminotetrazolo [1 , 5- b] pyridazin-6-yl.

L is a linking group or a bond covalently linking one monomer, [P1 -P2-P3-P4] to the other monomer, [P1 '-P2'-P3'-P4']. Commonly, -L- links P2 to P2' position such as at R3 or P4 to P4' such as at M, G, Q, or R⁷, or both P2 to P2' and P4 to P4'. L, therefore, can be a single or double covalent bond or a contiguous chain, branched or unbranched, substituted or unsubstituted, of 1 to about 100 atoms, typically 1 to about 30 atoms, e.g., an optionally substituted alkylene, alkenylene, alkylyne, cycloalkyl, alkylcycloalkyl, alkylarylalkyl chain of 2 to 20 atoms optionally with 1 -4 heteroatoms selected from -0-, -NH-, and -S-. Illustrative examples of L are a single or double covalent bond, C1 -12 alkylene, substituted C1 -12 alkylene, C1 -12 alkenylene, substituted C1 -12 alkenylene, C1 -12 alkynylene, substituted C1 -12 alkynylene, X_n-phenyl-Y_n, or X_n-(phenyl)₂-Y_n, wherein X and Y are independently C1 -6 alkylene, substituted C1 -6 alkylene, C1 - 6 alkenylene, substituted C1 -6 alkenylene, C1 -6 alkynylene, substituted C1 -6 alkynylene, or S(0)₂. Illustrative P3/P3' groups include, without limitation:

wherein

R⁶ is -H, C1 -6 alkyl, substituted C1 -6 alkyl, C1 -6 alkoxy, substituted C1 -6 alkoxy, C1 -6 alkylsulfonyl, arylsulfonyl, cycloalkyi, substituted cycloalkyi,

heterocycloalkyi, substituted heterocycloalkyi, aryl, substituted aryl, heteroaryl, or substituted heteroaryl;

R⁴ , R⁵, and R¹² are, independently, -H, -OH, C1 -6 alkyl, 01 -6 heteroalkyl, 01 -6 alkoxy, aryloxy, cycloalkyi, heterocycloalkyi, aryl, 01 -6 alkyl aryl, or heteroaryl, or 01 -6 alkyl heteroaryl, optionally substituted in each case except when R⁴ is -H or -OH.

As mentioned, in certain illustrative embodiments, the Smac mimetic used in the practice of the invention is bivalent.

Compound 15, i.e., birinapant, is an example of a specific Smac mimetic. Other illustrative examples are:

In certain illustrative embodiments, a selected Smac mimetic is a small molecule that binds to a BIR domain of at least XIAP, clAP-1 , and clAP-2, leading to ubiquitination (also, referred to as ubiquitylation) and degradation of the clAPs. In illustrative embodiments, the Smac mimetic does not inhibit NOD signaling or, if it does, then it does so only poorly. Because NOD signaling is dependent upon the E3 ubiquitin ligase activity of XIAP and mediates NF-kB activation, such Smac mimetic can be characterized as (i) not inhibiting XIAP E3 ubiquitin ligase activity or as only poorly inhibiting XIAP E3 ubiquitin ligase activity, (ii) not inhibiting or poorly inhibiting NOD (i.e., NOD1/2) signaling, or (iii) not inhibiting or poorly inhibiting NOD-mediated NF-kB activation. See, e.g., US20140303090.

Inhibition of NOD1/2-mediated NF-kB activation can be measured, e.g., in cell- based assays including such assays in which the NF-kB promoter is linked to a reporter gene, e.g., green fluorescent protein or luciferase. For example, in a NOD1/2 signaling assay, such as the DAP-stimulated and MDP-stimulated luciferase reporter gene assays described below, a Smac mimetic useful in the present invention when contacted with cells as 10 uM will reduce photon emission by no more than 50%, or about 50%. As photon emission in this assay is a surrogate for NOD1/2 signaling, such useful Smac mimetics will inhibit NOD1/2 signaling by no more than about 50% when tested at 10 uM

concentration. In terms of XIAP E3 ubiquitin ligase activity and NOD1/2-mediated NF-kB activation, such useful Smac mimetics will inhibit XIAP E3 ubiquitin ligase activity or NOD1/2-mediated NF-kB activation by no more than about 50%.

In other illustrative embodiments, such inhibition of reporter gene expression, NOD1/2 signaling, XIAP E3 ubiquitin ligase activity, or NOD1/2-mediated NF-kB activation does not exceed about 35%, or even about 25%. (i.e., reporter gene expression, NOD1/2 signaling, XIAP E3 ubiquitin ligase activity and NOD1/2- mediated NF-kB activation in the treated cells are greater than about 65%, e.g., greater than about 75%, of the level of activity observed in untreated cells.)

In certain illustrative embodiments, a selected Smac mimetic derepresses XIAP- mediated caspase-3 repression. In illustrative embodiments, the Smac mimetic degrades clAP-1 not bound to TRAF2 (non TRAF2-bound, e.g., "cytoplasmic" clAP-1 or "free" clAP-1 ) as well as clAP1 bound to TRAF2 and/or degrades clAP- 2 bound to TRAF2 but does not degrade clAP-2 not bound to TRAF2 or weakly degrades clAP-2 not bound to TRAF2 relative to degradation of clAP-2 bound to TRAF2.

For purposes of describing this invention, lAP antagonists also include molecules that reduce the expression of an lAP gene, such as clAP1 or clAP2. Suitable antagonists that are capable of reducing the expression of an lAP gene would be known to persons skilled in the art. Examples include nucleic acid molecules, such as RNA or DNA molecules (including double-stranded or single-stranded), and peptides, such as antisense peptide nucleic acids, that interfere with the expression of the target gene.

Useful DNA molecules include antisense, as well as sense (e.g. coding and/or regulatory) DNA molecules. Antisense DNA molecules include short

oligonucleotides. Persons skilled in the art would be able to design suitable short oligonucleotides for use in accordance with the present invention. An example is the XIAP antisense oligonucleotide, AEG35156, as described by Carter et al. [Apoptosis, 201 1 Vol.16(1 ):67-74). Other examples of useful DNA molecules include those encoding interfering RNAs, such as shRNA and siRNA. Yet another example are catalytic DNA molecules known as DNAzymes.

Useful RNA molecules capable of reducing the expression of an lAP gene, also referred to herein as RNA interference molecules, include siRNA, dsRNA, stRNA, shRNA, and miRNA (e.g., short temporal RNAs and small modulatory RNAs) and ribozymes.

