Scientists have identified the cell-surface co-receptor, glypican 2 (GPC2) as a key immunotherapy target for high-risk neuroblastoma and other childhood cancers, and developed an antibody–drug conjugate (ADC). [ascan.org]
Scientists have identified the cell-surface co-receptor, glypican 2 (GPC2) as a key immunotherapy target for high-risk neuroblastoma and other childhood cancers, and developed an antibody–drug conjugate (ADC). [ascan.org]

Scientists in the U.S. and Canada have identified a key immunotherapy target for high-risk neuroblastoma and other childhood cancers, and developed an antibody-drug conjugate (ADC) targeting the cell surface co-receptor, glypican 2 (GLP2), which regressed tumors and prolonged life in a mouse model of neuroblastoma.

The researchers, led by John M. Maris, M.D., a pediatric oncologist at the Children’s Hospital of Philadelphia (CHOP), claim their work lays “firm groundwork” for the development of GPC2-directed immunotherapies against neuroblastoma, including ADCs but also potentially CAR T cell therapies.

Reporting on their studies in Cancer Cell, they conclude, “Additional preclinical efficacy and toxicity studies could lead to the introduction of an immunotherapeutic for children with high-risk neruoblastoma and other aggressive pediatric malignancies.” The team describe their work in a paper entitled, “Identification of GPC2 ad an Oncoprotein and Candidate Immunotherapeutic Target in High-Risk Neuroblastoma.”

Neuroblastoma is a cancer of the developing peripheral nervous system, and the most common infant cancer. Long-term survival is less than 50%, even following aggressive cytotoxic therapy. Although a monoclonal antibody targeting the cell surface disialoganglioside GD2 has been approved for treating neuroblastoma, other types of immunotherapy, including ADCs, haven’t been studied against this cancer, primarily because there is a lack of known differentially expressed cell-surface molecules coupled with targeted therapeutics, the researchers write  “… a major hurdle to the development of effective immune-based therapies for pediatric cancers is in identifying tumor-specific cell-surface molecules with limited expression on normal childhood tissues. Ideally these differentially expressed cell-surface molecules will also be required for tumor sustenance, a second criterion that is becoming recognized as necessary to produce a long-lasting tumor cytotoxic effect without immune escape.”

To identify proteins that might be suitable targets for neuroblastoma immunotherapy, the team compared RNA sequencing data from 126 high-risk primary neuroblastomas, with RNA sequences from samples across 31 normal tissue types. “Our rationale was to identify a cell-surface molecule that an immune-based therapy could target without damaging healthy tissues,” commented Kristopher R. Bosse, M.D., also at CHOPS, who is lead author of the published paper. They filtered an initial list of 296 differentially expressed genes to identify those that had the qualities of an optimal candidate immunotherapeutic target, “including potentially having extracellular epitopes susceptible to cell- or protein-based immunotherapeutics and being highly expressed in the majority of neuroblastoma cases …”.

Through this process the team narrowed down their search to the extracellular glycosylphosphatidylinositol (GPI)-anchored signalling co-receptor glypican 2 gene. GPC2 is one of a family of six (GPC1-6) GPI-anchored, extracellular proteoglycan signalling coreceptors that play diverse roles in growth factor signalling and cancer cell growth. Interestingly, the glypican family member glypican 3 (GPC3) has recently been validated as a bona fide immunotherapeutic target in hepatocellular carcinoma, they point out.

When the researchers looked for GPC2 expression in a number of different neuroblastoma tumor datasets they confirmed that high levels of cell surface GPC2 were associated with worse overall survival, and were also evident in the tumors of patients with high-risk neuroblastoma. Direct analysis of tissue samples and cultured cells identified elevated GPC2 in high- and intermediate-risk neuroblastoma primary tumors, patient-derived xenografts (PDXs) derived from high-risk human primary tumors, and the majority of neuroblastoma cell lines evaluated. The researchers then confirmed that GPC2 protein was only present at very low levels, if at all, in the cell surface membranes of a wide range of normal pediatric tissues, and neural- and neural-crest-derived tissues.

To find out whether GPC2 was required for neuroblastoma cell proliferation the team next depleted the protein in neuroblastoma cell lines that normally expressed GPC2 at high levels. This resulted in reduced cell growth and increased apoptosis. In contrast, stably overexpressing GPC2 in otherwise low GPC2-expression neuroblastoma cell lines resulted in increased cell proliferation. These results indicated that the protein is a key regulator of neuroblastoma cell growth, the researchers suggest. 

They next analyzed RNA sequencing and gene expression profiling datasets spanning a number of pediatric malignancies, and identified similarly high GPC2 expression in pediatric medulloblastoma and retinoblastoma, as well as in subsets of other childhood cancers. High level cell-surface GPC2 expression was also confirmed directly in the majority of diagnostic medulloblastoma and retinoblastoma tissues tested.

For a final set of experiments the team worked with colleagues at the National Cancer Institute to develop an ADC candidate, D3-GPC2-PBD, which targeted the cytotoxic drug pyrrolobenzodiazepine directly to cell surface GPC2. In vitro and in vivo tests showed that the ADC effectively killed GPC2-expressing cell lines, without damaging normal cells, but also led to tumor regression and increased the lifespan of a PDX neuroblastoma mouse model, without causing any discernible toxicity.

“These findings establish that this type of immunotherapy could be potentially safe and effective against neuroblastoma,” commented Dr. Maris. Added Dr. Bosse, “Given GPC2's critical role in the growth of neuroblastomas, we hope that tumors will not be able to simply downregulate this protein in order to escape recognition by our immunotherapies that target GPC2.”

“Our next steps will be to further evaluate this ADC and also develop other immune-based therapies directed against GPC2,” Dr. Maris stated. “Because other glypicans in addition to GPC2 are overexpressed in other childhood cancers, it may also be possible to apply this approach across various types of high-risk pediatric cancers.”

 








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