Preeclampsia (PE) is a hypertensive disorder of pregnancy that is one of the leading causes of maternal and fetal morbidity and mortality. PE is characterized by chronic inflammation consisting of activated B cells, producing agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA). This autoantibody contributes to vascular dysfunction, cerebral damage, and inflammation in PE. AT1-AA is also present in maternal circulation for years postpartum and could be involved in the increased risk for diseases such as hypertension or dementia in postpartum PE women. AT1-AA has been identified in COVID-19 patients. It is produced during the inflammatory response and plays a role in the dysregulation of the renin-angiotensin system in COVID-19 patients. Pregnant women who experience a COVID-19 infection are at higher risk of developing preeclampsia, and AT1-AA may be contributing to this increased risk. Therapies targeting AT1-AA during and after preeclampsia and/or COVID-19 may reduce the risks and burdens of cardiovascular and neurological disease.

Introduction Tumor necrosis factor alpha (TNF-α) is elevated 2-fold in women with preeclampsia. Preclinical investigation shows TNF-α blockade reduces maternal blood pressure in reduced uterine perfusion pressure (RUPP) rats and attenuates the reduction in fetal weight. However, the benefits versus harms of this therapy on fetal growth and underlying pathogenesis are unknown. Thus, this study tested the hypothesis that maternal treatment with the soluble TNF-α inhibitor Etanercept (Etan) during late gestation improves placental perfusion, nutrient transport, and morphology, thereby improving fetal growth in the RUPP model of preeclampsia compared with Sham controls. We further hypothesized that maternal Etan treatment is associated with improved blood pressure and inflammatory profiles in offspring. Methods Sham or RUPP surgery was performed at gestational day (GD) 14, with vehicle or Etan (0.4 mg/kg, s.c.) administered at GD18. Results Fetal weight (p = 0.0365) and survival (p = 0.0002) were reduced in RUPP (p = 0.0365) at GD20; only fetal weight was improved in Etan-RUPP (p = 0.0480). At GD20, uterine artery resistance index (UARI) was increased in RUPP (p = 0.0094), but attenuated in Etan-RUPP, indicating improved placental perfusion. Impaired placental transport and morphology were evident in RUPP, with no improvement in Etan-RUPP. Birthweight was improved in Etan-RUPP (p = 0.0312), although total NK cells (p = 0.0376) were increased in female Etan-RUPP offspring. Circulating AT1-AA activity was elevated in adult male and female RUPP offspring (p = 0.0013, p = 0.0006), but attenuated in female Etan-RUPP offspring. Discussion: These results suggest improved fetal growth in Etan-RUPP, independent of placental morphology or nutrient transporter expression, with sex-specific effects on inflammation in adult RUPP offspring.

Preeclampsia (PE) is new-onset hypertension and multi-organ dysfunction following the 20th week of gestation. The risk of developing PE has shown a familial connection, as the female children from PE pregnancies have an elevated risk of developing PE. An agonistic autoantibody to angiotensin II type 1 receptor (AT1-AA) is produced in PE women, which causes a PE phenotype in pregnant rats, and is present in postpartum maternal circulation. Introducing AT1-AA during pregnancy is known to cause hypertension in the female offspring (OS) as adults; however, the impact of perinatal AT1-AA exposure on blood pressure and immune activation during OS pregnancy is unknown. We hypothesize that perinatal AT1-AA exposure will lead to female OS developing a PE phenotype during pregnancy. On gestational day (GD) 14, we infused AT1-AA (1:40 in saline) and allowed the dams (F0) to deliver, then measured the birth weight within 12 hours. OS (F1) grew to 3 months. Control males and control or AT1-AA females were mated. On F1 GD18, carotid catheters were inserted. On F1 GD19, mean arterial pressure (MAP) was measured, and then blood and tissues were collected. Weights and litter size were measured. Placental and renal preproendothelin were quantified using RT-PCR. Immune cells were measured via flow cytometry. A t-test was used for statistical analysis. F1 AT1-AA pregnant OS had increased MAP and smaller litters compared to F1 NP OS (117±3 mmHg, p< 0.05 vs 105±2 mmHg) (7±2 pups, vs 13±1 pups, p< 0.05). Placental T helper cells were significantly increased