RNA interference (RNAi) is particularly useful for specifically inhibiting the production of a particular protein. Although not wishing to be limited by theory, Waterhouse et al. (1998) have provided a model for the mechanism by which dsRNA can be used to reduce protein production. This technology relies on the presence of dsRNA molecules that contain a sequence that is essentially identical to the mRNA of the gene of interest or part thereof, in this case an mRNA encoding a polypeptide according to the invention. Conveniently, the dsRNA can be produced from a single promoter in a recombinant vector or host cell, where the sense and anti-sense sequences are flanked by an unrelated sequence which enables the sense and anti-sense sequences to hybridize to form the dsRNA molecule with the unrelated sequence forming a loop structure. The design and production of suitable dsRNA molecules for the present invention is well within the capacity of a person skilled in the art, particularly considering Waterhouse et al. (1998), Smith et al. (2000), WO 99/32619, WO 99/53050, WO 99/49029, and WO 01/34815.

In one example, a DNA is introduced that directs the synthesis of an at least partly double stranded RNA product(s) with homology to the target gene to be inactivated. The DNA therefore comprises both sense and antisense sequences that, when transcribed into RNA, can hybridize to form the double-stranded RNA region. In a preferred embodiment, the sense and antisense sequences are separated by a spacer region that comprises an intron which, when transcribed into RNA, is spliced out. This arrangement has been shown to result in a higher efficiency of gene silencing. The double-stranded region may comprise one or two RNA molecules, transcribed from either one DNA region or two. The presence of the double stranded molecule is thought to trigger a response from the cell that destroys both the double stranded RNA and also the homologous RNA transcript from the target gene, efficiently reducing or eliminating the activity of the target gene.

The length of the sense and antisense sequences that hybridize should each be at least 19 contiguous nucleotides, preferably at least 30 or 50 nucleotides, and more preferably at least 100, 200, 500 or 1000 nucleotides. The full-length sequence corresponding to the entire gene transcript may be used. The lengths are most preferably 100-2000 nucleotides. The degree of identity of the sense and antisense sequences to the targeted transcript should be at least 85%, preferably at least 90% and more preferably 95-100%. The RNA molecule may of course comprise unrelated sequences which may function to stabilize the molecule. The RNA molecule may be expressed under the control of a RNA polymerase II or RNA polymerase III promoter. Examples of the latter include tRNA or snRNA promoters. Preferred small interfering RNA ('siRNA") molecules comprise a nucleotide sequence that is identical to about 19-21 contiguous nucleotides of the target mRNA. Preferably, the target mRNA sequence commences with the dinucleotide AA, comprises a GC-content of about 30-70% (preferably, 30-60%, more preferably 40-60% and more preferably about 45%-55%), and does not have a high percentage identity to any nucleotide sequence other than the target in the genome of the cell in which it is to be introduced, e.g., as determined by standard BLAST search.

Synthesis of RNAi molecules suitable for use with present invention can be effected by first scanning the mRNA sequence of the target downstream of the AUG start codon for AA dinucleotide sequences. Occurrence of each AA and the 3' adjacent 19 nucleotides is recorded as potential siRNA target sites.

Preferably, siRNA target sites are selected from the open reading frame.

Second, potential target sites are compared to an appropriate genomic database using any sequence alignment software, such as BLAST. Putative target sites which exhibit significant homology to other coding sequences are filtered out. Qualifying target sequences are selected as template for siRNA synthesis.

Preferred sequences are those including low G/C content as these have proven to be more effective in mediating gene silencing as compared to those with G/C content higher than 55%. Several target sites are preferably selected along the length of the target gene for evaluation.

MicroRNA regulation is a clearly specialized branch of the RNA silencing pathway that evolved towards gene regulation, diverging from conventional RNAi/PTGS. MicroRNAs are a specific class of small RNAs that are encoded in gene-like elements organized in a characteristic inverted repeat. When transcribed, microRNA genes give rise to stem-looped precursor RNAs from which the microRNAs are subsequently processed. MicroRNAs are typically about 21 nucleotides in length. The released miRNAs are incorporated into RISC-like complexes containing a particular subset of Argonaute proteins that exert sequence-specific gene repression (see, for example, Millar and

Waterhouse, 2005; Pasquinelli et al., 2005; Almeida and Allshire, 2005). DNAzymes are single-stranded polynucleotides which are capable of cleaving single and double stranded polynucleotides which are capable of cleaving both single and double stranded target sequences (Breaker, R. R. and Joyce, G. Chemistry and Biology 1995; 2:655; Santoro, S. W. & Joyce, G. F. Proc. Natl, Acad. Sci. USA 1997; 943:4262) A general model (the "10-23" model) for the DNAzyme has been proposed. "10-23" DNAzymes have a catalytic domain of 15 deoxynbonucleotides, flanked by two substrate-recognition domains of seven to nine deoxynbonucleotides each. This type of DNAzyme can effectively cleave its substrate RNA at purine:pyrimidine junctions (Santoro, S. W. & Joyce, G. F. supra; for rev of DNAzymes see Khachigian, L M. Curr Opin Mol Ther 4: 1 19-21 (2002).

Examples of construction and amplification of synthetic, engineered DNAzymes recognizing single and double-stranded target cleavage sites have been disclosed in U.S. Pat. No. 6,326,174 to Joyce et al.

The terms "double stranded RNA" or "dsRNA" refer to RNA molecules that are comprised of two strands. Double-stranded molecules include those comprised of a single RNA molecule that doubles back on itself to form a two-stranded structure. For example, the stem loop structure of the progenitor molecules from which the single-stranded miRNA is derived, called the pre-miRNA, comprises a dsRNA molecule.

Other suitable RNA interference molecules include unmodified and modified double stranded (ds) RNA molecules including, short-temporal RNA (stRNA), small interfering RNA (si RNA), short-hairpin RNA (shRNA), microRNA (miRNA) and double-stranded RNA (dsRNA). The dsRNA molecules (e.g. siRNA) also may contain 3' overhangs, such as 3'UU or 3TT overhangs.

In an embodiment, the siRNA molecules of the present invention have a double stranded structure. In an embodiment, the siRNA molecules of the present invention are double stranded for more than about 25%, more than about 50%, more than about 60%, more than about 70%, more than about 80%, more than about 90% of their length. As used herein, "gene silencing" induced by RNA interference refers to a decrease in the mRNA level in a cell for a target gene (e.g., clAP1 gene and/or clAP2 gene) by at least about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 99%, about 100% of the mRNA level found in the cell in the absence of RNA interference.

