Triple kinase inhibition for the treatment of type i interferon response associated disorders

Abstract

The invention describes a novel combination therapy strategy and screening approaches, comprising a Receptor-interacting serine / threonine-protein kinase 1 (RIPK1) inhibitor, an inhibitor (IκB) kinase epsilon (IKKε), and a TANK binding kinase 1 (TBK1) inhibitor for the treatment and / or prevention of diseases associated with a modulated immune response, such as disorders associated with a type I interferon response. The invention provides such inhibitory compounds and their specific combinations for use in medical applications, as well as pharmaceutical compositions comprising the compounds of the invention. The invention further pertains to a method of treatment and / or prevention of a proliferative disease in a subject, the method comprising administering to the subject as single treatments or one or more combinatorial treatments, either sequentially or concomitantly, a therapeutically effective amount of inhibitors of RIPK1, inhibitors of IKKε – signaling, and inhibitors of TBK1. Further provided are approaches for screening and testing candidate kinase inhibitors for their applicability in a triple combination in accordance with the invention.

Description

[0001] TRIPLE KINASE INHIBITION FOR THE TREATMENT OF TYPE I INTERFERON RESPONSE ASSOCIATED DISORDERS FIELD OF THE INVENTION [1] The invention describes a novel combination therapy strategy and screening approaches, 5 comprising a Receptor-interacting serine / threonine-protein kinase 1 (RIPK1) inhibitor, an inhibitor IκB kinase epsilon (IKKε), and a TANK binding kinase 1 (TBK1) inhibitor for the treatment and / or prevention of diseases associated with a modulated immune response, such as disorders associated with a type I interferon response. The invention provides such inhibitory compounds and their specific combinations for use in medical applications, as well as pharmaceutical 10 compositions comprising the compounds of the invention. The invention further pertains to a method of treatment and / or prevention of a proliferative disease in a subject, the method comprising administering to the subject as single treatments or one or more combinatorial treatments, either sequentially or concomitantly, a therapeutically effective amount of inhibitors of RIPK1, inhibitors of IKKε – signaling, and inhibitors of TBK1. Further provided are approaches for 15 screening and testing candidate kinase inhibitors for their applicability in a triple combination in accordance with the invention. DESCRIPTION [2] The IkappaB kinase (IKK)-related serine / threonine kinases TBK1 and IKKε elicit anti-viral type-I interferon (IFN) response downstream of nearly all DNA and RNA sensors (Fitzgerald et 20 al., 2003; Hemmi et al., 2004; Yamamoto et al., 2003). TBK1 induces IFN responses downstream of the pathogen recognition receptors (PRRs) RIG-I and MDA5; cyclic GMP-AMP (cGAMP) synthase (cGAS); and TLR3 and TLR4, which transduce signals through the adaptor proteins MAVS, STING and TRIF, respectively. Upon ligand engagement, TBK1 phosphorylates the adaptor proteins MAVS, STING and TRIF, which results in the recruitment of the transcription 25 factor IRF3. IRF3 is then phosphorylated by TBK1, leading to its homo-dimerization and nuclear translocation. This cascade of events culminates in the induction of the expression of type I interferons and proinflammatory cytokines downstream of these nucleic acid sensors. Recognition of viral and bacterial nucleic acids in the cytosol or endosomal compartments and activation of TBK1 / IKK ^-dependent interferon and cytokine expression is important for host 30 defence. However, in addition to their important role in anti-viral immunity, type I interferons and cytokines critically contribute to the pathogenesis of a broad range of diseases characterized by autoinflammation and autoimmunity such as systemic lupus erythematosus (SLE) and Aicardi- Goutières Syndrome (AGS). Such diseases are often associated with abnormal regulation of nucleic acid metabolism and / or sensing and are collectively described under the term of type I 35 interferonopathies. Recognition of both endogenous and exogenous nucleic acids by innate immune receptors can trigger inflammation, because the capacity of innate sensors to discriminate between foreign and host nucleic acids is only limited. Mice lacking genes involved in DNA degradation such as DNAsel or DNAse1l3 and DNAse3 (also known as Trex1) display systemic lupus erythematous-like (SLE) pathology, and inflammatory myocarditis with 5 autoimmune-like pathology, respectively. In humans, mutations in the DNaseI or DNaseIl3 and DNase2a genes are respectively associated to SLE and deforming arthropathy, while mutations in the DNase3 gene is linked to chilblain lupus erythematous and Aicardi-Goutieres Syndrome (AGS). Studies in genetic mouse models showed that the genetic deletion of STING, IFNAR1 and IRF3 – together with IRF7 in the case of DNAse2a-deficient mice - rescues the myocarditis 10 phenotype of DNAse3-deficient mice and the embryonic lethal phenotype of DNAse2-deficient mice. Moreover, gain of function mutations in STING resulting in ligand-independent constitutive activation of TBK1 / IKK ^-mediated interferon and cytokine production also cause the pathogenesis of autoinflammatory lupus-like pathologies (Liu et al., 2014). In addition to abnormal activation of the DNA sensing machinery, genetic defects in proteins regulating RNA sensing have 15 also been linked to the pathogenesis of autoinflammatory and autoimmune pathologies. Mutations in ADAR1, an enzyme that deaminates adenosine in RNA and thus prevents its recognition by MDA5, are found in patients with AGS and mice lacking ADAR1 or its deaminase activity develop inflammatory pathologies (Heraud-Farlow and Walkley, 2016). [3] Together, the prior art shows that abnormal activation of interferon and cytokine production 20 caused by mutations affecting the nucleic acid sensing signalling machineries induce the pathogenesis of autoinflammatory and autoimmune diseases such as type I interferonopathies. Because of their essential function in inducing type I interferon and cytokine responses downstream of nucleic acid sensors, TBK1 and IKKε are therefore attractive therapeutic targets for the treatment of interferonopathies and other diseases triggered by abnormal activation of the 25 nucleic acid sensing and signalling machineries. In addition, TBK1 has been implicated in the pathogenesis of diet-induced obesity and metabolic disease (Zhao et al., 2018), suggesting that TBK1 inhibition could have a therapeutic effect in obesity-associated diseases. [4] The roles of TBK1 and IKK ^ have been studied using mice engineered to lack these kinases by disrupting the respective genes (these are referred to as ‘knockout’ mice). Mice lacking 30 IKK ^ were reported to be viable and fertile and did not develop any apparent pathology under steady state conditions, showing that IKK ^ is not essential for normal mouse development and physiology (Hemmi et al., 2004; Tenoever et al., 2007). In contrast, Tbk1- / -mice died during embryonic development due to TNFR1-mediated death of foetal liver cells (Bonnard et al., 2000). [5] Therefore, there is a continued need in the art to provide treatment strategies to tackle 35 immunological pathologies such as type I interferonopathies. It is an object of the present invention to meet that need. BRIEF DESCRIPTION OF THE INVENTION [6] Generally, and by way of brief description, the main aspects of the present invention can be described as follows: [7] In a first aspect, the invention pertains to a compound or composition for use in the 5 treatment of a condition in a subject, wherein ^ The compound or composition when administered to the subject induces an inhibition of (x) RIPK1 and (y) Inhibitor of TANK binding kinase 1 (TBK1) and / or IκB kinase epsilon (IKKε); and ^ Wherein the subject suffers from a disease, preferably an immunological disorder, 10 such as one associated with increased type I interferon responses. [8] In a second aspect, the invention pertains a RIPK1 inhibitor for use in the treatment or prevention of an adverse effect of an immune disease therapy in a subject, wherein the immune disease therapy involves the administration of an IKKε- and / or TBK1-inhibitor to the subject suffering from the immune disease. 15 [9] In a third aspect, the invention pertains to a method of treatment and / or prevention of a disease in a subject, the method comprising one or more steps of administering to the subject a therapeutically effective amount of a RIPK1 inhibitor, an IKKε inhibitor, and a TBK1 inhibitor.

[0010] In a fourth aspect, the invention pertains to a pharmaceutical composition comprising the compound or composition recited in the first or second aspect. 20

[0011] In a fifth aspect, the invention pertains a method for screening an IKKε-Inhibitor suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses in a subject, (a) Providing a test cell, (b) Optionally providing one or more control cells; 25 (c) Contacting the test cell with (i) a candidate IKKε-Inhibitor, while the test cell is contacted with a known TBK1 inhibitor and a known RIPK1 inhibitor; or (ii) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a known RIPK1 inhibitor; or (iii) a candidate IKKε-Inhibitor and a candidate TBK1 inhibitor, while the cell is contacted with a known RIPK1 inhibitor; or (iv) a candidate IKKε-Inhibitor, a candidate TBK1 inhibitor and a candidate RIPK1 inhibitor; or 5 (v) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a candidate RIPK1 inhibitor; (d) Optionally, where the control cell is not contacted in (a) to (d) with any of the candidate inhibitors; or alternatively, contacting the control cell with the candidate inhibitors but not the known inhibitors; 10 (e) Determining in the test cell at least cell death and release of one or more cytokines before and after step (iii), or in comparison to the control cell or in comparison to a reference value; wherein if in step (v) the following condition is determined: no increased cell death and / or a reduced release of the one or more test cytokines, then the candidate inhibitors in one or more of 15 (a) to (d) are suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses, wherein the triple combination comprises the inhibitors of the respective scenario (a) to (d) for which the condition was determined. DETAILED DESCRIPTION OF THE INVENTION

[0012] In the following, the elements of the invention will be described. These elements are listed 20 with specific embodiments, however, it should be understood that they may be combined in any manner and in any number to create additional embodiments. The variously described examples and preferred embodiments should not be construed to limit the present invention to only the explicitly described embodiments. This description should be understood to support and encompass embodiments which combine two or more of the explicitly described embodiments or 25 which combine the one or more of the explicitly described embodiments with any number of the disclosed and / or preferred elements. Furthermore, any permutations and combinations of all described elements in this application should be considered disclosed by the description of the present application unless the context indicates otherwise.

[0013] In a first aspect, the invention pertains to a compound or composition for use in the 30 treatment of a condition in a subject, wherein ^ The compound or composition when administered to the subject induces an inhibition of (x) RIPK1 and (y) Inhibitor of TANK binding kinase 1 (TBK1) and / or IκB kinase epsilon (IKKε); and ^ Wherein the subject suffers from a disease, preferably an immunological disorder, 5 such as one associated with increased type I interferon responses.

[0014] The term “inhibitor of Receptor-interacting serine / threonine-protein kinase 1 (RIPK1)”, “RIPK1 inhibitor” or “inhibitor of RIPK1” or “antagonist of RIPK1” or any similar expressions shall in context of the present invention encompass any compound or combination of compounds that have an activity as a modulator of the expression, function and / or stability of RIPK1, or of a variant 10 of RIPK1.

[0015] The term “inhibitor IκB kinase epsilon (IKKε)”, “inhibitor of IKKε” or “IKKε inhibitor” or “antagonist of IKKε” or any similar expressions shall in context of the present invention encompass any compound or combination of compounds that have an activity as a modulator of the expression, function and / or stability of IKKε, or of a variant of IKKε. 15

[0016] The term “Inhibitor of TANK binding kinase 1 (TBK1)”, “TBK1 inhibitor” or “inhibitor of TBK1” or “antagonist of TBK1” or any similar expressions shall in context of the present invention encompass any compound or combination of compounds that have an activity as a modulator of the expression, function and / or stability of TBK1, or of a variant of TBK1.

[0017] Preferred inhibitors of the invention are however inhibitors of the enzymatic function of the 20 above-mentioned kinases (kinase inhibitors).

[0018] Further, in context of the present invention the term “receptor interacting serine / threonine kinase 1” or “RIPK1” pertains to a human gene encoding for a protein according to the amino acid sequence shown in SEQ ID NO: 1. RIPK1 is also known as "receptor (TNFRSF)-interacting serine-threonine kinase 1" or "receptor-interacting protein kinase 1" (RIP) (HUGO Gene 25 Nomenclature Committee symbol: HGNC:10019, database version of November 2017). The human RIPK1 gene is located 6p25.2, homologs are known from mouse (MGI:108212; NCBI Gene: 19766) and rat (Rat Genome Database (RGD) ID: 1310158).

