Leveraging Immune Biomarkers in the Blood to Guide Immunotherapy—Current Application and Future Prospects

Leveraging Immune Biomarkers in the Blood to Guide Immunotherapy—Current Application and Future Prospects

Immune cells and tumor cells represent two opposing forces. By the time a tumor reaches macroscopic proportions, these two cell subsets share a complicated history of antagonism, resistance, and suppression that has shaped the tumor microenvironment battleground. Immune biomarkers offer insight into the past and ongoing interactions between the immune system and cancer—key intelligence that researchers can then exploit to better enable the former to defeat the latter by appropriately applying select immunotherapies.

Several talks given during the General Session “Immune Biomarkers in the Blood” at the ASCO-SITC Clinical Immuno-Oncology Symposium focused on this theme, highlighting the many blood-based immune biomarkers available and how these are being applied in the clinic.

Immune Cell Subsets and Functional Markers

Dr. Jeffrey Schlom
Analyzing a primary tumor biopsy can provide valuable information regarding a tumor’s phenotype and the immune cells infiltrating the lesion. However, this approach has its limitations. A sample is not always available, the sample taken may reflect only a small segment of a heterogeneous tumor, or it may differ markedly in composition from metastatic lesions.

To overcome these limitations, analysis of the peripheral immunome—the genes, proteins, and cells comprising the immune system—may provide supplementary information representative of all possible sites of disease to help guide effective immunotherapy, according to presenter Jeffrey Schlom, PhD, of the Center for Cancer Research, National Cancer Institute, National Institutes of Health.

Analysis of the immunome can reveal the effects of more traditional anticancer agents on the immune system and how these effects might impact immunotherapy—an area that Dr. Schlom said is often overlooked. To illustrate this, Dr. Schlom’s team has identified an inverse correlation between the number of prior chemotherapies and the immune response to a tumor vaccine, suggesting that patients heavily pretreated with chemotherapy are not the best candidates for vaccine-based approaches.

Assessing the immunome can also lend insight into the effect of immunotherapies on peripheral immune cell subsets that are not easily identified in biopsies. Dr. Schlom showed data in which flow cytometry of 30 markers on peripheral blood mononuclear cells identified 123 unique immune cell subsets. This analytic ability was put to the test in the setting of avelumab treatment, a fully human monoclonal antibody targeting PD-L1 that was flagged as possibly harming PD-L1–expressing immune cells. Using the flow cytometry technique, very little effect, if any, on the immune cell subsets was observed at various time points after administration of avelumab, even on cells expressing PD-L1, indicating that the agent can be safely administered to patients with cancer.

Emerging studies are showing that, in certain cancers, it is possible to generate algorithms that yield a “peripheral immunoscore” based on the types of immune cells present in the peripheral blood. These scores, once validated, can be used to identify patients most likely to benefit from immunotherapy or combination therapies. 

Imaging Inflammation

Dr. I. Jolanda M. de Vries
I. Jolanda M. de Vries, PhD, and her colleagues at the Radboud Institute for Molecular Life Sciences, in the Netherlands, have been focusing their efforts on rapidly developing myeloid dendritic cell vaccines targeted against melanoma antigens using automated technology. Once synthesized, the first vaccine dose is injected into a patient’s lymph nodes—the training ground for naive T cells—a mere 48 hours after the dendritic cells were originally harvested from the patient.

To ensure that this vaccine approach triggers the necessary immune responses to generate clinical benefit in a disease state where there is little room to squander time, Dr. de Vries’ team has relied on several imaging strategies to visualize early immune processes in the body.

First, they have leveraged both scintigraphic and MRI to track the dendritic cells after injection. Co-labeling the cells with indium-111 for scintography and a superparamagnetic iron oxide formulation for MRI not only enabled the accuracy of vaccine delivery to be determined, but it also revealed the intranodal migration patterns of the dendritic cells.

The group has also employed PET using radiolabeled 3′-fluoro-3′-deoxy-thymidine (18F-FLT), a thymidine analog that incorporates into the DNA of proliferating cells at varying time points after vaccination to visualize immune responses in the lymph nodes. Dr. de Vries stated that increased 18F-FLT uptake by proliferating T cells in the lymph nodes in response to the antigen-loaded dendritic cells could be seen as early as 2 days and as long as 3 weeks after vaccination. In contrast, control lymph nodes not injected, injected with saline, or injected with unmodified dendritic cells showed no selective 18F-FLT uptake.

