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KU-Led Team Begins Clinical Validation of Microfluidic Device for Leukemia MRD Detection

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NEW YORK – A team led by University of Kansas researchers has launched a National Institutes of Health-funded validation study to see if a microfluidic cell-capture and phenotyping assay could be used instead of invasive bone marrow aspirates to identify early cases of hematological cancers. 

Led by Steven Soper, a chemistry and mechanical engineering professor at the University of Kansas, the team aims to identify acute lymphoblastic leukemia (ALL) biomarkers in circulating leukemia cells (CLCs). 

If the validation study is successful, the researchers will use the data to support an application with the US Food and Drug Administration with plans to commercialize a diagnostic assay for multiple types of leukemia through San Diego-based startup Biofluidica, which Soper cofounded.

Soper's team has developed  a microfluidic cartridge consisting of two units. A sinusoidal device separates enriched CLCs from 1 ml of a patient's peripheral blood, while the other unit is an imaging chip that immunophenotypes and performs FISH on the enriched CLCs.

In the first module, B-ALL and T-ALL cells bind to specific antibodies (anti-CD19 and anti-CD7 respectively) that are immobilized on the module's capture surface by a photocleavable linker.

After non-CLCs are washed away, the module photolitically releases the target cells from the capture surface into the second module. There, CLC-specific biomarkers, including terminal deoxynucleotidyl transferase, trap the CLCs in a microtrap, which allows automated cell staining and phenotyping. The team can then perform immunophenotyping and FISH analysis directly on the chip. 

Soper noted that B-ALL patients can have a variety of mutations — including hyper-diploidy, MLL translocations, and iAMP21 chromosomal abnormalities — that clinicians can use to determine the most effective treatment strategies. 

"Because of the sensitivity of the assay, we can detect signs of MRD from patients who have been deemed negative by bone marrow aspirate," Soper said. "Rather than categorizing patients as MRD-negative, we can still continue to look for signs of relapse, leading to better outcomes in the patients." 

Sopher also highlighted that the assay, from sample extraction to completing FISH analysis, only requires about six hours, compared to the three days minimum for bone marrow aspirate samples. 

Previously applying the technology to detect multiple myeloma, Soper's team started using the modules to identify CLCs in AML and ALL pediatric patients last July. He explained that the process for different cell types is very similar, and that users only need to switch out specific antibodies on the chip depending on the type of cancer cell.  

Working with Children's Mercy Hospital in Kansas City, Kansas in 2017, Soper's team initially performed a pilot study that involved collecting peripheral blood samples from 20 pediatric B-ALL patients and analyzing them for cases of relapse. Funded by the Midwest Cancer Alliance, the study applied the two-module microfluidic platform, which Soper originally built for solid cancers at Louisiana State University.

"We [followed] MRD in the patients over an 85-day period during their chemotherapy," Soper said. "We were able to stratify patients into low and high risk in terms of the number of CLCs they possessed, which agreed with clinical observations." 

Because of the pilot trial's success, Soper's team applied for NIH funding earlier this year to see if the platform could indicate the onset of MRD relapse in a larger cohort of ALL patients, and received a National Cancer Institute grant in September. The researchers anticipate receiving nearly $725,000 over the three years of the study and have begun enrolling pediatric ALL patients who have undergone chemotherapy at Children's Mercy Hospital. The group will test patients as they undergo induction and consolidation therapy, monitoring them over a period of about three months. 

While the group initially collected B-type CLCs in the pilot study, Soper expects to also collect samples containing T-type CLCs in the new cohort of 50 ALL patients. The researchers will perform CLC FISH analysis on the chip for high-risk patients to gather information on chromosomal abnormalities, which they believe will provide information for precision-based treatment. 

"Cytogenic abnormalities detected at diagnosis or generated during chemotherapy constitute important prognostic factors, [since] the detection of specific chromosome aberrations … are used to assign B-ALL patients to specific targeted therapies," Soper said. "Most of the … FISH-based assays are predicated on the use of bone marrow, which is enriched in leukemia cell content."

If the validation study is successful, Soper's team will use the results to initiate discussions with the FDA as part of a 510(k) application for the blood-based CLC detection platform. Biofluidica is currently developing the technology to detect and isolate mutated cells in patients with clonal plasma cell disorders, but the startup has also partnered with Soper's team to use the technology on the pediatric ALL study. 

Soper, who is chief technology officer at Biofluidica, and his group will partner with the company to potentially commercialize the microfluidic technology for early detection and prognosis of hematological cancers. He believes that the two-module platform will translate well to clinical hematology because of its use of blood samples rather than bone marrow aspirates. In addition, he noted that the platform also involves minimal hands-on time; a user only needs to load the blood sample onto the chip and wait for the process to finish before performing FISH or other downstream analyses. 

Soper believes that if clinicians could identify disease relapse and chromosomal defects from peripheral blood samples, "painful bone marrow biopsies could be avoided, and [doctors] could obtain information in near real-time and potentially implement changes in treatment to affect better outcomes for B-ALL patients."