enitociclib (VIP152)

Enitociclib potently inhibited in vivo tumor growth of cell line-derived and patient-derived xenografts having therapeutic resistance. Our data demonstrate the potency of enitociclib in overcoming therapeutic resistance in MCL models and provide evidence in favor of its clinical investigation.

P1/2, N=130, Recruiting, National Cancer Institute (NCI) | Trial primary completion date: Mar 2024 --> Mar 2025

P1, N=6, Terminated, Vincerx Pharma, Inc. | N=54 --> 6 | Trial completion date: Feb 2026 --> May 2023 | Recruiting --> Terminated | Trial primary completion date: Feb 2026 --> May 2023; Slow enrollment

An unbiased analysis of the genes affected by CDK9 inhibition in both cell lines demonstrates that RNA polymerase II and transcription pathways are primarily affected and novel enitociclib targets such as PHF23 and TP53RK are discovered. These findings are recapitulated in blood samples from enitociclib-treated patients; while MYC downregulation is most robust with enitociclib treatment, other CDK9 regulated targets may be MYC independent delivering a transcriptional downregulation via RNA polymerase II.

Moreover, VIP152 markedly inhibited the growth of three patient-derived xenograft tumor models, overcoming BTKi resistance (P<0.001), BTKi-venetoclax dual resistance (P<0.01), and BTKi-CAR-T dual resistance (P<0.001). Conclusion These data showed that CDK9 was a promising target for treating MCL and using VIP152 to specifically target it was efficacious in overcoming a variety of therapeutic resistances in MCL.

In this study, we established an AML cell line with acquired resistance to a highly selective CDK9 inhibitor BAY1251152...To overcome the drug resistance mediated by the CDK9-L156F mutation, we discovered a compound, IHMT-CDK9-36 which showed potent inhibition activity both for CDK9 WT and L156F mutant. Together, we report a novel resistance mechanism for CDK9 inhibitors and provide a novel chemical scaffold for the future development of CDK9 inhibitors.

In its first in-human trial, enitociclib demonstrated clinical activity in a small cohort of patients with high grade B lymphoma with MYC and BCL2 and/or BCL6 rearrangements, inducing complete responses in 2 of 7 subjects (29%) in monotherapy. These data suggest CDK9 inhibitors could play a role in the treatment of hematological diseases and could be a great ally when combined with other therapeutic approaches.

NVP-2, MC180295, fadraciclib, KB-0742, LZT-106, and 21e have been developed mainly for treating solid tumors, and most of them work only on certain genotypes of solid tumors. Only VIP152 has been proven to benefit the patients with advanced high-grade lymphoma (HGL) and solid tumors in clinical trials. Continued efforts to explore the molecular mechanisms underlying the inhibitory effects, and to identify suitable tumor genotypes and combination treatment strategies, are crucial to demonstrate the efficacy of selective CDK9 inhibitors and degraders in tumor therapy.

SynergyFinder found synergistic effects of enitociclib with irinotecan, carfilzomib, etoposide, bortezomib, selinexor or topotecan tested at clinically relevant concentrations in the aRMS cell line Rh41. Our results suggest that CDK9 inhibition is potentially clinically relevant for MYCN-amplified solid tumors such as aRMS and NBL due to its significant antitumor activity and pharmacologic targetability. The data provide essential information on the distinct targets, biomarkers of activity and clinically feasible drug combinations for the development of enitociclib in clinical trials addressing an unmet need in pediatric oncology.

P1/2, N=130, Recruiting, National Cancer Institute (NCI) | Not yet recruiting --> Recruiting

Combining BAY1251152 with Cisplatin synergistically overcame chemoresistance of Caspase-8-deficient cervical cancer cells. Therefore, Caspase-8 expression could be a marker in chemoresistant cervical tumors, suggesting CDK9 inhibitor treatment for their sensitization to Cisplatin-based chemotherapy.

Finally, immune competent mice engrafted with CLL-like cells of Eµ-MTCP1 over-expressing mice and treated with VIP152 demonstrated reduced disease burden and improvement in overall survival compared to vehicle-treated mice. These data suggest that VIP152 is a highly selective inhibitor of CDK9 that represents an attractive new therapy for CLL.

Our studies present the first proof-of-concept evidence that enitociclib has significant antitumor activity against several MM cell lines and provides specific pharmacological targetability of several key oncogenic pathways involving proteins such as MYC, MCL1 and PCNA, leading to growth inhibition and apoptosis. Taken together, the data provide the rationale and biological reasoning for further optimization studies of CDK9 inhibitors for clinical application to improve MM patient outcomes.

Enitociclib has a favorable safety profile with evidence of clinical activity in DH-DLBCL and perhaps, with longer follow up, in other B-cell malignancies. Also, we show that enitociclib downregulates the MYC transcriptional program and is unlikely to abrogate vaccine efficacy, an important feature given the current COVID-19 pandemic.