Dr. de Vries has used the PET technique enough times now to know that the absence of 18F-FLT uptake by a patient injected with the dendritic cell vaccine portends a lack of tumor response, allowing her and her team to quickly change course in terms of melanoma treatment.

PD-L1 Expression Within Exosomes

Dr. Theresa L. Whiteside
Presenter Theresa L. Whiteside, PhD, MDhc, of the University of Pittsburgh Cancer Institute, framed her talk with this question: Are PD-L1/PD-1 exosomes in plasma biomarkers of tumor immune evasion? The short answer is possibly. But first, a bit of background is in order.

Exosomes are small membrane-bound vesicles made and released by nearly all cells, including tumor cells. These vesicles are decorated with a variety of lipids and proteins taken from their parent cells, while the lumen holds pieces of DNA and other nucleic acids, along with a bevy of cytosolic proteins from the parent cells. Exosomes travel through the systemic circulation and may serve as vehicles of communication between cells.

Use of immunologic techniques (e.g., Western blot) to view the molecular cargo within exosomes isolated from an individual’s plasma reveals that tumor-derived exosomes (TEX) carry a unique payload that reflects, in part, the molecular and genetic content of the parent tumor, like a liquid tumor biopsy. Within the exosome mélange in patients with active disease are an abundance of PD-1, PD-L1, TRAIL, TGF-b, CTLA4, and COX2, among other molecules.

Because “all cells are bathed in a sea of exosomes” in the peripheral blood, according to Dr. Whiteside, this enables tumors the opportunity to interact with immune cells outside the tumor microenvironment. More specifically, this affords TEX the ability to block immune-cell functions by delivering immunosuppressive signals such as PD-L1, TGF-b, and TRAIL directly to effector T cells, while also promoting the differentiation and function of regulatory T cells. This is supported by bench research showing that exosomes isolated from patients with cancer can induce apoptosis of activated CD8+ T cells, suppress the proliferation of activated CD4+ T cells, and increase the frequency of CD4+CD39+ regulatory T cells. 

Dr. Whiteside presented the idea that just as TEX can serve as surrogates for a tumor biopsy, immune cell-derived exosomes (IEX) may be able to serve as surrogates that reflect a patient’s immune status. Dr. Whiteside envisions that, similar to TEX, IEX carry molecular cargo derived from immune cells that may signal immune dysfunction and potential tumor immune escape. Her laboratory is currently trying to tease apart TEX from IEX within patient plasma to identify biomarkers pertinent to the tumor and pertinent to the immune competence of the patient.

Serologic Markers

Dr. Sacha Gnjatic
Presenter Sacha Gnjatic, PhD, of the Icahn School of Medicine at Mount Sinai, concluded the session by discussing serologic markers of immune responses, with a focus on circulating antibodies with specificity to tumor antigens. He said that the serum represents a global view of events in the body, including those occurring locally, but that by sifting out specific markers from the serum, clinicians may be able to obtain a snapshot of events occurring at the tumor site.

Antibodies continuously develop as tumors grow and shed antigens. “Antibodies spontaneously developing to tumors are a sign of immunocompetence,” Dr. Gnjatic said, and, because of this, they can be leveraged as diagnostic tools to inform patient prognosis, predict the response to treatment, and monitor treatment response.

As an example, Dr. Gnjatic showed that anti–NY-ESO-1 antibody titers from the sera of patients with non–small cell lung cancer correlates with tumor stage, where the higher the titer the more advanced the disease, and clinical outcomes, with the presence of NY-ESO-1 antibodies heralding shorter overall survival. He also presented research suggesting that a positive NY-ESO-1 antibody status in patients with melanoma prior to treatment predicts a more favorable response to ipilimumab.

Dr. Gnjatic offered these take-home messages regarding the use of serologic markers in immunotherapy:

  • Changes in serum analytes and antibody responses can be useful for understanding the mechanistic aspects of anticancer treatment.
  • Although antibody responses may offer utility as pharmacodynamic biomarkers, they are not always related to clinical benefit.
  • Unlike with targeted therapies, it is unlikely that a single predictive serum biomarker will be found for immuno-oncology.
  • High-dimensional high-throughput assays, or seromics, can detect antigen-specific changes in autoantibody profiles, providing complex signatures that may be mined for relevant signals.

– Kara Nyberg, PhD