This study provides proof concept for the efficacy and safety of Ciapavir and indicates that Smac mimetics can constitute a crucial element of a safe and efficacious treatment technique to get rid of the latent HIV-1 reservoir

This study provides proof concept for the efficacy and safety of Ciapavir and indicates that Smac mimetics can constitute a crucial element of a safe and efficacious treatment technique to get rid of the latent HIV-1 reservoir. are protein kinase C (PKC) agonists, such as ingenol and bryostatin.9 However, this class of compounds is connected with systemic T?cell activation,10 and undesireable effects have already been reported in clinical studies.11 Additionally, a recently available study discovered that blockade of checkpoint proteins programmed loss of life-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) using the agonist vesatolimod, proposed as latency-reversing treatment previously,12, 13, 14 didn’t influence viral rebound kinetics following Artwork interruption in simian immunodeficiency trojan (SIV)-contaminated macaques.15 Therefore, a secure and efficient modality for HIV-1 latency reversal is constantly on the represent a crucial unmet therapeutic want. The inhibitor of apoptosis (IAP) protein family is a functionally and structurally related band of proteins that primarily serve as cellular inhibitors of programmed cell loss of life, or apoptosis.16,17 Smac mimetics certainly are a course of small-molecule peptidomimetics produced from a conserved binding theme of Smac (second mitochondria-derived activator of caspases), an endogenous proteins inhibitor of IAPs, such as XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics were made to focus on XIAP to modulate apoptosis originally; however, in addition they antagonize cIAP1 and various other members of the proteins family members to varying levels. ingenol.9 However, this class of compounds is connected with systemic T?cell activation,10 and undesireable effects have already been reported in Methyl linolenate clinical studies.11 Additionally, a recently available study discovered that blockade of checkpoint proteins programmed loss of life-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) using the agonist vesatolimod, previously proposed as latency-reversing treatment,12, 13, 14 didn’t influence viral rebound kinetics following Artwork interruption in simian immunodeficiency trojan (SIV)-contaminated macaques.15 Therefore, a effective and safe modality for HIV-1 latency reversal is constantly on the represent a crucial unmet therapeutic need. The inhibitor of apoptosis (IAP) proteins family members is normally a functionally and structurally related band of proteins that mainly serve as mobile inhibitors of designed cell loss of life, or apoptosis.16,17 Smac mimetics certainly are a course of small-molecule peptidomimetics produced from a conserved binding theme of Smac (second mitochondria-derived activator of caspases), an endogenous proteins inhibitor of IAPs, such as XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics had been originally made to focus on XIAP to modulate apoptosis; nevertheless, in addition they antagonize cIAP1 and various other members of the proteins family members to varying levels. cIAP1, an E3 ubiquitin member and ligase from the IAP family members, regulates the activation from the non-canonical nuclear aspect B (ncNF-B) pathway, generating expression of a particular group of genes that govern immune system function.24, 25, 26, 27 We previously reported that Smac mimetic substances that may focus on the inhibitor of apoptosis proteins cIAP1 (Birc2) harbor LRA activity.28 Specifically, this previous research revealed that genetic or pharmacological antagonism of cIAP1 promoted ncNF-B-dependent activation from the HIV-1 long terminal repeat (LTR), a task that was present to potently reactivate latent HIV-1 also. Predicated on this preliminary work, we have now survey the preclinical characterization and advancement of a bivalent next-generation Smac mimetic substance, Ciapavir (SBI-0953294), that was particularly optimized to improve LRA activity and drug-like properties for reversal from the latent HIV-1 tank. Outcomes Bivalent Smac Mimetics Harbor Greater Strength as LRAs Than Monovalent Substances We’ve previously confirmed that latency reversal of HIV-1 could be marketed in and systems through pharmacological manipulation from the non-canonical NF-B pathway using the Smac