Optimization of Cervical Collections in Pregnancy

The study will be non-interventional and designed for prospective sample collection and concurrent analysis. The primary endpoint will be a determination of the number of extravillous trophoblast (EVT) cells (placental cells that migrate into the reproductive tract) obtained by each device and procedure. The number of EVT cells, adjusted for purity (expression of a fetal marker, e.g., cytokeratin 7) directly determines the amount of fetal DNA that will be available for prenatal genetic testing. Secondary endpoints will include the total number of cells in the initial cervical samples and the amount of fetal DNA, RNA and/or protein isolated from the EVT cells. Where fetal DNA is obtained, we will sequence the fetal and maternal DNA for comparison of single nucleotide variant (SNV) and short tandem repeat (STR) loci to assess the purity of the fetal DNA (fetal fraction) or perform whole genome sequencing. Cell-free biomolecules present in cervical fluid will also be assessed for proteins or RNAs associated with adverse pregnancy outcomes, or cell-free fetal DNA. These parameters will be compared between the various collection methods to describe an optimal collection protocol for the purpose of analyzing fetal cells and their components.

Cohort: We will follow a group of pregnant individuals over time, collecting cervical secretions at various stages throughout pregnancy and then comparing these data to the pregnancy outcomes.

Control: We will collect cervical fluid from non-pregnant, non-menstruating women of reproductive age for comparison to the cohort participant samples. It is expected that cells or biomolecules produced by the placenta will be absent and that any signals detected would define background levels.

Descriptive: The study aims to observe and describe the properties of cervical secretions throughout pregnancy and their association with pregnancy outcomes or fetal genotype.

Establishing a registry/database: The researchers need to create a database to organize and analyze the information collected from participants, such as the properties of their cervical secretions and the outcomes of their pregnancies.

Pregnant women (N=2000) will be enrolled in this study because it is an investigation of a method for prenatal testing and the endpoint is detection of EVT cells or their products that are of fetal origin, specifically from the placenta. The age range of 18 to 45 years was chosen because this is the principal range for reproductive activity in women. Nonpregnant women (N=600) will also be enrolled to control for EVT cells in pregnant samples and determine the respective number of cells obtained in the study protocol when no EVT cells are present.

There will be no enrollment restriction based on racial or ethnic origins. The aim is to include sufficient diversity in enrollment to ensure that the benefits and burdens of research participation are distributed in an equitable manner. It is also desirable that the findings are reproducible across all racial and ethnic groups.

If the sample is taken at the time of a Pap smear, the healthcare professional will follow recommended procedures. The initial excess mucus from the canal and ectocervix, will be deposited in the collection containers provided by the PI rather than being discarded. If sample collection is conducted at a different time than the pregnancy cervical cytology, a cervical brush in addition may be obtained. Sample collection kits using devices used for Pap smears (including during pregnancy) and other methods to be investigated will be provided to the clinics. These kits will contain all devices for sample collection and shipment back to the PI. Kits will also contain a participant data sheet and instructions for sample collection. All kit materials will be coded to identify each participant. The key to the codes will be retained by each clinical site collaborator in a locked cabinet in their office so that personnel working with the PI will only have the de-identified code for each participant. To link laboratory data to clinical data, the clinical site collaborator will conduct the analysis using the key to the codes. No information from the analysis of the cervical fluid will be returned to the participant or their doctor since the study is designed to evaluate the procedures for sample procurement, not the test results.