NEW YORK –Diagnostic developer Entopsis is taking its first steps into the cancer space by adapting its multi-analyte detection platform for prostate cancer screening and partnering with national laboratory Genetics Institute of America (GIA).
The Miami -based firm's platform, called OpsisDx, uses a glass slide with 70 hydrogel structures that bind to molecules in a patient's liquid sample and create a colorimetric signature when heated that reflect biometric changes in a patient's body.
Obdulio Piloto, CEO and cofounder of Entopsis, began envisioning the concept with cofounder Ian Cheong while working on their PhDs at Johns Hopkins Medical School. Aware that several colleagues were aiming to link specific RNA, DNA, and protein alterations in liquid samples to cancers for early detection, the team chose to instead create a tool that could diagnose any disorder using the same approach in liquid samples.
"The [other] approach is likely to only offer insights to select cancers, where alterations are accessible in the early stages of the diseases," Piloto said. "This approach is laborious and costly, requiring the identification of alterations that are associated with each cancer."
After receiving an initial amont of undisclosed funding from Peter Thiel's Breakout Labs, Piloto's team launched Entopsis in 2012 to develop a platform that would capture disease-specific immune, metabolic, and proteomic changes in biofluids, starting with urine, as a means of "gaining insight into a person's health and disease state."Â
"Instead of looking for specific biomarkers, like DNA and RNA, we are creating a molecular profile of a person using their liquid sample," Piloto said. "We're trying to capture and sample a representative cross-section, like a snapshot, of molecules in that person's biofluid."
Piloto explained that OpsisDx relies on a method called "nanoscale unbiased textured capture" (NuTec), in which a 3 square-inch glass slide containing 70 hydrogel surfaces act as little sponges when submerged in a liquid sample. Each hydrogel contains holes with different shapes and chemistries that bind to molecules such as proteins and protein complexes.
"Since there's a limited number of binding pockets, there's competition between the different molecules in the liquid sample. Different molecules could also bind to a spot because there are different affinities," Piloto explained. "Therefore, not only are we capturing molecules, but we are also looking at changes in concentrations between simple molecules at each spot."
In order to visualize the bound analytes, the OpsisDx platform heats the NuTec slide, creating a colorimetric profile for analysis. Piloto noted that each surface has a different number of molecules in it that, when heated, induces reactions to bound analytes, creating a variety of colors.
"Each spot is effectively a mini signature that looks slightly different with different analytes bound to it, and each NuTec slide is a signature composed of 70 visible colorimetric signatures," Piloto said. "The readout therefore gives an overall molecular profile of the patient's liquid sample."
In order to develop a disease-specific assay, however, Piloto's team begins by using the platform to analyze clinical samples from patients with a confirmed condition, such as urine samples from prostate cancer patients previously confirmed by tissue biopsy.
The firm uses a cloud-based machine learning algorithm to identify visual differences and create molecular profiles of positive and negative patient cohorts, eventually training the algorithm to create what Piloto describes as "a unique fingerprint" for the disease.
Piloto explained that a disease-specific assay running on the OpsisDx platform would need 4 to 5 ml of a patient's urine sample, the molecular profile of which is compared to Entopsis' growing disease database. Piloto said the database currently contains clinical data from over 1,500 unique patient donors, with cancers including lung, colorectal, prostate, breast, and liver, as well as hepatitis B viral infections.
Piloto said the algorithm provides a score as to the likelihood of a patient having a condition, based on the several training sets the firm has built over time. He also noted that the platform can theoretically produce results from a single sample in under an hour.
Commercial aspirations
By signing a commercialization agreement with GIA, a CLIA-certified, CAP-accredited medical laboratory in Delray, Florida, Piloto hopes to develop a test that will help urologists and family practice clinicians determine if their patients do not require a risky and invasive tissue biopsy for prostate cancer detection.
As part of the commercialization agreement, GIA has begun an IRB-approved prospective validation study on the OpsisDx platform in male patients scheduled for prostate biopsies. Rob Cardwell, chief strategy officer of GIA, explained that the lab is collecting urine samples from patients suspected of having prostate cancer before they undergo tissue biopsy.
Cardwell said that GIA has already begun receiving samples from a mix of 10 different academic and private urological practices from New England, Florida, New Mexico, and Arkansas. Aiming to gather a wide demographic mix, GIA anticipates validating the assay on about 300 samples.
