Insulin-like Growth Factor Binding Protein 3 Increases Mouse Preimplantation Embryo Cleavage Rate by Activation of IGF1R and EGFR Independent of IGF1 Signalling.

The viability of embryos cultured in vitro is poor compared to those that develop in vivo. The lack of maternally derived growth factors in vitro may contribute to this problem. Insulin-like growth factor binding protein 3 (IGFBP3) is one such growth factor that has been identified in the maternal reproductive system. This study examined the role of autocrine and exogenous IGFBP3 in mouse preimplantation embryos. Embryos expressed IGFBP3 across all stages of preimplantation development, and addition of exogenous IGFBP3 to embryo culture media increased the rate of development to the 2-, 4-, 5-, and 8-cell stages. Addition of inhibitors of the IGF1 and EGF receptors prevented this IGFBP3-mediated improvement in developmental rate, but the effect was not cumulative, indicating that both receptors are transactivated downstream of IGFBP3 as part of the same signalling pathway. Acute exposure to IGFBP3 increased phosphorylation of Akt and rps6 in 4-8 cell embryos, suggesting activation of the PI3-kinase/Akt pathway downstream of the IGF1 and EGFR receptors to promote cell proliferation and survival. In conclusion, addition of IGFBP3 to embryo culture media increases early cleavage rates independent of IGF1 signalling and therefore, IGFBP3 addition to IVF culture media should be considered.

In vitro and in vivo drug screens of tumor cells identify novel therapies for high-risk child cancer.

Biomarkers which better match anticancer drugs with cancer driver genes hold the promise of improved clinical responses and cure rates. We developed a precision medicine platform of rapid high-throughput drug screening (HTS) and patient-derived xenografting (PDX) of primary tumor tissue, and evaluated its potential for treatment identification among 56 consecutively enrolled high-risk pediatric cancer patients, compared with conventional molecular genomics and transcriptomics. Drug hits were seen in the majority of HTS and PDX screens, which identified therapeutic options for 10 patients for whom no targetable molecular lesions could be found. Screens also provided orthogonal proof of drug efficacy suggested by molecular analyses and negative results for some molecular findings. We identified treatment options across the whole testing platform for 70% of patients. Only molecular therapeutic recommendations were provided to treating oncologists and led to a change in therapy in 53% of patients, of whom 29% had clinical benefit. These data indicate that in vitro and in vivo drug screening of tumor cells could increase therapeutic options and improve clinical outcomes for high-risk pediatric cancer patients.

Optimization of a clofarabine-based drug combination regimen for the preclinical evaluation of pediatric acute lymphoblastic leukemia.

BACKGROUND: The aim of this study was to improve the predictive power of patient-derived xenografts (PDXs, also known as mouse avatars) to more accurately reflect outcomes of clofarabine-based treatment in pediatric acute lymphoblastic leukemia (ALL) patients. PROCEDURE: Pharmacokinetic (PK) studies were conducted using clofarabine at 3.5 to 15 mg/kg in mice. PDXs were established from relapsed/refractory ALL patients who exhibited good or poor responses to clofarabine. PDX engraftment and response to clofarabine (either as a single agent or in combinations) were assessed based on stringent objective response measures modeled after the clinical setting. RESULTS: In naïve immune-deficient NSG mice, we determined that a clofarabine dose of 3.5 mg/kg resulted in systemic exposures equivalent to those achieved in pediatric ALL patients treated with clofarabine-based regimens. This dose was markedly lower than the doses of clofarabine used in previously reported preclinical studies (typically 30-60 mg/kg) and, when scheduled consistent with the clinical regimen (daily × 5), resulted in 34-fold lower clofarabine exposures. Using a well-tolerated clofarabine/etoposide/cyclophosphamide combination regimen, we then found that the responses of PDXs better reflected the clinical responses of the patients from whom the PDXs were derived. CONCLUSIONS: This study has identified an in vivo clofarabine treatment regimen that reflects the clinical responses of relapsed/refractory pediatric ALL patients. This regimen could be used prospectively to identify patients who might benefit from clofarabine-based treatment. Our findings are an important step toward individualizing prospective patient selection for the use of clofarabine in relapsed/refractory pediatric ALL patients and highlight the need for detailed PK evaluation in murine PDX models.

Lymphocyte-Specific Chromatin Accessibility Pre-determines Glucocorticoid Resistance in Acute Lymphoblastic Leukemia.

Glucocorticoids play a critical role in the treatment of lymphoid malignancies. While glucocorticoid efficacy can be largely attributed to lymphocyte-specific apoptosis, its molecular basis remains elusive. Here, we studied genome-wide lymphocyte-specific open chromatin domains (LSOs), and integrated LSOs with glucocorticoid-induced RNA transcription and chromatin modulation using an in vivo patient-derived xenograft model of acute lymphoblastic leukemia (ALL). This led to the identification of LSOs critical for glucocorticoid-induced apoptosis. Glucocorticoid receptor cooperated with CTCF at these LSOs to mediate DNA looping, which was inhibited by increased DNA methylation in glucocorticoid-resistant ALL and non-lymphoid cell types. Our study demonstrates that lymphocyte-specific epigenetic modifications pre-determine glucocorticoid resistance in ALL and may account for the lack of glucocorticoid sensitivity in other cell types.