[0019] The terms “RIPK1-protein” or “protein of RIPK1” as used in context of the herein disclosed invention shall pertain to a protein (such as a full-length protein, fusion protein or partial protein) 30 comprising a sequence as shown in SEQ ID NO: 1. The terms shall also refer to a protein comprising the amino acid sequence according to SEQ ID NO: 1 with any protein modifications. Such protein modifications preferably do not alter the amino acid sequence of the polypeptide chain, but constitute a functional group, which is conjugated to the basic amino acid polymer chain. Protein modifications in context of the invention may be selected from a conjugation of 35 additional amino acid sequences to the RIPK1 amino acid chain, such as ubiquitination, sumoylation, methylation, or similar small protein conjugates. Other protein modifications include, but are not limited to, glycosylation, methylation, lipid-conjugation, or other natural or artificial post-translational modifications known to the skilled person. The terms “protein of a variant of RIPK1” and the like, shall have the corresponding meaning with respect to a variant of RIPK1. 5

[0020] The terms “RIPK1-mRNA” or “mRNA of RIPK1” as used in context of the herein disclosed invention shall pertain to a messenger ribonucleic acid (such as a full-length mRNA, fusion mRNA or partial mRNA, and / or splice-variants thereof) comprising a region encoding for an amino acid sequence as shown in SEQ ID NO: 1. The terms shall also refer to an mRNA comprising a region encoding for the amino acid sequence according to SEQ ID NO: 1 with any codon or nucleotide 10 modifications. Such modifications preferably would not alter the amino acid sequence of the encoded polypeptide chain. The terms “mRNA of a variant of RIPK1” and the like, shall have the corresponding meaning with respect to a variant of RIPK1.

[0021] A variant of RIPK1 is, in some embodiments, a protein comprising an amino acid sequence having at least 60%, 70%, 80%, 90%, preferably at least 80% such as at least 90%, 15 sequence identity to SEQ ID NO: 1, and most preferably at least 95% (such as at least 98%) sequence identity to SEQ ID NO: 1 (the human RIPK1 amino acid sequence). In one preferred embodiment of the invention, the variant of RIPK1 comprises an amino acid sequence with at least 80% sequence identity to the amino acid sequence shown in SEQ ID NO: 1.

[0022] As used herein, the terms “identical” or percent “identity”, when used anywhere herein in 20 the context of two or more nucleic acid or protein / polypeptide sequences, refer to two or more sequences or subsequences that are the same or have (or have at least) a specified percentage of amino acid residues or nucleotides that are the same (i.e., at, or at least, about 60% identity, preferably at, or at least, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93% or 94%, identity, and more preferably at, or at least, about 95%, 96%, 97%, 98%, 99%, or higher identity over a 25 specified region - preferably over their full length sequences - , when compared and aligned for maximum correspondence over the comparison window or designated region) as measured using a sequence comparison algorithms, or by manual alignment and visual inspection (see, e.g., NCBI web site).

[0023] Preferred RIPK1 inhibitors of the invention are small molecular compounds. Such30 compounds are preferably selected from necrostatin-1 stable (5-((7-chloro-1 H-indol-3-yl)methyl)- 3-methyl-2,4-imidazolidinedione), necrostatin-1 (5-(1H-indo1-3-ylmethyl)-3-methyl-2-thioxo-4- imidazolidinone), necrostatin-2 (5-[(7-chloro-1H-indol-3-yl)methyl]-3-methyl-2,4- imidazolidinedione), necrostatin-3 (3-phenyl-3,3a,4,5-tetrahydro-2H-benz[g]indazole, 3-phenyl- 2,3,3a,4-tetrahydro[1]benzopyrano[4,3-c]pyrazole, 3-phenyl-2,3,3a,4-35 tetrahydro[1]benzothiopyrano[4,3-c]pyrazole, 5,5-dioxo-3-phenyl-2,3,3a,4- tetrahydro[1]benzothiopyrano[4,3-c]pyrazole]), necrostatin-4 ((S)-N-(1-[2-chloro-6- fluorophenyl]ethyl)-5-cyano-1-methyl-1H-pyrrole-2-carboxamide), necrostatin-5 (2-[[3,4,5,6,7,8- hexahydro-3-(4-methoxyphenyl)-4-oxo[1]benzothieno[2,3-d]pyrimidin-2-yl]thio]-acetonitrile), necrostatin-7 (5-((3-(4-fluorophenyl)-1H-pyrazol-4-yl)methylene)-2-imino-3-(thiazol-2- yl)thiazolidin-4-one), necrostatin-21, (S)-5-benzyl-N-(7,9difluoro-2-oxo-2,3,4,5-tetrahydro-1H- 5 benzo[b]azepin-3-yl)1H-1,2,4-triazole-3-carboxamide (GSK3145095), N-benzyl-N-hydroxy-2,2- dimethylbutanamide (RIPA-56), DNL747 (SAR 443060), (S)-N-(5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-4,5,6,7-tetrahydro-1H-indazole-3-carboxamide, (S)-N-(5- methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-4H-1,2,4-triazole-3-carboxamide, (R)-5-methyl-N-((S)-5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-4,5,6,7-10 tetrahydro-1H-indazole-3-carboxamide, (R)-5-methyl-N-((S)-5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-4,5,6,7-tetrahydro-1H-indazole-3-carboxamide, (S)-6,6- dimethyl-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-4,5,6,7-tetrahydro- 1H-indazole-3-carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3- yl)-1',4',5',7'- tetrahydrospiro[cyclopropane-1,6'-indazole]-3'-carboxamide, N-((S)-5-methyl-4-15 oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-1,4,4a,5,5a,6- hexahydrocyclopropa[f]indazole-3-carboxamide, N-((S)-5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-3,4,5,5a,6,6a-hexahydrocyclopropa[e]indazole-1- carboxamide, (S)-5,5-dimethyl-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)- 4,5,6,7-tetrahydro-1H-indazole-3-carboxamide, (R)-N-((S)-5-methyl-4-oxo-2,3,4,5-20 tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5- (trifluoromethyl)-4,5,6,7-tetrahydro-1H-indazole-3- carboxamide, (R)-5-(tert-butyl)-N-((S)-5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin- 3-yl)-4,5,6,7-tetrahydro-1H-indazole-3-carboxamide, N-((S)-5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-phenyl-4,5,6,7- tetrahydro-1H-indazole-3-carboxamide, N-((S)-5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-(1H-pyrazol-1-yl)-4,5,6,7-25 tetrahydro-1H-indazole-3-carboxamide, (R)-1-methyl-N-((S)-5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-(trifluoromethyl)-4,5,6,7-tetrahydro-1H-indazole-3- carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5- (trifluoromethyl)-1H-indazole-3-carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-(trifluoromethyl)-1H-pyrazolo[3,4-b]pyridine-3-30 carboxamide, (R)-N-((S)-5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5- (trifluoromethyl)-4,5,6,7-tetrahydro-[1,2,3]triazolo [1,5-a]pyridine-3-carboxamide, (S)-N-(5- methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-7-(trifluoromethyl)imidazo[1,5- a]pyridine-1-carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)- 6- (trifluoromethyl)-[1,2,4]triazolo[4,3-b]pyridazine-3-carboxamide, (S)-5-isopropyl-N-(5-methyl- 35 4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-1H-pyrazolo[3,4-b]pyridine-3-carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-(perfluoroethyl)-1H- pyrazolo[3,4-b]pyridine-3-carboxamide, N-((S)-5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-5-(perfluoroethyl)-4,5,6,7-tetrahydro-[1,2,3]triazolo[1,5- a]pyridine-3-carboxamide, (S)-N-(5-methyl-4-oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)- 6-(perfluoroethyl)-[1,2,4]triazolo[4,3-b]pyridazine-3-carboxamide, (S)-5-benzyl-N-(5-methyl-4- oxo-2,3,4,5-tetrahydrobenzo[b][1,4]oxazepin-3-yl)isoxazole-3-carboxamide (GSK481), 1-[(5S)- 5 4,5-Dihydro-5-phenyl-1H-pyrazol-1-yl]-2,2-dimethyl-1-propanone, 2,2-Dimethyl-1-(5(S)-phenyl- 4,5-dihydro-pyrazol-1-yl)-propan-1-one (GSK963), (S)-5-benzyl-N-(5-methyl-4-oxo-2,3,4,5- tetrahydrobenzo[b][1,4]oxazepin-3-yl)-1H-1,2,4-triazole-3-carboxamide (GSK2982772), 5-(indol- 3-ylmethyl)-3-methyl-2-thio-hydantoin), and their derivatives or analogues. In a particularly preferred embodiment, the RIPK1 inhibitor is necrostatin-1 stable (Nec-1s). However, any other 10 known RIPK1 inhibitor may be used in context of the invention.

[0024] Further preferred RIPK1 inhibitors of the invention are small molecular compounds as disclosed in WO 2017 / 136727. In particular, a RIPK1 inhibitor useful in context of the present invention may be a compound selected from those compounds in Table 1, 2, 3 or 4 in WO 2017 / 136727 (such compounds of tables 1, 2, 3 and 4 in WO 2017 / 136727 are incorporated 15 herein by reference), or stereoisomers and mixtures of stereoisomers thereof. Also included within the disclosure is a compound selected from Table 1, 2, 3 or 4 in WO 2017 / 136727, as a pharmaceutically acceptable salt thereof.

[0025] Further preferred RIPK1 inhibitors of the invention are small molecular compounds as disclosed in WO 2018 / 109097. In particular, a RIPK1 inhibitor useful in context of the present 20 invention may be a compound selected from those compounds as listed beginning on page 4 line 3 and ending on page 6 line 25 of WO 2018 / 109097 - the listing of compounds is incorporated herein by reference.

[0026] Further, in context of the present invention the term “Inhibitor of nuclear factor kappa-B kinase subunit epsilon” or “IKKε” pertains to a human gene encoding for a protein according to 25 the amino acid sequence shown in SEQ ID NO: 2. IKKε is encoded by a gene also known as "IKBKE" (HUGO Gene Nomenclature Committee symbol: HGNC: 14552, database version of February 2023). The human IKKε gene is located 1q32.1.

[0027] The terms “IKKε -protein” or “protein of IKKε” as used in context of the herein disclosed invention shall pertain to a protein (such as a full-length protein, fusion protein or partial protein) 30 comprising a sequence as shown in SEQ ID NO: 2. The terms shall also refer to a protein comprising the amino acid sequence according to SEQ ID NO: 2 with any protein modifications. Such protein modifications preferably do not alter the amino acid sequence of the polypeptide chain, but constitute a functional group, which is conjugated to the basic amino acid polymer chain. Protein modifications in context of the invention may be selected from a conjugation of 35 additional amino acid sequences to the IKKε amino acid chain, such as ubiquitination, sumoylation, methylation, or similar small protein conjugates. Other protein modifications include, but are not limited to, glycosylation, methylation, lipid-conjugation, or other natural or artificial post-translational modifications known to the skilled person. The terms “protein of a variant of IKKε” and the like, shall have the corresponding meaning with respect to a variant of IKKε.

[0028] The terms “IKKε -mRNA” or “mRNA of IKKε” as used in context of the herein disclosed 5 invention shall pertain to a messenger ribonucleic acid (such as a full-length mRNA, fusion mRNA or partial mRNA, and / or splice-variants thereof) comprising a region encoding for an amino acid sequence as shown in SEQ ID NO: 2. The terms shall also refer to an mRNA comprising a region encoding for the amino acid sequence according to SEQ ID NO: 2 with any codon or nucleotide modifications. Such modifications preferably would not alter the amino acid sequence of the 10 encoded polypeptide chain. The terms “mRNA of a variant of IKKε” and the like, shall have the corresponding meaning with respect to a variant of IKKε.

[0029] A variant of IKKε is, in some embodiments, a protein comprising an amino acid sequence having at least 60%, 70%, 80%, 90%, preferably at least 80% such as at least 90%, sequence identity to SEQ ID NO: 2, and most preferably at least 95% (such as at least 98%) sequence 15 identity to SEQ ID NO: 2 (the human IKKε amino acid sequence). In one preferred embodiment of the invention, the variant of IKKε comprises an amino acid sequence with at least 80% sequence identity to the amino acid sequence shown in SEQ ID NO: 2.

[0030] Further, in context of the present invention the term “TANK binding kinase 1” or “TBK1” pertains to a human gene encoding for a protein according to the amino acid sequence shown in 20 SEQ ID NO: 3. TBK1is encoded by a gene also known as "TBK1" (HUGO Gene Nomenclature Committee symbol: HGNC: 11584, database version of February 2023). The human TBK1 gene is located on 12q14.2.

[0031] The terms “TBK1-protein” or “protein of TBK1” as used in context of the herein disclosed invention shall pertain to a protein (such as a full-length protein, fusion protein or partial protein) 25 comprising a sequence as shown in SEQ ID NO: 3. The terms shall also refer to a protein comprising the amino acid sequence according to SEQ ID NO: 3 with any protein modifications. Such protein modifications preferably do not alter the amino acid sequence of the polypeptide chain, but constitute a functional group, which is conjugated to the basic amino acid polymer chain. Protein modifications in context of the invention may be selected from a conjugation of 30 additional amino acid sequences to the TBK1 amino acid chain, such as ubiquitination, sumoylation, methylation, or similar small protein conjugates. Other protein modifications include, but are not limited to, glycosylation, methylation, lipid-conjugation, or other natural or artificial post-translational modifications known to the skilled person. The terms “protein of a variant of TBK1” and the like, shall have the corresponding meaning with respect to a variant of TBK1.

