Placental nanoparticle-mediated IGF1 gene therapy corrects fetal growth restriction in a guinea pig model

Abstract

Fetal growth restriction (FGR) caused by placental insufficiency is a major contributor to neonatal morbidity and mortality. There is currently no in utero treatment for placental insufficiency or FGR. The placenta serves as the vital communication, supply, exchange, and defense organ for the developing fetus and offers an excellent opportunity for therapeutic interventions. Here we show efficacy of repeated treatments of trophoblast-specific human insulin-like 1 growth factor (IGF1) gene therapy delivered in a non-viral, polymer nanoparticle to the placenta for the treatment of FGR. Using a guinea pig maternal nutrient restriction model (70% food intake) of FGR, nanoparticle-mediated IGF1 treatment was delivered to the placenta via ultrasound guidance across the second half of pregnancy, after establishment of FGR. This treatment resulted in correction of fetal weight in MNR + IGF1 animals compared to sham treated controls on an ad libitum diet, increased fetal blood glucose and decreased fetal blood cortisol levels compared to sham treated MNR, and showed no negative maternal side-effects. Overall, we show a therapy capable of positively impacting the entire pregnancy environment: maternal, placental, and fetal. This combined with our previous studies using this therapy at mid pregnancy in the guinea pig and in two different mouse model and three different human in vitro/ex vivo models, demonstrate the plausibility of this therapy for future human translation. Our overall goal is to improve health outcomes of neonates and decrease numerous morbidities associated with the developmental origins of disease.

Fig. 1. Effects of maternal nutrient restriction (MNR) and repeated nanoparticle-mediated IGF1 delivery (MNR + IGF1) on plasmid-specific human IGF1 (hIGF1) and endogenous guinea pig Igf1 (gpIgf1).

A hIGF1 mRNA was present within directly injected and indirectly exposed placentas from dams treated with the hIGF1 nanoparticle, although indirectly exposed placentas had less. B Endogenous gpIgf1 was lower in the sham treated MNR placentas of male fetuses compared to sham treated Control; gpIgf1 levels in the MNR + IGF1 groups were comparable to Control. C In placentas of female fetuses, there was no difference in endogenous gpIgf1 between Control, MNR or MNR + IGF1. Control (+ sham treatment): n = 6 Dams (8 female and 11 male fetuses), MNR (+ sham treatment): n = 6 Dams (5 female and 11 male fetuses), MNR + IGF1: n = 5 Dams (6 female and 10 male fetuses). Data are estimated marginal means ± 95% confidence interval. P values calculated using generalized estimating equations with Bonferroni post hoc analysis. *P ≤ 0.05; **P ≤ 0.01.

Fig. 2. Effect of maternal nutrient restriction (MNR) diet, and repeated IGF1 delivery on placenta IGF1 signaling genes.

A Insulin like growth factor 2 (Igf2) was unaltered among all male placental groups. B Females in the MNR and MNR + IGF1 groups had increased Igf2 placental expression of compared to controls. C Insulin like growth factor 1 Receptor (Igf1R) levels were unaltered between placentas of male fetuses. D Igf1R expression was increased in MNR + IGF1 indirectly exposed female placentas compared to controls. E Insulin like growth factor binding partner 3 (IgfBP3) was unaltered between placentas of male fetuses. F IgfBP3 expression was increased in MNR + IGF1 indirectly exposed female placentas compared to control and MNR. Control (+ sham treatment): n = 6 dams (8 female and 11 male fetuses), MNR (+ sham treatment): n = 6 dams (5 female and 11 male fetuses), MNR + IGF1: n = 5 dams (6 female and 10 male fetuses). Data are estimated marginal means ± 95% confidence interval. P values calculated using generalized estimating equations with Bonferroni post hoc analysis. *P ≤ 0.05; **P ≤ 0.01, ***P ≤ 0.001.

Fig. 3. Effect of maternal nutrient restriction (MNR) diet, and repeated IGF1 delivery on fetal and placental weight.

A and B MNR fetuses weighed significantly less than controls in both sexes. MNR + IGF1 (direct injection and indirect exposed) fetuses showed comparable weights to controls in males and females with full correction from MNR with direct IGF1 treatment in males. C Delivered placenta weight showed no changes among any groups in male fetuses. D Delivered placenta weight decreased in MNR + IGF1 compared to controls in females. E Delivered placental efficiency showed no changes among males in any group. F Delivered placental efficiency decreased with MNR in females compared to controls but was corrected in the MNR + IGF1 group. Control (+ sham treatment): n = 6 (8 female and 11 male), MNR (+ sham treatment): n = 6 (5 female and 11 male), MNR + IGF1: n = 5 (6 female and 10 male). Data are estimated marginal means ± 95% confidence interval. P values calculated using generalized estimating equations with Bonferroni post hoc analysis. *P ≤ 0.05; **P ≤ 0.01, ***P ≤ 0.001.

Fig. 4. Effect of maternal nutrient restriction (MNR) diet, and repeated IGF1 delivery on fetal blood glucose, lactate, and cortisol.

A Male MNR fetuses had lower blood glucose levels compared to controls. MNR + IGF1 direct injection and indirect exposure increased blood glucose back to control levels. B There were no changes in female fetal blood glucose levels. C Blood lactate levels were unaltered in male fetuses. D Female fetuses had increased blood lactate with MNR and MNR + IGF1. E, F There were increased blood cortisol levels with MNR in both males and females. MNR + IGF1 male and female fetuses had blood cortisol levels comparable to control. Control (+ sham treatment): n = 6 dams (8 female and 11 male fetuses), MNR (+ sham treatment): n = 6 dams (5 female and 11 male fetuses), MNR + IGF1: n = 5 dams (6 female and 10 male fetuses). Data are estimated marginal means ± 95% confidence interval. P values calculated using generalized estimating equations with Bonferroni post hoc analysis. *P ≤ 0.05; **P ≤ 0.01, ***P ≤ 0.001.

Fig. 5. Effect of maternal nutrient restriction (MNR) diet, and repeated nanoparticle-mediated IGF1 delivery on maternal blood biochemical measures.

A Maternal blood glucose was unaltered among groups. B Maternal blood lactate was unaltered among groups. C Maternal blood sodium was unaltered among groups. D Maternal blood potassium was unaltered among groups. E Cortisol was increased with MNR but corrected to control levels with MNR + IGF1. F Maternal blood progesterone was unaltered among groups. Dotted lines denote normal ranges for each measure in non-pregnant guinea pigs. Control (+ sham treatment) n = 6, MNR (+ sham treatment) n = 6, MNR + IGF1 n = 5. Data are estimated marginal means ± 95% confidence interval. P values calculated using generalized linear models with Bonferroni post hoc analysis. *P ≤ 0.05; **P ≤ 0.01, ***P ≤ 0.001.