in F1 AT1-AA OS (34±3 % gated) compared to F1 NP OS (16±3 % gated), with decreased T regulatory cells in F1 AT1-AA OS (0.44±0.12 % gated) compared to F1 NP OS (1.27±0.40 % gated). AT1-AA female F1 OS got pregnant earlier than normal pregnant (NP) female F1 OS (14±2 weeks, vs 16±3 weeks, p< 0.05). F1 AT1-AA pregnant OS were smaller at gestational day 19 than F1 NP OS (293±5g, vs 338±7g, p< 0.0001). Placental PPET and renal PPET were significantly increased 9.5±2.0 fold, 7.5±2.5 fold in F1 AT1-AA dams compared to F1 NP dams. Perinatal AT1-AA exposure predisposes F1 female OS to have elevated blood pressure and inflammation during pregnancy, supporting the hypothesis that AT1-AA may be a component contributing to PE familial risk. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

OBJECTIVE:Gestational diabetes mellitus (GDM) refers to glucose intolerance, insulin sensitivity, and beta islet cell dysfunction during pregnancy. GDM pathogenesis is associated with hypertension, impaired placental and renal function, oxidative stress, and increased circulating CD4+ T cells. There are limited animal models to explore GDM pathology and treatment. This study sought to determine a role for GDM placental CD4+ T cells to parallel manifestations of the GDM phenotype in pregnant athymic nude rats. METHODS:GDM placental CD4+ T cells (GDM T cells) were isolated upon delivery and injected into pregnant nude rats on gestational day (GD) 12. Mean arterial pressure and markers of renal injury, proteinuria, kidney injury molecule-1, and neutrophil gelatinase-associated lipocalin, were assessed on GD19. Glucose, insulin tolerance, and glucose tolerance tests were also performed. Renal and pancreatic tissues were stained using Periodic acid Schiff and hematoxylin and eosin, respectively. A one-way ANOVA was used for statistical analysis. RESULTS:Adoptive transfer of GDMT cells increased blood pressure (120.8 ± 2.2 mmHg, p < 0.05) compared to controls (105.4 ± 2.8 mmHg) and normotensive Tcell recipients (96.3 ± 3.9 mmHg). Metformin or MitoTEMPO attenuated this response. GDM T cell recipients had elevated blood glucose (p < 0.05) and impaired glucose tolerance and insulin sensitivity, which improved with metformin or MitoTEMPO treatment. Renal injury was more severe in GDM T cell recipients, but attenuated with metformin or MitoTEMPO. Pancreatic morphology showed reduced beta islet numbers in GDM T cell recipients. CONCLUSION:GDM CD4+ T cells contribute to hypertension, glucose intolerance, and renal dysfunction, improved byMitoTEMPO. These findings supports optional therapeutics that support mitochondrial function during pregnancy.

Preeclampsia (PE) is new-onset hypertension with chronic inflammation and systemic endothelial dysfunction after the 20th week of pregnancy. We’ve shown that CD4+ T helper cells contribute to PE phenotype through production of proinflammatory cytokines such as TNF-α. We have previously shown that etanercept, a soluble TNF-α inhibitor, improves the PE phenotype in RUPP rats. Therefore, we hypothesize that etanercept will improve blood pressure, endothelial function, and inflammation in response to the adoptive transfer of placental PE CD4+ T cells into pregnant athymic nude rats. Placental CD4+ T cells were isolated from normal pregnant (NP) and PE patients following delivery. One million CD4+ T cells were injected I.P. into athymic nude rats on gestational day (GD) 12. Etanercept (0.4 mg/kg) was given as an I.P. injection on GD 15. Carotid catheters were implanted on GD 18. Blood pressure (MAP) was measured on GD 19. Preproendothelin expression, measured by RT-PCR, and ET-1 and TNF-α were measured with ELISA. A one-way ANOVA was used for statistical analysis. MAP increased with PE CD4+ T cells to 120±3 mmHg, n=17, p< 0.05, compared to NP CD4+ T cells (102±4 mmHg, n=12), which was blunted with Etanercept (110±3 mmHg, n=8). Renal ET-1 was lower in Etanercept treated rats (1.6±034, n=7) compared to PE recipients (3.4±0.4 pg/mg protein). Placental ET-1 was significantly increased with PE CD4+T cells (20.4±0.7, n=4) compared to NP CD4+ T cells (13.6±2.4, n=5) or Etanercept treated rats (10.8±1.0 pg/mg protein, n=5). Renal TNF-α was significantly increased with PE CD4+ T cells (3.9±0.3, n=7) compared to NP recipients (3.1±0.2 pg/mg protein, n=6) and PE CD4+ T cells recipients treated with Etanercept (1.3±0.2 pg/mg protein, n=8). TNF-α blockade with Etanercept lowers blood pressure and multiorgan ET-1, a marker of endothelial dysfunction and a hallmark of PE, in response to adoptive transfer of preeclamptic CD4+ T cells. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Pre-eclampsia (PE), new onset hypertension during pregnancy in association with multi-organ dysfunction, immune activation, agonistic autoantibodies to the angiotensin II type 1 receptor (AT1-AA), and endothelial dysfunction with increased vasoconstrictor endothelin (ET-1). AT1-AA exposure during pregnancy contributes to hypertension, immune activation, and vascular dysfunction by stimulating endothelin-1 during pregnancy. The AT1-AA is produced 7 years postpartum suggesting a memory immune response mediated by memory B cells in the pathophysiology of PE. Although we have shown the role of CD19+ B cells in causing hypertension during pregnancy, the role of CD20+ B cells, a marker expressed on most B cells excluding CD19+ plasma cells but is retained on memory B cells, has not been examined. Thus, we hypothesize that placental CD20+ B cells from PE patients will induce a PE phenotype in pregnant athymic nude rats possibly through AT1-AA stimulated pathways, such as ET-1. The objective of this study was to determine if the adoptive transfer of placental CD20+ B cells isolated from PE women at delivery causes hypertension and endothelial dysfunction via the production of AT1-AA, similarly to CD19+ B cells, in pregnant recipient rats. Three hundred thousand placental CD20+ B cells from normal pregnant (NP) or PE patients were isolated and injected i.p. into nude athymic rats on gestational day (GD) 12. Mini-osmotic pumps containing the AT1-AA activity-blocking peptide, ‘n7AAc’, at a dose of 24 µg/day were inserted on GD 14. On GD 18, carotid catheters were implanted. Mean arterial pressure (MAP) was measured and blood and tissues were collected on GD 19. Renal preproendothelin (PPET) was quantified using Real Time-PCR. AT1-AA was quantified using a cardiomyocyte bioassay. A one-way ANOVA was used for statistical analysis. Participants had similar age, BMI, and gestational age (36±1 (PE) vs 39±0 (NP) weeks) at delivery. PE participants had higher MAP at delivery than controls (p<0.05). MAP was 109±3 mmHg (n=9) in the recipient rats of Normal Pregnant (NP) CD20+ B cells which increased to 123±2 mmHg (n=8, p<0.01) in the recipients of PE B cells. This increase in MAP was lower, 110±3 mmHg (n=6, p<0.05), with ‘n7AAc’. Renal PPET expression increased by 2.60±0.61 fold (ΔΔct) in PE B cell recipients (p<0.05) compared to NP B cell recipients and was significantly lower in ‘n7AAc’ treated rats 0.79±0.15 fold (p<0.01). AT1-AA activity was elevated in the PE B cell recipients (14±2 ΔBPM) compared to NP B cell (1±1 ΔBPM, p<0.01) and was lowered with ‘n7AAc’ (2±3 ΔBPM, p<0.05). CD20+ B cells from placentas of PE women cause hypertension and ET-1 through AT1-AA mediated pathways during pregnancy, thereby supporting an important role for CD20+, possibly memory B cells, in the pathophysiology of PE. This study was supported by NIH grants RO1HL170622 (BL), F31HD110230 (NC), and P20GM121334 (BL). This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Preeclampsia (PE), new-onset hypertension during pregnancy, is associated with chronic inflammation both in the placenta and systemically. PE is characterized by placental ischemia, which then results in the production and release of anti-angiogenic factors and inflammatory mediators. Inflammation in PE leads to placental, renal, and vascular damage, which contribute to the phenotype of hypertension and organ dysfunction during pregnancy. T cells, B cells, Natural Killer cells, and macrophages have all been shown to play a role in the inflammation present in the disease. T helper cells contribute to the chronic inflammation in PE. They also activate B cells, which produce agonistic autoantibodies against the angiotensin II type 1 receptor. Natural Killer cells are activated in PE and shift away from decidual Natural killer cells, which produce angiogenic factors, and toward cytotoxic Natural Killer cells, which contribute to tissue damage. Macrophages are polarized towards proinflammatory subtypes and contribute to tissue damage and inflammatory signaling in PE patients. As the immune system plays a role in the pathophysiology of the disease, it may be a potential target for therapeutic intervention to improve maternal and fetal outcomes during and following a PE pregnancy.

