Imaging mass cytometry can ‘aid precision immunology’ in cancer research
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Imaging mass cytometry appeared feasible and allowed for a more comprehensive overview of triple-negative breast cancer heterogeneity at a single-cell level, according to study results presented at San Antonio Breast Cancer Symposium.
The next-generation technology also provided insights into the value of adding the anti-PD-L1 monoclonal antibody atezolizumab (Tecentriq, Genentech) to neoadjuvant chemotherapy for patients with early high-risk and locally advanced triple-negative breast cancer, results showed.
“We are experiencing a revolution in the technologies available for characterizing the molecular complexity of tumors,” Giampaolo Bianchini, MD, head of the breast cancer group in the department of clinical oncology at IRCCS Ospedale San Raffaele in Italy, said in a press release. “Among these, imaging mass cytometry allows us to collect unprecedented information about the heterogeneity of tumors and their surrounding microenvironment.”
Imaging mass cytometry provides high-dimensional tissue imaging at subcellular resolution, which yields information about cell type composition, functional status and spatial organization, according to study background.
This approach — which combines the principles of flow cytometry and mass spectrometry — allows for simultaneous analysis of more than 40 markers in a single tissue section. This helps determine the proteins present on individual cells, as well as their precise location within tissue.
Prior studies established the effectiveness of immune checkpoint inhibitors for early and advanced triple-negative breast cancer, and the anti-PD-1 antibody pembrolizumab (Keytruda, Merck) is approved in the United States in combination with neoadjuvant chemotherapy in this setting. However, only a small percentage of patients derive benefit.
“Unfortunately, one size does not fit all patients,” Bianchini said. “It is possible that some of them may have responded to chemotherapy alone, while others who originally benefited from immunotherapy will eventually relapse.
“In addition, although immunotherapy is overall well-tolerated, some rare but potentially serious immune-related side effects have been reported,” Bianchini added. “For these reasons, biomarkers are urgently needed to help us identify the patients who will benefit the most from the addition of immunotherapy — potentially leading to chemotherapy de-escalation or [chemotherapy]-free strategies — and those who will do well just with chemotherapy.”
Bianchini and colleagues assessed whether imaging mass cytometry analysis — performed in the context of the phase 3 NeoTRIPaPDL1 trial — could determine which patients may derive the most benefit from this treatment strategy.
The NeoTRIPaPDL1 trial included 280 patients with early high-risk and locally advanced triple-negative breast cancer.
Researchers randomly assigned patients to eight cycles of neoadjuvant nab-paclitaxel (Abraxane, Bristol Myers Squibb) and carboplatin with or without atezolizumab. All patients underwent surgery within 6 weeks of treatment completion.
Bianchini and colleagues used pretreatment biopsies from 243 evaluable patients to assess 43 proteins expressed on more than 1 million single cells.
“[Formalin-fixed paraffin-embedded] samples were labeled with antibodies conjugated to isotopically pure rare earth metal reports and profiled at one micron resolution by imaging mass cytometry,” researchers wrote. “For each sample, we ... generated three high-dimensional images that encompass the tumor, tumor-stroma interface and adjacent stroma.”
Researchers evaluated the association of protein expression — assessed separately for tumor microenvironment and epithelial cells, cell phenotypes and spatial architectures — and pathologic complete response.
They determined PD-L1-positive tumors, high stromal tumor-infiltrating lymphocytes and triple-negative breast cancer type exhibited extreme heterogeneity, as well as unique cell-type and spatial tumor microenvironment composition.
Researchers identified several biomarkers that exhibited a significant test for interaction.
With regard to protein expression, these included GATA3 and CD20 in the tumor microenvironment, HLA-DR in epithelial cells, and Ki67 assessed both in epithelial cells and the tumor microenvironment (P < .05 for all).
High expression of all of these biomarkers appeared associated with increased pathologic complete response rate with atezolizumab. Among patients with lower expression, researchers reported similar pathologic complete response rates with or without atezolizumab.
Results showed two cell phenotypes — PD-L1-positive plus indoleamine 2,3-dioxygenase (IDO)-positive antigen-presenting cells, and CD56-positive neuroendocrine epithelial cells — had a significant test for interaction.
Greater expression of these biomarkers appeared associated with increased likelihood of pathologic complete response among patients assigned chemotherapy plus atezolizumab, but not among those who received chemotherapy alone.
High degree of spatial connectivity between specific tumor microenvironment cells and epithelial cells also appeared associated with a significantly higher pathologic complete response rate among atezolizumab-treated patients. In contrast, patients with lower expression of these markers achieved similar pathologic complete response rates regardless of whether they received atezolizumab-chemotherapy or chemotherapy alone.
“Our results demonstrated that spatial data on the interactions among specific cells in the [tumor microenvironment] might be very informative about the benefit provided by an immune checkpoint inhibitor such as atezolizumab in addition to chemotherapy,” Bianchini said. “This type of information can only be provided by technologies that allow us to simultaneously characterize the single cells and their spatial localization with precision.”
The majority of interaction tests remained significant after researchers adjusted for stromal tumor-infiltrating lymphocytes and PD-L1 status. However, none of the seven proliferation-related signatures or 61 gene expression-based immune-related pathways demonstrated a significant test of interaction.
The results of this study must be validated independently, and the applicability and reproducibility of this technology outside the research setting still must be established, researchers acknowledged.
However, the results show the imaging mass cytometry can be applied successfully to samples prospectively collected in large trials, suggesting the potential for broad implementation in oncology research “to aid precision immunology,” Bianchini said.
“Overall, we demonstrated that imaging mass cytometry is feasible in a large, randomized trial and provides independent predictive information on immune checkpoint inhibitors’ benefit to PD-L1, [tumor-infiltrating lymphocyte] and gene-expression profiles,” Bianchini and colleagues wrote.
Perspective
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The KEYNOTE-522 trial established that pembrolizumab, when added to neoadjuvant chemotherapy and then adjuvant therapy, increased complete pathologic response in triple-negative breast cancer irrespective of PDL-1 status. The pathologic complete response rate with pembrolizumab was 65%, a clinically meaningful improvement over neoadjuvant chemotherapy alone (51%).
Pembrolizumab is now the standard of care in localized triple-negative breast cancer. However, what about the 35% of patients who do not achieve pathologic complete response?
Investigators have piloted a new technology, imaging mass cytometry, in the context of a randomized trial in the neoadjuvant triple-negative breast cancer setting.
Imaging mass cytometry is designed to do single-cell analyses of breast cancer cells and cells in the surrounding tumor microenvironment by combining flow cytometry techniques to identify biomarkers and mass spectrometry to evaluate 43 proteins. The investigators created 3D maps that delineated critical relationships between the tumor cells and cells in the tumor microenvironment (eg, immune cells and other cells). These relationships led to predictions from the initial tumor biopsies about higher — or lower — likelihood of pathologic complete response and response to atezolizumab.
Imaging mass cytometry allows a “'deeper dive” into the determinants of pathologic complete response and more significant insights into the biology of triple-negative breast cancer by allowing an appreciation of the spatial relationships between the tumor cells and surrounding tumor microenvironment. It seems that this technology could extend to other subsets of breast cancers and different cancer sites. It will take a while before imaging mass cytometry has relevance for routine clinical practice, but this is a great start.
Icahn School of Medicine at Mount Sinai
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Bianchini G, et al. Abstract GS1-00. Presented at: San Antonio Breast Cancer Symposium; Dec. 7-10, 2021; San Antonio.
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