The RNA interference molecules also include modified RNA molecules having one or more non-natural nucleotides; that is, nucleotides other than adenine "A", guanine "G", uracil "U", or cytosine "C". A modified nucleotide residue or a derivative or analog of a natural nucleotide may also be used. Any modified residue, derivative or analog may be used to the extent that it does not eliminate or substantially reduce (by at least 50%) RNAi activity of the molecule. Examples of suitable modified residues include aminoallyl UTP, pseudo-UTP, 5-l-UTP, 5-I- CTP, 5-Br-UTP, alpha-S ATP, alpha-S CTP, alpha-S GTP, alpha-S UTP, 4-thio UTP, 2-thio-CTP, 2'NH₂ UTP, 2'NH₂ CTP, and 2'F. UTP. Suitable modified nucleotides also include aminoallyl uridine, pseudo-uridine, 5-l-uridine, 5-I- cytidine, 5-Br-uridine, alpha-S adenosine, alpha-S cytidine, alpha-S guanosine, alpha-S uridine, 4-thio uridine, 2-thio-cytidine, 2'NH₂ uridine, 2'NH₂ cytidine, and 2'F uridine, including the free pho (NTP) RNA molecules, as well as all other useful forms of the nucleotides.

RNA interference molecules may also contain modifications in the ribose sugars, as well as modifications in the phosphate backbone of the nucleotide chain. For example, siRNA or miRNA molecules containing a-D-arabinofuranosyl structures in place of the naturally-occurring a-D-ribonucleosides found in RNA can be used as RNA interference molecules according to the present invention. Other examples include RNA molecules containing the o-linkage between the sugar and the heterocyclic base of the nucleoside, which confers nuclease resistance and tight complementary strand binding to the oligonucleotidesmolecules similar to the oligonucleotides containing 2'-0-methyl ribose, arabinose and particularly a-arabinose. Phosphorothioate linkages can also be used to stabilize the siRNA and miRNA molecules. An "siRNA" refers to a nucleic acid that forms a double stranded RNA, which double stranded RNA has the ability to reduce or inhibit expression of a gene or target gene when the siRNA is expressed in the same cell as the gene or target gene. "siRNA" thus refers to the double stranded RNA formed by the

complementary strands. The complementary portions of the siRNA that hybridize to form the double stranded molecule typically have substantial or complete identity. In an embodiment, an siRNA refers to a nucleic acid that has substantial or complete identity to a target gene and forms a double stranded siRNA. The sequence of the siRNA can correspond to the full length target gene, or a subsequence thereof.

In an embodiment, the siRNA is at least about 15-50 nucleotides in length (e.g., each complementary sequence of the double stranded siRNA is about 15-50 nucleotides in length, and the double stranded siRNA is about 15-50 base pairs in length, preferably about 19-30 base nucleotides, preferably about 20-25 nucleotides in length, e.g., 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, or 30

nucleotides in length).

Suitable siRNAs also include small hairpin (also called stem loop) RNAs

(shRNAs). In an embodiment, the shRNA comprises short, e.g. about 19 to about 25 nucleotide, antisense strand, followed by a nucleotide loop of about 5 to about 9 nucleotides, and the analogous sense strand. Alternatively, the sense strand may precede the nucleotide loop structure and the antisense strand may follow.

In an embodiment, the antagonist of IAP is siRNA, shRNA or miRNA.

Specific RNA interference molecules, such as siRNA, shRNA and miRNA molecules, can be easily designed by one skilled in the art having regard to the sequence of the target gene.

In an embodiment, the siRNA, shRNA or miRNA is targeted against a sequence selected from the group consisting of NCBI Reference Sequence: NM_001 166.4, NCBI Reference Sequence: NM_001256163.1 , NCBI Reference Sequence: NM_001256166.1 , GenBank: DQ068066.1 , NCBI Reference Sequence:

NM_001 165.4, NCBI Reference Sequence: NM_182962.2, GenBank: BC037420.1 , NCBI Reference Sequence: NM_001 167.3, NCBI Reference Sequence: NM_001204401.1 , NCBI Reference Sequence: NR_037916.1 , and NCBI Reference Sequence: NG_007264.1.

Other RNA molecules which are single stranded, or are not considered to be RNA interference molecules, may also be useful as therapeutic agents in accordance with the present invention, including messenger RNAs (and the progenitor pre-messenger RNAs), small nuclear RNAs, small nucleolar RNAs, transfer RNAs and ribosomal RNAs.

Gene therapy techniques may also be used to antagonize lAPs by introducing coding sequences for Smac and thereby amplifying expression of endogenous Smac, e.g., full or partial Smac cDNA molecules. An illustrative example of an IAP antagonist that employs gene therapy is the recombinant vaccinia virus carrying a partial Smac gene disclosed in Pan et al., "SMAC-armed vaccinia virus induced both apoptosis and necroptosis and synergizes the efficiency of vinblastine in HCC," Hum Cell. 2014 Oct; 27(4): 162-71 . doi: 10.1007/s13577- 014-0093-z. Epub 2014 Apr 26.

A TRAIL receptor agonist, or TRAIL agonist, is an agent that binds to a TRAIL receptor, such as TRAIL receptor 1 (TRAIL R1 , also known as "death receptor 4" or DR4), TRAIL receptor 2 (TRAIL R2, also known as "death receptor 5" or DR5), or both DR4 and DR5, and leads to apoptosis in at least one mammalian (e.g., human) cell type (such as a TRAIL-sensitive tumor cell line) when used in an amount effective to induce apoptosis under physiological conditions. TRAIL receptor agonists used in the present invention preferably do not bind to TRAIL decoy receptors.

TRAIL has received considerable attention recently because of the finding that many cancer cell types are sensitive to TRAIL-induced apoptosis, while most normal cells appear to be resistant to this action of TRAIL. TRAIL-resistant cells may arise by a variety of different mechanisms including loss of the receptor, presence of decoy receptors, or overexpression of FLIP which competes for zymogen caspase-8 binding during DISC formation. In TRAIL resistance, the compounds or compositions that are used in the method of the present invention may increase tumor cell sensitivity to TRAIL leading to enhanced cell death, the clinical correlations of which are expected to be increased apoptotic activity in TRAIL resistant tumors, improved clinical response, increased response duration, and ultimately, enhanced patient survival rate. In support of this, reduction in XIAP levels by in vitro antisense treatment has been shown to cause sensitization of resistant melanoma cells and renal carcinoma cells to TRAIL (Chawla-Sarkar, et al., 2004). The Smac mimetic compounds used in the method of the present invention bind to lAPs and inhibit their interaction with caspases, therein potentiating TRAIL-induced apoptosis.

TRAIL agonists include, for example, an antibody, such as mapatumumab or lexatumumab (Human Genome Sciences), HGS-TR2J (Human Genome

Sciences), Apomab (Genentech), CS-1008 (Daiichi Sankyo), LBY135 (Novartis), APG350 (Apogenix), conatumumab (Amgen), and TRA-8 (University of Alabama- Birmingham). The antibody can be a DR4 antibody, a DR5 antibody, or an antibody that binds to both DR4 and DR5. In other embodiments the TRAIL receptor agonist is recombinant human TRAIL or a soluble TRAIL polypeptide or a Fc-TRAIL fusion peptidobody. Fc-TRAIL fusion proteins are disclosed, e.g., in US20130064838. TRAIL fusion proteins comprising a collectin trimerization domain are disclosed, e.g., in US 20130178604, US20140171622, and

US8383774, among others. Other TRAIL agonists include those disclosed in, e.g., US6284236; US69981 16; US7915245.

