The scenario of cancer during pregnancy is constantly growing due to the increasing incidence of cancer in young populations. Safety data on long-term effects of in utero exposure are limited to early adolescence and focus on neurocognitive and cardiac function; however, there is a lack of data regarding other organs. The clinical literature indicates that chemotherapy minimally passes through the placenta, yet evidence relies on measuring chemotherapy levels in amniotic fluid extracted after amniocentesis of pregnant cancer patients after chemotherapy treatment, though not immediately. We have established a preclinical model of pregnant mice to follow the short- and long-term effects of in utero exposure to chemotherapy and aimed to assess chemotherapy placental pass and potential induced methylation in the offspring as a short-term sequel indicating epigenetic changes.

Pregnant ICR mice were treated once with DXR (10 mg/kg) or saline on pregnancy day E12.5, which is equivalent to the 2nd trimester in women. DXR concentration in each individual amniotic fluid was evaluated via amniocentesis of every amniotic sac at 10 minutes-48 hours post administration to the mother. In another cohort of treated mothers, following delivery, blood will be drawn at D2 from pups and mothers (DXR-treated and controls), and Reduced Representation Bisulfite (RRBS) analysis will be performed to examine the methylation changes that take place following exposure to DXR in utero.

Doxorubicin was measurable in a high proportion of fetuses as early as 10m post-injection, reaching 92% exposure by 30m. It remained measurable at 24 h but diminished by 48 h post maternal treatment. RRBS revealed a high-resolution, genome-wide DNA methylation maps for in-utero exposed cohort and controls and indicated a significantly differential methylation pattern between in utero exposed mice and controls

Our study is the first to systematically evaluate chemotherapy placental pass and early fetal impact. Our results indicate that DXR passes through the placenta and reaches the amniotic fluid and fetal compartment and induces differential methylation patterns in exposed embryos that may reflect early epigenetic changes.