In the United States, turning 45 brings with it a rather unpleasant passage ritual: the beginning of regular colonics, in which an endoscope equipped with light and a camera is used to visually check the colon for signs of cancer. Colorectal cancer, which is slow-growing, can often be treated surgically if caught early. However, it becomes more difficult to treat the longer it lies undetected, making it the fourth leading cause of cancer-related deaths in the country.
Despite the availability of this advanced visual screening process, treatment decisions for individual patients are still largely guided by traditional histology – pathologists assess colorectal cancer by examining slides of tumor samples under a microscope.
Now, a team at Harvard Medical School has combined histology with cutting-edge single-cell imaging technologies to create large-scale 2D and 3D spatial maps of colorectal cancer. The maps, described in celllayering extensive molecular information on top of histological features to provide new information about the structure of cancer, as well as how it forms, progresses, and interacts with the immune system.
“Our approach provides a molecular window into 150 years of diagnostic pathology — and reveals that many of the elements and structures traditionally thought to be discrete are interconnected in unexpected ways,” said co-author senior Peter Sorger, the Otto Krayer Professor of Systems Pharmacology at the Blavatnik Institute at HMS. “The analogy is that we were only looking at the tail or the bottom of the elephant, but now, for the first time, we can start to see the whole elephant at once.”
The maps are part of the team’s broader efforts to create atlases for different types of cancer that will be readily available to the scientific community as part of the National Cancer Institute’s Human Tumor Atlas Network. Previously, the researchers used a similar approach to create depth maps of early-stage melanoma, and maps for other cancers are already being developed. Ultimately, the team hopes that these cancer atlases will stimulate research and improve diagnosis and treatment.
Knotted old and new
Histology has long been the cornerstone of cancer diagnosis and treatment: Pathologists examine a tumor sample stained with hematoxylin and eosin (H&E) under a microscope and select key features to determine the grade and stage of cancer. Oncologists use this information to develop a treatment plan, which usually involves some combination of surgery, drugs and radiation. H&E-based histology is relatively simple, cheap, fast, and can reveal a lot about a tumor.
“Our existing maps of colorectal cancer are from pathology — over a period of 150 years, the most important S&E features to diagnose a patient,” said co-senior author Sandro Santagata, HMS associate professor of systems biology and associate professor in pathology at Brigham and Women’s Hospital.
However, traditional histology has limitations – that is, it does not capture the molecular makeup or physical structure of cancer, making it difficult to take full advantage of the knowledge that cancer researchers have gained over the past 50 years.
“Histology is extremely powerful, but we often don’t know what it means in modern molecular terms,” said Sorger.
In the new paper, the researchers combined histology with single-cell molecular imaging data obtained through a multiplexed imaging technique called cyclic immunofluorescence, or CyCIF. They used this information to create detailed 2D maps of large regions of colorectal cancer. First author Jia-Ren Lin, platform director in the Laboratory of Pharmacology at HMS, led an effort to stitch these maps together to create a large-scale 3D reconstruction of a tumor.
“Our maps include information on nearly 100 million cells from large pieces of tumor, and provide a relatively unprecedented look at colorectal cancer,” Santagata said. They allow researchers to begin asking key questions about differences between normal and tumor tissue and variations within a tumor, he said, and reveal “exciting architectural features that have never been observed before, as well as molecular changes associated with these features.” “
The maps showed that a single tumor can have more and less invasive sections, and regions that appear more or less malignant — resulting in histological and molecular gradients where one part of the tumor moves into the next one.
“Within each tumor, there’s a wide range of colorectal cancer symptoms—we see many different regions and neighborhoods with distinct characteristics, as well as the transitions between them,” Santagata said. From here, he said, scientists can now explore what drives these differences within individual strains.
For example, the maps showed that there was great variation in immune environments within a single tumor.
“They were so different among tumors alone and among tumors – which is important because there are tumor-immune interactions that you want to target with immunotherapy,” said Sorger. Similar to their discovery in melanoma, the researchers observed that the T cells tasked with fighting the cancer were not directly suppressed by tumor cells, but by other immune cells in the environment surrounding the tumor.
“This gives us a new appreciation of how diverse and plastic tumor environments are—they’re rich communities, and we’re now better able to learn how they evolve,” Santagata said.
The maps also provided new insight into the architecture of the tumors. For example, scientists previously identified what they thought were 2D pools of a mucus-like substance called mucin with clusters of cancer cells floating within. However, in the new study, the 3D reconstruction showed that these mucin pools are, in fact, a series of cavities interconnected by channels, with finger-like projections of cancer cells.
“It’s a wild new look at these tumor structures that we’ve never really appreciated before,” Santagata said. “Because we can see them in 3D, we have a clear view of the structures, and we can now study why they exist, how they form, and how they shape the evolution of tumors.”
Ultimately, the goal of these colorectal cancer maps is the same as for all the cancer atlases the team is developing: to advance research and improve diagnosis and treatment. Precision medicine, which involves tailoring therapy to an individual patient’s cancer, is becoming an increasingly important part of treatment, according to Sorger, but it can only go so far with pathology and genetics alone.
“The big translational story here is developing the knowledge to make precision medicine practical for most patients,” he said. “We are currently working with Brigham and Women’s and the Dana-Farber Cancer Institute to determine how our methods can be used in a clinical setting.”
“This allows us to extract a whole additional set of molecular and structural features that we think will provide diagnostic and prognostic information and improve our ability to target these cancers,” added Santagata.
Now, the researchers want to further refine their ability to create 3D reconstructions of tumors and continue to integrate new imaging technologies into their maps. They also want to build a larger cohort of colorectal cancer samples for mapping and explore the underlying biology of the disease highlighted in their maps.
For Sorger, the project represents an unusual collaboration between pathologists, engineers, and computational scientists: As the imaging data was fed in, the computational scientists used machine learning to identify interesting findings that they presented to the pathologists, and the pathologists pointed out the main aspects that had to be done. parsing with machine learning.
“This was a very close conversation between the computational group and the pathology group, going back and forth between the rich history of medicine known to pathologists and modern machine learning methods.” Sorger said. “I think it’s an exciting insight into how these computational methods can be used in medicine in the future, where you will tightly connect biologists and physicians with computing, rather than seeing them as substitutes for each other.”
The team chose melanoma and colorectal cancer as a starting point because they are common cancers with unmet medical needs that include large solid tumors and require important treatment decisions. Next, the researchers plan to tackle breast cancer and brain cancer. They also want to train other scientists to use the imaging technologies to build their own cancer maps, paving the way for more atlases to be created.
“A new era in molecular pathology is beginning, and this deep look at a tumor shows us how amazing the results can be,” said Santagata.
Authority, funding, disclosure
Additional authors include Shu Wang, Yu-An Chen, Clarence Yapp, Madison Tyler, and Maulik Nariya of HMS; Shannon Coy of HMS and Brigham and Women’s; and Cody Heiser and Ken Lau of Vanderbilt University School of Medicine.
The research was supported by the National Institutes of Health (U54-CA225088; U2C-CA233280; U2C-CA233262; U2C-CA233291; R01-DK103831; T32-GM007748, P30-CA06516), the Gray Cancer Research Foundation. and the David Liposarcoma Research Initiative.
Sorger is on the board of directors of Glencoe Software and Applied BioMath, the scientific advisory board for RareCyte, NanoString, and Montai Health and is a consultant for Merck. Chen is a consultant for RareCyte.