Background

CDK4/6 inhibitors are being used to treat a variety of human malignancies. In well-differentiated (WDLS) and dedifferentiated (DDLS) liposarcoma we had previously suggested that their clinical promise might be associated with their ability to down-regulate the MDM2 protein. In cultured cell lines, the down-regulation of MDM2, after cells left the cell cycle following CDK4 inhibition, induces them to progress from quiescence into senescence. Today, we will present further evidence supporting the idea that senescence after growth arrest is a mechanism that can account for the activity of CDK4 inhibitors, at least in WD/DDLS.

Results

We have found that PDLIM7 physically associates with MDM2 and prevents MDM2 turnover in quiescent cells induced by treatment with palbociclib (PD0332991). However, if PDLIM7 is sequestered by an intracellular cadherin, CDH18, MDM2 turnover occurs following treatment with PD0332991, driving the quiescent cell into the senescent state. CRISPR knockout of CDH18 can prevent both MDM2 degradation and senescence in cells challenged with CDK4 inhibitor. Establishing the clinical relevancy of this pathway, CDH18 expression is associated with clinical outcome and histologic subtype in patients with advanced WDLS and DDLS from our previous phase II trials. 92% of WDLS and 57% of DDLS had high CDH18 expression judged by immunohistochemistry. High expression of CDH18 in DDLS was associated with improved clinical outcomes.

Conclusions

This supports the hypothesis that the transition from quiescence to senescence has clinical relevance as a mechanism for this class of drugs.

Legal entity responsible for the study

Memorial Sloan Kettering Cancer Center

Funding

National Cancer Institute, USA

Disclosure

All authors have declared no conflicts of interest.

Background

D-type cyclins (D1, D2 and D3) together with their associated cyclin-dependent kinases CDK4 and CDK6 are components of the core cell cycle machinery that drives cell proliferation. Inhibitors of CDK4 and CDK6 are currently in clinical trials for patients with several cancer types, with promising results. We demonstrated that cyclin D3-CDK6 phosphorylates and inhibits the catalytic activity of two key enzymes in the glycolytic pathway, 6-phosphofructokinase and pyruvate kinase M2. This re-directs the glycolytic intermediates into the pentose phosphate (PPP) and serine pathways. Inhibition of cyclin D3-CDK6 in tumor cells reduces PPP and serine pathway flows, thereby depleting anti-oxidants NADPH and glutathione. This, in turn, elevates the levels of reactive oxygen species and causes tumor cell apoptosis. The pro-survival function of cyclin D-associated kinase operates in tumors expressing high levels of cyclin D3-CDK6 complexes. We propose that measuring the levels of cyclin D3-CDK6 in human cancers might help to identify tumor subsets that undergo cell death and tumor regression upon CDK4/6-inhibition. Cyclin D3-CDK6, through its ability to link cell cycle and cell metabolism represents a particularly powerful oncogene that affects cancer cells at several levels, and this property can be exploited for anti-cancer therapy.

Targeting immune checkpoints such as the one mediated by programmed cell death protein 1 (PD-1) and its ligand PD-L1 have been approved for treating multiple types of human cancers with durable clinical benefit. However, many cancer patients fail to respond to anti-PD-1/PD-L1 treatment, and the underling mechanism(s) is not well understood. Recent studies revealed that response to PD-L1 blockade might correlate with PD-L1 expression levels on tumor cells. Hence, it is important to mechanistically understand the pathways controlling PD-L1 protein expression and stability, which can offer a molecular basis to improve the clinical response rate and efficacy of PD-1/PD-L1 blockade in cancer patients. We found that PD-L1 protein abundance is regulated by cyclin D-CDK4, and that inhibition of CDK4/6 in vivo elevates PD-L1 protein levels. Combining CDK4/6 inhibitor treatment with anti-PD-1 immunotherapy enhanced tumor regression and dramatically improved overall survival rates in mouse tumor models. Our study uncovered a novel molecular mechanism for regulating PD-L1 protein stability by a cell cycle kinase and revealed the potential for using combination treatment with CDK4/6 inhibitors and PD-1/PD-L1 immune checkpoint blockade to enhance therapeutic efficacy for human cancers.

Legal entity responsible for the study

Dana-Farber Cancer Institute

Disclosure

P. Sicinski: Receive a research grant from Novartis.

Funding

NIH

Background

While immune checkpoint inhibitors (ICI) are efficacious and lead to durable responses in patients with various cancers, only a minority of patients respond. An approach to increase the efficacy of ICI is to combine them with chemotherapy, in order to enhance immunogenic cell death and “prime” the immune system. However, chemotherapy itself can cause damage to various cell types of the immune system, potentially diminishing the activity of the ICI combination. Using trilaciclib, a short-acting IV CDK4/6 inhibitor that preserves hematopoietic stem cells and enhances immune system function during chemotherapy, we have developed a novel approach to preserve immune system function during chemotherapy to allow optimal activity of ICI.

Methods

To evaluate the efficacy of trilaciclib added to chemotherapy/ICI combinations, tumor-bearing immunocompetent mice (MC38 or CT26) were treated with trilaciclib, chemotherapy (oxaliplatin or 5-FU), or ICIs (anti-PD-L1 or anti-PD-1) alone or in combination, and tumor size was measured for up to 100 days. To gain insight into the effect of transient exposure of trilaciclib on the tumor microenvironment, we are examining the cellular composition, proliferation status and gene expression of tumor immune infiltrates from MC38 tumor-bearing mice post-treatment.

Results

The addition of trilaciclib to chemotherapy/ICI significantly improved anti-tumor activity and prolonged overall survival. Potential mechanisms by which transient trilaciclib exposure enhances anti-tumor activity include: reduction of Treg populations, preservation of T lymphocyte function, and changes in gene expression leading to enhanced lymphocyte activation and a pro-inflammatory tumor microenvironment.

Conclusions

These findings suggest that transient trilaciclib-induced G1 cell-cycle arrest in tumor immune infiltrates prior to chemotherapy/ICI combination treatment creates a favorable tumor microenvironment resulting in increased antitumor activity. Based on these findings, a Phase 2 study to assess the safety and efficacy of trilaciclib or placebo with carboplatin, etoposide, and atezolizumab in first-line extensive stage SCLC patients is ongoing (NCT03041311).

Legal entity responsible for the study

G1 Therapeutics

Disclosure

P.J. Roberts, A.Y. Lai, J.A. Sorrentino, R.K. Malik: All authors are employees of G1 Therapeutics.

Funding

G1 Therapeutics