mimetic substance SBI-0637142.28 This molecule induced HIV-1 latency in CD4+ T modestly?cells from ART-suppressed aviremic HIV-infected sufferers as an individual agent; however, solid activity was noticed when administered in conjunction with the HDACi panobinostat. This recommended that SBI-0637142 likely possesses suboptimal efficacy to mediate latency reversal as an individual agent effectively?modeling predicated on the crystal structure of cIAP1 BIR3.22 From these investigations, we figured a previously described linker in the P4 placement21 could possibly be employed to make a dimeric edition of SBI-0637142 with enhanced strength (Body?1D). Predicated on this, we synthesized and designed a bivalent Smac mimetic, SBI-0953294, which we’ve termed Ciapavir (cIAP1 antagonist for viral reactivation; Body?1D). An evaluation of the initial- and second-generation substances in the 2D10 Jurkat latency model verified the fact that bivalent molecule Ciapavir displays substantially greater strength and efficiency as an LRA, inducing equivalent degrees of reversal at concentrations 10- to at least one 1 latency,000-fold less than the first-generation molecule SBI-0637142 (Body?1E), lacking any upsurge in cytotoxicity (Body?S1B). Ciapavir reached >65% from the LRA activity of phorbol 12-myristate 13-acetate (PMA)/ionomycin treatment (Body?S1C). Furthermore, hereditary ablation of NF-B-inducing kinase (NIK), a kinase needed for ncNF-B activation, was enough to invert Ciapavir LRA activity in this technique (Body?1F), indicating that second-generation molecule is mediating LRA activity through activation from the ncNF-B pathway, in keeping with the?defined mechanism reported by our group previously.28 Ciapavir Synergizes with Epigenetic Regulators.Three from the nine mice treated with Ciapavir (Figure?4C) exhibited boosts in plasma RNA on the 48-h necropsy period point, although non-e of the various other animals (like the 9 automobile control mice) had quantifiable plasma viremia. efficacious and secure treatment technique to get rid of the latent HIV-1 reservoir. are proteins kinase C (PKC) agonists, which include bryostatin and ingenol.9 However, this class of compounds is associated with systemic T?cell activation,10 and adverse effects have been reported in clinical trials.11 Additionally, a recent study found that blockade of checkpoint protein programmed death-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) with the agonist vesatolimod, previously proposed as latency-reversing treatment,12, 13, 14 did not impact viral rebound kinetics following ART interruption in simian immunodeficiency virus (SIV)-infected macaques.15 Therefore, a safe and effective modality for HIV-1 latency reversal continues to represent a critical unmet therapeutic need. The inhibitor of apoptosis (IAP) protein family is a functionally and structurally related group of proteins that primarily serve as cellular inhibitors of programmed cell death, or apoptosis.16,17 Smac mimetics are a class of small-molecule peptidomimetics derived from a conserved binding motif of Smac (second mitochondria-derived activator of caspases), an endogenous protein inhibitor of IAPs, which include XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics were originally designed to target XIAP to modulate apoptosis; however, they also antagonize cIAP1 and other members of this protein family to varying degrees. cIAP1, an E3 ubiquitin ligase and member of the IAP family, regulates the activation of the non-canonical nuclear factor B (ncNF-B) pathway, driving expression of a specific set of genes that govern immune function.24, 25, 26, 27 We previously reported that Smac mimetic compounds that can target the inhibitor of apoptosis protein cIAP1 (Birc2) harbor LRA activity.28 Specifically, this previous study revealed that genetic or pharmacological antagonism of cIAP1 promoted ncNF-B-dependent activation of the HIV-1 long terminal repeat (LTR), an activity that was also found to potently reactivate latent HIV-1. Based on this initial work, we now report the preclinical development and characterization of a bivalent next-generation Smac mimetic compound, Ciapavir (SBI-0953294), that was specifically optimized to enhance LRA activity and drug-like properties for reversal of the latent HIV-1 reservoir. Results Bivalent Smac Mimetics Harbor Greater Potency as LRAs Than Monovalent Compounds We have previously demonstrated that latency reversal of HIV-1 can be promoted