The reference compound enitociclib (BAY 1251152) was used for experimental validation. Although several cytotoxic molecules that inhibit CDK9 are in the therapeutic pipeline, many show cross-reactivity to other CDK family members, leading to toxicity and tolerability issues. GFH009 is a potent and highly selective CDK9 inhibitor with the ability to reduce expression of downstream oncogenes required for rapid cellular division and protein expression through specific, short-lived inhibition of CDK9. We postulate that this mechanism drives GFH009's inhibition of cellular division, as tumor stabilization and shrinkage appear to be dose-dependent.

To demonstrate the utility of a selective CDK9 inhibitor for treatment of hematologic patients with an unmet medical need such as high-grade B cell lymphoma (HGBL) and chronic lymphocytic leukemia who relapse or are refractory to ibrutinib and venetoclax (R/R CLL)...Differential expression (DE) analysis from VIP152, atuveciclib and KB-0742 treatment of 2 lymphoma cell lines by RNA seq is compared to DE of samples from 7 VIP152 treated HGBL patients...With high ATP, VIP152, AZD4573 and alvocidib maintain potency in low nM range, while fadraciclib and KB-0742 have 760 nM-1.7 µM IC50s...Selective transcriptional downregulation is observed with 30mg IV weekly. Samples from patients with DLBCL or CLL are sensitive to VIP152 and updated PK, PD and ctDNA dynamics from the ongoing phase 1 trials (NCT02635672, NCT04978779) will be presented.

P1/2, N=130, Not yet recruiting, National Cancer Institute (NCI)

VIP152 monotherapy, administered intravenously once weekly, demonstrated a favorable safety profile and evidence of clinical benefit in patients with advanced HGL and solid tumors.

furthermore, we assessed its in vivo efficacy using patient-derived xenograft (PDX) models derived from MCL patients including one with Dual-R.Results The expression of MYC oncogene associates with ibrutinib resistance and Dual-R, and poor clinical outcome...In addition, BAY-1251152 also significantly suppressed tumor growth in the PDX models derived from a BTKi-R patient (p=0.00015) and a BTKi-venetoclax dual-resistant patient (p=0.009).Conclusion Our findings showed that targeting CDK9 by its specific inhibitor BAY-1251152 led to potent in vitro anti-MCL activity...BAY-1251152 is also potent in inhibiting tumor growth in PDX models. These data support that CDK9 is a promising target to overcome BTKi-CAR T dual resistance in MCL, which is in urgent need.

VIP152 demonstrates sensitivity in gynecologic cell lines independent of cisplatin sensitivity. An interim gene signature that is associated with VIP152 sensitivity was defined and we plan to optimize this signature with a larger cell line panel. Dose-dependent tumor growth inhibition in an in vivo xenograft model is demonstrated.

P1, N=54, Recruiting, Vincerx Pharma, Inc. | Not yet recruiting --> Recruiting | Trial completion date: Apr 2024 --> Feb 2026 | Initiation date: Aug 2021 --> Dec 2021 | Trial primary completion date: Apr 2024 --> Feb 2026

CDK9 inhibition with VIP152 selectively depletes short half-life transcripts including MYC, a known driver of HGBL. Clearance of MYC, MCL1 and PCNA protein demonstrates VIP152 control of multiple parallel oncogenic pathways in the clinic and in HGBL preclinical models. Dose-dependent regression and tumor-outgrowth control demonstrates that VIP152 QW treatment can drive antitumor efficacy.

Broad inhibition of cyclin dependent kinases (CDK) and associated alternative target enzymes with agents such as flavopiridol or dinaciclib have demonstrated significant clinical activity in CLL but are hindered by a relatively narrow therapeutic window...Kinase profiling revealed the IC 50 of VIP152 was lowest for CDK9/Cyclin T1 and CDK9/Cyclin T2 with close similarity to dinaciclib and greater than 1 log superiority over KB-0742... Our data demonstrate VIP152 to be a highly selective and potent CDK9 inhibitor that disrupts the CDK9 nuclear complex and mediates significant preclinical activity against CLL cell lines and primary CLL cells. VIP152 also demonstrates predictable and new pharmacodynamic markers to assess target engagement. Collectively, these data support the recently initiated CLL clinical trial (NCT04978779).

