Unusual mechanism of CDK4/6 inhibitor resistance found, may be reversible

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Breast cancer cells can acquire resistance to the relatively new drugs known as CDK4/6 inhibitors by producing higher amounts of CDK6, a cell cycle protein, a new study has found, and those cells can transfer their resistant traits to other cancer cells via molecular carriers called exosomes.

This drug resistance mechanism, identified by researchers at Dana-Farber Cancer Institute, is highly unusual, they said, and laboratory studies suggest it may be reversible. Breast cancer cells that developed resistance to a CDK4/6 inhibitor, palbociclib, became sensitive to the drug again after a “treatment holiday” of several weeks when no palbociclib was administered, they said in an article in Cell Reports.

CDK4/6 inhibitors, which include palbociclib, ribociclib, and abemaciclib, are designer drugs that thwart cancer cell growth by blocking two cell cycle protein kinases, CDK4 and CDK6, which control when cells divide. Overactivity of cell cycle kinases is common in many forms of cancer, allowing uncontrolled proliferation of cancer cells.  Since 2015, Food and Drug Administration approvals of these agents have been granted for their use in combination with hormonal treatments for women with estrogen receptor (ER)-positive, HER2-negative advanced or metastatic breast cancer. Abemaciclib is also approved as a single agent for patients with advanced or metastatic disease that has progressed following prior endocrine therapy and chemotherapy.

The CDK4/6 inhibitor drugs have demonstrated benefit in advanced breast cancer by extending the period before the disease worsens, and the drugs are being tested in an array of other cancer types. But their effectiveness is limited by the inevitable development of resistance, and why this happens is an area of critical research need, said the Dana-Farber investigators, led by Geoffrey Shapiro, MD, PhD, and Liam Cornell, PhD.

To search for molecular events leading to CDK4/6 drug resistance, the scientists treated breast cancer cells with palbociclib until the drug no longer was able to halt the cells’ growth – a period of 12 weeks. The cells were then considered treatment-resistant. The concentration of palbociclib was increased until the cells became resistant to these higher doses as well. Through a series of analytic experiments, the scientists linked the development of resistance to an increase in expression of the CDK6 (but not CDK4) protein in the cancer cells. The specific mechanism by which CDK6 expression was dialed up turned out to be the suppression of a cell signaling pathway, the TGF-beta pathway, by a particular microRNA, or miRNA. miRNAs are tiny pieces of genetic material that act to tweak gene expression.  Additionally, high levels of the relevant miRNA were found in primary tumor samples from patients who did not respond to CDK4/6 inhibitors or whose tumors developed acquired resistance.

What caught the investigators’ attention was that, unlike in many instances of cancer cells evolving resistance to drugs, all the breast cancer cells in a laboratory dish were found to acquire resistance at a similar time. It’s more typical, they said, for resistance to emerge in a clone of identical cancer cells because of a genetic mutation, and that clone of resistant cells becomes dominant.

Further studies revealed that the palbociclib-resistant cells did not contain a mutation, but instead became resistant and spread that trait among all neighboring cells via exosomes – tiny bubble-like particles that can ferry proteins and genetic material between cells. This hypothesis “was further supported by the reversibility of resistance, which could not occur had resistance arisen due to a permanent genetic event,” said the investigators.

Experiments showed that cancer cells’ resistance to palbociclib could be reversed by stopping treatment with the drug for seven weeks. The treatment “holiday” was tested in mice as well: palbociclib-resistant tumor material was implanted into animals, who were treated with the drug until resistant tumors were established. The drug was then discontinued for 28 days, and after treatment was started again, the tumors shrank.

The discovery that resistance to a CDK4/6 inhibitor is transmitted to tumor cells via exosomes has a potential benefit for patient management, the scientists said. “We may be able to examine patients’ exosomes [by means of a blood test] and see if they are becoming resistant earlier than would be apparent because the cancer progressed on radiographic scans,” said Shapiro.

Seth Wander, MD, PhD, and Nikhil Wagle, MD, of DFCI’s Center for Cancer Precision Medicine and the Broad Institute, collaborated on analysis of primary tumors in the project.   The research was supported by the Dana-Farber/Harvard Cancer Center Specialized Program of Research Excellence in Breast Cancer; National Institutes of Health grant P50 CA168504; a Susan G. Komen Career Catalyst Research Grant CCR15333343; AACR NextGen Grant for Transformative Cancer Research 1620-38-WAGL, as well as grants from the V Foundation and The Cancer Couch Foundation.

Shapiro has received research funding from Eli Lilly, Merck KGaA/EMD-Serono, Merck, and Sierra Oncology. He has served on advisory boards for Pfizer, Eli Lilly, G1 Therapeutics, Roche, Merck KGaA/EMD-Serono, Sierra Oncology, Bicycle Therapeutics, Fusion Pharmaceuticals, Cybrexa Therapeutics, Astex, Almac, Ipsen, Bayer, Angiex and Daiichi Sankyo.

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