Raikar Laboratory

The overall goal of the Raikar laboratory is to develop novel therapeutics for pediatric blood cancers through (i) development of innovative cellular immunotherapies utilizing chimeric antigen receptors, (ii) production of novel recombinant protein-based therapeutics such as L-asparaginase and (iii) identifying new therapeutic targets in rare leukemias utilizing advanced bioinformatic tools. The specific diseases we investigate include acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML) and mixed phenotype acute leukemia (MPAL). While the survival of pediatric leukemia patients has greatly improved with the intensification of chemotherapy, relapsed disease still accounts for high rate of mortality among childhood cancer patients. Thus, a need exists to develop novel alternative approaches to target relapsed disease with lesser side effects.

In recent years, chimeric antigen receptor (CAR) T-cell immunotherapy for relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL) has been a revolutionary breakthrough in pediatric cancer. In this form of therapy, the patient's own immune cells (T cells) are genetically modified to express receptors called CARs, which enable them to specifically target B cells. While CAR T-cell therapy is extremely successful in targeting B-cell cancers, the same approach has not been successful in targeting T-cell cancers such as T-cell acute lymphoblastic leukemia (T-ALL), which has a higher rate of relapse and is more difficult to cure compared to B- ALL. One of our main research aims is centered on adapting this novel CAR technology to T-cell disease. Given the lack of a tumor specific antigen in T-cell malignancies, utilizing CAR-based immunotherapy in this disease has been extremely challenging as it can result in (i) fratricide (self-killing) of CAR T cells, (ii) long-standing immunosuppression from T-cell aplasia and (iii) product contamination from accidental transduction of malignant T cells. We are exploring several different approaches to overcome these challenges, including the use of unique immune cells such as natural killer cells and gamma delta T cells. As an extension of this work, we are now also exploring the use of cellular therapy in acute myeloid leukemia (AML), a more aggressive form of childhood blood cancer with survival around 60-70%.

The Raikar lab also has interest in developing novel chemotherapeutics through protein engineering. A current ongoing project is the development of a novel humanized L-asparaginase drug candidate utilizing an innovative protein drug discovery and optimization platform called ancestral sequence reconstruction (ASR). L-asparaginase has been a critical component of the chemotherapy armamentarium used to treat ALL for several decades. However, current L-asparaginases are bacterial in origin, derived from either Escherichia coli and Erwinia chrysanthemi, and hence are highly immunogenic with reactions ranging from silent inactivation to severe anaphylaxis. Additionally, these drugs have a high level of liver and pancreatic toxicity, thus limiting its widespread use. Recent data has shown that discontinuation of L-asparaginase results in poorer prognostic outcomes in ALL. Thus, development of a more humanized and less immunogenic asparaginase is essential to overcome the major deficiencies of the current bacterial L-asparaginase products. We are currently utilizing the ASR platform to identify and characterize potential less toxic L-asparaginase candidates.

Finally, we have a specific interest in studying the biology of mixed phenotype acute leukemia (MPAL), a rare leukemia accounting for only ~2-3% of all pediatric leukemias. MPAL has features of both AML and ALL and can be broadly divided into two major subtypes - B/myeloid and T/myeloid MPAL. While recent literature has suggested that this disease responds better to ALL therapy, there remains no clear consensus as to how to treat this disease. We recently published the first study describing the single cell transcriptomic landscape of pediatric MPAL. We are now expanding this study to include genomic, epigenetic and proteomic analysis and hope to identify novel therapeutic targets to treat MPAL through this work.

• Developing chimeric antigen receptor (CAR) based strategies to target T-ALL and AML
• Utilizing gamma delta T cells in combination with chemotherapeutic agents to target AML and T-ALL
• Utilizing microfluidic mechanotransfection as a means to deliver CRISPR/Cas9 editing molecules for developing gene-edited CAR T-cells (collaboration with Todd Sulchek, PhD)
• Developing a novel humanized L-asparaginase with reduced toxicity utilizing ancestral sequence reconstruction (collaboration with Christopher Doering, PhD)
• Leveraging single cell multiomic analysis to study the biology of MPAL (collaboration with Manoj Bhasin, PhD)
• Studying the effects of the obese microenvironment on CAR T-cell function (collaboration with Curtis Henry, PhD and Sarwish Rafiq, PhD)