Researchers Identify Factors Associated With CAR-T Resistance in B-ALL

Rimas Orentas, PhD, is a professor in the Department of Pediatrics at the University of Washington School of Medicine and a researcher at Seattle Children’s Research Institute, where he also serves as Director of the Ben Towne Center for Childhood Cancer Research. Dr Orentas also serves as Director of Scientific Integration for CureWorks, a network of like-minded academic institutions looking to bring novel clinical trials to more children, faster.

In an interview with Oncology Learning Network, Dr Orentas discusses CAR-T resistance in patients with B-cell acute lymphoblastic leukemia (B-ALL) and results from an analysis presented at the 2019 ASH Annual Meeting.

What existing data led you and your co-investigators to conduct this research?

Seattle Children’s has one of the largest immunotherapy portfolios for children with cancer in the country. The size and breadth of our clinical experience in using chimeric antigen receptor (CAR) modified T-cells. to treat leukemia led to a series of unique observations. These included published studies by Drs Rebecca Gardner and Michael Jensen describing T-cell attributes and disease characteristics, such as the amount of B-ALL present in the marrow, that correspond to good responses to CAR-T therapy. 

While many patients with B-ALL do have phenomenal responses to CAR-T therapy, we found a set of patients who appeared to be inherently resistant to this treatment. These CAR-T resistant patients were the focus of our study.

Please briefly describe your study and its findings. Were any of the outcomes particularly surprising?

Using bone marrow samples from CAR-T resistant patients, collected and organized by Drs Olivia Finn and Ashley Wilson of our correlative studies lab, we initiated one of the broadest genomic analyses of CAR-T patient leukemia to date. When we complied, data used to stratify patient risk, we found that standard genetic attributes of the disease in patients (such as the presence of the Philadelphia chromosome, that result of a translocation and associated with poor outcome) did not correlate with CAR-T susceptibility or resistance. 

Therefore in association with Dr Javed Khan of the Genetics Branch of the NCI (National Cancer Institute, NIH, Bethesda, MD) we carried out RNA-sequencing (bulk), single-cell RNA sequencing, whole-exome sequencing, DNA-methylation analysis, analysis of chromatin structure by ATAC-Seq, gene enhancer analysis by H3K27ac ChIP Seq, analysis of VDJ rearrangements in the leukemia, and CYTOF advanced proteomic analysis. 

We are still analyzing these large data sets and we have made some important preliminary observations.  The CAR-T resistant disease appears to have an inherently more “open” chromatin structure, meaning that more genes are able to be expressed in a more fluid manner at any one time. The term for this is “genetic plasticity.” We have also described unique fusion genes and mutations that may be responsible for this plasticity.

Usually we think of leukemia as evolving over time, or mutating specific genes in response to therapy, including CAR-T therapy. But in these inherently resistant cases, the disease may be already capable of escape from therapy, even before treatment.

What are the possible real-world applications of these findings in clinical practice?

There are 2 ways in which these studies will impact our future clinical practice. First, we want to be able to identify CAR-T patients that are resistant prior to initiating therapy. As we nail down a specific genetic signature, we will identify those patients that we know should immediately be referred to alternative approaches to single agent CAR-T. Secondly, our data will set the stage for new therapeutic interventions, potentially based on addressing the epigenetic variability that we are now quantifying, and which is likely responsible for the plasticity we observe.

Do you and your co-investigators intend to expand upon this research? If so, what are/will be your next steps?

The advanced genomic analysis we have initiated creates so much data that it takes our bioinformatics team months to analyze it. As this analysis is refined, we will continue to gain insight as to the specific disease attributes that are associated with resistance to CAR-T therapy. We will expand on this research by identifying other B-ALL patients who are resistant to CAR-T in order to see if our findings hold up. We will also begin to design CAR-T approaches that may be able to target the resistant population as well as small molecule approaches designed to limit epigenetic plasticity.  

Is there anything else pertaining to your research and findings that you would like to add?

This research is not possible without the brave patients and their families who consented to these studies, and who have inspired us to keep searching for new cures and less toxic cures for B-ALL. We can’t do this without them, and they are paving the way forward for new treatments for patients we will see in the future. They are always the story behind the story.