With the PacBio Beyond the Bench video series now live, we’re feeling the enthusiasm surrounding a personal look at scientists’ lives and the rituals that anchor their work. This series offers a chance to step beyond the data and hear directly about the habits and motivations that drive their research. Here we take a closer look at the science of one of the featured researchers and the ideas guiding his approach to cancer biology.
Understanding immune cell behavior in cancer with RNA sequencing
Dr. Santosha Vardhana is a physician scientist at Memorial Sloan Kettering whose lab focuses on how immune cells behave in cancer. At the center of his work is a fundamental puzzle: Immune cells often recognize tumors and even infiltrate them, yet they are often unable to mount an effective response. Understanding why that happens requires looking closely at how immune cells adapt, change, and sometimes malfunction in response to cancer.
A key theme running through Dr. Vardhana’s research is cellular heterogeneity. Immune cells, particularly T cells, do not fall into a small number of fixed categories. Instead, they exist along a continuum of states as they respond to metabolic constraints and cancer signals of the tumor microenvironment, which makes RNA sequencing essential for capturing how molecular programs differ from cell to cell. Capturing that complexity has shaped how his lab approaches questions about immune dysfunction and cancer progression.
How single-cell RNA sequencing reveals immune cell diversity
This is where single-cell transcriptomics enters the picture. In Santosha’s interview, he describes how early in his work it became clear that looking at transcripts at the single-cell level is critical for his research. He uses a simple analogy to explain the difference: Looking at bulk RNA data is like drinking a smoothie. You can identify which fruits are in the smoothie, but you are unable to gain information about each fruit individually, for example, if the blueberries are fresh or frozen. Single-cell approaches, by contrast, are more like examining a fruit salad, where each component can be inspected on its own. That distinction matters when subtle changes can have outsized effects on cell behavior.
HiFi long-read RNA sequencing for cancer transcriptomics
This is where HiFi RNA sequencing comes into play. By sequencing full-length RNA transcripts, researchers are able to preserve complete transcript structures, making it possible to directly observe RNA isoforms without assembly. In cancer research, this allows scientists to examine gene expression, alternative splicing, fusion transcripts, and expressed mutations together, offering a more complete picture of RNA regulation.
RNA isoforms are key for understanding cancer biology. Alternative splicing can give rise to different RNA isoforms that alter protein function, disrupt regulatory networks, or generate oncogenic fusion transcripts, sometimes without any corresponding change in the underlying DNA. When transcript structure is not fully resolved, these events can be misinterpreted or missed entirely. Full-length RNA sequencing provides clarity into how transcripts differ between healthy and diseased cells and how those differences contribute to cancer development and progression.
HiFi long-read RNA sequencing has been applied across a wide range of cancer studies, from characterizing isoforms associated with survival outcomes in gastric cancer to identifying novel isoforms that might serve as vaccine candidates in colorectal cancer. Researchers have used it to uncover previously unannotated isoforms, characterize complex fusion transcripts, and define cellular states based on RNA structure rather than expression levels alone. In single-cell studies, pairing cell-level resolution with long-read sequencing has revealed layers of regulation that were difficult to access with short-read technologies.
Advancing cancer research through long-read RNA sequencing
These insights reflect a broader shift in cancer research toward understanding disease as an interplay of multiple molecular processes. DNA mutations, RNA processing, metabolism, and immune function are tightly linked. Technologies that can capture these layers together allow researchers to ask more nuanced questions and uncover mechanisms that might otherwise remain hidden.
With Beyond the Bench now released and shining a light on the people and ideas driving modern biology, Santosha’s work offers a compelling example of how new sequencing approaches can open up long standing questions in cancer biology. By looking closely at immune cell diversity and RNA isoforms, his research sheds light on why immune responses falter and how we might better understand immune responses in the ongoing effort to improve cancer outcomes.