In some embodiments of the invention, pharmaceutical compositions comprising an IAP antagonist and, optionally, a TRAIL agonist, alone or in combination with one or more other active pharmaceutical ingredients, are administered to a human or veterinary subject. The pharmaceutical compositions typically comprise at least one pharmaceutically acceptable excipient, e.g., a carrier or diluent, and can be administered in the conventional manner by routes including systemic, subcutaneous, topical, or oral routes. Administration may be by intravenous injection, either as a bolus or infusion, but other routes of

administration, including, among others, subcutaneous or oral administration, are not precluded. An intravenous formulation can contain, e.g., from 1 mg/mL up to and including 5 mg/mL of the IAP antagonist, such as specifically birinapant, in sterile 0.05M citrate buffered PBS, pH 5. Formulation may be by immediate release or prolonged release. Specific modes of administration and formulation will depend on the indication and other factors including the particular compound being administered. The amount of compound to be administered is that amount which is therapeutically effective, i.e., the amount that ameliorates the disease symptoms, i.e., that slows cancer progression or causes regression, without serious adverse effects relative to the disease being treated or causes

improvement of the infectious disease or the autoimmune disease. Put another way, an effective dose is one that over the course of therapy, which may be, e.g., 1 or more weeks, e.g., multiple courses of 3 weeks on/1 week off, results in treatment of the proliferative disorder, i.e., a decrease in the rate of disease progression, termination of disease progression, or regression or remission, or results in a decrease in infectious burden or resolution of symptoms of an infectious or autoimmune disease.

The phrase "pharmaceutical composition" refers to a composition suitable for administration in medical use.

The dosage to be administered will depend on the characteristics of the subject being treated, e.g., the particular patient treated, age, weight, health, types of concurrent treatment, if any and the specific disease or disorder that is being treated. Frequency of treatments can be easily determined by one of skill in the art (e.g., by the clinician).

The TRAIL agonist, when utilized, can also be administered intravenously. A composition and administration of an anti-DR5 antibody is disclosed, e.g., in US20140004120.

The term "antibody" includes reference to isolated forms of both glycosylated and non-glycosylated immunoglobulins of any isotype or subclass, including any combination of: 1 ) human (e.g., CDR-grafted), humanized, and chimeric antibodies, 2) monospecific (e.g., DR5) or multi-specific antibodies (e.g., DR4 and DR5), and 3) monoclonal, polyclonal, or single chain (scFv) antibodies, irrespective of whether such antibodies are produced, in whole or in part, via immunization, through recombinant technology, by way of in vitro synthetic means, or otherwise. Thus, the term "antibody" is inclusive of those that are prepared, expressed, created or isolated by recombinant means, such as (a) antibodies isolated from an animal (e.g., a mouse) that is transgenic for human immunoglobulin genes or a hybridoma prepared therefrom, (b) antibodies isolated from a host cell transfected to express the antibody (e.g., from a transfectoma), (c) antibodies isolated from a recombinant, combinatorial antibody library, and (d) antibodies prepared, expressed, created or isolated by any other means that involve splicing of immunoglobulin gene sequences to other DNA sequences. In some embodiments the antibodies of the present invention are monoclonal antibodies, such as humanized or fully-human monoclonal antibodies.

This also encompasses molecules with TRAIL agonist activity that may be fragments of antibodies, peptides, recombinant forms of the endogenous TRAIL, or small molecules, that effectively engage the receptor and thereby trigger TRAIL-receptor signaling. Accordingly, in addition to recombinant antibodies, new classes of therapeutic proteins are being developed and include

recombinant protein scaffolds (e.g. DARPINs, anticalins, affibodies, fibronectin domains), in which binding is mediated by surface diversity that interacts with targets which include TRAIL receptors DR4 and DR5 (e.g. Veesler, D., et al., J. Biol. Chem. 284(44):30718-30726. (2009).

The dose of the TRAIL agonist when given in combination with an IAP antagonist in accordance with this invention is expected to be the same as it would be were it administered alone or with another additional chemotherapeutic agent.

However in some situations the dose of the TRAIL agonist, or the dose of the Smac mimetic required when used together or in combination, may be less than the dose of either agent when used alone. While it is possible to combine an IAP antagonist and a TRAIL agonist into a single dosage unit, e.g., a sterile solution for intravenous administration, in practice, it may be preferable to administer each agent separately, e.g., using a separate pharmaceutical dosage unit, including by administering the separate dosage units according to a different dosing regimen.

The NSAID is typically administered in accordance with dosing regimens approved for use as a monotherapy for the treatment of pain or inflammation. Anti-inflammatory therapy can be initiated one or more days, e.g., 1 to 7 days, 1 to 3 days, or 1 to 2 days, prior to initiation of treatment with the IAP antagonist. Or, initiation of anti-inflammatory therapy can begin simultaneously with initiation of treatment with the IAP antagonist. Anti-inflammatory therapy can be

maintained throughout the course of treatment with the IAP antagonist.

While it is possible to combine an IAP antagonist and an NSAID into a single dosage unit, e.g., a sterile solution for intravenous administration, in practice, it may be preferable to administer each agent separately, e.g., using a separate pharmaceutical dosage unit, including by administering the separate dosage units according to a different dosing regimen.

Pharmaceutical compositions to be used comprise a therapeutically effective amount of the active pharmaceutical ingredients as described above with one or more pharmaceutically acceptable excipients. The phrase "pharmaceutical composition" refers to a composition suitable for administration in medical or veterinary use. It should be appreciated that the determinations of proper dosage forms, dosage amounts, and routes of administration for a particular patient are within the level of ordinary skill in the pharmaceutical and medical arts.

Compositions suitable for parenteral administration conveniently comprise a sterile aqueous preparation of the active pharmaceutical ingredients, which is preferably isotonic with the blood of the recipient. This aqueous preparation may be formulated according to known methods using suitable dispersing or wetting agents, emulsifying and suspending agents. Various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, and sorbic acid also may be included. The sterile injectable preparation also may be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1 ,3-butane diol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono-or di-glycerides. In addition, fatty acids such as oleic acid may be used in the preparation of injectables. Prolonged absorption of the injectable pharmaceutical form can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin. Carrier formulation suitable for subcutaneous, intravenous, intramuscular, etc. administrations can be found in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA.