Introduction: Preeclampsia (PE) is characterized by new onset hypertension after 20 weeks of gestation. It affects 5-7% of all pregnancies in the U.S.A, and is associated with activated CD4+T cells, inflammation, autoantibodies to angiotensin II type 1 receptor (AT1-AA), reduced fetal weight and hypertension. The mechanisms responsible for the pathogenesis of PE are unclear and currently the only treatment is early delivery of the feto-placental unit. We have previously shown that CD4+ T cells adoptive transfer causes a PE phenotype in athymic nude rats. Objective: Determine whether progesterone, in the form of 17-hydroxyprogesterone caproate (17-OHPC), attenuates the effect of placental PE CD4+ T cells to cause signs of PE in athymic nude rats. Methods: One million of placental CD4+ T cells from normal pregnant (NP) or PE participants were isolated by magnetic separation and injected I.P. into pregnant nude athymic rats on gestational day (GD) 12. On GD 15, 17-OHPC (3.32mg/kg) was administered I.P. On GD18, carotid catheters were inserted. On GD19, mean arterial blood pressure, blood and tissues were collected. Participants whose placental CD4+ T cells were used in this study had matched gestational age. A student’s t-test or one-way ANOVA were used for statistical analysis. All data are expressed as mean ± standard error means (SEM). Results: Blood pressure was 116+/-4 in NP (n=8), 147 +/- 4 mmHg in PE group (n=19) and significantly reduced to 136 +/- 3 mmHg in PE+17-OHPC group (n=14, p<0.05). CD4+ T cells were 2+/- 1 % gate in NP, 6 +/- 1 in PE, and 3 +/-1 % gate in PE+17-OHPC group (n= 5). TNF-alpha were 21 +/- 4 pg/mL in NP(n=6),41 +/-8 in PE group (n=8), and reduced to 21+/-5 in PE+17-OHPC (n=6, p<0.05). AT1-AA levels were 23 +/- 5 ΔBPM (beats per minute) in PE group (n=4) and 10/-2 in PE+17-OHPC group (n=8). MAP was 98 +/- 2 mmHg in NP CD4+ T cells rats (n=5), 81+/-2 in NP CD4+ Tcells+17-OHPC (n=5), 124 +/- 5 mmHg in PE CD+4 T cells (n=9), which decreased to 103 +/- 4 mmHg in PE CD4+ T cells+17-OHPC (n=10). Pup weight was 1.8+/- 1 grams in PE CD+4 T cells, which increased to 2.2 +/- 1 grams mmHg in PE CD4+ T cells+17-OHPC (p<0.05). AT1-AA were -3 +/- 2 Δ beats per minute (BPM) in NP CD4+ T cells group, 8+/-2 ΔBPM in PE CD+4 T cells (n=4). 17-OHPC treatment significantly decreased to -6 +/- 1 ΔBPM (n=3, p<0.05). Conclusion: CD4+ T cells play an important role in the pathophysiology of preeclampsia and 17-OHPC reduces the signs of PE in pregnant nude-athymic rats. Supported by NIH P20GM121334 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Preeclampsia (PE), new-onset hypertension during pregnancy, is accompanied by organ dysfunction, in addition to placental dysfunction, and is associated with chronic inflammation and fetal growth restriction. Agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA) are produced in PE women and induce a PE-like phenotype when infused into pregnant rats. PE offspring are at higher risk for cardiovascular disease and hypertension; however, AT1-AA's role in these risks is unknown. Therefore, we hypothesized that AT1-AA exposure during pregnancy contributes to hypertension in adult offspring. We infused AT1-AA (1:40) starting on gestational day (GD) 14, allowed the dams to give birth, and birth weight was measured within 12 h. Offspring were aged to 4 months, one male and one female per litter were randomly selected to undergo carotid catheterization, mean arterial pressure (MAP) measurement, and blood and tissue collection. Female AT1-AA offspring had elevated MAP compared to control female offspring. Female AT1-AA offspring also had elevated progesterone, measured by mass spectroscopy, and renal endothelin-1, measured by RT-PCR and ELISA, compared to female control offspring. Male and female AT1-AA offspring had increased circulating AT1-AA, measured by cardiomyocyte bioassay, compared to male and female control offspring. These data demonstrate that AT1-AA infusion during pregnancy can predispose female offspring to have elevated blood pressure and persistently increased renal ET-1 and AT1-AA in adulthood.