in and systems through pharmacological manipulation of the non-canonical NF-B pathway using the Smac mimetic compound SBI-0637142.28 This molecule modestly induced HIV-1 latency in CD4+ T?cells from ART-suppressed aviremic HIV-infected patients as a single agent; however, robust activity was observed when administered in combination with the HDACi panobinostat. This suggested that SBI-0637142 likely possesses suboptimal efficacy to effectively mediate latency reversal as a single agent?modeling based on the crystal structure of cIAP1 BIR3.22 From these investigations, we concluded that a previously described linker in the P4 position21 could be employed to create a dimeric version of SBI-0637142 with enhanced potency (Figure?1D). Based on this, we designed and synthesized a bivalent Smac mimetic, SBI-0953294, which we have termed Ciapavir (cIAP1 antagonist for viral reactivation; Figure?1D). A comparison of the first- and second-generation compounds in the 2D10 Jurkat latency model confirmed that the bivalent molecule Ciapavir exhibits substantially greater potency and efficacy as an LRA, inducing comparable levels of latency reversal at concentrations 10- to 1 1,000-fold lower than the first-generation molecule SBI-0637142 (Figure?1E), without an increase in cytotoxicity (Figure?S1B). Ciapavir reached >65% of the LRA activity of phorbol 12-myristate 13-acetate (PMA)/ionomycin.Cells were then incubated at 4?C for 20?min, washed with PBS, and resuspended in 2% paraformaldehyde. HIV-1 Methyl linolenate reservoirs in a bone marrow, liver, thymus (BLT) humanized mouse model without mediating systemic T?cell activation. This study provides proof of concept for the efficacy and safety of Ciapavir and shows that Smac mimetics can constitute a critical component of a safe and efficacious treatment strategy to eliminate the latent HIV-1 reservoir. are protein kinase C (PKC) agonists, which include bryostatin and ingenol.9 However, this class of compounds is associated with systemic T?cell activation,10 and adverse effects have been reported in clinical tests.11 Additionally, a recent study found that blockade of checkpoint protein programmed death-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) with the agonist vesatolimod, previously proposed as latency-reversing treatment,12, 13, 14 did not effect viral rebound kinetics following ART interruption in simian immunodeficiency disease (SIV)-infected macaques.15 Therefore, a safe and effective modality for HIV-1 latency reversal continues to represent a critical unmet therapeutic need. The inhibitor of apoptosis (IAP) protein family is definitely a functionally and structurally related group of proteins that primarily serve as cellular inhibitors of programmed cell death, or apoptosis.16,17 Smac mimetics are a class of small-molecule peptidomimetics derived from a conserved binding motif of Smac (second mitochondria-derived activator of caspases), an endogenous protein Methyl linolenate inhibitor of IAPs, which include XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics were originally designed to target XIAP to modulate apoptosis; however, they also antagonize cIAP1 and additional members of this protein family to varying degrees. cIAP1, an E3 ubiquitin ligase and member of the IAP family, regulates the activation of the non-canonical nuclear element B (ncNF-B) pathway, traveling expression of a specific set of genes that govern immune function.24, 25, 26, 27 We previously reported that Smac mimetic compounds that can target the inhibitor of apoptosis protein cIAP1 (Birc2) harbor LRA activity.28 Specifically, this previous study revealed that genetic or pharmacological antagonism of cIAP1 promoted ncNF-B-dependent activation of the HIV-1 long terminal repeat (LTR), an activity that was also found to potently reactivate latent HIV-1. Based on this initial work, we now statement the preclinical development and characterization of a bivalent next-generation Smac mimetic compound, Ciapavir (SBI-0953294), that was specifically optimized to enhance LRA activity and drug-like properties for reversal of the latent HIV-1 reservoir. Results Bivalent Smac Mimetics Harbor Greater Potency as LRAs Than Monovalent Compounds We have previously shown that latency reversal of HIV-1 can be advertised in and systems through pharmacological manipulation of the non-canonical NF-B pathway using the Smac mimetic compound SBI-0637142.28 This molecule modestly induced HIV-1 latency in