May 1, 2018

Story by Nadir Balba

The Paper of the Month for April 2018 is "LSD1 activates a lethal prostate cancer gene network independently of its demethylase function" published in the Proceedings of the National Academy of Sciences. This project was led by Drs. Archana Sehrawat and Joshi Alumkal, Wayne D. Kuni & Joan E. Kuni Foundation Endowed Chair for Prostate Cancer Research, associate professor of medicine and molecular and medical genetics, OHSU School of Medicine, and member of the OHSU Knight Cancer Institute. The study is the result of an extensive multi-site collaboration between six different institutions and the work of almost 30 different contributing authors (see full author list below).

Fighting back against therapy-resistant cancers

One in nine men in the United States will be diagnosed with prostate cancer, which is the second leading cause of cancer death behind lung cancer. One of the most common treatments for prostate cancer is androgen-deprivation therapy. This involves lowering male hormones, or androgens, in patients in order to block the function of the androgen receptor (AR).

AR activation is important for prostate cancer cell growth, and blocking AR can temporarily control tumors. However, many tumors become resistant to androgen deprivation therapy and become incurable and lethal. Therefore, there is an urgent need to identify the drivers of this treatment-resistant form of prostate cancer. With this in mind, Dr. Alumkal's lab focuses on identifying the mechanisms by which prostate cancer progresses to its lethal form and developing new treatments to overcome lethal prostate cancer.

The role of LSD1

Lysine-specific demethylase 1, or LSD1, is an enzyme and key regulator of gene expression related to cell growth, differentiation, and migration in cancers, including prostate cancer. Due to its known interaction with the AR protein, LSD1 was thought to merely be a partner in crime. With this in mind, the team sought to answer two major questions: does LSD1 promote survival of lethal prostate cancer cells, and, if so, how is that effect mediated?

In this breakthrough study, the team first determined that LSD1 is highly expressed in lethal prostate cancer tumors resistant to drugs that block the AR, suggesting that LSD1 overexpression is selected for in the transition into this deadly form of prostate cancer. 

Next, they tested whether LSD1's effect on cancer cell survival was AR-dependent through an elaborate series of experiments, which manipulated LSD1 protein expression using RNA interference (RNAi) in cells with or without AR expression. Surprisingly, they demonstrated that LSD1 promotes cancer cell survival independently of AR, dispelling the hypothesis that LSD1 was simply an AR cofactor. They also looked at gene expression after LSD1 RNAi to identify critical pathways controlled by LSD1. They found that the embryonic stem cell and cell cycle pathways – two pathways that are enriched in lethal human tumors – were key LSD1 targets.

Histone modification and gene expression

Their next step was figuring out how LSD1 promotes cancer cell survival. One potential mechanism, supported by previous research, was through histone modifications. Histones are spherical-shaped proteins that act as spools around which DNA strands wrap themselves.

LSD1, and other histone demethylases, can remove methyl groups from histones causing them to "open" and unravel DNA strands. This allows transcription factors to convert these genes into RNA, which eventually leads to protein synthesis. It was previously unknown if this process was how LSD1 manipulated gene expression and enabled cancer growth, or if LSD1 was involved in other processes that facilitated gene expression and the survival of cancer cells.

To determine if histone modifications were critical for regulating gene expression, the team performed chromatin immunoprecipitation sequencing (ChIP-seq) for histone marks LSD1 has the capacity to modulate. These experiments led to another surprise and demonstrated for the first time that LSD1 regulated cell survival and gene expression independently from histone modifications. These results were in direct conflict with what many in the field thought to be true: that LSD1's demethylase function was critical for its effects on gene regulation.

Discovering a new target

The team was forced to think outside the box, which led them to their hypothesis that LSD1 acts as a scaffold to cooperate with other proteins to activate lethal prostate cancer gene networks. Using affinity immunopurification, a technique that helps identify relevant LSD1 interacting proteins, they discovered that a key LSD1 binding partner was the protein ZNF217.

