NEW YORK – A team of researchers in the US and Brazil is developing a noninvasive approach to assess risk for preterm birth by measuring levels of D-lactic acid and TIMP-1 in vaginal secretions.
Their work, published recently in mBio, represents a first step toward development of an ELISA-based point-of-care diagnostic test that may be used in the future as an alternative to current invasive testing, said Larry Forney, a principal investigator in the study and professor of biological sciences at the University of Idaho in Moscow, Idaho.
Using the new approach, women at risk of preterm birth in low-resource settings who don't have access to current technology could be identified and receive appropriate early intervention and treatment for the first time, Forney said.
"In the study, we developed an approach that is an alternative to transvaginal ultrasound, an invasive procedure used to directly measure cervical length, and also provides an alternative to labor-intensive methods to identify bacterial composition in the vagina," Forney said.
Such an approach is still years away from potentially being commercialized, but it may be possible following integration of the newly discovered biomarkers into an affordable point-of-care assay, the researchers said.
Clinicians use transvaginal ultrasound to measure cervical length because women with shortened cervixes can have ascending infections from the vagina into the uterus, and those infections can elicit inflammation and trigger contractions and preterm births.
In countries where people have access to medical technology, most pregnant women undergo a transvaginal ultrasound at 18- to 24-weeks' gestation to determine cervical length, and a woman with a short cervix is often treated with progesterone to reduce the likelihood of a premature delivery. In low-resource countries, the equipment and expertise to perform a transvaginal ultrasound or characterize the composition of vaginal bacteria are usually unavailable.
In the mBio study, a prospective trial funded by the Bill and Melinda Gates Foundation, the researchers set out to identify biomarkers that might be used to predict bacteria that dominate the vaginal microbiome and indicate the presence of a shortened cervix.
The study participants were mid-trimester pregnant women who were undergoing a routine vaginal ultrasound to assess cervical length at an obstetrical outpatient clinic in Sao Paulo. The patient mix comprised women at low risk for a preterm birth and those with an identified characteristic that placed them at elevated risk, including short cervical length, history of preterm birth or spontaneous miscarriage, vaginal bleeding in the first trimester, or obesity.
Forney along with Steven Witkin, a professor emeritus of infection and immunology in obstetrics and gynecology at Weill Cornell Medicine in New York, and Antonio Moron, a professor in the department of obstetrics at the Federal University of Sao Paulo, were the principal investigators of the study.
"We amplified the 16S ribosomal RNA gene from the samples and sequenced the gene using high-throughput sequencing," Forney said, adding that based on differences in gene sequences, they were able to classify bacteria in the samples and quantify their abundance.
By analyzing vaginal fluid samples from 340 pregnant women, they found that concentrations of TIMP-1, short for tissue inhibitor of matrix metalloproteinases-1, and D-lactic acid, age, and race were correlated with cervical length, a predictor of preterm birth risk. Additionally, levels of TIMP-1 and D-lactic acid was predictive of which bacterium was dominant in the vagina, Witkin noted.
There is a clear unmet need for an effective diagnostic test for this indication, Kelle Moley, chief scientific officer of March of Dimes, said in an interview, adding that recent concerns about the effectiveness of preterm birth therapies have amplified health hurdles for patients at risk of preterm birth.
She noted that a US Food and Drug Administration committee recently met to better understand and interpret a confirmatory clinical trial for AMAG Pharmaceuticals' Makena hydroxyprogesterone caproate injection, a treatment that had received accelerated approval for use to reduce preterm birth in pregnant women who have had a prior spontaneous preterm birth. Nine out of sixteen advisory committee members voted to recommend that the FDA pursue withdrawal of approval for Makena.
"Now we are not just racing to find a test, we are also racing to find a treatment for this condition, and that's why deaths and the prematurity rate keeps going up," Moley said.
Arlington, Virginia-based, March of Dimes, a nonprofit organization that works to improve the health of mothers and babies, provides funding to organizations developing preterm birth tests and therapies. Its 2019 March of Dimes Report Card on maternal and infant health released Monday noted that each year in the US alone about 1 in 10 babies is born prematurely.
"We have a huge societal cost of preterm birth in this country and at this point we have no effective diagnostic tools for the indication," Moley said. "The investigators in the mBio study reported that the use of TIMP-1 and D-lactic acid in vaginal secretions might be a straightforward way to assess the risk for preterm birth, but they will obviously have to validate that in a larger setting."
Forney and his colleagues aim to develop a point-of-care diagnostic test that people can use in a clinic and that doesn't require expensive instrumentation. For that, the researchers are looking to collaborate with a diagnostic company interested in integrating the biomarkers into a point-of-care platform.
"This study is a promising initial step in identifying biomarkers that correlate with a short cervix, a risk factor in preterm birth," said Adam Woolley, a professor in the department of chemistry and biochemistry at Brigham Young University, Provo Utah.
Woolley, who is not associated with the current study, is developing an integrated microfluidic test that uses a multistep process to capture and identify peptides and may one day detect the risk of pregnant women having preterm births.
He said with reference to the mBio study that "this approach could be used to screen for women at risk for a preterm birth, allowing treatment to potentially avert associated complications and high infant mortality," but only "if larger patient cohort studies corroborate this initial finding, and if an inexpensive and simple diagnostic test can be designed for the identified biomarkers."
An accurate evaluation of how well the current biomarkers predict preterm birth will indeed require more expansive studies and may include women in different stages of pregnancy as well as additional populations of women, Forney said.
A future larger study may also include use of samples from women in the first trimester so that if the findings are similar to the current study, monitoring and possible treatment could begin earlier in gestation, he added.
Moley noted that a number of companies have completed studies similar to that published in mBio. The studies use a variety of technologies to address the unmet need associated with diagnosing the risk of preterm birth. Their approaches include analyzing vaginal secretion metabolites and vaginal bacterial signatures that predict preterm birth as well as developing proteomic and cell-free RNA tests.
Many of these companies are at a similar stage of test development to the mBio investigators and are "taking a next step to do larger clinical studies looking to further validate their tests," she said, adding that March of Dimes has sponsored clinical studies for some tests.
Among companies developing tests for preterm birth risk, NX Prenatal has entered a collaboration with Indiana University School of Medicine to further validate its early pregnancy biomarker panels that predict spontaneous preterm birth and to continue developing its biomarker panels for preeclampsia.
Thermo Fisher Scientific and Milu Labs recently announced that they plan to collaborate to develop clinical diagnostic technologies for women's health in China. Under a memorandum of understanding, the companies plan to explore opportunities to use Thermo Fisher's liquid chromatography-mass spectrometry instrumentation to develop tools to determine the risk for adverse pregnancy outcomes.