RB, or Not RB: That is the Question!

I just returned from a tremendously insightful meeting at the National Cancer Institute on the lineage plasticity of prostate cancer. The focus of the meeting was the myriad changes that systemic treatment induces in prostate cancer over time – most notably the emergence of neuroendocrine/small cell prostate cancer. I’ll address this entity in a subsequent entry. Before that, though, the striking focal point of discussion at the meeting was a single molecule -RB  ( or, the retinoblastoma gene) and its pivotal role in prostate cancer progression. I predict that within the not too distant future, clinicians will be wanting to know the “RB status” of the tumors of the patients they are treating. Thus, in my latest effort to educate you ( and myself) about the key molecular drivers of prostate cancer progression, metastasis and how it becomes lethal, I have chosen to focus on Rb. It is likely that aberrations in Rb account for much of the misery that afflicts those with mCRPC as it is one of the key molecular drivers in the disease. One day, hopefully soon, knowing this status will drive treatment decisions.

  1. What does it do?
Rb is the classic tumor suppressor gene. Its name arose from the fact that it was first discovered that loss of both copies of this gene was prevalent in children with rare retinoblastoma tumors. It was hypothesized that the children would inherit one defective copy and acquire a second ‘hit’ leading to the development of this tumor. Published in SCIENCE in 1984,  it became one of the first fully elucidated tumor suppressors and a landmark observation in cancer genetics.  We now know that loss of Rb is implicated in the aggressiveness of a wide variety of tumors beyond retinoblastoma, including breast and prostate cancer.

Rb is a large protein with a “pocket” region that sequesters and thereby inactivates other smaller proteins that are involved in the cell cycle and a host of other mechanisms. During the cell’s G phase, when it is not synthesizing DNA or undergoing mitosis,  Rb binds to elongation factor 2 ( e2F) proteins, sequestering them from pushing the cell into S phase. Thus, a normally functioning Rb keeps the cell cycle turned off. This is far from its only function as Rb can bind to and regulate up to 100 known proteins in the cell, regulating a number of factors.

If a cell is in S phase ( and thus synthesizing the DNA needed for replication) Rb can still regulate it by binding to histones as they regulate DNA unraveling. In this context, its main role is to bind to and therefore inhibit, the transcription factors that drive the cell from the G1 phase to S phase.

  1. How common are aberrations in Rb?
They are very common. Rb can be inactivated in tumors through a variety of process, loss of the gene through mutation and deletion as well as phosphorylation of the protein.  In an important analysis,  Barry Taylor and colleagues at Memorial Sloan Kettering Cancer Center looked comprehensively at the genomics of both primary and metastatic prostate cancer. They found that Rb was altered in about a quarter of primary tumors but about three-quarters of metastatic lesions1. Loss of a functioning Rb is another example of treatment mediated selection pressure – tumor cells that sporadically lose both copies have a growth and metastasis advantage and thus, by the time we are measuring metastatic tumors, the population is likely to be enriched by them.

Rb loss drives many other mechanisms beyond engaging the cell cycle, including enabling the process that drives metastasis in animal models but this is probably the key one to understand in order to grasp its importance in tumor biology.

Rb loss in EGFR mutated non-small cell lung cancer is associated with the transformation of NSCLC to emergent small cell lung cancer – this transformation itself is fascinating and challenges our notion that ‘small cell’ and ‘Non-small cell” lung cancer are mutually exclusive entities.

  1. How can we use this knowledge to treat CRPC?

So, it is possible that the combination of molecular defects, as they occur in mCRPC, may drive the ‘evolution’ of prostate cancer to the resistant form of the disease we identify as ‘anaplastic CRPC”  or “treatment-emergent small cell neuroendocrine cancer”

Interestingly, in vitro models, high levels of androgen can inactivate prostate cancer tumor growth, this forms the basis for clinical trials in which patients with CRPC are being given testosterone. It is likely that this ‘repressive’ function of high levels of androgens occurs through the recruitment of Rb. Thus, if Rb is absent, this repression can’t occur.2

Some laboratory work suggests that for a metastatic CRPC cell to develop full-on resistance to Enzalutamide, both p53 and Rb need to be deleted. Deleting one or the other only has a mild effect.3

A mechanism that brings the issue to prostate cancer is the role of AR signaling in suppressing Rb. Through a variety of intermediate steps (cyclin D1, mTOR and CDK4/6) high levels of androgen receptor activation, as we would see in CRPC (where the androgen receptor is frequently amplified), results in the phosphorylation (and thus inactivation) of Rb.

This latter point is critical for targeting patients with Rb loss. While there isn’t really a way to target loss of Rb directly (it’s pretty hard to replace a complex protein that is missing) it may be possible to target some of the pathways that are overactivated by an underactivated Rb. The most likely candidate in this regard is CDK4/6, which is heavily activated in the context of Rb loss. This pathway has already been validated as a therapeutic target in breast cancer. Palbociclib and Ribociclib and Abemaciclib are drugs that are approved for breast cancer and can be used alone or in combination with hormonal therapies. They are all being tested in prostate cancer. 

A number of CDK4/6 inhibitors are being tested in prostate cancer in combination with docetaxel, abiraterone, and even as stand-alone therapies. Theoretically, these agents would be more effective in the setting of an intact RB, because blocking CDK4/6 would be expected to suppress the inactivation of RB, thus essentially turning it back on. One major challenge is developing a reliable test that can be used on tumor and circulating tumor cell samples that can help us know if Rb is present or not. More work is needed in that area as the drugs continue their clinical development.

Keep an eye on this space.

So, with apologies to Shakespeare, we may one day paraphrase Hamlet by asking our pathologists: “Rb, or not RB”?


Written by: Charles J. Ryan, MD

References:

1.Taylor B et al. (2010). Integrative Genomic Profiling of Human Prostate Cancer, Cancer Cell, Volume 18, (Issue 1), 11-22.
2. Gao G et Al. (2016) Androgen Receptor Tumor Suppressor Function Is Mediated by Recruitment of Retinoblastoma Protein. Cell Reports, Volume 17 (Issue 4), 966-976.
3. Mu P et al. (2017), SOX2 promotes lineage plasticity and antiandrogen resistance in TP53- and RB1-deficient prostate cancer, Science, Volume 355, (Issue 6320), 84-88.

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