CD4+ T?cells from ART-suppressed aviremic HIV-infected individuals as a single agent; however, powerful activity was observed when administered in combination with the HDACi panobinostat. This suggested that SBI-0637142 likely possesses suboptimal effectiveness to efficiently mediate latency reversal as a single agent?modeling based on the crystal structure of cIAP1 BIR3.22 From these investigations, we concluded that a previously described linker in the P4 position21 could be employed to create a dimeric version of SBI-0637142 with enhanced potency (Number?1D). Based on this, we designed and synthesized a bivalent Smac mimetic, SBI-0953294, which we have termed Ciapavir (cIAP1 antagonist for viral reactivation; Number?1D). A comparison of the 1st- and second-generation compounds in the 2D10 Jurkat latency model confirmed the bivalent molecule Ciapavir exhibits substantially greater potency and effectiveness as an LRA, inducing similar levels of latency reversal at concentrations 10- to 1 1,000-fold lower than the first-generation molecule SBI-0637142 (Number?1E), without an increase in cytotoxicity (Number?S1B). Ciapavir reached >65% of the LRA activity of phorbol 12-myristate 13-acetate (PMA)/ionomycin treatment (Number?S1C). Furthermore, genetic ablation of NF-B-inducing kinase (NIK), a kinase essential for ncNF-B activation, was adequate to reverse Ciapavir LRA activity in this system (Number?1F), indicating that this second-generation molecule is mediating LRA activity through activation of the ncNF-B pathway, consistent with the?previously described mechanism reported by our group.28 Ciapavir Synergizes with Epigenetic Regulators to Enhance HIV-1 Latency Reversal Much like combinatorial ART, effective latency reversal as part of? a curative therapy may ultimately require the combination of multiple LRAs to maximize efficacy. 45 We previously decided that this Smac mimetic SBI-0637142 synergizes with the HDACis? vorinostat and panobinostat.28 Here, we evaluated combinations of Ciapavir with two other well-established classes of LRAs, bromodomain and extraterminal domain inhibitors (BETi), and PKC agonists (PKCas) (Figures 1G and S1D). We observed potent Bliss synergy46 of Ciapavir with the BETi JQ1 and I-BET151, with extra over Bliss (EOB) values.Despite the capacity of this compound to induce expression of latent HIV-1 and may therefore show useful in “shock and kill” approaches to HIV-1 cure. Open in a separate window Figure?4 Viral Loads and RNA Expression in Humanized Mice Treated with Ciapavir Mice were infected with HIV-1, treated with ART to suppress viral loads, and then administered with Ciapavir or vehicle control. (A and B) Viral loads of control (A) and Ciapavir-treated (B) BLT mice. (C) 3 responding mice showing increases in viral loads 2?days after administration of Ciapavir. (D) Bone marrow HIV RNA levels, with responding mice 1, 5, and 8 indicated. (ECH) Mice described were subjected to flow cytometry analysis at necropsy to analyze immune activation. malignancy. Critically, this molecule induced activation of HIV-1 reservoirs in a bone marrow, liver, thymus (BLT) humanized mouse model without mediating systemic T?cell activation. This study provides proof of concept for the efficacy and security of Ciapavir and indicates that Smac mimetics can constitute a critical component of a safe and efficacious treatment strategy to eliminate the latent HIV-1 reservoir. are protein kinase C (PKC) agonists, which include bryostatin and ingenol.9 However, this class of compounds is associated with systemic T?cell activation,10 and adverse effects have been reported in clinical trials.11 Additionally, a recent study found that blockade of checkpoint protein programmed death-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) with the agonist vesatolimod, previously proposed as latency-reversing treatment,12, 13, 14 did not impact viral rebound kinetics following ART interruption in simian immunodeficiency computer virus (SIV)-infected macaques.15 Therefore, a safe and effective modality for HIV-1 latency reversal continues to represent a critical unmet therapeutic need. The inhibitor of apoptosis (IAP) protein family is usually a functionally and structurally Rabbit polyclonal to EPHA4 related group of proteins that primarily serve as cellular inhibitors of programmed cell death, or apoptosis.16,17 Smac mimetics are a class of small-molecule peptidomimetics derived from a conserved binding motif of Smac (second mitochondria-derived activator of caspases), an endogenous protein inhibitor of IAPs, which include XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics were originally designed to target XIAP to modulate apoptosis; however, they also antagonize cIAP1 and other members of this protein family to varying degrees. cIAP1, an E3 ubiquitin ligase and member of the IAP family, regulates the activation of the non-canonical nuclear factor B (ncNF-B) pathway, driving expression of a specific set of genes that govern immune function.24, 25, 26, 27 We previously reported that Smac mimetic compounds that can target the inhibitor of apoptosis protein cIAP1 (Birc2) harbor LRA activity.28 Specifically, this previous study revealed that genetic or pharmacological antagonism of cIAP1 promoted ncNF-B-dependent activation of the HIV-1 long terminal repeat (LTR), an activity that was also found to potently reactivate latent HIV-1. Based on this initial work, we now statement the preclinical development and characterization of a bivalent next-generation Smac mimetic compound, Ciapavir (SBI-0953294), that was specifically optimized to enhance LRA activity and drug-like properties for reversal of the latent HIV-1 reservoir. Results Bivalent Smac Mimetics Harbor Greater Strength as LRAs Than Monovalent Substances We’ve previously confirmed that latency reversal of HIV-1 could be marketed in and systems through pharmacological manipulation from the non-canonical NF-B pathway using the Smac mimetic substance SBI-0637142.28 This molecule modestly induced HIV-1 latency in CD4+ T?cells from ART-suppressed aviremic HIV-infected sufferers as an individual agent; however, solid activity was noticed when administered in conjunction with the HDACi panobinostat. This recommended that SBI-0637142 most likely possesses suboptimal efficiency to successfully mediate latency reversal as an individual agent?modeling predicated on the crystal structure of cIAP1 BIR3.22 From these investigations, we figured a previously described linker in the P4 placement21 could possibly be employed to make a dimeric edition of SBI-0637142 with enhanced strength (Body?1D). Predicated on this, we designed and synthesized a bivalent Smac mimetic, SBI-0953294, which we’ve termed Ciapavir (cIAP1 antagonist for viral reactivation; Body?1D). An evaluation of the initial- and second-generation substances in the 2D10 Jurkat latency model verified the fact that bivalent molecule Ciapavir displays substantially greater strength and efficiency as an LRA, inducing equivalent degrees of latency reversal at concentrations 10- to at least one 1,000-fold less than the first-generation molecule SBI-0637142 (Body?1E), lacking any upsurge in cytotoxicity (Body?S1B). Ciapavir reached >65% from the LRA activity of phorbol 12-myristate 13-acetate (PMA)/ionomycin treatment (Body?S1C). Furthermore, hereditary ablation of NF-B-inducing kinase (NIK), a kinase needed for ncNF-B activation, was enough to invert Ciapavir LRA activity.Cytokine amounts were determined using the LEGENDplex Mouse Irritation Panel (BioLegend). Humanized mouse research Humanized mice had been contaminated with an X4-tropic strain of HIV-1 predicated on NL4-3 customized to add HSA instead of vpr53, then?further modified to add HA instead of HSA52 with random 21 nucleotide series inserted in non-coding area beside HSA gene (M.D. tumor. Critically, this molecule induced activation of HIV-1 reservoirs within a bone tissue marrow, liver organ, thymus (BLT) humanized mouse model without mediating systemic T?cell activation. This research provides proof idea for the efficiency and protection of Ciapavir and signifies that Smac mimetics can constitute a crucial element of a secure and efficacious treatment technique to get rid of the latent HIV-1 tank. are proteins kinase C (PKC) agonists, such as bryostatin and ingenol.9 However, this class of compounds is connected with systemic T?cell activation,10 and undesireable effects have already been reported in clinical studies.11 Additionally, a recently available study discovered that blockade of checkpoint proteins programmed loss of life-1 (PD-1) using the antibody nivolumab and activation of Toll-like receptor 7 (TLR7) using the agonist vesatolimod, previously proposed as latency-reversing treatment,12, 13, 14 didn’t influence viral rebound kinetics following Artwork interruption in simian immunodeficiency pathogen (SIV)-contaminated macaques.15 Therefore, a effective