NEW YORK (GenomeWeb) – Vanderbilt University researchers have developed a targeted proteomics assay that could prove useful in the development of cancer immunotherapies.
Described in a study published last week in Molecular & Cellular Proteomics, the assay uses parallel-reaction monitoring mass spectrometry to measure levels of eight proteins involved in immune checkpoint regulation and, said Daniel Liebler, senior author on the paper, and highlights potential gaps in existing antibody-based tests used to guide immune-oncology treatment.
Cancer immunotherapies aim to use the body's immune defenses to attack cancer. One main approach clinicians have taken in this effort is targeting immune checkpoint proteins like programmed cell death 1 (PD-1), which is expressed on CD8 T cells, and its ligand programmed cell death 1 ligand 1 (PD-L1), which is expressed in certain tumors.
When PD-1 binds PD-L1, it protects tumor cells from the body's immune response, and so, the thinking goes, inhibiting this binding could eliminate this protection and expose tumor cells to the immune system. Such approaches have emerged in recent years as a hot area of cancer drug research with more than 1,000 clinical trials launched for immune checkpoint therapies, and inhibitors of PD-1-PD-L1 binding have proved effective in cancers including melanoma, renal clear cell carcinoma, non-small cell lung cancer.
At the same time, response rates vary significantly. According to the MCP authors, between 15 percent and 45 percent of solid tumor patients benefit from immunotherapies. This, they note, suggests that both a better understanding of the mechanics underlying this response and better biomarkers and assays for predicting it are needed.
Mass spec could provide help in this regard, said Liebler, formerly professor of biochemistry at Vanderbilt's School of Medicine. He left this position at the end of last year to found Protypia, a mass spec firm focused on cancer proteogenomics and cancer drug development.
Currently, PD-L1 is the most commonly measured marker in cancer immunotherapy with patient levels typically assessed using immunohistochemistry. However, Liebler said, there are a number of questions around the performance and standardization of these tests.
"There's a lot of literature and a lot of activity on immunohistochemistry for PD-L1," he said. "And there are multiple competing commercial tests and they all have different alliances with different drug companies and different drugs that are in development. So the picture there is very complicated."
According to Lieber and his co-authors, clinical trials have found that up to 50 percent of tumors positive for PD-L1 by IHC fail to respond to immunotherapy, while around 15 percent of PD-L1-negative tumors do respond.
"They're trying to harmonize these tests, but they use different antibodies and different cutoffs for interpreting PD-L1 positive versus negative tissues and so on. It's kind of a mess," he added. So I thought that this was an area that could benefit a lot from targeted mass spec."
Additionally, while PD-L1 is the most commonly used marker for guiding immunotherapy, additional proteins might also be useful, and mass spec could enable improved detection and quantitation of these proteins, Liebler said.
He noted in particular the protein PD-L2, which like PD-L1 can protect tumor cells from the immune response by binding PD-1, but which has received less attention due in part to the perception that it is not commonly expressed in solid tumors.
"People have assumed or people have concluded that it's not present in solid tumors," he said. "But it's also known that there really aren't good antibodies to detect this protein in immunohistochemistry."
Using mass spec, Liebler and his colleagues, demonstrated in a set of 22 melanoma samples that, in fact, PD-L2 is present at levels comparable to PD-L1, suggesting, he said, "that it may be contributing to the silencing of T cells in melanoma, and that you can't ignore [it]."
The researchers also compared in the melanoma samples the results of their mass spec-based assays to IHC tests for PD-1 and PD-L1, finding that while the two approaches provided largely similar results, the mass spec measurements offered potential insights into the performance of the IHC test. Specifically, they identified glycosylated forms of PD-L1 that the IHC test had difficulty detecting, suggesting that some tumors that test negative for PD-L1 may, in fact, have substantial levels of glycosylated forms of the protein.
They also looked at several other proteins — IDO1, LAG3, HAVCR2 (TIM-3), C10orf54 (VISTA) and CD40 — known to regulate interactions between tumors and the immune system, finding that each was present at different levels across the samples, none of which could be predicted by a sample's levels of PD-1, PD-L1, or PD-L2, indicating that they provide additional independent information around tumor-immune interactions.
The Vanderbilt researchers are not alone in adopting a mass spec-based approach to studying cancer immunotherapy. For instance, molecular diagnostics firm Biodesix said last month that it plans this year to launch a MALDI mass spec-based test for guiding immunotherapy in non-small cell lung cancer patients.
Liebler said, however, that unlike Biodesix, he doesn't see mass spec in the short term becoming a clinical platform for guiding immunotherapy. Rather, he said he envisions mass spec assays like that presented in the MCP paper as tools for further refining IHC-based tests.
"I don't have the opinion that mass spec will replace immunohistochemistry and so forth," he said. "I don't see that's realistic in any reasonably near-term future."
"I think it's quite likely that this kind of platform could help refine antibody based approaches to diagnostics," Liebler said. "It could help in the choice of reagents. It would also, perhaps, give you a better idea of the limitations of what your antibody can see."
For instance, the researchers noted, in the case of patients negative for PD-L1 by IHC but who nonetheless responded to treatment, mass spec could help test whether IHC failed to detect PD-L1 due to post-translational modifications like glycosylation or whether expression of PD-L2 compensated for a lack of PD-L1.
"I think that targeted mass spec would be something that immunohistochemistry providers would want to use to help improve and refine their platforms," Liebler said.