AZD5991 is a highly selective BH3-mimetic that demonstrates high in vitro potency in MCL1-dependent cell lines with an IC50 <1 nM.7 We have demonstrated that direct MCL1 inhibition with AZD5991 disrupts survival of neoplastic B-cells in lymph-node mimicking conditions, induces mitochondrial dysfunction and cooperates with BCL2/X inhibitors in vitro and in vivo.8 A phase 1 clinical trial of AZD5991 alone or in combination with venetoclax in hematologic malignancies is ongoing (NCT03218683)...AMG-176 demonstrated synergy with venetoclax in AML models; however, it also caused cytopenias.9 AMG-176 was shown to induce apoptosis of CLL cells independent of prognostic markers and overcame the protective effect of stromal microenvironment.10 However, suppressive effects on hematopoiesis will likely become the dose- limiting factor in clinical trials of MCL1-targeting agents. The effect of MCL1 inhibition on cord blood-derived CD34+ cells was studied using a small-molecule inhibitor, S63845,11 resulting in almost complete depletion of human hematopoietic stem and progenitor cells, while mature blood cells survived normally...We have shown that SYK inhibition with entospletinib leads to downmodulation of MCL1 protein in CLL both in vitro and in the clinic.12,13 Treatment of CLL cells with entospletinib, but not other BCR-signaling inhibitors, led to a disruption of BAFF-BCR cross-talk and downmodulation of MCL1 mRNA and protein, thus implicating SYK in transduction of multiple pro-survival signals emanating from the tumor microenvironment.13 Entospletinib has shown promising clinical activity in CLL.12,14 It is currently being developed in myeloid malignancies.15 A novel dual SYK/BTK inhibitor luxeptinib also was shown to downregulate MCL1 and other pro-survival BCL2 proteins in CLL.16 Transcriptional cyclin-dependent kinases (CDK7/9) regulate activity of RNA polymerase, thereby controlling production of mRNA. Downmodulation of MCL1 has been considered a key mechanistic event accounting for the pro-apoptotic activity of CDK inhibitors in neoplastic B-cells.17,18 Pan-CDK inhibitors (flavopiridol, dinaciclib) demonstrate clinical efficacy in lymphoid malignancies. We and others reported that preclinical efficacy of AZ5576, a selective inhibitor of CDK9, and its clinical congener AZD4573 in lymphoid tumors depended on rapid downmodulation of MCL1, BFL1, and MYC.19–21 CDK9 inhibition can increase efficacy of BH3-mimetics, but safety of this combination will need to be explored.22,23 CDK9 inhibitors in development, in addition to AZD4573, include VIP152, CYC065, and voruciclib...Navitoclax is a small-molecule inhibitor of BCL2, BCLX, and BCL2L2; however, its use in CLL has been plagued by thrombocytopenia.24 AZD4320 is an alternative agent that co-targets BCL2/X...ABBV-155, an antibody-drug conjugate targeting BCLX, where BCLX is being used as a payload to avoid “off-tumor” effects, demonstrated promise in solid tumor models.26 Finally, as many microenvironment-driven pro-survival pathways converge on NFB, leading to induction of chemokines, cell cycle regulators, and BCL2 family proteins themselves, this transcription factor continues to be an attractive target in CLL...When combined with ibrutinib in murine CLL models, VAY-736 produced prolonged survival compared with either therapy alone.28 Alternative targets for therapeutic antibodies include ROR1 (cirmtuzumab), CD19 (tafasitamab), and others.29 As resistance to novel agents is becoming an unmet need in CLL, exploration of novel targets is of paramount importance...Multiple CDK inhibitors are currently being explored and have anti-tumor effects not restricted to MCL1 inhibition. CAR T cells and bi-specific antibodies have exceptional efficacy in lymphoid malignancies and thus are of high relevance in CLL.

P1, N=54, Not yet recruiting, Vincerx Pharma, Inc.

In fact, the Phase I clinical trials of the latest, highly specific CDK9 inhibitor BAY1251152, against different solid tumors have shown good anti-tumor and on-target activities and pharmacokinetics, combined with manageable safety profile while the phase I and II clinical trials of another inhibitor AT-7519 have been undertaken or are undergoing. To enhance the effectiveness and target diversity and reduce potential drug-resistance, the future of CDK9 inhibition would likely involve combining CDK9 inhibitors with inhibitors like those against BRD4, SEC, MYC, MCL-1 and HSP90.

P1, N=107, Recruiting, Vincerx Pharma, Inc. | Active, not recruiting --> Recruiting | N=37 --> 107 | Trial completion date: Jun 2021 --> Dec 2024 | Trial primary completion date: Jun 2021 --> Dec 2024

VIP152 had a manageable safety profile, on-target pharmacodynamic activity and signs of durable monotherapy antitumor activity in patients with DHL . These encouraging results warrant further evaluation of VIP152 in patients with MYC-driven lymphoma and solid tumors.

P1; N=33; Active, not recruiting; Sponsor: Bayer; Trial completion date: Apr 2020 --> Oct 2020; Trial primary completion date: Apr 2020 --> Oct 2020

P1; N=33; Active, not recruiting; Sponsor: Bayer; Trial completion date: Oct 2019 --> Apr 2020; Trial primary completion date: Oct 2019 --> Apr 2020

Conclusion s: Treatment of AML patients with BAY did not result in objective responses despite achieving drug levels in the expected therapeutic range and evidence of target engagement. Given the good tolerability and on-target activity, BAY 1251152 might be a good candidate for future combination therapies.

P1; N=33; Active, not recruiting; Sponsor: Bayer; Recruiting --> Active, not recruiting; N=120 --> 33

P1; N=120; Recruiting; Sponsor: Bayer; Trial primary completion date: Sep 2018 --> Oct 2019