Cardwell said that the envisioned assay will help clinicians and patients avoid unnecessary biopsies and potential complications, thereby cutting healthcare costs and allowing doctors to better triage their patients.Â
In order to ensure that the clinical database is comprehensive and can distinguish between different conditions, Piloto explained that his team routinely runs numerous internal validation studies to assess the uniqueness of each disease training set. However, he noted that the overlap between training sets — such as liver cancer and colorectal cancer — is minimal and that they show up on the platform as distinct diseases.
In addition, Entopsis also ensures that each disease training set is created to address a specific issue. The positive and negative donors used to create the training set, therefore, are carefully selected for the problem. For example, he explained that the prostate cancer assay that GIA is developing is specifically geared to find out whether a tumor will be found if the biopsy is actually performed.
"The training set for this test only encompasses men who are going to be biopsied, and the pathology report will let us know which men actually had prostate tumors," Piloto said. "NuTec profiles from these men make up the 'positive' arm of the training set, while those with no tumors make up the 'negative' arm, [allowing] our algorithm to learn the difference between these two groups of men."
However, Piloto emphasized that the training set will not be used to screen men in the general population, as that would require a training set that "reflects the ultimate goal of the test."
Once the team completes the validation study, GIA plans to commercialize the LDT out of its laboratory. Cardwell believes that urologists could potentially request the screening test after identifying certain risk factors such as an elevated PSA in their patients.
"If a male patient comes in, has an elevated PSA, and a doctor finds something odd during a digital rectal examination, they'd then order this [screening] test as the next step," Cardwell explained. "If positive, the doctor would schedule a time for tissue biopsy and imaging. But if the result is negative, they'd avoid the unnecessary surgery and its costs."
According to Holly Magliochetti, CEO of GIA, the firm has begun discussions with selected healthcare payors to "educate" them about the envisioned LDT. However, she noted that GIA could not disclose pricing specifics other than that it "will be more cost-efficient than a tissue biopsy."
Branching out
While prostate cancer is Entopsis' initial focus area, Piloto said the firm is also examining the platform's potential for other cancers and conditions, such as autoimmune disorders and infectious disease.
The firm has submitted a peer-reviewed manuscript for publication later this year, which Piloto claims  will demonstrate the platform's ability to sensitively detect five cancers: lung, prostate, colorectal, breast, and liver cancer, as well as hepatitis B in liquid samples such as urine. Piloto highlighted that in a cohort of about 1,000 samples, the team successfully identified a subset of patients who were hepatis B positive and liver cancer negative, and were able to use the platform to observe the transition of hepatitis B to liver cancer in certain patients.
Piloto noted that Entopsis has filed for two "large, comprehensive patents," with the US Patent and Trademark Office and has been issued more than 10 global patents for the platform.
Acknowledging that Entopsis has a relatively small team with limited financial resources, Piloto noted that the firm is partnering with larger labs like GIA to validate and later commercialize tests on the OpsisDx platform. Â
Entopsis is also collaborating with three clinical groups in Beijing, China — the Chinese Academy of Medical Sciences, Peking Union Medical College, and the Second Affiliated Hospital of Chongqing Medical University — to screen for lung, colorectal, breast, and liver cancers, as well as hepatitis B viral infections. Piloto noted that Entopsis has encountered several technical and financial hurdles while developing the OpsisDx platform. One major barrier the team has dealt with is collecting appropriate clinical samples at reasonable prices in order to establish training sets.
Piloto also acknowledged that his team has struggled explaining to investors that OpsisDx does not rely on a specific biomarker, but instead captures a sample's molecular profile and identifies alterations in its biochemical composition.
"Our hypothesis is that the body would identify diseases before there's any circulating tumor DNA or circulating tumor cells in the bloodstream," Piloto said. "It's not one protein, one set of DNA, or tumor we're looking at, but instead [we] are trying to observe whether the changes are metabolic or immune [in nature]."
Piloto believes that an LDT using OpsisDx would be able to identify prostate cancer at an earlier timepoint, deliver results faster, and potentially be less expensive than ctDNA- or CTC-based tests.
Piloto also suggested that Entopsis could simultaneously provide other health metrics besides the potential presence of cancer.
"For example, we could find out if it was a slow-growing versus an aggressive prostate cancer, which could help with treatment decisions, or if a patient would benefit from a given treatment," Piloto said. "If a database has been validated, created, and tested for prostate, liver, colorectal, or whatnot, then a patient with an unknown profile could also be evaluated for the different cancers, and we could see how confident the AI is with that result."
Having raised about $2 million in seed funding since launching in 2012, Entopsis is now aiming to raise an additional $5 million in an ongoing financing round, which it will use to validate OpsisDx for additional diseases and establish new commercialization partners.