and safe modality for HIV-1 latency reversal is constantly on the represent a crucial unmet therapeutic need. The inhibitor of apoptosis (IAP) proteins family is certainly a functionally and structurally related band of proteins that mainly serve as mobile inhibitors of designed cell loss of life, or apoptosis.16,17 Smac mimetics certainly are a course of small-molecule peptidomimetics derived from a conserved binding motif of Smac (second mitochondria-derived activator of caspases), an endogenous protein inhibitor of IAPs, which include XIAP, cIAP1, cIAP2, ILP2, BRUCE/Apollon, survivin, NAIP, and ML-IAP.18, 19, 20, 21, 22, 23 Smac mimetics were originally designed to target XIAP to modulate apoptosis; however, they also antagonize cIAP1 and other members of this protein family to varying degrees. cIAP1, an E3 ubiquitin ligase and member of the IAP family, regulates the activation of the non-canonical nuclear factor B (ncNF-B) pathway, driving expression of a specific set of genes that govern immune function.24, 25, 26, 27 We previously reported that Smac mimetic compounds that can target the inhibitor of apoptosis protein cIAP1 (Birc2) harbor LRA activity.28 Specifically, this previous study revealed that genetic or pharmacological antagonism of cIAP1 promoted ncNF-B-dependent activation of the HIV-1 long terminal repeat (LTR), an activity that was also found to potently reactivate latent HIV-1. Based on this initial work, we now report the preclinical development and characterization of a bivalent next-generation Smac mimetic compound, Ciapavir (SBI-0953294), that was specifically optimized to enhance LRA activity and drug-like properties for reversal of the latent HIV-1 reservoir. Results Bivalent Smac Mimetics Harbor Greater Potency as LRAs Than Monovalent Compounds We have previously demonstrated that latency reversal of HIV-1 can be promoted in and systems through pharmacological manipulation of the non-canonical NF-B pathway using the Smac mimetic compound SBI-0637142.28 This molecule modestly induced HIV-1 latency in CD4+ T?cells from ART-suppressed aviremic HIV-infected patients as a Methyl linolenate single agent; however, robust activity was observed when administered in combination with the HDACi panobinostat. This suggested that SBI-0637142 likely possesses suboptimal efficacy to effectively mediate latency reversal as a single agent?modeling based on the crystal structure of cIAP1 BIR3.22 From these investigations, we concluded that a previously described linker in the P4 position21 could be employed to create a dimeric version of SBI-0637142 with enhanced potency (Figure?1D). Based on this, we designed and synthesized a bivalent Smac mimetic, SBI-0953294, which we have termed Ciapavir (cIAP1 antagonist for viral reactivation; Figure?1D). A comparison of the first- and second-generation compounds in the 2D10 Jurkat latency model confirmed that the bivalent molecule Ciapavir exhibits substantially greater potency and efficacy as an LRA, inducing comparable levels of latency reversal at concentrations 10- to 1 1,000-fold lower than the first-generation molecule SBI-0637142 (Figure?1E), without an increase in cytotoxicity (Figure?S1B). Ciapavir reached >65% of the LRA activity of phorbol 12-myristate 13-acetate (PMA)/ionomycin treatment (Figure?S1C). Furthermore, genetic ablation of NF-B-inducing kinase (NIK), a kinase essential for ncNF-B activation, was sufficient to reverse Ciapavir LRA activity in this system (Figure?1F), indicating that this second-generation molecule is mediating LRA activity through activation of the ncNF-B pathway, consistent with the?previously described mechanism reported by our group.28 Ciapavir Synergizes with Epigenetic Regulators to Enhance HIV-1 Latency Reversal Similar to combinatorial ART, effective latency reversal as part of?a curative therapy may ultimately require the combination of multiple LRAs to maximize efficacy.45 We previously determined that the Smac mimetic SBI-0637142 synergizes with the HDACis?vorinostat and panobinostat.28 Here, we evaluated combinations of Ciapavir with two other well-established classes of LRAs, bromodomain and extraterminal domain inhibitors (BETi), and PKC agonists (PKCas) (Figures 1G and S1D). We observed potent Bliss synergy46 of Ciapavir with the BETi JQ1 and I-BET151, with excess over Bliss (EOB) values greater than 0.6. Combinations of Ciapavir with the PKCas bryostatin-1 or ingenol-3-angelate, by contrast, exhibited concomitant.