A pharmaceutical composition in intravenous unit dose form may comprise, e.g., a vial or pre-filled syringe, or an infusion bag or device, each comprising an effective amount or a convenient fraction of an effective amount such that the contents of one vial or syringe are administered at a time.

An effective dose is one that over the course of therapy, which may be, e.g., 1 or more weeks, e.g., multiple courses of 3 weeks on/1 week off, results in treatment of the disorder, e.g., a decrease in the rate of disease progression, termination of disease progression, or regression or remission.

Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active pharmaceutical ingredient(s) are admixed with at least one inert pharmaceutically acceptable excipient such as (a) fillers or extenders, as for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid, (b) binders, as for example, carboxymethylcellulose, alignates, gelatin, polyvinylpyrrolidone, sucrose, and acacia, (c) humectants, as for example, glycerol, (d) disintegrating agents, as for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate, (e) solution retarders, as for example paraffin, (f) absorption accelerators, as for example, quaternary ammonium compounds, (g) wetting agents, as for example, cetyl alcohol, and glycerol monostearate, (h) adsorbents, as for example, kaolin and bentonite, and (i) lubricants, as for example, 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 dosage forms such as tablets, dragees, capsules, pills, and granules also can be prepared with coatings and shells, such as enteric coatings and others well known in the art. The solid dosage form also may contain opacifying agents, and can also be of such composition that they release the active compound or compounds in a certain part of the intestinal tract in a delayed manner. Examples of embedding compositions which can be used are polymeric substances and waxes. The active compounds can also be in microencapsulated form, if appropriate, with one or more of the above-mentioned excipients. Such solid dosage forms may generally contain from 1 % to 95% (w/w) of the active compounds. In certain embodiments, the active compounds generally range from 5% to 70% (w/w).

Since one aspect of the present invention contemplates the treatment of the disease/conditions with a combination of pharmaceutically active agents that may be administered separately, the invention further relates to combining separate pharmaceutical compositions in kit form. The kit comprises two separate pharmaceutical compositions: one composition contains the IAP antagonist used in the method of the present invention, and a second composition contains the NSAID. The kit comprises a container for containing the separate compositions such as a divided bottle or a divided foil packet. Additional examples of containers include syringes, e.g., pre-filled syringes, boxes and bags. Typically, the kit comprises directions for the use of the separate components. The kit form is particularly advantageous when the separate components are preferably administered in different dosage forms (e.g., oral and parenteral), are administered at different dosage intervals, or when titration of the individual components of the combination is desired by the prescribing physician or veterinarian.

An example of such a kit is a so-called blister pack. Blister packs are well known in the packaging industry and are being widely used for the packaging of pharmaceutical unit dosage forms (tablets, capsules, and the like). Blister packs generally consist of a sheet of relatively stiff material covered with a foil of a preferably transparent plastic material. During the packaging process recesses are formed in the plastic foil. The recesses have the size and shape of the tablets or capsules to be packed. Next, the tablets or capsules are placed in the recesses and the sheet of relatively stiff material is sealed against the plastic foil at the face of the foil which is opposite from the direction in which the recesses were formed. As a result, the tablets or capsules are sealed in the recesses between the plastic foil and the sheet. Preferably the strength of the sheet is such that the tablets or capsules can be removed from the blister pack by manually applying pressure on the recesses whereby an opening is formed in the sheet at the place of the recess. The tablet or capsule can then be removed via said opening.

It may be desirable to provide a memory aid on the kit, e.g., in the form of numbers next to the tablets or capsules whereby the numbers correspond with the days of the regimen which the tablets or capsules so specified should be ingested. Another example of such a memory aid is a calendar printed on the card, e.g., as follows "First Week, Monday, Tuesday, . . . etc . . . Second Week, Monday, Tuesday, . . . " etc. Other variations of memory aids will be readily apparent. A "daily dose" can be a single tablet or capsule or several pills or capsules to be taken on a given day. Also, a daily dose of a substance of the present invention can consist of one tablet or capsule, while a daily dose of the second substance can consist of several tablets or capsules and vice versa. The memory aid should reflect this variety and aid in correct administration of the active agents. In another specific embodiment of the invention, a dispenser designed to dispense the daily doses one at a time in the order of their intended use is provided. Preferably, the dispenser is equipped with a memory-aid, so as to further facilitate compliance with the regimen. An example of such a memory-aid is a mechanical counter which indicates the number of daily doses that has been dispensed. Another example of such a memory-aid is a battery-powered microchip memory coupled with a liquid crystal readout, or audible reminder signal which, for example, reads out the date that the last daily dose has been taken and/or reminds one when the next dose is to be taken.

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

The compounds and compositions used in the method of the present invention also may benefit from a variety of delivery systems, including time-released, delayed release or sustained release delivery systems. Such option may be particularly beneficial when the compounds and composition are used in conjunction with other treatment protocols as described in more detail below.

Many types of controlled release delivery systems are available and known to those of ordinary skill in the art. They include polymer base systems such as poly(lactide-glycolide), copolyoxalates, polycaprolactones, polyesteramides, polyorthoesters, polyhydroxybutyric acid, and polyanhydrides. Microcapsules of the foregoing polymers containing drugs are described in, for example, U.S. Pat. No. 5,075, 109. Delivery systems also include non-polymer systems that are: lipids including sterols such as cholesterol, cholesterol esters and fatty acids or neutral fats such as mono-di-and tri-glycerides; hydrogel release systems; sylastic systems; peptide based systems; wax coatings; compressed tablets using conventional binders and excipients; partially fused implants; and the like. Specific examples include, but are not limited to: (a) erosional systems in which the active compound is contained in a form within a matrix such as those described in U.S. Pat. Nos. 4,452,775, 4,667,014, 4,748,034 and 5,239,660 and (b) diffusional systems in which an active component permeates at a controlled rate from a polymer such as described in U.S. Pat. Nos. 3,832,253, and 3,854,480. In addition, pump-based hardware delivery systems can be used, some of which are adapted for implantation.

Use of a long-term sustained release implant may be desirable. Long-term release, as used herein, means that the implant is constructed and arranged to deliver therapeutic levels of the active compounds for at least 30 days, and preferably 60 days. Long-term sustained release implants are well-known to those of ordinary skill in the art and include some of the release systems described above.

The compounds used in the method of the present invention and pharmaceutical compositions comprising compounds used in the method of the present invention can be administered to a subject suffering from cancer, an autoimmune disease or another disorder where a defect in apoptosis is implicated. In connection with such treatments, the patient can be treated prophylactically, acutely, or chronically using the compounds and compositions used in connection with the method of the present invention, depending on the nature of the disease. Typically, the host or subject in each of these methods is human, although other mammals may also benefit from the present invention.