Preeclampsia (PE), new-onset hypertension during pregnancy in association with other-organ dysfunction including the placenta. It is associated with inflammatory cytokines and fetal growth restriction (FGR) which itself is associated with increased risk for hypertension and cardiovascular disease in adult offspring. PE women have increased tumor necrosis factor alpha (TNF-α) and agonistic autoantibodies to the angiotensin II type 1 receptor (AT1-AA). Moreover, female PE children are at elevated risk for PE during pregnancy indicating a familial connection with PE. The reduced uterine perfusion pressure (RUPP) model of PE recaptures many of the characteristics of PE including hypertension, inflammatory activation, and fetal growth restriction. Growth-restricted RUPP offspring develop hypertension as adults and we have recently shown that adult RUPP offspring also produce AT1-AA. Importantly, RUPP rats have increased TNF-α and we have shown that TNF-α blockade with etanercept improves maternal blood pressure. The use of etanercept has not resulted in negative effects for mothers with autoimmune disorders that are kept on their treatment throughout pregnancy, however, its effect on fetal weight and long-term health is still being investigated. We hypothesize that etanercept could improve cardiovascular and chronic inflammatory outcomes in adult RUPP offspring. Pregnant rats were divided into the following groups: Sham, Sham + Etanercept, RUPP, RUPP + Etanercept. RUPP surgeries were performed on gestational day 14 and Etanercept (400 mcg/kg) was injected i.p. on gestational day 18. The dams were allowed to deliver naturally and birth weights were recorded within 12 hours. At 4 months of age mean arterial pressure (MAP) was recorded via carotid catheters, and blood and tissues were collected. Immune cells were measured by flow cytometry. AT1-AA was measured using a cardiomyocyte bioassay. A two-way ANOVA was used for statistical analysis. Etanercept improves birth weight in male RUPP offspring (5.2±0.2 g, n=7 (RUPP) vs 6.0±0.3g, n=9, p<0.05 (RUPP+Etanercept)), but did not affect adult blood pressure or inflammatory factors. Although Etanercept did not improve female birth weight, Etanercept trended to improve MAP in RUPP female offspring (130±5 mmHg, n=6, (RUPP) vs 123±4 mmHg, n=7 (RUPP+Etanercept)). In addition, there was a trending decrease in circulating B cells in the RUPP female offspring compared to sham female offspring which was normalized in the RUPP + Etanercept female offspring. (4.24±0.81 % gated, n=5 (RUPP); vs 9.56±2.07 % gated, n=7 (Sham), vs 9.04±1.42 % gated, n=6, (RUPP+Etanercept)). RUPP + Etanercept female offspring also had decreased in circulating AT1-AA which was elevated in RUPP female offspring (16±3 ΔBPM (RUPP), n=5, 7±2 ΔBPM, n=4 (RUPP+Etanercept), -1±2 ΔBPM, n=5, (Sham)). Overall, perinatal Etanercept treatment did not worsen fetal outcomes. Importantly, Etanercept may have a more positive effect to improve overall health outcomes of female offspring as seen by the trend to reduce blood pressure, normalize immune cells, and reduce circulating AT1-AA in adult females. Blocking TNF-α during PE may be a potential therapeutic strategy to improve both maternal and female fetal outcomes. This study was supported by NIH grants R01HL170622 (BL), R01HL143459 (BTA), F31HD110230 (NC), P20GM104357 (BTA), P30GM149404 (BTA), and P20GM121334 (BL, BTA). This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Preeclampsia (PE) is new-onset hypertension and multi-organ dysfunction following the 20 th week of gestation. Female PE children are at elevated