[0032] The terms “TBK1-mRNA” or “mRNA of TBK1” as used in context of the herein disclosed invention shall pertain to a messenger ribonucleic acid (such as a full-length mRNA, fusion mRNA or partial mRNA, and / or splice-variants thereof) comprising a region encoding for an amino acid sequence as shown in SEQ ID NO: 3. The terms shall also refer to an mRNA comprising a region 5 encoding for the amino acid sequence according to SEQ ID NO: 3 with any codon or nucleotide modifications. Such modifications preferably would not alter the amino acid sequence of the encoded polypeptide chain. The terms “mRNA of a variant of TBK1” and the like, shall have the corresponding meaning with respect to a variant of TBK1.

[0033] A variant of TBK1 is, in some embodiments, a protein comprising an amino acid sequence 10 having at least 60%, 70%, 80%, 90%, preferably at least 80% such as at least 90%, sequence identity to SEQ ID NO: 3, and most preferably at least 95% (such as at least 98%) sequence identity to SEQ ID NO: 3 (the human IKKε amino acid sequence). In one preferred embodiment of the invention, the variant of IKKε comprises an amino acid sequence with at least 80% sequence identity to the amino acid sequence shown in SEQ ID NO: 3. 15

[0034] A preferred TBK1 inhibitor may be selected from any one of the following compounds: MRT67307, BX795, TBK1-protac, TBK1 / IKKε-IN-1, TBK1 / IKKε-IN-2, GSK8612, Amlexanox, AZ13102909, MPI-0485520 (TBK1 / IKK dual inhibitor) and BAY-985.

[0035] Further TBK1 inhibitors usable in context of the invention are disclosed in RICHTERS ET AL: "Identification and Further Development of Potent TBK1 Inhibitors", ACS CHEMICAL 20 BIOLOGY, vol.10, no.1, 16 January 2015 (2015-01-16), pages 289 - 298, XP055397470, ISSN: 1554-8929, DOI: 10.1021 / cb500908d; DOUGLAS W. THOMSON ET AL: "Discovery of GSK8612, a Highly Selective and Potent TBK1 Inhibitor", ACS MEDICINAL CHEMISTRY LETTERS, vol.10, no.5, 11 March 2019 (2019-03-11), US, pages 780 - 785, XP055611911, ISSN: 1948-5875, DOI: 10.1021 / acsmedchemlett.9b00027 and BERGE, J. PHARM. SCI., vol.66, 1977, pages 1 – 19 25 (which are included herein by reference in their entirety.)

[0036] WO / 2019 / 233891 discloses TBK1 inhibitory compounds which are included herein by reference, any one of compounds 1 to 14 shown on pages 7-9 of WO / 2019 / 233891.

[0037] As used herein, the term “multi-target inhibitor” or “multi-target agent” refers to a single molecule having the capacity to interact with two different proteins (such as IKK epsilon) and at 30 least one other protein target such as TBK1 in vitro or in vivo including the capacity to inhibit the activity or normal function of said targets, e.g., to inhibit binding and / or enzymatic activity, such as a kinase activity of IKK epsilon and TBK1.

[0038] As used herein, the term “dual inhibitor” refers to a single molecule that interacts with and / or inhibits the activity or normal function, preferably a kinase activity, of two different target 35 proteins, for example, IKK epsilon and TBK1 in vivo or in vitro.

[0039] In context of the invention in some embodiments it is preferred that in (i) the compound or composition when administered to the subject induces an inhibition of the kinase activity of (x) and (y) in cells associated with the disorder.

[0040] A treatable disease of the invention is preferably a condition associated with increased 5 type I interferon responses (“interferonopathies”).

[0041] The compounds and compositions of the invention are expected to be useful in the treatment of interferonopathies. Diseases grouped under the term ‘interferonopathies’ are diseases having in common upregulation of type I interferon and / or increased expression of interferon-stimulated genes (ISG) profiles. Examples of ‘interferonopathies’ include familial 10 chilblain lupus, Aicardi-Goutières Syndrome, spondyloenchondromatosis, Singleton-Merten Syndrome, and SAVI (STING-associated vasculopathy with onset in infancy). Also, the compounds of the invention are expected to be useful in the treatment of cancer, specifically, in the treatment of patient populations in which the disease is associated with IKKε and / or TBK1 activity. IKKε has been implicated in breast cancer, including tamoxifen resistant breast cancer, 15 ovarian cancer, including cis-platin resistant ovarian cancer, cancer in which tumour growth and / or survival is dependent upon IKKε kinase activity, cancers harbouring Ras mutations and Ras- dependant tumours, and cancers involving amplification of the 1q32 gene locus. TBK1 has been implicated in cancers which harbour K-ras mutation and K-ras dependent tumours, cancers which harbour Ras mutations and cancers which are Ras-dependent, breast cancer, lung cancer, 20 particularly non-small cell lung cancer (NSCLC), ovarian cancer, prostate cancer, myeloma, leukemia, oral cancer, pancreatic cancer, bowel cancer and skin cancer.

[0042] In addition to cancer, specifically IKKε and / or TBK1 associated cancers, the compounds of the invention are expected to be particularly useful in the treatment and prevention of obesity (in which IKKε is implicated); and diseases in which hypoxia-induced angiogenesis is important, 25 for example the treatment and prevention of septic shock, and primary open angle glaucoma (in all of which TBK1 is implicated). Also viral

[0043] Preferably, the type I interferon-associated disease is characterized by a pathological increased type I interferon activity and / or accumulation and / or increased expression of interferon- stimulated genes (ISGs). 30

[0044] In some embodiments it is preferred that the compound or composition when administered to the subject selectively induces the inhibition of the kinase activity of (x) and (y).

[0045] In other alternative or additional embodiments, the compound or composition when administered to the subject induces inhibition (of the targets as mentioned herein) selectively in the myeloid compartment.

[0046] A preferred embodiment pertains to a compound or composition of the invention which is a combination of at least two compounds, wherein the first compound is an inhibitor of (x) and wherein the second compound is one or two compounds which is (are) an inhibitor of (y); or, wherein the compound or composition is a dual inhibitor having a first inhibitory activity towards 5 (x) and a second inhibitory activity towards (y), such as a fused compound comprising two structural elements of which one can bind to and inhibit (x) and one can bind to and inhibit (y). Preferably, the compound or composition involves in (y) an inhibition of both IKKε and TBK1.

[0047] In a second aspect, the invention pertains a RIPK1 inhibitor for use in the treatment or prevention of an adverse effect of an immune disease therapy in a subject, wherein the immune 10 disease therapy involves the administration of an IKKε- and / or TBK1-inhibitor to the subject suffering from the immune disease.

[0048] In a third aspect, the invention pertains to a method of treatment and / or prevention of a disease in a subject, the method comprising one or more steps of administering to the subject a therapeutically effective amount of a RIPK1 inhibitor, an IKKε inhibitor, and a TBK1 inhibitor. 15

[0049] Preferably, the immune disease therapy involves sequential or concomitant inhibition of IKKε and TBK1 in the subject suffering from the immune disease.

[0050] Diseases to be treated with the combinations of the invention are provided herein above in context of the first aspect.

[0051] In particular preferred embodiments, the invention relates in some of the herein disclosed 20 aspects and embodiments to a use of the combinations of the invention in the treatment of patients that carry mutations in the genes for human Trex1, Adar1 and / or Dnase2a, and any disease or disorder associated with these genese, or associated with mutated versions of these genes. For example, the invention pertains to diseases or disorders in subjects having one or more mutations in any one human Trex1, Adar1 and / or Dnase2, such as a type I 25 interferonopathies.

[0052] In another aspect, the invention further pertains to a RIPK1 inhibitor for use in the treatment of a IKKε / TBK1 inhibitor treatment related condition in a subject, wherein the subject is receiving a IKKε / TBK1 inhibitor treatment.

[0053] The IKKε / TBK1 inhibitor treatment related condition is preferably an adverse effect or 30 result of the IKKε / TBK1 inhibitor treatment that would otherwise result in a reduced IKKε / TBK1 inhibitor dosing and / or discontinuing of IKKε / TBK1 inhibitor treatment.

[0054] In this additional aspect, an adverse effect or result of the IKKε / TBK1 inhibitor treatment that would otherwise result in a reduced IKKε / TBK1 inhibitor dosing and / or discontinuing of IKKε / TBK1 inhibitor treatment relates to an adverse effect caused by the IKKε / TBK1 inhibitor 35 treatment that without any combinatorial treatments of the present invention would result in a change of treatment of the subject, namely discontinuing or reducing the dosage in order to avoid or reduce said adverse effect. The RIPK1 inhibitor used as suggested here therefore has the effect of increasing the therapeutic window of IKKε / TBK1 inhibitor treatment a subject. Alternatively, the RIPK1 is used in order to reduce and / or prevent adverse effects of IKKε / TBK1 5 inhibitor treatment in a subject.

[0055] In a fourth aspect, the invention pertains to a pharmaceutical composition comprising the compound or composition recited in the first or second aspect.

[0056] Whilst a compound of the invention may be used as the sole active agent, it is also possible for the compound to be used in combination with one or more further active agents 10 (which may also be referred to as “additional active agents”). Such combinations of compounds are preferred in context of the present invention. Such further active agents may be further compounds according to the invention, or they may be different therapeutic agents, for example agents targeting one of the diseases mentioned above, and particularly the same disease as that targeted by the compound of the invention. The compound of the invention may be co-formulated 15 with the one or more further active agent, or it may be formulated separately and administered consecutively, simultaneously or sequentially with the one or more further active agent.

[0057] The amount of active agent which is required to achieve a therapeutic effect will, of course, vary with the particular compound, the route of administration, the subject under treatment, including the type, species, age, weight, sex, and medical condition of the subject and the renal 20 and hepatic function of the subject, and the particular disorder or disease being treated, as well as its severity. An ordinarily skilled physician or veterinarian can readily determine and prescribe the effective amount of the drug required to prevent, counter or arrest the progress of the condition.

[0058] Advantageously, compounds of the present invention may be administered in a single 25 daily dose, or the total daily dosage may be administered in divided doses of two, three or four times daily.

[0059] The pharmaceutical formulations according to the invention include those suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous (bolus or infusion), and intraarticular), inhalation (including fine particle dusts or mists which may be generated by 30 means of various types of metered doses pressurized aerosols), nebulizers or insufflators, rectal, intraperitoneal and topical (including dermal, buccal, sublingual, and intraocular) administration, although the most suitable route may depend upon, for example, the condition and disorder of the recipient. Preferred pharmaceutical formulations according to the invention are those suitable for oral and parenteral administration; and more preferably those suitable for oral administration. 35 In another embodiment, compounds of the present invention can be administered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using those forms of transdermal skin patches well known to those of ordinary skill in the art. To be administered in the form of a transdermal delivery system, the dosage administration will, of course, be continuous rather than intermittent throughout the dosage regimen. 5

[0060] The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. All methods include the step of bringing the active agent into association with the carrier which constitutes one or more accessory ingredients. In general the formulations are prepared by uniformly and intimately bringing into association the active agent with liquid carriers or finely divided solid carriers or both 10 and then, if necessary, shaping the product into the desired formulation.

[0061] Oral dosages of the present invention, when used for the indicated effects, will range between about 0.01 mg per kg of body weight per day (mg / kg / day) to about 100 mg / kg / day, preferably 0.01 mg per kg of body weight per day (mg / kg / day) to 10 mg / kg / day, more preferably 0.1 to 5.0 mg / kg / day, and even more preferably 0.5 to 3 mg / kg / day for adult humans. An example 15 of a daily total oral dosage may be from 1 mg to 300 mg, more preferably from about 10 mg to about 250 mg, and even more preferably from about 75 mg to about 200 mg (for example 100 mg). For oral administration, the compositions are preferably provided in the form of tablets or other forms of presentation provided in discrete units containing 0.01, 0.05, 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0, 50.0, 100, and 500 milligrams of the active agent for the symptomatic adjustment 20 of the dosage to the patient to be treated. A medicament typically contains from about 0.01 mg to about 500 mg of the active agent, preferably from about 1 mg to about 300 mg of active agent, and more preferably from about 10 mg to about 250 mg of active agent, and most preferably from about 75 mg to about 200 mg of active agent (for example 100 mg of active agent). Intravenously, the most preferred doses will range from about 0.1 to about 10 mg / kg / minute during a constant 25 rate infusion.

[0062] Formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets, pills or tablets each containing a predetermined amount of the active agent; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid 30 emulsion. The active agent may also be presented as a bolus, electuary or paste.