As described in US 7,244,851 , IAP antagonists can be used for the treatment of all cancer types which fail to undergo apoptosis. Thus, compounds used in the method of the present invention can be used to provide a therapeutic approach to the treatment of many kinds of solid tumors, including but not limited to carcinomas, sarcomas including Kaposi's sarcoma, erythroblastoma, glioblastoma, meningioma, astrocytoma, melanoma and myoblastoma. Treatment or prevention of non-solid tumor cancers such as leukemia is also contemplated by this invention. Indications may include, but are not limited to brain cancers, skin cancers, bladder cancers, ovarian cancers, breast cancers, gastric cancers, pancreatic cancers, colon cancers, blood cancers, lung cancers and bone cancers. Examples of such cancer types include neuroblastoma, intestine carcinoma such as rectum carcinoma, colon carcinoma, familial adenomatous polyposis carcinoma and hereditary non- polyposis colorectal cancer, esophageal carcinoma, labial carcinoma, larynx carcinoma, hypopharynx carcinoma, tongue carcinoma, salivary gland carcinoma, gastric carcinoma, adenocarcinoma, medullary thyroid carcinoma, papillary thyroid carcinoma, renal carcinoma, kidney parenchymal carcinoma, ovarian carcinoma, cervix carcinoma, uterine corpus carcinoma, endometrium carcinoma, chorion carcinoma, pancreatic carcinoma, prostate carcinoma, testis carcinoma, breast carcinoma, urinary carcinoma, melanoma, brain tumors such as glioblastoma, astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermal tumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, acute lymphatic leukemia (ALL), chronic lymphatic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), adult T-cell leukemia lymphoma, hepatocellular carcinoma, gall bladder carcinoma, bronchial carcinoma, small cell lung carcinoma, non-small cell lung carcinoma, multiple myeloma, basal cell carcinoma, teratoma, retinoblastoma, choroidea melanoma, seminoma, rhabdomyo sarcoma, craniopharyngeoma, osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma, Ewing sarcoma and plasmocytoma.

The IAP antagonists suitable for use in the method of the present invention will be active for treating human malignancies including, but not limited to, such human malignancies in which clAP1 and clAP2 are over-expressed (e.g., lung cancers, see Dai et al, Hu. Molec. Genetics, 2003 v 12 pp791 -801 ; leukemias (multiple references), and other cancers (Tamm et al, Clin Cancer Res, 2000, v 6, 1796-1803). Also, the IAP antagonists suitable for use in the method of the present invention may be active in disorders that may be driven by inflammatory cytokines such as TNFa playing a pro-survival role (for example, there is a well defined role for TNFa acting as a survival factor in ovarian carcinoma, similarly for gastric cancers (see Kulbe, et al, Cancer Res 2007, 67, 585-592).

In addition to apoptosis defects found in tumors, defects in the ability to eliminate self-reactive cells of the immune system due to apoptosis resistance are considered to play a key role in the pathogenesis of autoimmune diseases. Autoimmune diseases are characterized in that the cells of the immune system produce antibodies against its own organs and molecules or directly attack tissues resulting in the destruction of the latter. A failure of those self-reactive cells to undergo apoptosis leads to the manifestation of the disease. Defects in apoptosis regulation have been identified in autoimmune diseases such as systemic lupus erythematosus or rheumatoid arthritis.

Examples of such autoimmune diseases include collagen diseases such as rheumatoid arthritis, systemic lupus erythematosus, Sharp's syndrome, CREST syndrome (calcinosis, Raynaud's syndrome, esophageal dysmotility, telangiectasia), dermatomyositis, vasculitis (Morbus Wegener's) and Sjogren's syndrome, renal diseases such as Goodpasture's syndrome, rapidly-progressing glomerulonephritis and membrano-proliferative glomerulonephritis type II, endocrine diseases such as type-l diabetes, autoimmune polyendocrinopathy- candidiasis-ectodermal dystrophy (APECED), autoimmune parathyroidism, pernicious anemia, gonad insufficiency, idiopathic Morbus Addison's, hyperthyroidosis, Hashimoto's thyroiditis and primary myxedema, skin diseases such as pemphigus vulgaris, bullous pemphigoid, herpes gestationis, epidermolysis bullosa and erythema multiforme major, liver diseases such as primary biliary cirrhosis, autoimmune cholangitis, autoimmune hepatitis type-1 , autoimmune hepatitis type-2, primary sclerosing cholangitis, neuronal diseases such as multiple sclerosis, myasthenia gravis, myasthenic Lambert-Eaton syndrome, acquired neuromyotony, Guillain-Barre syndrome (Muller-Fischer syndrome), stiff-man syndrome, cerebellar degeneration, ataxia, opsoklonus, sensoric neuropathy and achalasia, blood diseases such as autoimmune hemolytic anemia, idiopathic thrombocytopenic purpura (Morbus Werlhof), infectious diseases with associated autoimmune reactions such as AIDS, Malaria and Chagas disease.

The present invention can be carried out in conjunction with other treatment approaches, e.g., in combination with a biologic or chemotherapeutic agent or with chemoradiation. As discussed above, embodiments of the invention also include a method of treating a patient afflicted with cancer by the

contemporaneous or concurrent administration of a biological or

chemotherapeutic agent additional to the Smac mimetic, such as birinapant, and an NSAID. Such biological or chemotherapeutic agents include but are not limited to the chemotherapeutic agents described in "Modern Pharmacology with Clinical Applications", Sixth Edition, Craig & Stitzel, Chpt. 56, pg 639-656 (2004). The chemotherapeutic agent can be, but is not limited to, alkylating agents, antimetabolites, anti-tumor antibiotics, plant-derived products such as taxanes, enzymes, hormonal agents, miscellaneous agents such as cisplatin, monoclonal antibodies, glucocorticoids, mitotic inhibitors, topoisomerase I inhibitors, topoisomerase II inhibitors, immunomodulating agents such as interferons, cellular growth factors, cytokines, cellular growth factors and kinase inhibitors. Other suitable classifications for chemotherapeutic agents include mitotic inhibitors, and anti-estrogenic agents.

Specific examples of suitable biological and chemotherapeutic agents include, but are not limited to, carboplatin, cisplatin, carmustine (BCNU), 5-fluorouracil (5- FU), cytarabine (Ara-C), gemcitabine, methotrexate, daunorubicin, doxorubicin, dexamethasone, irinotecan, topotecan, etoposide, paclitaxel, docetaxel, vincristine, tamoxifen, TNFa, TRAIL and other members, i.e., other than TRAIL and TNFa, of the TNFa superfamily of molecules., interferon (in both its alpha and beta forms), thalidomide, thalidomide derivatives such as lenalidomide, melphalan, and PARP inhibitors. Other specific examples of suitable chemotherapeutic agents include nitrogen mustards such as cyclophosphamide, alkyl sulfonates, nitrosoureas, ethylenimines, triazenes, folate antagonists, purine analogs, pyrimidine analogs, anthracyclines, bleomycins, mitomycins,

dactinomycins, plicamycin, vinca alkaloids, epipodophyllotoxins, taxanes, glucocorticoids, L-asparaginase, estrogens, androgens, progestins, luteinizing hormones, octreotide actetate, hydroxyurea, procarbazine, mitotane,

hexamethylmelamine, carboplatin, mitoxantrone, monoclonal antibodies, levamisole, interferons, interleukins, filgrastim and sargramostim.