risk for PE during pregnancy, indicating a familial connection. Agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA) are produced in PE women, induce a PE phenotype when infused into pregnant rats, and are present in postpartum maternal circulation. We have previously shown that perinatal exposure to AT1-AA causes adult female first generation (F1) offspring (OS) to have elevated blood pressure. However, the role of perinatal AT1-AA exposure to increase the risk for hypertension and immune activation during pregnancy is unknown. Thus, we hypothesize that perinatal exposure to AT1-AA would cause F1 AT1-AA female OS to develop a PE phenotype during pregnancy. We infused AT1-AA (1:40) starting on gestational day (GD)14 and allowed the dams (F0) to give birth. Birth weight was measured within 12 hours. OS were aged to 3 months. Control males and control or AT1-AA females were mated. Pregnancy was confirmed by vaginal smear. On GD18 carotid catheters were inserted. On GD19, mean arterial pressure (MAP) was measured and blood and tissues were collected. Weights and litter size were measured. Placental and renal preproendothelin were measured by RT-PCR. Immune cells were measured with flow cytometry. A student’s t-test was used for statistical analysis. F1 AT1-AA pregnant OS had increased MAP and smaller litters compared to F1 NP OS (117±3 mmHg, p<0.05 vs 105±2 mmHg) (7±2 pups, vs 13±1 pups, p<0.05). Placental T helper cells were significantly increased in F1 AT1-AA OS (34±3 % gated) compared to F1 NP OS (16±3 % gated) with decreased T regulatory cells in F1 AT1-AA OS (0.44±0.12 % gated) compared to F1 NP OS (1.27±0.40 % gated). AT1-AA female F1 OS got pregnant earlier than normal pregnant (NP) female F1 OS (14±2 weeks, vs 16±3 weeks, p<0.05). F1 AT1-AA pregnant OS were smaller at gestational day 19 than F1 NP OS (293±5g, vs 338±7g, p<0.0001). Placental PPET and renal PPET were significantly increased 9.5±2.0 fold; 7.5±2.5 fold in F1 AT1-AA dams compared to F1 NP dams. These data demonstrate that perinatal AT1-AA exposure predisposes F1 female OS to have elevated blood pressure and inflammation during pregnancy supporting the hypothesis that AT1-AA may be a factor contributing to the familial risk for PE.

Circulating tumor DNA (ctDNA) is an emerging biomarker for the treatment of early breast cancer (EBC). We sought to evaluate a highly sensitive tumor-informed ctDNA assay in a real-world cohort of patients receiving neoadjuvant therapy (NAT) to assess clinical validity and explore prognostic outcomes. ctDNA is detected in 77.2% (88/114) of participants at baseline, with 18/88 (20.5%) having a baseline estimated variant allele frequency (eVAF) of <0.01%. Persistent detection of ctDNA, measured midway through NAT (mid-NAT), is associated with disease recurrence in all participants, reaching statistical significance in those with HER2-negative disease. Stratified analyses demonstrate that ctDNA detected mid-NAT enhances the prognostic accuracy of the residual cancer burden (RCB) score for disease recurrence. Postoperative or follow-up detection of ctDNA demonstrates a 100% positive predictive value for disease recurrence, with a median lead time of 374 days (range: 13-1010 days). These data suggest that assays with high analytical sensitivity may improve baseline ctDNA detection in patients with EBC. The ability to replicate the prognostic association of ctDNA dynamics in a real-world cohort supports further investigation. Prospective trials incorporating ctDNA testing are warranted to assess and develop the clinical utility of ctDNA-guided treatment strategies.