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Custom NanoString Assay Detects Bowel Cancer Subtypes, May Help Guide Therapies

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NEW YORK (GenomeWeb) – Scientists at London's Institute of Cancer Research (ICR) are developing a gene expression assay that they anticipate may facilitate prospective validation of bowel cancer subtypes in clinical trials.

In a study published in Scientific Reports on Tuesday, the scientists described a 38-gene proof-of-concept test that they have used to detect gene expression patterns associated with each type of bowel cancer

The researchers developed the assay on the nCounter, a US Food and Drug Administration-cleared platform from NanoString Technologies that the ICR scientists have been using to conduct diagnostics research and development work for the past five years.

The platform and assays are highly applicable to clinically available FFPE samples and could be applied in colorectal cancer clinical trials that require low-cost, reproducible, and rapid assays to prospectively validate bowel cancer subtypes, the researchers wrote.

"We wanted a technology platform that speeds turnaround time in selecting patients with specific bowel cancer subtypes," said Anguraj Sadanandam, team leader in systems and precision cancer medicine at the Institute of Cancer Research in London and a study author. "We worked with the manufacturer to bring down the cost of using its platform by 30 percent and then evaluated how it could be useful with archived samples because that is what is used in the clinic."

As a consequence, the cost of testing using the ICR assay is between 40 and 45 percent less than costs associated with using NGS to identify bowel cancer subtypes, and the turnaround time has been reduced to about a week from potentially more than a month, he said.

Such advances are important because the group wanted to demonstrate a more affordable assay that could be used to identify patients and cancer subtypes that could respond to available treatment regimens, Sadanandam said.

Brad Gray, president and CEO of NanoString, said that as the treatment options for cancer expand and target distinct molecular pathways within tumors, it is important to have diagnostic assays that will be able to characterize the intrinsic subtypes of cancer so that those therapies can be directed to the patient population that is most likely to benefit.

"The work of [the ICR group] provides an important tool for the colorectal cancer research community because it demonstrates that the NanoCRCA assay is an accurate and robust solution for transcriptional profiling to identify the consensus molecular subtypes of colorectal tumors," Gray added.

Existing targeted therapies for colorectal cancers, including anti-EGFR antibodies and anti-angiogenic and immunotherapy agents, can extend survival up to 30 months in selected patients. However, identifying patients who will benefit from different standard-of-care treatment regimens remains challenging.

Predictive biomarkers are required to spare patients from unnecessary toxicities, improve outcomes, and increase cost-effectiveness of treatments, the researchers said.

The ICR investigators previously published research in Nature Medicine on a 786-gene signature that they had established leveraging microarray technology to classify colorectal cancers into five subtypes with different prognoses and potential treatment responses.

Subsequently other groups independently identified between three and six molecularly distinct colorectal cancer subtypes based on expression profiles, the researchers noted.

To address ambiguity caused by independent studies, they aggregated the findings to establish consensus molecular subtypes, including inflammatory, enterocyte, goblet-like, stem-like, and a mixed subtypes. They also implemented machine learning to reduce the relevant gene set to 38 in the current study.

The group began the newly published study in 2014, setting out "to validate the robustness of the test and determine if it could be moved closer to the clinic," Sadanandam said, by testing its performance on six sets of archived samples from 355 patients and comparing it with other technologies such as microarrays and sequencing.

When the investigators examined the subtypes identified in a cohort of fresh frozen samples, they found that the enterocyte subtype was the most prevalent, followed by goblet-like, stem-like, and inflammatory.

Importantly, the team demonstrated "through discovery of the five bowel cancer subtypes that we could show not only prognostic differences but also predicted differences for treatment responses," Sadanandam said.

As a prognostic biomarker test it provides information about a patient's overall cancer outcome regardless of therapy, while as a predictive biomarker test it provides information about the effect of a therapeutic intervention.

The results of the current study require further validation in additional clinical cohorts, but a CE marked assay and lab-developed test for use in a CLIA-certified laboratory in the US could be available in about three years, Sadanandam said.

He noted that the group selected the NanoString platform in part because it had been used to develop the FDA-cleared Prosigna Breast Cancer Prognostic Gene Signature Assay, which predicts the risk of recurrence in treated patients.

The platform measures gene expression in the form of discrete counts of barcoded mRNAs, and requires no amplification step. It uses capture and reporter probes to measure gene expression through fluorescence and by counting barcoded mRNAs, Sadanandam said. The method provides the sensitivity of a qPCR assay but can more effectively identify the cancer subtypes because it can analyze the presence of a significantly larger number of targets. Microarrays and sequencing can capture and analyze tens of thousands of genes, but the level of sensitivity using those methods is far lower than that of the NanoString platform, he said.

The next step for the ICR team is to conduct additional validation studies and prospective trials, he said. For that, the firm is looking to collaborate with diagnostic industry partners as well as seek public, government-backed funding.

While the group is first targeting use of the test with cancer subtypes for which therapies currently exist, the method has potential to be extended for use with newly developed therapies as they come online for specific cancer subtypes, he said.

The investigators noted that oncology researchers have made progress with targeting bowel cancer subtypes with therapies. Researchers have reported that the addition of oxaliplatin to fluorouracil-leucovorin in early-stage disease revealed an advantage in disease-free survival in the enterocyte bowel cancer subtype compared to the other subtypes. However, the prospective validation of these findings into routine clinical practice "remains challenging, mainly due to the lack of a fit-for-purpose assay that can classify patient samples into subtypes within a clinically relevant turnaround time and reasonable costs using formalin-fixed paraffin-embedded (FFPE) samples," they wrote in their paper.