Targeting Alternative RNA Splicing in Pediatric Cancers

RNA splicing, the enzymatic process of removing segments of premature RNA to produce mature RNA, is a key regulator of gene expression. Recent studies have identified variations in the transcriptomes of tumors due to alternative splicing changes, in addition to mutations in splicing factors and regulatory signals in most tumor types. Some of these alterations have been linked to tumor growth, metastasis and therapy resistance.

Dawn Chandler, PhD, is a principal investigator in the Center for Childhood Cancer and Blood Diseases at the Abigail Wexner Research Institute at Nationwide Children’s. Her lab studies the regulation of pre-mRNA splicing and how disruption of this regulation can lead to pediatric cancers.

“We know that the tumor suppressor protein p53 is critically important in a number of different cancers,” says Dr. Chandler. “We also know that the oncogene MDM2 is one of the two main regulators of that tumor suppressor protein. When MDM2 is overexpressed, it can turn off p53 and basically turn off all its tumor suppressor activities.”

In response to genotoxic stress, MDM2 undergoes alternative splicing to generate splice variants that are unable to regulate p53 expression, contributing to tumor formation. Dr. Chandler and her colleagues have previously shown that a protein called SRSF1 is a negative regulator of MDM2 splicing that induces the formation of an alternative splice variant called MDM2-ALT1 in response to genotoxic stress.

To study the alternative splicing of MDM2, Dr. Chandler and her colleagues developed a damage-inducible minigene system, a shortened representation of MDM2 that allowed them to home in on how it is regulated. They reported their findings in the journal Molecular Cancer Research.

Dr. Chandler and her colleagues identified a protein called SRSF2 as a positive splicing factor of MDM2. Mutation of the SRSF2-binding sites or targeting them with splice-switching oligonucleotides promoted expression of the alternatively spliced transcript MDM2-ALT1 and p53 protein expression. Overexpression of SRSF2 reduced the generation of MDM2-ALT1 under genotoxic stress, whereas SRSF2 knockdown induced the expression of MDM2-ALT1 in the absence of genotoxic stress.

“This really opens up a whole array of new targets that can be identified and pursued with these methodologies,” says Dr. Chandler, who is also an associate professor in the Department of Pediatrics at The Ohio State University College of Medicine.

“We think we can use splicing modifications to treat a multitude of different diseases, including cancer.”

MDM2 is overexpressed in many types of cancer, such as osteosarcoma, and these cancers would benefit from persistent alternative splicing of MDM2 to downregulate MDM2 expression, reactivate p53, and sensitize these tumors to current therapies.

Having demonstrated these changes in tissue culture cells, Dr. Chandler and her colleagues next want to investigate these effects in tumors within the body. She says a big challenge for the future is controlling splicing in targeted cells in the body. Another goal of Dr. Chandler’s team is to investigate genes in addition to MDM2 that could be potentially targeted with this same methodology.

“People might be intimidated by splicing and the complexity it lends to the already complicated transcriptome in cancer cells,” says Dr. Chandler. “Even if it’s complex, it may actually be targetable and understanding these processes on a more global level may be attainable.”

Reference:

Comiskey Jr., DF, Montes M, Khurshid S, Singh RK, and Chandler DS. SRSF2 regulation of MDM2 reveals splicing as a therapeutic vulnerability of the p53 pathway. Molecular Cancer Research. Oct 29 2019. Doi: 10.1158/1541-7786.MCR-19-0541.