[0063] The present compounds can, for example, be administered in a form suitable for immediate release or extended release. Immediate release or extended release can be achieved by the use of suitable pharmaceutical compositions comprising the present compounds, or, particularly in the case of extended release, by the use of devices such as subcutaneous implants 35 or osmotic pumps.

[0064] Exemplary compositions for oral administration include suspensions which can contain, for example, microcrystalline cellulose for imparting bulk, alginic acid or sodium alginate as a suspending agent, methylcellulose as a viscosity enhancer, and sweeteners or flavoring agents such as those known in the art; and immediate release tablets which can contain, for example, 5 microcrystalline cellulose, dicalcium phosphate, starch, magnesium stearate, calcium sulfate, sorbitol, glucose and / or lactose and / or other excipients, binders, extenders, disintegrants, diluents and lubricants such as those known in the art. Suitable binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, 10 waxes and the like. Disintegrators include without limitation starch, methylcellulose, agar, bentonite, xanthan gum and the like. The compounds of the invention can also be delivered through the oral cavity by sublingual and / or buccal administration. Molded tablets, compressed tablets or freeze-dried tablets are exemplary forms which may be used. Exemplary compositions include those formulating the present compound(s) with fast dissolving diluents such as mannitol, 15 lactose, sucrose and / or cyclodextrins. Also included in such formulations may be high molecular weight excipients such as celluloses (avicel) or polyethylene glycols (PEG). Such formulations can also include an excipient to aid mucosal adhesion such as hydroxy propyl cellulose (HPC), hydroxy propyl methyl cellulose (HPMC), sodium carboxy methyl cellulose (SCMC), maleic anhydride copolymer (e.g., Gantrez), and agents to control release such as polyacrylic copolymer 20 (e.g. Carbopol 934). Lubricants, glidants, flavors, coloring agents and stabilizers may also be added for ease of fabrication and use. Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like. For oral administration in liquid form, the oral drug components can be combined with any oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, 25 and the like.

[0065] Formulations for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non- aqueous sterile suspensions which may include suspending agents and thickening agents. The 30 formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example saline or water-for-injection, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. Exemplary compositions for parenteral 35 administration include injectable solutions or suspensions which can contain, for example, suitable non-toxic, parenterally acceptable diluents or solvents, such as mannitol, 1,3-butanediol, water, Ringer's solution, an isotonic sodium chloride solution, or other suitable dispersing or wetting and suspending agents, including synthetic mono- or diglycerides, and fatty acids, including oleic acid, or Cremophor.

[0066] Exemplary compositions for nasal, aerosol or inhalation administration include solutions in saline, which can contain, for example, benzyl alcohol or other suitable preservatives, 5 absorption promoters to enhance bioavailability, and / or other solubilizing or dispersing agents such as those known in the art.

[0067] Formulations for rectal administration may be presented as a suppository with the usual carriers such as cocoa butter, synthetic glyceride esters or polyethylene glycol. Such carriers are typically solid at ordinary temperatures, but liquefy and / or dissolve in the rectal cavity to release 10 the drug.

[0068] Formulations for topical administration in the mouth, for example buccally or sublingually, include lozenges comprising the active agent in a flavoured basis such as sucrose and acacia or tragacanth, and pastilles comprising the active agent in a basis such as gelatin and glycerine or sucrose and acacia. Exemplary compositions for topical administration include a topical carrier 15 such as Plastibase (mineral oil gelled with polyethylene).

[0069] In a fifth aspect, the invention pertains a method for screening an IKKε-Inhibitor suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses in a subject, (f) Providing a test cell, 20 (g) Optionally providing one or more control cells; (h) Contacting the test cell with (i) a candidate IKKε-Inhibitor, while the test cell is contacted with a known TBK1 inhibitor and a known RIPK1 inhibitor; or (ii) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with 25 a known RIPK1 inhibitor; or (iii) a candidate IKKε-Inhibitor and a candidate TBK1 inhibitor, while the cell is contacted with a known RIPK1 inhibitor; or (iv) a candidate IKKε-Inhibitor, a candidate TBK1 inhibitor and a candidate RIPK1 inhibitor; or (v) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a candidate RIPK1 inhibitor; (i) Optionally, where the control cell is not contacted in (a) to (d) with any of the candidate inhibitors; or alternatively, contacting the control cell with the candidate 5 inhibitors but not the known inhibitors; (j) Determining in the test cell at least cell death and release of one or more cytokines before and after step (iii), or in comparison to the control cell or in comparison to a reference value; wherein if in step (v) the following condition is determined: no increased cell death and / or a 10 reduced release of the one or more test cytokines, then the candidate inhibitors in one or more of (a) to (d) are suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses, wherein the triple combination comprises the inhibitors of the respective scenario (a) to (d) for which the condition was determined.

[0070] Preferably, the known RIPK1 inhibitor is selected from the group consisting of IPK1-IN-7, 15 GNE684, GSK3145095, GSK298277259, GSK'547, GSK3502421, DHP76, DHP77, AV123, MBM105, ZB-R-55, PN10, RIPK1 kinase inhibitor 38, GSK′963, DNL758, DNL788, Necrostatin- 1s (Nec1s) or GSK2982772.

[0071] The known TBK1 inhibitor may be, preferably, selected from MRT67307, BX795, TBK1- protac, TBK1 / IKKε-IN-1, TBK1 / IKKε-IN-2, GSK8612, Amlexanox, AZ13102909, MPI-0485520 20 (TBK1 / IKK dual inhibitor) and BAY-985.

[0072] In some embodiments, the screening method of the invention may comprise a further step of contacting a nonhuman test animal with the so identified triple combination.

[0073] In some embodiments, the test cell and / or the control cell is a bone marrow derived macrophage. 25

[0074] In some preferred embodiments, the one or more test cytokines are IL-1alpha and / or IL- 1beta.

[0075] Preferably, in certain embodiments of the invention, candidate inhibitors are selective inhibitors of the kinase activity of their target, and not inhibitors that reduce overall protein activity of other protein functions, protein expression and / or protein stability. 30

[0076] Preferably, the candidate inhibitor is a small molecular compound suspected to harbour an activity as a kinase inhibitor.

[0077] The term “kinase inhibitor” generally refers to molecules that antagonize or inhibit phosphorylation-dependent cell signaling and / or growth pathways in a cell. Kinase inhibitors may be naturally occurring or synthetic and include small molecules that have the potential to be administered as oral therapeutics. Kinase inhibitors have the ability to rapidly and specifically 5 inhibit the activation of the target kinase molecules. Protein kinases are attractive drug targets, in part because they regulate a wide variety of signaling and growth pathways and include many different proteins.

[0078] A “small molecular compound” or “small molecular inhibitor” shall refer to a generally oranic molecule of less than 5000 D. More preferably the compound has less than 2500 D or less 10 than, preferably 1000 D.

[0079] The term "candidate compound" or "candidate substrate" is meant to refer to any compound wherein the characterization of the compound's susceptibility to enzymatic metabolism is desirable, in particular towards an inhibition of the respective target or targets (which in context of the invention is one or mor a combination of RIPK1, TBK1 and IKK epsilon). Exemplary 15 candidate compounds or substrates include xenobiotics such as drugs and other therapeutic agents, carcinogens and environmental pollutants, as well as endobiotics such as steroids, fatty acids and prostaglandins, or other small molecular compounds. A candidate compound may be in particular derived from a screening library of potential kinase inhibitors. Such libraries are well known to the skilled artisan. 20

[0080] The invention in certain aspects and embodiments as defined herein above, may relate to the following specific itemized embodiments: Item 1: A compound or composition for use in the treatment of a condition in a subject, wherein (a) The compound or composition when administered to the subject induces an inhibition of 25 (x) RIPK1 and (y) Inhibitor of TANK binding kinase 1 (TBK1) and / or IκB kinase epsilon (IKKε); (b) Wherein the subject suffers from a disease associated with increased type I interferon responses. Item 2: The compound or composition for use of item 1, wherein the disease is a condition associated with increased type I interferon responses. 30 Item 3: The compound or composition for use of item 1 or 2, wherein the type I interferon- associated disease is characterized by a pathological increased type I interferon activity and / or accumulation and / or increased expression of interferon-stimulated genes (ISGs). Item 4: The compound or composition for use of any one of items 1 to 3, wherein the compound or composition is a combination of at least two compounds, wherein the first compound is an inhibitor of (x) and wherein the second compound is one or two compounds which is (are) an inhibitor of (y); or, wherein the compound or composition is a dual inhibitor having a first 5 inhibitory activity towards (x) and a second inhibitory activity towards (y), such as a fused compound comprising two structural elements of which one can bind to and inhibit (x) and one can bind to and inhibit (y). Item 5: The compound or composition for use of any one of items 1 to 4, wherein the condition is an autoimmune or inflammatory disease, infection or cancer, preferably wherein the 10 autoimmune or inflammatory disease or cancer is associated with increased type I interferon responses. Item 6: A RIPK1 inhibitor for use in the treatment or prevention of an adverse effect of an immune disease therapy in a subject, wherein the immune disease therapy involves the administration of an IKKε- and / or TBK1-inhibitor to the subject suffering from the immune disease. 15 Item 7: The RIPK1 inhibitor for use of item 6, wherein the immune disease therapy involves sequential or concomitant inhibition of IKKε and TBK1 in the subject suffering from the immune disease. Item 8: The RIPK1 inhibitor for use of item 6 or 7, wherein the immune disease is an autoimmune disease or cancer, or is an inflammatory disease, or viral disease. 20 Item 9: A pharmaceutical composition comprising the compound or composition of any one of items 1 to 5, together with a pharmaceutically acceptable carrier and / or excipient. Item 10: A method for screening an IKKε-Inhibitor suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses in a subject, 25 (a) Providing a test cell, (b) Optionally providing one or more control cells; (c) Contacting the test cell with (i) a candidate IKKε-Inhibitor, while the test cell is contacted with a known TBK1 inhibitor and a known RIPK1 inhibitor; or (ii) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a known RIPK1 inhibitor; or (iii) a candidate IKKε-Inhibitor and a candidate TBK1 inhibitor, while the cell is contacted with a known RIPK1 inhibitor; or 5 (iv) a candidate IKKε-Inhibitor, a candidate TBK1 inhibitor and a candidate RIPK1 inhibitor; or (v) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a candidate RIPK1 inhibitor; (d) Optionally, where the control cell is not contacted in (a) to (d) with any of the candidate 10 inhibitors; or alternatively, contacting the control cell with the candidate inhibitors but not the known inhibitors; (e) Determining in the test cell at least cell death and release of one or more cytokines before and after step (iii), or in comparison to the control cell or in comparison to a reference value; Wherein if in step (v) the following condition is determined: no increased cell death and / or a 15 reduced release of the one or more test cytokines, then the candidate inhibitors in one or more of (a) to (d) are suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses, wherein the triple combination comprises the inhibitors of the respective scenario (a) to (d) for which the condition was determined. Item 11: The method of item 10, wherein the known RIPK1 inhibitor is selected from the 20 group consisting of IPK1-IN-7, GNE684, GSK3145095, GSK298277259, GSK'547, GSK3502421, DHP76, DHP77, AV123, MBM105, ZB-R-55, PN10, RIPK1 kinase inhibitor 38, GSK′963, DNL758, DNL788, Necrostatin-1s (Nec1s) or GSK2982772. Item 12: The method of item 10 or, wherein the known TBK1 inhibitor is selected from MRT67307, BX795, TBK1-protac, TBK1 / IKKε-IN-1, TBK1 / IKKε-IN-2, GSK8612, Amlexanox, 25 AZ13102909, MPI-0485520 (TBK1 / IKK dual inhibitor) and BAY-985. Item 13: The method of any one of items 10 to 12, wherein the type I interferon response is in context of a disease selected from an auto inflammatory disease, infectious or auto immune disease. Item 14: The method of any one of items 10 to 13, wherein the candidate inhibitors are selective inhibitors of the kinase activity of their target, and not inhibitors that reduce overall protein activity of other protein functions, protein expression and / or protein stability. Item 15: The method of any one of items 10 to 14, wherein the candidate inhibitor is a small 5 molecular compound suspected to harbour an activity as a kinase inhibitor.

[0081] The terms “of the [present] invention”, “in accordance with the invention”, “according to the invention” and the like, as used herein are intended to refer to all aspects and embodiments of the invention described and / or claimed herein.