In the case of treatment of infection, e.g., chronic intracellular infection, the method of treatment of the invention can be carried concomitantly with other therapies, e.g., antiviral, antibacterial, antifungal, or antiprotozoal agents.

To the extent that co-administration of a TRAIL agonist with an IAP antagonist exacerbates the inflammation-related side effects observed in some cases of treatment with an IAP antagonist alone (e.g., symptoms of VI cranial nerve palsy or Bell's palsy), co-administration of an anti-inflammatory other than a TNFa inhibitor would lessen the side effects and thereby improve the tolerability of the IAP antagonist + TRAIL agonist therapy.

EXAMPLE

Adverse event (AE) data from two studies of birinapant (TL3271 1 ), Study 007 and Study 0078, were analyzed to identify correlations between cranial nerve palsy including Bell's Palsy and certain variables.

One of these studies, Study 007 (NCT00993239), was a Phase 1 open-label, non-randomized dose escalation study. The purpose of this study was to determine the maximum tolerated dose (MTD) and characterize the safety and tolerability of TL3271 1 when administered as a 30 minute intravenous infusion once weekly for three weeks per repeated 4 week intervals in subjects with refractory solid tumors or lymphoma. Additionally, the study was designed to assess anti-tumor activity, pharmacokinetics, and exploratory biomarkers as a measurement of pharmacodynamic effects. The doses of birinapant administered to patients were in the range of 0.18 mg/m² to 63 mg/m² See, Amaravadi et al., 2015, Mol Cancer Ther 14(11 ): 1 -7 (PubMed ID: 26333381 ).

The second of these studies, Study 0078 (NCT01 188499), was a dose escalation safety study of TL3271 1 in combination with chemotherapy. The purpose of this study was to determine the safety and maximum tolerated dose of TL3271 1 as a 30 minute intravenous infusion once a week, for 2 consecutive weeks, when combined with standard regimens of chemotherapy in subjects with advanced or metastatic solid tumors. Additionally, the study was designed to assess antitumor activity, pharmacokinetics, and exploratory biomarkers as a measurement of pharmacodynamic effects. The doses of birinapant administered to patients were in the range of 2.8 mg/m² to 47 mg/m². See, Amaravadi et al., 2013, J Clin Oncol 31 (suppl; abstr 2504)

226 Patients collectively completed both studies. Nine Cranial nerve palsy, including Bell's palsy, adverse events (4%) were reported.

Of the 226 patients who completed the studies, 52 received NSAID therapy while being treated with birinapant and 174 did not; of the 52, 38 also received dexamethasone therapy.

Of the 226 patients, 154 received dexamethasone therapy while being treated with birinapant and 72 did not; of the 154, 38 also received NSAID therapy.

NSAID therapy comprised

□ ibuprofen - 20 patients received between 200 mg and 600 mg taken as needed for pain

□ naproxen - 12 patients received between 220 and 500 mg taken twice daily for pain

□ celecoxib, etodolac, or meloxicam - 8 patients received 200 mg of

celecoxib, 400 mg twice daily of etodolac or 15 mg of meloxicam once daily for arthritis

□ aspirin products (Excedrin(R), ) - 8 patients 325mg - 500 mg once or twice daily for pain, fever, headache □ diclofenac - 4 patients received this topically for pain.

Dexamethasone therapy comprised 4mg, 8mg, 10 mg or 20 mg given prior to chemotherapy for chemo-prophylaxis. There were 14 patients (of the 154 who received dexamethasone while being treated with birinapant) that received only one dose. 8 patients received dexamethasone for reasons other than chemo- prophylaxis (edema, itching, nausea, vomiting)

The results of the AE analysis are reported in the following table.

These results indicate that co-administration of an NSAID with a Smac mimetic reduces the incidence of Cranial Nerve palsy including Bell's palsy. Surprisingly, the antiinflammatory agent, dexamethasone, did not have a similar effect. These data also support co-administration of an NSAID during Smac mimetic therapy to reduce the incidence of Cranial Nerve palsy.

Both Dexamethasone and NSAID were given to patients who received birinapant across a range of doses from 2.8 mg/m² to 47 mg/m² Further, the development of Cranial Nerve palsy with birinapant occurred across a range of administered doses of birinapant. Thus the failure to develop cranial nerve palsy was not attributed to different doses of birinapant.

83 Patients who received prophylactic antiviral therapy also received dexamethasone. Of these 4 developed a cranial nerve palsy. 17 Patients who received prophylactic antiviral therapy received NSAIDs. Of these 17 patients, 16 also received dexamethasone. None of these patients developed a cranial nerve palsy.

The inability of dexamethasone to prevent cranial nerve palsy, even in the presence of an antiviral agent, indicates that dexamethasone is not protective for cranial nerve palsy.

It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and the scope of the appended claims. All patent and literature references cited herein are incorporated by reference herein as though fully set forth.

Claims

Claims:

1. A method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an IAP antagonist, said method comprising coadministering to the patient an NSAID.

2. The method of claim 1 wherein the patient is being treated for a cancer, a benign proliferative disorder, an autoimmune disorder, an angiogenesis- associated disorder or an intracellular infection and wherein the cranial nerve palsy is VI cranial nerve palsy or Bell's palsy.

