NEW YORK – As part of its push to develop a range of liquid biopsy assays for cancer detection, Lucence Diagnostics has completed two studies that demonstrate the ability of its amplicon-based next-generation sequencing platform, AmpliMark, to detect structural rearrangements (SR) in lung cancer as well as common cancer mutations in breast patients.
Lucence believes the studies support the clinical utility of using AmpliMark for detecting actionable mutations in the different cancer types, allowing for faster and more accurate testing than current approaches.
In the near term, however, the Singapore-based firm will focus on launching a laboratory-developed test version of its LiquidHallmark late-stage lung cancer treatment selection assay out of its lab in Palo Alto, California.
Lucence Dx CEO and Founder Min-Han Tan explained that the AmpliMark platform combines molecular barcoding and its SunTzu.AI clinical analytics engine to target clinically relevant genomic alterations in blood samples.
AmpliMark uses two different types of barcoded primers: one binds to fragments comprising the defined target region, while another "separation" primer binds to fragments containing a partly defined region in a cancer patient's cfDNA. The primers that anneal to the defined region are separated from the primers that bind to the partly defined region.
"This is important because management of the well-defined region often pinpoints actionable SNPs or point mutations, while the other helps [with identifying variants] such as fusions, which may not necessarily [have well-defined] partners," Tan explained. "We want to make sure that for the well-defined region's [product], we have a reverse primer, and for the [partly-defined] region's product, we have a double-stranded oligo that's ligated to the end of the fragment."
AmpliMark then amplifies both barcoded products and downstream NGS.
While noting that the technology is platform-agnostic, Tan said that the firm has been using Illumina's NextSeq 550 instrument. He highlighted that the overall process takes about eight days to produce actionable results for clinicians.
At the virtual annual meeting of the 2020 American Society of Clinical Oncology this week, Lucence presented two study abstracts detailing the use of AmpliMark in different cancer cohorts.
In the first study, the firm collaborated with researchers at the National Cancer Center Singapore and applied AmpliMark to identify PD-L1 structural rearrangements from tumor tissue samples in NK/T-cell lymphoma patients. The team validated the platform using mixtures of fragmented genomic DNA from an ALK SR-positive cell line, commercial standards containing RET and ROS1 SR, and synthetic PD-L1 SR gene constructs. Here the researchers found that they could detect SR down to an allele frequency limit of detection of 0.5 percent, with a clinical sensitivity of 90 percent and specificity of 100 percent.
The team then assessed the tool's clinical performance in cfDNA samples from 374 lung cancer cases and tumor tissue DNA samples from natural killer (NK) T-cell lymphoma patients.
In the lung cancer samples, the group detected actionable SR for ALK, RET, and ROS1 in nine samples, with an overall detection rate of 3 out of 186 samples when limited to pre-treatment lung cancer cases. In the NK/T-cell lymphoma cohort, 9 of 29 samples were positive for PD-L1 SR using AmpliMark, which was confirmed by whole-genome sequencing, targeted sequencing, or Sanger sequencing.
"The limit of detection of fusions is one of the biggest challenges in liquid biopsy," Tan said. "While ALK, ROS1, and RET were classic key rearrangements we needed to detect, we also detected PD-L1, a rearrangement observed in several cancers, which highlights the validity of our tech."
The researchers noted that they have begun studies to further evaluate AmpliMark's performance and utility across a larger number of clinical samples to detect SR in the study's list of genes, as well as other cancer-linked SR involving NTRK1/2/3, FGFR2/3, and TMPRSS2.
In the second study, researchers from the University of Hong Kong tested AmpliMark's ability to detect gene mutations in hormone receptor-positive, HER2-negative breast cancers.
Collecting plasma cfDNA from a cohort of 113 breast cancer patients (83 metastatic), the team analyzed genetic alterations in the cfDNA using AmpliMark. The team then correlated the presence of PIK3CA and other breast cancer mutations to molecular subtypes and treatment histories.
Identifying at least one mutation in 80 cancer patients, the researchers saw that mutations appeared more frequently in metastatic cancer cases compared to non-metastatic cases. Across all patients, the team also saw that the most frequent mutations occurred in PIK3CA, TP53, ESR1, GATA3, and ERRB2.
Noting that PIK3CA mutations appeared the most often in HR+, HER2- patients, Tan said that several PIK3CA mutations occurred in about 18 percent of the PIK3CA mutant cases. The group identified a link between PIK3CA mutations and prior treatment with CDK4/6 inhibitors or mTOR inhibitors, with 58 percent of PIK3CA-mutant cases having received the treatment previously compared to only 20 percent of PIK3CA wild types.
Tan emphasized that the team successfully identified similar PIK3CA mutation frequencies between AmpliMark and prior tumor tissue testing in breast cancer. In addition, the group successfully observed similar frequencies of other driver mutations in external tissue studies, suggesting high sensitivity as the primary reason for performance.
"Together with the lung cancer report, we show promising data in both breast and some interesting fusions that are common in lung and lymphoma," Tan said. "We believe the use of AmpliMark for lung cancer will allow for quicker and more accurate testing than current approaches."
While Lucence will continue exploring avenues in breast and blood-based cancers, Tan declined to discuss specific development timelines for commercial assays based on the new applications.
However, Tan noted that the Lucence will focus on commercially launching an LDT version of its LiquidHallmark late-stage lung cancer treatment selection assay in Q3 2020.
Emphasizing that the lung cancer assay contains all of the US Food and Drug Administration-approved lung cancer genetic biomarkers, Tan said the firm plans to offer the LDT from its lab in Palo Alto. The firm expects to complete construction of the lab this summer, followed by an application for CLIA-certification and CAP-accreditation.
Lucence is currently working out its pricing approach for the assay, as negotiations with private and public payors are ongoing, but Tan noted that the firm is committed to having the assay being affordable to lung cancer patients.