[0082] As used herein, the term “comprising” is to be construed as encompassing both “including” 10 and “consisting of”, both meanings being specifically intended, and hence individually disclosed embodiments in accordance with the present invention. Where used herein, “and / or” is to be taken as specific disclosure of each of the two specified features or components with or without the other. For example, “A and / or B” is to be taken as specific disclosure of each of (i) A, (ii) B and (iii) A and B, just as if each is set out individually herein. In the context of the present invention, 15 the terms “about” and “approximately” denote an interval of accuracy that the person skilled in the art will understand to still ensure the technical effect of the feature in question. The term typically indicates deviation from the indicated numerical value by ±20%, ±15%, ±10%, and for example ±5%. As will be appreciated by the person of ordinary skill, the specific such deviation for a numerical value for a given technical effect will depend on the nature of the technical effect. 20 For example, a natural or biological technical effect may generally have a larger such deviation than one for a man-made or engineering technical effect. As will be appreciated by the person of ordinary skill, the specific such deviation for a numerical value for a given technical effect will depend on the nature of the technical effect. For example, a natural or biological technical effect may generally have a larger such deviation than one for a man-made or engineering technical 25 effect. Where an indefinite or definite article is used when referring to a singular noun, e.g. "a", "an" or "the", this includes a plural of that noun unless something else is specifically stated.

[0083] It is to be understood that application of the teachings of the present invention to a specific problem or environment, and the inclusion of variations of the present invention or additional features thereto (such as further aspects and embodiments), will be within the capabilities of one 30 having ordinary skill in the art in light of the teachings contained herein.

[0084] Unless context dictates otherwise, the descriptions and definitions of the features set out above are not limited to any particular aspect or embodiment of the invention and apply equally to all aspects and embodiments which are described.

[0085] All references, patents, and publications cited herein are hereby incorporated by reference in their entirety. BRIEF DESCRIPTION OF THE FIGURES AND SEQUENCES

[0086] The figures show:

[0087] Figure 1: Mice expressing kinase inactive TBK1D138N die during embryogenesis and are rescued by crossing to mice expressing kinase inactive RIPK1D138N. A) Generation of mice with Tbk1D135Nalleles using CRISPR / Cas9-mediated genome editing. The genome map is depicted together with the corresponding Sanger DNA sequencing chromatogram of a PCR product of genomic DNA of F1 progeny obtained by crossing a Tbk1wt / D135Nfounder with C57BL / 6 mice. B) Expected and observed frequency of mice of the indicated genotypes in offspring from the intercross of Tbk1D135N / D135N; Ripk1WT / D138Nparents. Note that Tbk1D135N / D135Nmice are only born in the presence of heterozygous or homozygous RIPK1D138N expression.

[0088] Figure 2: Mice expressing kinase inactive alleles of both TBK1 (Tbk1D135N / D135N) and IKKe (IkkeK38A / K38A) die during embryogenesis and are rescued by crossing to mice expressing kinase inactive RIPK1D138N. A) Generation of mice with Tbk1D135Nalleles using CRISPR / Cas9-mediated genome editing. The genome map is depicted together with the corresponding Sanger DNA sequencing chromatogram of a PCR product of genomic DNA from an IkkeK38A / K38Amouse. B) Expected and observed frequency of mice of the indicated genotypes in offspring from the intercross of IkkeK38A / K38A; Tbk1WT / D135N; Ripk1WT / D138Nparents. Note that Tbk1D135N / D135N; IkkeK38A / K38Amice are only born in the presence of heterozygous or homozygous RIPK1D138N expression.

[0089] Figure 3: Mice expressing kinase inactive alleles of both TBK1 (Tbk1D135N / D135N) and IKKe (IkkeK38A / K38A) die during embryogenesis and are rescued by crossing to mice expressing kinase inactive RIPK1D138N. A) Generation of mice with Tbk1D135Nalleles using CRISPR / Cas9-mediated genome editing. The genome map is depicted together with the corresponding Sanger DNA sequencing chromatogram of a PCR product of genomic DNA from an IkkeK38A / K38Amouse. B) Expected and observed frequency of mice of the indicated genotypes in offspring from the intercross of IkkeK38A / K38A; Tbk1WT / D135N; Ripk1WT / D138Nparents. Note that Tbk1D135N / D135N; IkkeK38A / K38Amice are only born in the presence of heterozygous or homozygous RIPK1D138N expression.

[0090] Figure 4: Combined inhibition of TBK1 and IKKe kinase activity causes liver damage in a RIPK1-kinase activity dependent manner. Graph showing serum alanine amino transferase values in 8-12 week-old mice with the indicated genotypes. ALT levels were measured calorimetrically using a Roche Cobas C111 analyser. Ordinary one-way ANOVA with Tukey’s multiple comparisons test were used for statistical assessments. (ns=non-significant, ****P < 0.0001). Dots indicate individual mice. Data is displayed as mean ± SD.

[0091] Figure 5: Genetic inhibition of TBK1 and IKKε kinase activity completely abrogates IRF3 phosphorylation and IFN-β production induced by LPS stimulation in bone marrow derived macrophages. RIPK1D138N / WTand IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / WTbone marrow-derived macrophages were stimulated with medium or LPS (100ng / ml). (A) Cell lysates were immunoblotted for phosho-IRF3, IRF3, IKKε and TBK1 after indicated times. (B) After 24 hours of incubation, the supernatant was assayed for IFN-β using ELISA.

[0092] Figure 6: Schematic depicting the generation of mice with myeloid cell-specific inhibition of TBK1-kinase kinase activity using the Cre / lox recombination system. Tbk1FL / D135Nmice, which harbour one kinase inactive TBK1D135N allele and one loxP- flanked TBK1 allele, were crossed with Cx3Cr1-Cre mice expressing Cre recombinase specifically in myeloid cells. The expression of Cre recombinase leads to the deletion of the loxP-flanked Tbk1 exons, culminating in the expression of only kinase inactive TBK1- D135N in myeloid cells.

[0093] Figure 7: Combined inhibition of TBK1 and IKKe kinase activities in myeloid cells causes in neutrophilia and splenomegaly that depend on RIPK1 kinase activity.

[0094] (A) Spleen and body weight were measured and the ratio of spleen to body weight are shown for 8-12 week-old mice of the indicated genotypes. (B) Granulocyte numbers in peripheral blood, measured using a blood analyser, of 8-12 week-old mice with the indicated genotypes are shown. Dots indicate individual mice. Ordinary one-way ANOVA with Tukey’s multiple comparisons test were used for statistical assessments. (ns=non- significant, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001). Data is displayed as mean ± SD.

[0095] Figure 8: Treatment of BMDMs with the TBK1 / IKKe kinase inhibitor BX795 and MRT67307 causes IL-1β release in a RIPK1-kinase activity-dependent manner. (A-B) Bone marrow derived macrophages (BMDMs) from wild-type (Ripk1wt / wt) and Ripk1D138N / D138Nmice were treated with 10μM, 5 μM, 2,5 μM, 1,25 μM and 0,6 μM MRT67307. (A) After 24 hours the supernatants were collected and assayed for IL-1α and IL-1β using specific ELISA assays. (B) Cells were imaged for 24 hours in an Incucyte imaging system and cell death was quantified using Diyo-1, a cell impermeable DNA intercalating dye. (C-D) BMDMs from wild-type mice were treated with 10μM, 5 μM, 2,5 μM and 1,25 μM BX795 or MRT67307. (C) After 24 hours the supernatants were collected and assayed for IL-1β using specific ELISA assays. (D) Cells were imaged for 20 hours in an Incucyte imaging system and cell death was quantified using Diyo-1. Data are pooled from three-independent experiments, with n = 1 mice per experiment. Data is displayed as mean ± SD.

[0096] Figure 9: Combined genetic inhibition of TBK1 (Tbk1D135N / D135N) and RIPK1 (Ripk1D138N / D138N) activities strongly inhibit the development of myocarditis in Trex1KO / KOmice. Heart sections of 9-10 week old mice of the indicated genotypes were immunostained with an antibody against the immune cell markers CD45 and CD3 (brown color indicates positively stained cells), or were stained with Masson’s Trichrome and TUNEL staining. Collagen stained blue and cytoplasm red with Masson’s Trichrome staining. Nuclei were counterstained with DAPI (blue) in TUNEL staining. TUNEL positive cells, which have nuclear fragmentation, are displayed in green. Pictures shown are representative of Trex1KO / KO(n=6), Trex1KO / KO; Tbk1D135N / D135N; Ripk1D138N / D138N(n=7), Trex1KO / KO; Ripk1D138N / D138N(n=4)Trex1KO / WT; Tbk1D135N / D135N; Ripk1D138N / D138N(n=4).

[0097] Figure 10: Combined genetic inhibition of TBK1 and RIPK1 kinase activity prolongs the survival of Adar1 mZa / - mice. Kaplan–Meier survival graph shows the survival of mice with the indicated genotypes.1: Control (n=36); 2: Adar1mZa / -(n=22); 3: Adar1mZa / -(n=22) Tbk1D135N / D135NRipk1D318N / D138N.

[0098] The sequences show:

[0099] SEQ ID NOs.1: (RIPK1 isoform 1, human – canonical sequence):

[0100] MQPDMSLNVIKMKSSDFLESAELDSGGFGKVSLCFHRTQGLMIMKTVYKGPNCIEHNE ALLEEAKMMNRLRHSRVVKLLGVIIEEGKYSLVMEYMEKGNLMHVLKAEMSTPLSVKGRIILEIIE GMCYLHGKGVIHKDLKPENILVDNDFHIKIADLGLASFKMWSKLNNEEHNELREVDGTAKKNGG TLYYMAPEHLNDVNAKPTEKSDVYSFAVVLWAIFANKEPYENAICEQQLIMCIKSGNRPDVDDIT EYCPREIISLMKLCWEANPEARPTFPGIEEKFRPFYLSQLEESVEEDVKSLKKEYSNENAVVKR MQSLQLDCVAVPSSRSNSATEQPGSLHSSQGLGMGPVEESWFAPSLEHPQEENEPSLQSKLQ DEANYHLYGSRMDRQTKQQPRQNVAYNREEERRRRVSHDPFAQQRPYENFQNTEGKGTAYS SAASHGNAVHQPSGLTSQPQVLYQNNGLYSSHGFGTRPLDPGTAGPRVWYRPIPSHMPSLHNI PVPETNYLGNTPTMPFSSLPPTDESIKYTIYNSTGIQIGAYNYMEIGGTSSSLLDSTNTNFKEEPA AKYQAIFDNTTSLTDKHLDPIRENLGKHWKNCARKLGFTQSQIDEIDHDYERDGLKEKVYQMLQ KWVMREGIKGATVGKLAQALHQCSRIDLLSSLIYVSQN

[0101] SEQ ID NOs.2: (IKK epsilon isoform 1, human – canonical sequence):

[0102] MQSTANYLWHTDDLLGQGATASVYKARNKKSGELVAVKVFNTTSYLRPREVQVREFEV LRKLNHQNIVKLFAVEETGGSRQKVLVMEYCSSGSLLSVLESPENAFGLPEDEFLVVLRCVVAG MNHLRENGIVHRDIKPGNIMRLVGEEGQSIYKLTDFGAARELDDDEKFVSVYGTEEYLHPDMYE RAVLRKPQQKAFGVTVDLWSIGVTLYHAATGSLPFIPFGGPRRNKEIMYRITTEKPAGAIAGAQR RENGPLEWSYTLPITCQLSLGLQSQLVPILANILEVEQAKCWGFDQFFAETSDILQRVVVHVFSL SQAVLHHIYIHAHNTIAIFQEAVHKQTSVAPRHQEYLFEGHLCVLEPSVSAQHIAHTTASSPLTLF STAIPKGLAFRDPALDVPKFVPKVDLQADYNTAKGVLGAGYQALRLARALLDGQELMFRGLHW VMEVLQATCRRTLEVARTSLLYLSSSLGTERFSSVAGTPEIQELKAAAELRSRLRTLAEVLSRCS QNITETQESLSSLNRELVKSRDQVHEDRSIQQIQCCLDKMNFIYKQFKKSRMRPGLGYNEEQIH KLDKVNFSHLAKRLLQVFQEECVQKYQASLVTHGKRMRVVHETRNHLRLVGCSVAACNTEAQ GVQESLSKLLEELSHQLLQDRAKGAQASPPPIAPYPSPTRKDLLLHMQELCEGMKLLASDLLDN NRIIERLNRVPAPPDV

[0103] SEQ ID NOs.3: (TBK1 isoform 1, human – canonical sequence):