3. The method of claim 2 wherein

the cancer is selected from:

pancreatic cancer, ovarian cancer, breast cancer, mesothelioma, peripheral neuroma, glioblastoma, melanoma, adrenocortical carcinoma, AIDS-related lymphoma, anal cancer, bladder cancer, meningioma, glioma, astrocytoma, cervical cancer, chronic myeloproliferative disorders, colon cancer, endometrial cancer, ependymoma, esophageal cancer, Ewing's sarcoma, extracranial germ cell tumors, extrahepatic bile duct cancer, gallbladder cancer, gastric cancer, gastrointestinal carcinoid tumors, gestational trophoblastic tumors, hairy cell leukemia, Hodgkin lymphoma, non-Hodgkin lymphoma, hypopharyngeal cancer, islet cell carcinoma, Kaposi sarcoma, laryngeal cancer, leukemia, lip cancer, oral cavity cancer, liver cancer, male breast cancer, malignant mesothelioma, medulloblastoma, Merkel cell carcinoma, metastatic squamous neck cell carcinoma, multiple myeloma and other plasma cell neoplasms, mycosis fungoides and the Sezary syndrome, myelodysplasia syndromes, nasopharyngeal cancer, neuroblastoma, non-small cell lung cancer, small cell lung cancer, oropharyngeal cancer, bone cancers, including

osteosarcoma and malignant fibrous histiocytoma of bone, paranasal sinus cancer, parathyroid cancer, penile cancer, pheochromocytoma, pituitary tumors, prostate cancer, rectal cancer, renal cell cancer, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, small intestine cancer, soft tissue sarcoma, supratentorial primitive neuroectodermal tumors, pineoblastoma, testicular cancer, thymoma, thymic carcinoma, thyroid cancer, transitional cell cancer of the renal pelvis and ureter, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, and Wilm's tumor and other childhood kidney tumors;

the benign proliferative disorder is selected from:

psoriasis, benign prostatic hypertrophy, and restenosis;

the autoimmune disorder is selected from:

rheumatoid arthritis, psoriasis, autoimmune proliferative

glomerulonephritis, lymphoproliferative autoimmune responses, systemic lupus erythematosus, psoriasis, and idiopathic thrombocytopenic purpura (Morbus Werlhof);

the angiogensis-associated disorder is selected from:

macular degeneration, diabetic retinopathy, retinopathy of prematurity, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubcosis, Osier-Webber Syndrome, myocardial angiogenesis, plaque neovascularization, telangiectasia, angiofibroma, wound granulation, intestinal adhesions, atherosclerosis, scleroderma, or hypertrophic scarring;

the intracellular infection is by:

a virus, bacterium, fungus, yeast, or protozoa.

4. The method of claim 2 wherein the infection is caused by:

a virus selected from the group consisting of Human papillomaviruses, Herpes viruses including herpes simplex 1/2, varicella zoster, EBV, CMV, HHV-6/7, HTLV, Human papovaviruses, including JC virus and BK virus, adeno and parvoviruses, HIV, HBV and HCV. bacteria selected from the group consisting of Salmonella spp., Ehrlichia spp., Mycobacteria spp., Spirochetes, Legionella spp., Listeria spp., Rickettsia spp., Chlamydia spp., Mycoplasma spp., Coxiella spp., Yersinia spp., Francisella spp., Brucella spp., Neisseria spp, and Nocardia spp.. fungus or yeast selected from the group consisting of Histoplasma spp., Aspergillus spp., Cryptococcus spp., and Pneunocystis jirovecii, protozoa selected from the group consisting of Trypanosomatids (e.g., Leishmania spp.), Apicomplexans, including liver forms of Plasmodium spp., Toxoplasma spp., and Cryptosporidium spp.

5. The method of any of the preceding claims wherein the lAP antagonist is a Smac mimetic.

6. The method of claim 5 wherein the Smac mimetic is a bivalent Smac mimetic.

7. The method of claim 5 wherein the Smac mimetic is characterized as (i) not inhibiting XIAP E3 ubiquitin ligase activity or as only poorly inhibiting XIAP E3 ubiquitin ligase activity, (ii) not inhibiting or poorly inhibiting NOD (i.e., NOD1/2) signaling, or (iii) not inhibiting or poorly inhibiting NOD-mediated NF-kB activation.

8. The method of claim 5 wherein the Smac mimetic is birinapant.

9. The method of claim 5 wherein the NSAID is administered within 48 hours prior to or concurrently with first administration of the lAP antagonist.

10. The method of claim 9 wherein the lAP antagonist and the NSAID are first administered concurrently.

1 1 . The method of any of the preceding claims wherein the patient is selected for coadministration of an NSAID because the patient is pre-disposed to symptoms of a cranial nerve palsy.

12. A method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising internally coadministering to the patient an IAP antagonist and an NSAID.

13. The method of claim 12 wherein the patient is pre-disposed to symptoms of a cranial nerve palsy.

14. A method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an IAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the IAP antagonist, coadministering an NSAID to the patient, or both.

15. An IAP antagonist for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising coadministering to the patient an IAP antagonist and an NSAID.

16. An NSAID for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising coadministering to the patient an IAP antagonist and an NSAID.

17. An IAP antagonist for use in a method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an IAP antagonist, said method comprising coadministering to the patient an NSAID.

18. An NSAID for use in a method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an IAP antagonist, said method comprising coadministering to the patient an NSAID.

19. An IAP antagonist for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an lAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the lAP antagonist, coadministering an NSAID to the patient, or both.

20. An NSAID for use in a method of treating a disorder that is amenable to therapy with an lAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an lAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the lAP antagonist, coadministering an NSAID to the patient, or both.

21 . A product comprising an lAP antagonist and an NSAID for simultaneous, separate or sequential use in a method of treating a disorder that is amenable to therapy with an lAP antagonist in a patient suffering from such disorder.

22. A product comprising an lAP antagonist and an NSAID for simultaneous, separate or sequential use in a method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an lAP antagonist.

23. A product comprising an lAP antagonist and an NSAID for simultaneous, separate or sequential use in a method of treating a disorder that is amenable to therapy with an lAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an lAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the lAP antagonist, coadministering an NSAID to the patient, or both.

24. A pharmaceutical composition comprising an lAP antagonist, an NSAID, and a pharmaceutically acceptable excipient.

25. Use of an lAP antagonist for the manufacture of a medicament for use in a method of treating a disorder that is amenable to therapy with an lAP antagonist in a patient suffering from such disorder, said method comprising coadministering to the patient an IAP antagonist and an NSAID.

26. Use of an NSAID for the manufacture of a medicament for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising coadministering to the patient an IAP antagonist and an NSAID.

27. Use of an IAP antagonist for the manufacture of a medicament for use in a method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an IAP antagonist, said method comprising coadministering to the patient an NSAID.

28. Use of an NSAID for the manufacture of a medicament for use in a method of preventing or reducing manifestation of symptoms of a cranial nerve palsy in a patient being treated by administration of an IAP antagonist, said method comprising coadministering to the patient an NSAID.

29. Use of an IAP antagonist for the manufacture of a medicament for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an IAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the IAP antagonist, coadministering an NSAID to the patient, or both.

30. Use of an NSAID for the manufacture of a medicament for use in a method of treating a disorder that is amenable to therapy with an IAP antagonist in a patient suffering from such disorder, said method comprising administering to the patient an IAP antagonist and monitoring the patient for manifestation of symptoms of a cranial nerve palsy and, if such symptoms are manifested, then reducing the dose of the IAP antagonist, coadministering an NSAID to the patient, or both.