Mary Heinricher, Ph.D., associate dean for basic research, OHSU School of Medicine, was fascinated by the study's many unexpected results, which led to this new therapeutic target. 

"What was really intriguing about this translational paper was that the group targeted this molecule because of a known role as a demethylase, and indeed, found it to be important. But it was important for a different reason than the one that made them look at it in the first place."

The team then aimed to block LSD1 function and prevent cancer proliferation using several commercially available compounds. In line with their earlier results, the team found that drugs that block LSD1's demethylase function had no effect. However, they did find success using a drug called SP-2509, which they determined interfered with LSD1's interaction with ZNF217.

SP-2509 reduced prostate cancer cell survival in vitro and reduced growth of prostate cancer tumors in mice without toxicity or causing weight loss. This work provides a new roadmap for targeting LSD1 in cancer – by blocking key protein-protein interactions or LSD1 protein stability. The fact that that SP-2509 was effective, without causing serious sides effects in mice, means there is potential that LSD1 inhibitors like SP-2509 could be a new weapon in the fight against prostate cancer. The team's results have led Salarius Pharmaceuticals, which makes a drug similar to SP-2509, to consider moving their drug into a Phase I clinical trial in men with prostate cancer.

Dr. Alumkal will discuss his research at an upcoming Marquam Hill Lecture on Thursday, May 17, at 7 p.m., in the Collaborative Life Sciences Building, Portland, Ore.

Authors 

The authors, in descending order, are Drs. Archana Sehrawat and Lina Gao, OHSU Knight Cancer Institute; Yuliang Wang, Department of Biomedical Engineering, and Computational Biology Program, OHSU School of Medicine; Dr. Armand Bankhead, Department of Computational Medicine and Bioinformatics, University of Michigan; Dr. Shannon McWeeney, OHSU Knight Cancer Institute, Department of Medical Informatics and Clinical Epidemiology, OHSU School of Medicine; Carly King, Department of Biomedical Engineering, OHSU School of Medicine; Jacob Schwartzman and Joshua Urrutia, OHSU Knight Cancer Institute; Dr. William Bisson, Department of Environmental and Molecular Toxicology, Oregon State University; Drs. Daniel Coleman, Sunil Joshi, Daehwan Kim, and David Sampson, OHSU Knight Cancer Institute; Dr. Shelia Weinmann, Kaiser Permanente Center for Health Research; Dr. Bhaskar Kallakur, Department of Pathology, Georgetown University; Dr. Deborah Berry, Lombardi Comprehensive Cancer Center, Georgetown University; Dr. Reina Haque and Stephen Van Den Eeden, Kaiser Permanente; Dr. Sunil Sharma and Jared Bearss, Huntsman Cancer Institute, University of Utah; Drs. Tomasz Beer and George Thomas, OHSU Knight Cancer Institute; Dr. Laura Heiser, Department of Biomedical Engineering, OHSU School of Medicine ; and Dr. Joshi Alumkal.

Resources

• Read the paper
• Read the commentary from the same issue
  
• Cracking the code of advanced prostrate cancer (OHSU Cancer Translated blog)
  

Citation

Sehrawat, Archana, Lina Gao, Yuliang Wang, Armand Bankhead, Shannon K. McWeeney, Carly J. King, Jacob Schwartzman et al. "LSD1 activates a lethal prostate cancer gene network independently of its demethylase function." Proceedings of the National Academy of Sciences (2018): 201719168.

Ellis, Leigh and Loda, Massimo. "LSD1: A single target to combat lineage plasticity in lethal prostate cancer." Proceedings of the National Academy of Sciences (2018)

Pictured above, from left to right, Dr. Joshi Alumkal, Chelsea Jenkins, Jacob Schwartzman, Archana Sehrawat, Daehwan Kim, David Sampson, Daniel Coleman.