[0104] MQSTSNHLWLLSDILGQGATANVFRGRHKKTGDLFAIKVFNNISFLRPVDVQMREFEVL 5 KKLNHKNIVKLFAIEEETTTRHKVLIMEFCPCGSLYTVLEEPSNAYGLPESEFLIVLRDVVGGMNH LRENGIVHRDIKPGNIMRVIGEDGQSVYKLTDFGAARELEDDEQFVSLYGTEEYLHPDMYERAV LRKDHQKKYGATVDLWSIGVTFYHAATGSLPFRPFEGPRRNKEVMYKIITGKPSGAISGVQKAE NGPIDWSGDMPVSCSLSRGLQVLLTPVLANILEADQEKCWGFDQFFAETSDILHRMVIHVFSLQ QMTAHKIYIHSYNTATIFHELVYKQTKIISSNQELIYEGRRLVLEPGRLAQHFPKTTEENPIFVVSR 10 EPLNTIGLIYEKISLPKVHPRYDLDGDASMAKAITGVVCYACRIASTLLLYQELMRKGIRWLIELIK DDYNETVHKKTEVVITLDFCIRNIEKTVKVYEKLMKINLEAAELGEISDIHTKLLRLSSSQGTIETSL QDIDSRLSPGGSLADAWAHQEGTHPKDRNVEKLQVLLNCMTEIYYQFKKDKAERRLAYNEEQI HKFDKQKLYYHATKAMTHFTDECVKKYEAFLNKSEEWIRKMLHLRKQLLSLTNQCFDIEEEVSK YQEYTNELQETLPQKMFTASSGIKHTMTPIYPSSNTLVEMTLGMKKLKEEMEGVVKELAENNHI 15 LERFGSLTMDGGLRNVDCL

[0002] EXAMPLES

[0105] Certain aspects and embodiments of the invention will now be illustrated by way of example and with reference to the description, figures and tables set out herein. Such examples of the methods, uses and other aspects of the present invention are representative only, and should not be taken to limit the scope of the present invention to only such representative examples. Material and Methods

[0106] Mice (sequences are shown in sequence listing in SEQ ID Nos: 4 to 16)

[0107] The generation of Ripk1D138Nmice were previously described (Polykratis et al., 2014). Trex1ko, Tbk1D135N, and IkbkeK38Amice were generated using CRISPR / Cas9 technology. Of note, Trex1 is a single exon gene. For Trex1komice, a short-guide (sg)RNA (5’- TTCCAGGTCTAAGAAGATGA -3’) and (5’- CCTGGGCAGTAAGTCAA GAG, for Tbk1D135Nmice, a short-guide (sg)RNA (5′- TGTTGCCTGGCTTGATAT CT - 3′) and long single stranded(ss) oligonucleotide (5′- AAATCCGTGAGTTTGTACACAGACTGGCCGTCCTCCCCTATGACGCGCATGATG TTGCCTGGCTTGATATtTCGaTGCACTATGCCGTTCTCTCGGAGATGATTCATCC CGCCCACTTCAGCAATAGGTACAAAACAAAGG -3′), and for IkbkeK38Amice a sgRNA (5′- CGGGGAGGTGGTTGC TGTAA-3′) and long ss oligonucleotide (5′- GCAGAGGTGGTCCCCATTGCTTACTCTCGCCCTGTGCCCACGCTAGAAATCCG GGGAGGTGGTTGCTGTAgctGTCTTCAACTCAGCCAGCTATCGGCGACCTCCTG AGGTTCAGGTGAGGGAGTTTGAGGTCCTGCGGAGGC -3′) and co-injected into fertilized wild- type oocytes together with Cas9 protein and mRNA.

[0108] The deletion of Trex1 exon was confirmed using the following primers: (5’- ATCCCAC TAGAACAACCCTGCC -3’) and (5’- TTCAGACTCCGCACCCTCATTT - 3’). For genotyping of IkbkeK38Amice, 330bp region spanning the mutation site of Ikbke were amplified using Universal Forward (5′- ACC CTT GAG GGA CAT CAG GT -3′) and Universal Reverse (5’- CGATGTTCTGGTGATTCAGCCT -3’), and WT (259bp) and mutant (107bp) sequence were respectively amplified using WT Reverse (5’- GCTGGCTGAGTTG AAGACCTTT -3’) and (5’- GGA GGT GGT TGC TGT AGC TG -3’) primers in a single PCR reaction. TBK1- D135N mutation destroyed the EcoRV restriction site on this genomic region. For genotyping of the Tbk1D135Nallele, the amplicon generated with primers (5’- ACTCCACATAGAACGTGCCTC - 3’) and (5’ ACACACTTGTGCCTGA GGATT -3’) were incubated with EcoRV-HF (Neb, cat no R3195) restriction enzyme for 90 minutes at 37°C. PCR amplicons were visualized on 2.0% agarose gels stained with ethidium bromide.

[0109] Mice were maintained at the specific-pathogen-free (SPF) animal facilities of the Institute for Genetics and the CECAD Research Center of the University of Cologne. All mouse procedures were conducted in accordance with national and institutional guidelines, and protocols were approved by the responsible local authorities in Germany (Landesamt für Natur, Umwelt und 5 Verbraucherschutz Nordrhein- Westfalen). Mice of the indicated genotype were assigned at random to groups and experiments were not blinded. Both male and female mice are included in all groups.

[0110] Blood analysis

[0111] Blood-cell-count analysis was performed in 40µl of peripheral blood using an Abacus 10 Junior Vet analyser according to the manufacturer's instructions.

[0112] Isolation and culture of bone marrow derived macrophages

[0113] For the preparation of BMDMs, bone marrow cells were plated on 145 mm bacterial Petri dishes (Greiner, cat no 639161_120) in DMEM medium (Invitrogen, cat no 41965062) supplemented with 20ng / ml mM-CSF (ImmunoTools, cat no.12343113), 10% fetal calf serum, 15 penicillin / streptomycin and sodium pyruvate. The cells were plated in tissue-coated dishes at Day 6 of culture. The experiments were performed on Day 7 of culture. The BMDMs were stimulated with MRT67307 (Selleckchem, cat no. S7948) or 100ng / ml ultrapure LPS (Enzo Life Sciences, cat no. ALX-581–010- L002) before the collection of samples.

[0114] Cytokine analysis 20

[0115] The cytokine levels in the supernatants from macrophage cultures were determined by ELISA kits for IL-1α (Thermo Scientific, cat no.88–5019–22), IL-1β (Thermo Scientific, cat. no. 88–7013–88), and IFNβ (Biolegend, cat no. BLD-43907) according to the manufacturers' instructions.

[0116] Immunoblotting 25

[0117] Cell lysates were denatured in 2×Laemmli buffer (Biorad, cat no 34095) supplement with 4% β-mercaptoethanol and 10mM Dithiothreitol. The protein samples were subsequently boiled at 95 °C for 4 minutes and separated by NuPageTM(Thermo Scientific, cat no. NP0336BOX). Separated proteins were transferred to PVDF membranes and were blocked using SuperBlockTMblocking buffer (Thermo Scientific, cat no.37515). Incubation with secondary antibody and 30 washing of the membrane were done in TBS supplemented with 0.1% Tween-20 (v / v) and 5% (w / v) non-fat dry milk. Incubation of the membranes with primary antibodies was performed in TBS supplemented with 0.1% Tween-20 (v / v), 5% (w / v) BSA and 0.01% sodium azide.

[0118] For re-blotting, membrane was stripped using Restore™ Western Blot Stripping

[0119] Buffer (Thermo, cat no.21059). The immunoblots were incubated overnight with primary antibodies against p-IRF3 Ser396 (Thermo Scientific, cat no MA5-14947), IRF3 (CST, cat no. 4302), IKKe (CST, cat no.3416), and TBK1 (CST, cat no.3013). anti-rabbit secondary antibody (Amersham Pharmacia, NA934V) was used to detect protein using chemiluminescence (Thermo 5 Scientific, cat no 34578 or Thermo Scientific cat no. 34095). Chemiluminescent signal on membranes was measured using Vilber FX system.

[0120] Biochemical serum analysis

[0121] Alanine aminotransferase (ALT) were measured in blood serum using a standard assay in a Cobas C111 biochemical analyzer (Roche, Mannheim, Germany). 10

[0122] Cell death assay

[0123] BMDMs were seeded in 96-well plate (3x105cells per well) one day before the treatment in the presence of 20ng / ml M-CSF (ImmunoTools, cat no.12343113).

[0124] Cells were treated with different concentrations of MRT67307 (Selleckchem, cat no. S7948) or BX795 (Medchemexpress, cat no. MCE-HY-10514) in combination with 10μM Nec1s 15 (Selleckchem, cat no. S8641) or 10μM GSK2982772 (Selleckchem, cat no. S8484) in the presence of 0.25 μM Diyo-1 (AAT Bioquest, cat no. 143413-85-8) and 20ng / ml M-CSF (ImmunoTools, cat no. 12343113). Cells were imaged for 24 hours using IncuCyte imaging system. Cell death was measured by the incorporation of Diyo-1 (AAT Bioquest, cat no.143413- 85-8) using Incucyte image analysis software. 20

[0125] Immunohistochemistry and histology

[0126] Heart samples from mice were fixed in 4 % PFA and embedded in paraffin. Sections of 5 µm were subjected to histological analysis by H&E staining and Masson’s Trichrome staining. For immunohistochemical analysis, slides were rehydrated and incubated with peroxidase blocking buffer (0.04 M NaCitrate, 0.121 M Na2HPO2, 0.03M NaN3, 3% H2O2) for 15 minutes. Slides 25 were washed and antigen-retrieval was performed by digestion with proteinase K (10 µg / ml) for 5 minutes in TEX-buffer (50 mM Tris, 1 mM EDTA, 0.5 % Triton X-100, pH 8.0) for CD45 staining or in citrate TRIS buffer (pH 6) in a pressure cooker for CD3 staining. Sections were blocked for 60 minutes and incubated with the primary antibody for CD45 (clone 30 F-11, eBioscience 14- 0451) or CD3 (clone CD3-12, Biorad MCA1477) at 4°C overnight. A biotinylated secondary anti- 30 rat antibody (Jackson Immuno Research, 112-065-003) was incubated on the sections for 60 minutes. Stainings were visualized using the ABC Vectastain Elite Kit (Vector Laboratories) and DAB substrate (Dako and Vector Laboratories). Sections were then counterstained with hematoxylin for staining of the nuclei, dehydrated and mounted with Entellan.

[0127] TUNEL staining

[0128] TUNEL stain was performed according to the protocol of the DeadEndTMFluorometric TUNEL Kit from Promega (G3250). In brief, sections were rehydrated and washed with 0.85 % NaCl solution and PBS. Tissue sections were then fixed in 4 % PFA for

[0129] 15 minutes and washed with PBS. Digestion with Proteinase K (20 µg / ml) was performed for 5 minutes and sections were washed with PBS. For TUNEL staining, sections were equilibrated with equilibration buffer (provided by the kit) and incubated with rTdT incubation buffer (provided by the kit) for 60 minutes at 37 °C covered with coverslips. The reaction was terminated by immersing the slides in 2x SSC buffer (provided by the kit) for 15 minutes. Sections were washed with PBS and mounted with Vectashield + DAPI (Vector Laboratories) to stain nuclei. Samples were analyzed under a fluorescence microscope.

[0130] The examples show:

[0131] To investigate the role of the kinase activity of TBK1 and IKK ^ in vivo the inventors used CRISPR / Cas9-mediated gene targeting to generate knock-in mice expressing kinase inactive TBK1 (TBK1D135N) (Figure 1A). This approach allows to study the specific function of the kinase activity of TBK1, thus simulating the effect of small molecules that inhibit its enzymatic activity without affecting any kinase-independent scaffold functions. Tbk1D135N / D135Nmice were embryonic lethal similarly to Tbk1- / -mice (Figure 1B), demonstrating that lack of TBK1 kinase activity, and not of any kinase-independent scaffold functions, causes embryonic lethality. The embryonic lethality of Tbk1D135N / D135Nmice was dependent on RIPK1 kinase activity, as crossing to Ripk1D138N / D138Nmice expressing kinase-inactive RIPK1 allowed the survival of Tbk1D135N / D135Nmice (Figure 1B). Tbk1D135N / D135N; Ripk1D138N / D138Nbut also Tbk1D135N / D135N; Ripk1D138N / WTmice reached adulthood without showing apparent pathology (Figure 1B and data not shown), demonstrating that even one RIPK1 kinase inactive allele was sufficient to rescue the embryonic lethality of Tbk1D135N / D135Nmice. These results demonstrated that genetic inhibition of TBK1 kinase activity leads to embryonic lethality in a RIPK1 kinase activity-dependent manner.

[0132] To study the role of the enzymatic activity of IKK ^ and to address its functional redundancy with TBK1 the inventors generated mice that express kinase inactive IKK ^ (IKKeK38A) (Figure 2A). IkkeK38A / K38Amice were viable and fertile and showed no apparent pathology, similarly to IKK ^- deficient mice (Hemmi et al., 2004; Tenoever et al., 2007). To assess possible functional redundancy between IKK ^ and TBK1, the inventors examined whether RIPK1 inhibition also prevents embryonic lethality in mice expressing kinase inactive versions of both IKK ^ and TBK1. The inventors found that Tbk1D135N / D135N; IkkeK38A / K38Amice also died during embryogenesis but were rescued by crossing to Ripk1D138N / D138Nmice (Figure 2B). Also heterozygous expression of kinase inactive RIPK1D138N was sufficient to rescue the embryonic lethality of these animals, with IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / WTmice surviving to adulthood. However, IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / WTmice developed neutrophilia and splenomegaly (Figure 3A-B), and showed liver damage as indicated by elevated ALT levels (Figure 4). Homozygous expression of kinase inactive RIPK1D138N conferred strong protection from splenomegaly and neutrophilia, with only a small fraction of Tbk1D135N / D135N; IkkeK38A / K38A; Ripk1D138N / D138Nmice showing mildly increased numbers of neutrophils and spleen size (Figure 3). Importantly IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / D138Nmice did not show liver damage (Figure 4). Immunoblot analysis of bone marrow derived macrophages (BMDMs) from IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / WTmice showed a complete lack of IRF3 phosphorylation in response to LPS stimulation, demonstrating that the introduced mutations fully inhibited TBK1 and IKK ^ kinase activity (Figure 5A). In addition, IkkeK38A / K38A; Tbk1D135N / D135N; Ripk1D138N / WTBMDMs did not produce IFN ^ in response to LPS stimulation, further confirming the full suppression of TBK1 / IKK ^-dependent interferon responses (Figure 5B). Therefore, inhibition of RIPK1 kinase activity rescued the embryonic lethality, suppressed splenomegaly and neutrophilia and prevented liver damage induced by the combined inhibition of the kinase activities of TBK1 and IKK ^.

[0133] To further investigate the mechanisms by which inhibition of TBK1 and IKK ^ cause neutrophilia in a RIPK1-kinase dependent manner, the inventors studied the role of TBK1 and IKK ^ in myeloid cells by employing a genetic mouse model allowing the inhibition of TBK1 kinase activity in myeloid cells. Specifically, the inventors generated mice containing one kinase inactive TBK1D135N allele with one loxP-flanked TBK1 allele (Tbk1FL / D135N) that were crossed with Cx3Cr1-Cre mice expressing Cre recombinase specifically in myeloid cells. Cre recombinase expression deletes the loxP-flanked Tbk1 allele resulting in the expression exclusively of the kinase inactive TBK1-D135N in myeloid cells (Figure 6). These Tbk1FL / D135NCx3Cr1-Cre mice did not develop neutrophilia or splenomegaly (Figure 7). To assess whether IKK ^ might compensate for the lack of TBK1 kinase activity, the inventors then generated IkkeK38A / K38ATbk1FL / D135NCx3Cr1- Cre, which lack IKK ^ kinase activity in all cells and in addition lack TBK1 kinase activity in myeloid cells. Analysis of these animals revealed that they developed prominent neutrophilia associated with splenomegaly (Figure 7). These results showed that the combined inhibition of IKK ^ and TBK1 in myeloid cells caused systemic pathology manifesting with neutrophilia and splenomegaly. This phenotype was reminiscent of the neutrophilia developing in mice with myeloid cell specific inhibition of IKK2 kinase activity (Ikk2FL / K44ACx3Cr1-Cre), which the inventors found is dependent on RIPK1 kinase activity as it is fully prevented by genetic inhibition of RIPK1 in Ikk2FL / K44ACx3Cr1-Cre Ripk1D138N / D138Nmice (WO2019 / 110832 incorporated herein by reference). The inventors therefore generated IkkeK38A / K38A; Tbk1FL / D135N; Cx3Cr1-Cre; Ripk1D138N / D138Nmice and found that inhibition of RIPK1 kinase activity prevented the development of splenomegaly and neutrophilia induced by the combined inhibition of TBK1 and IKK ^ in myeloid cells (Figure 7). Notably, heterozygous expression of kinase inactive RIPK1D138N had a partial effect considerably suppressing but not fully preventing splenomegaly and neutrophilia in IkkeK38A / K38ATbk1FL / D135N; Cx3Cr1-Cre; Ripk1WT / D138Nmice (Figure 7). Therefore, inhibition of RIPK1 kinase activity prevented the development of splenomegaly and neutrophilia caused by 5 combined inhibition of TBK1 and IKK ^ kinase activities. Moreover, the inventors found that pharmacological inhibition of TBK1 and IKK ^ kinase activity by treatment with MRT67307, a small molecule inhibitor of TBK1 and IKK ^ (Clark et al., 2011), caused cell death and the spontaneous release of IL-1 ^ and IL-1 ^ in BMDMs derived from wild type mice but not from Ripk1D138N / D138Nmice (Figure 8A, B). Pharmacological inhibition of RIPK1 using two highly specific RIPK1 kinase 10 inhibitors, namely Necrostatin-1s (Nec1s) and GSK2982772, also prevented IL-1 ^ release and cell death in BMDMs from wild type mice. In addition, treatment of wild type BMDMs with another TBK1 / IKK ^ inhibitor, BX795, also caused cell death and IL-1 ^ release that were fully prevented by pharmacological inhibition of RIPK1 with Nec1s or GSK2982772. Taken together, these results showed that inhibition of RIPK1 kinase activity prevents cell death, and IL-1 ^ and IL-1 ^ release 15 in BMDMs as well as the development of neutrophilia and splenomegaly in mice with myeloid cell specific inhibition of TBK1 and IKK ^ kinase activities.

[0134] The in vivo and in vitro findings showed that inhibition of RIPK1 kinase activity prevents embryonic lethality and supresses the development of neutrophilia, splenomegaly and liver damage in adult mice induced by combined inhibition of the kinase activities TBK1 and IKKε. 20 Based on these findings, the inventors envision that RIPK1 kinase inhibition will prevent the adverse effects caused by pharmacological inhibition of TBK1 / IKK ^, thus allowing the safe application of TBK1 / IKK ^ inhibitors for the treatment of human inflammatory and autoimmune diseases associated with elevated type I interferon signalling. To experimentally assess the applicability of combined inhibition of RIPK1 together with TBK1 / IKK ^ for the treatment of 25 diseases, the inventors performed a proof-of-concept study using Trex1-deficient mice as a mouse model of AGS, one of a group of diseases characterised as type I interferonopathies (Lee- Kirsch et al., . To investigate whether the combined inhibition of TBK1 and RIPK1 kinase activity can prevent the pathology induced by Trex1 deficiency, the inventors generated Trex1- / -; Tbk1D135N / D135N; Ripk1D138N / D138Nmice. Indeed, the inventors found that the combined genetic 30 inhibition of TBK1 and RIPK1 kinase activities, but not of RIPK1 kinase activity alone, strongly alleviated the development of pathology in Trex1- / -mice, as shown by strongly reduced infiltration of CD45+immune cells and particularly CD3+lymphocytes, reduced collagen deposition as well as reduced numbers of TUNEL+dying cells in the hearts of Trex1-deficient mice (Figure 9). Taken together, these results in a relevant mouse model of AGS provide experimental evidence 35 demonstrating that combined inhibition TBK1 and RIPK1 is well tolerated and effective for the therapy of type I interferonopathies.

[0135] In conclusion, this invention describes the combined application of RIPK1 inhibitors together with TBK1 / IKK ^ inhibitors for the treatment of human diseases associated with increased TBK1 / IKK ^-mediated responses such as type I interferon-associated diseases (e.g. example SLE and AGS), as well as metabolic diseases. The inventors provide evidence that RIPK1 inhibition 5 prevents the adverse effects caused by TBK1 / IKK ^ inhibition (neutrophilia, splenomegaly, liver damage), therefore RIPK1 inhibitors are predicted to allow the safe therapeutic application of TBK1 / IKK ^ inhibitors for the treatment of human diseases. In cases where both RIPK1-mediated cell death and inflammatory signalling and TBK1 / IKK ^-mediated responses contribute to disease pathogenesis, combination treatment with RIPK1 and TBK1 / IKK ^ inhibitors is expected to exhibit 10 synergistic therapeutic effects.

[0136] The inventors furthermore conducted cell culture experiments, which show that with increasing concentrations of TBK1 / IKKe inhibitor primary cells die / release IL-1B, which is prevented by RIPK1 kinase inhibition. Hence, the experiment shows that RIPK1 inhibitors can be used for extending the therapeutic window of TBK1 / IKKe inhibitors. 15

[0137] Combined inhibition of TBK1 and RIPK1 kinase activity prolongs survival of mice carrying Adar1 mutations, an enzyme that deaminates adenosine in RNA and thus prevents its recognition by MDA5, are found in patients with Aicardi–Goutières syndrome and mice lacking ADAR1 or its deaminase activity develop inflammatory pathologies. Particularly ADAR1 mutations mapping to the Zα domain combined with alleles resulting in loss of ADAR1 or specifically its p150 isoform 20 were shown to cause AGS and bilateral striatal necrosis (BSN) in human patients and severe MDA5-MAVS-mediated type I interferon-dependent pathology in mice. To model this condition, the inventors previously generated Adar1mZa / - mice and found that they developed a severe phenotype characterized by reduced body weight and early postnatal lethality. Messenger RNA expression analysis of lung, brain and spleen tissue of Adar1 mZa / - mice revealed increased 25 expression of several genes, the majority of which were linked to type I interferon responses. To assess whether combined inhibition of TBK1 and RIPK1 kinase activity could provide an effective approach for the treatment of this type I interferon-associated pathology the inventors crossed Adar1mZα / – mice with mice expressing kinase inactive TBK1-D135N and RIPK1-D138N resulting in Adar1mZα / –Tbk1D135N / D135N Ripk1D138N / D138N mice. The genetic inhibition of TBK1 and 30 RIPK1 kinase activities prolonged the survival of Adar1mZα / – mice, demonstrating that the inhibition of both TBK1 and RIPK1 kinase activities can be used effectively to treat a type I interferon-driven pathology caused by mutations in Adar1 (Figure 10).

[0138] Combined inhibition of TBK1 and RIPK1 kinase activity prevents the embryonic lethality of Dnase2a-deficient mice. Humans carrying mutations in Dnase2a, a lysosomal DNA degrading 35 enzyme, develop polyarthritis. Dnase2a-deficient mice display type I interferon receptor 1 (IFNAR1)-dependent embryonic lethality. However, with age mice lacking both Dnase2a and type I interferon receptor 1 (IFNAR1) develop polyarthritis that can be ameliorated by blocking TNF, IL1, or IL-6. Notably, the genetic abrogation of STING rescues the embryonic lethal and arthritis phenotype of Dnase2a-deficient mice, demonstrating that STING-induced inflammatory signaling drives these phenotypes in the absence of Dnase2a. A mutation, which disturbs the capacity of 5 STING to induce TNF production and TBK1 / IKKe-activation, can protect Dnase2a-deficient mice from both embryonic lethality and polyarthritis development. Dnase2a-deficient patient blood shows a specific genetic pattern related to blood disorders and an increased presence of developing red blood cells, indicating a potential unique impact of interferon on blood cell production. To assess whether combined inhibition of TBK1 and RIPK1 kinase activity could 10 provide an efficacious approach for treating the type I interferon-driven pathologies in these patients, the inventors crossed mice with mice lacking TBK1 and RIPK1 kinase activities resulting in Dnase2a- / - Tbk1D135N / D135N Ripk1D138N / D138N mice. Our data showed that the genetic inhibition of the kinase activities of both Tbk1 and Ripk1, but not Ripk1 alone, rescues the embryonic lethality of Dnase2a-deficient mice, indicating that the inhibition of both TBK1 and 15 RIPK1 kinase activities can effectively treat a pathology driven by type I interferon due to mutations in Dnase2a (Table A). Table A: The combined genetic inhibition of TBK1 and RIPK1 kinase activities rescue the embryonic lethality of Dnase2a-deficeint mice. Mendelian ratio of progeny from the intercross of Tbk1D135N / wtRipk1D138N / D138NDnase2a- / -mice is displayed. 20 REFERENCES

[0139] The references are: Ahn, J., Gutman, D., Saijo, S., and Barber, G.N. (2012). STING manifests self DNA-dependent inflammatory disease. Proceedings of the National Academy of Sciences of the United States of America 109, 19386-19391. 5 Al-Mayouf, S.M., Sunker, A., Abdwani, R., Abrawi, S.A., Almurshedi, F., Alhashmi, N., Al Sonbul, A., Sewairi, W., Qari, A., Abdallah, E., et al. (2011). Loss-of-function variant in DNASE1L3 causes a familial form of systemic lupus erythematosus. Nat Genet 43, 1186-1188. Barrat, F.J., and Su, L. (2019). A pathogenic role of plasmacytoid dendritic cells in autoimmunity and chronic viral infection. J Exp Med 216, 1974-1985. 10 Bonnard, M., Mirtsos, C., Suzuki, S., Graham, K., Huang, J., Ng, M., Itie, A., Wakeham, A., Shahinian, A., Henzel, W.J., et al. (2000). Deficiency of T2K leads to apoptotic liver degeneration and impaired NF-kappaB-dependent gene transcription. The EMBO journal 19, 4976-4985. Clark, K., Peggie, M., Plater, L., Sorcek, R.J., Young, E.R., Madwed, J.B., Hough, J., McIver, E.G., 15 and Cohen, P. (2011). Novel cross-talk within the IKK family controls innate immunity. The Biochemical journal 434, 93-104. Fitzgerald, K.A., McWhirter, S.M., Faia, K.L., Rowe, D.C., Latz, E., Golenbock, D.T., Coyle, A.J., Liao, S.M., and Maniatis, T. (2003). IKKepsilon and TBK1 are essential components of the IRF3 signaling pathway. Nat Immunol 4, 491-496. 20 Gao, D., Li, T., Li, X.D., Chen, X., Li, Q.Z., Wight-Carter, M., and Chen, Z.J. (2015). Activation of cyclic GMP-AMP synthase by self-DNA causes autoimmune diseases. Proceedings of the National Academy of Sciences of the United States of America 112, E5699-5705. Hemmi, H., Takeuchi, O., Sato, S., Yamamoto, M., Kaisho, T., Sanjo, H., Kawai, T., Hoshino, K., Takeda, K., and Akira, S. (2004). The roles of two IkappaB kinase-related kinases in 25 lipopolysaccharide and double stranded RNA signaling and viral infection. J Exp Med 199, 1641-1650. Heraud-Farlow, J.E., and Walkley, C.R. (2016). The role of RNA editing by ADAR1 in prevention of innate immune sensing of self-RNA. Journal of molecular medicine 94, 1095-1102. Lafont, E., Draber, P., Rieser, E., Reichert, M., Kupka, S., de Miguel, D., Draberova, H., von 30 Massenhausen, A., Bhamra, A., Henderson, S., et al. (2018). TBK1 and IKKepsilon prevent TNF-induced cell death by RIPK1 phosphorylation. Nature cell biology 20, 1389- 1399. Lee-Kirsch, M.A., Wolf, C., Kretschmer, S., and Roers, A. (2015). Type I interferonopathies--an expanding disease spectrum of immunodysregulation. Seminars in immunopathology 37, 349-357. Liu, S., Cai, X., Wu, J., Cong, Q., Chen, X., Li, T., Du, F., Ren, J., Wu, Y.T., Grishin, N.V., et al. 5 (2015). Phosphorylation of innate immune adaptor proteins MAVS, STING, and TRIF induces IRF3 activation. Science 347, aaa2630. Liu, Y., Jesus, A.A., Marrero, B., Yang, D., Ramsey, S.E., Sanchez, G.A.M., Tenbrock, K., Wittkowski, H., Jones, O.Y., Kuehn, H.S., et al. (2014). Activated STING in a vascular and pulmonary syndrome. The New England journal of medicine 371, 507-518. 10 Medzhitov, R. (2008). Origin and physiological roles of inflammation. Nature 454, 428-435. Morita, M., Stamp, G., Robins, P., Dulic, A., Rosewell, I., Hrivnak, G., Daly, G., Lindahl, T., and Barnes, D.E. (2004). Gene-targeted mice lacking the Trex1 (DNase III) 3'-->5' DNA exonuclease develop inflammatory myocarditis. Molecular and cellular biology 24, 6719- 6727. 15 Napirei, M., Karsunky, H., Zevnik, B., Stephan, H., Mannherz, H.G., and Moroy, T. (2000). Features of systemic lupus erythematosus in Dnase1-deficient mice. Nat Genet 25, 177- 181. Okabe, Y., Kawane, K., and Nagata, S. (2008). IFN regulatory factor (IRF) 3 / 7-dependent and - independent gene induction by mammalian DNA that escapes degradation. European 20 journal of immunology 38, 3150-3158. Rice, G., Newman, W.G., Dean, J., Patrick, T., Parmar, R., Flintoff, K., Robins, P., Harvey, S., Hollis, T., O'Hara, A., et al. (2007). Heterozygous mutations in TREX1 cause familial chilblain lupus and dominant Aicardi-Goutieres syndrome. Am J Hum Genet 80, 811-815. Rodero, M.P., Tesser, A., Bartok, E., Rice, G.I., Della Mina, E., Depp, M., Beitz, B., Bondet, V., 25 Cagnard, N., Duffy, D., et al. (2017). Type I interferon-mediated autoinflammation due to DNase II deficiency. Nat Commun 8, 2176. Sisirak, V., Sally, B., D'Agati, V., Martinez-Ortiz, W., Ozcakar, Z.B., David, J., Rashidfarrokhi, A., Yeste, A., Panea, C., Chida, A.S., et al. (2016). Digestion of Chromatin in Apoptotic Cell Microparticles Prevents Autoimmunity. Cell 166, 88-101. 30 Stetson, D.B., Ko, J.S., Heidmann, T., and Medzhitov, R. (2008). Trex1 prevents cell-intrinsic initiation of autoimmunity. Cell 134, 587-598. Tenoever, B.R., Ng, S.L., Chua, M.A., McWhirter, S.M., Garcia-Sastre, A., and Maniatis, T. (2007). Multiple functions of the IKK-related kinase IKKepsilon in interferon-mediated antiviral immunity. Science 315, 1274-1278. Thomson, D.W., and Bergamini, G. (2021). Recent progress in small molecule TBK1 inhibitors: a patent review (2015- 2020). Expert Opin Ther Pat, 1-10. Xu, D., Jin, T., Zhu, H., Chen, H., Ofengeim, D., Zou, C., Mifflin, L., Pan, L., Amin, P., Li, W., et al. (2018). TBK1 Suppresses RIPK1-Driven Apoptosis and Inflammation during Development 5 and in Aging. Cell 174, 1477-1491 e1419. Yamamoto, M., Sato, S., Hemmi, H., Hoshino, K., Kaisho, T., Sanjo, H., Takeuchi, O., Sugiyama, M., Okabe, M., Takeda, K., et al. (2003). Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science 301, 640-643. Yasutomo, K., Horiuchi, T., Kagami, S., Tsukamoto, H., Hashimura, C., Urushihara, M., and 10 Kuroda, Y. (2001). Mutation of DNASE1 in people with systemic lupus erythematosus. Nat Genet 28, 313-314. Zhang, C., Shang, G., Gui, X., Zhang, X., Bai, X.C., and Chen, Z.J. (2019). Structural basis of STING binding with and phosphorylation by TBK1. Nature 567, 394-398. Zhao, B., Du, F., Xu, P., Shu, C., Sankaran, B., Bell, S.L., Liu, M., Lei, Y., Gao, X., Fu, X., et al. 15 (2019). A conserved PLPLRT / SD motif of STING mediates the recruitment and activation of TBK1. Nature 569, 718-722. Zhao, P., Wong, K.I., Sun, X., Reilly, S.M., Uhm, M., Liao, Z., Skorobogatko, Y., and Saltiel, A.R. (2018). TBK1 at the Crossroads of Inflammation and Energy Homeostasis in Adipose Tissue. Cell 172, 731-743 e712. 20

Claims

CLAIMS 1. A compound or composition for use in the treatment of a condition in a subject, wherein (a) The compound or composition when administered to the subject induces an 5 inhibition of (x) RIPK1 and (y) Inhibitor of TANK binding kinase 1 (TBK1) and / or IκB kinase epsilon (IKKε); (b) Wherein the subject suffers from a disease associated with a mutation in one of the genes selected from human Trex1, Adar1, and Dnase2a.

2. The compound or composition for use of claim 1, wherein the disease is a condition associated with increased type I interferon responses.

3. The compound or composition for use of claim 1 or 2, wherein the type I interferon- associated disease is characterized by a pathological increased type I interferon activity and / or accumulation and / or increased expression of interferon-stimulated genes (ISGs).

4. The compound or composition for use of any one of claims 1 to 3, wherein the compound or composition is a combination of at least two compounds, wherein the first compound is an inhibitor of (x) and wherein the second compound is one or two compounds which is (are) an inhibitor of (y); or, wherein the compound or composition is a dual inhibitor having a first inhibitory activity towards (x) and a second inhibitory activity towards (y), such as a fused compound comprising two structural elements of which one can bind to and inhibit (x) and one can bind to and inhibit (y).

5. The compound or composition for use of any one of claims 1 to 4, wherein the condition is an autoimmune or inflammatory disease, infection or cancer, preferably wherein the autoimmune or inflammatory disease or cancer is associated with increased type I interferon responses.

6. A RIPK1 inhibitor for use in the treatment or prevention of an adverse effect of an immune disease therapy in a subject, wherein the immune disease therapy involves the administration of an IKKε- and / or TBK1-inhibitor to the subject suffering from the immune disease.

7. The RIPK1 inhibitor for use of claim 6, wherein the immune disease therapy involves sequential or concomitant inhibition of IKKε and TBK1 in the subject suffering from the immune disease.

8. The RIPK1 inhibitor for use of claim 6 or 7, wherein the immune disease is an 5 autoimmune disease or cancer, or is an inflammatory disease, or viral disease.

9. A pharmaceutical composition comprising the compound or composition of any one of claims 1 to 5, together with a pharmaceutically acceptable carrier and / or excipient.

10. A method for screening an IKKε-Inhibitor suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses in a subject, (a) Providing a test cell, (b) Optionally providing one or more control cells; (c) Contacting the test cell with (i) a candidate IKKε-Inhibitor, while the test cell is contacted with a known TBK1 inhibitor and a known RIPK1 inhibitor; or (ii) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a known RIPK1 inhibitor; or (iii) a candidate IKKε-Inhibitor and a candidate TBK1 inhibitor, while the cell is contacted with a known RIPK1 inhibitor; or (iv) a candidate IKKε-Inhibitor, a candidate TBK1 inhibitor and a candidate RIPK1 inhibitor; or (v) a candidate IKKε / TBK1 dual inhibitor where the test cell is contacted with a candidate RIPK1 inhibitor; (d) Optionally, where the control cell is not contacted in (a) to (d) with any of the candidate inhibitors; or alternatively, contacting the control cell with the candidate inhibitors but not the known inhibitors;(e) Determining in the test cell at least cell death and release of one or more cytokines before and after step (iii), or in comparison to the control cell or in comparison to a reference value; Wherein if in step (v) the following condition is determined: no increased cell death and / or 5 a reduced release of the one or more test cytokines, then the candidate inhibitors in one or more of (a) to (d) are suitable for use in a therapy involving a triple inhibition of IKKε, TBK1- and RIPK1 for the treatment of type I interferon responses, wherein the triple combination comprises the inhibitors of the respective scenario (a) to (d) for which the condition was determined.

11. The method of claim 10, wherein the known RIPK1 inhibitor is selected from the group consisting of IPK1-IN-7, GNE684, GSK3145095, GSK298277259, GSK'547, GSK3502421, DHP76, DHP77, AV123, MBM105, ZB-R-55, PN10, RIPK1 kinase inhibitor 38, GSK′963, DNL758, DNL788, Necrostatin-1s (Nec1s) or GSK2982772.

12. The method of claim 10 or, wherein the known TBK1 inhibitor is selected from MRT67307, BX795, TBK1-protac, TBK1 / IKKε-IN-1, TBK1 / IKKε-IN-2, GSK8612, Amlexanox, AZ13102909, MPI-0485520 (TBK1 / IKK dual inhibitor) and BAY-985.

13. The method of any one of claims 10 to 12, wherein the type I interferon response is in context of a disease selected from an auto inflammatory disease, infectious or auto immune disease.

14. The method of any one of claims 10 to 13, wherein the candidate inhibitors are selective inhibitors of the kinase activity of their target, and not inhibitors that reduce overall protein activity of other protein functions, protein expression and / or protein stability.

15. The method of any one of claims 10 to 14, wherein the candidate inhibitor is a small molecular compound suspected to harbour an activity as a kinase inhibitor.

16. A RIPK1 inhibitor for use in the treatment of a IKKε / TBK1 inhibitor treatment related condition in a subject, wherein the subject is receiving a IKKε / TBK1 inhibitor treatment.

17. The RIPK1 inhibitor for use of claim 16, wherein the IKKε / TBK1 inhibitor treatment related condition is an adverse effect or result of the IKKε / TBK1 inhibitor treatment that would otherwise result in a reduced IKKε / TBK1 inhibitor dosing and / or discontinuing of IKKε / TBK1 inhibitor treatment.

18. The RIPK1 inhibitor for use of claim 16 or 17, wherein the inhibitors are as defined in any one of the preceding claims.

Images