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Duvelisib, an oral dual inhibitor of PI3K-δ and PI3K-γ, is in phase III trials for the treatment of chronic lymphocytic leukemia (CLL) and indolent non-Hodgkin's lymphoma. In CLL, duvelisib monotherapy is associated with high iwCLL (International Workshop on Chronic Lymphocytic Leukemia) and nodal response rates, but complete remissions are rare. To characterize the molecular effect of duvelisib, we obtained samples from CLL patients on the duvelisib phase I trial. Gene expression studies (RNAseq, Nanostring, Affymetrix array and real-time RT-PCR) demonstrated increased expression of BCL2 along with several BH3-only pro-apoptotic genes. In concert with induction of transcript levels, reverse phase protein arrays and immunoblots confirmed increase at the protein level. The BCL2 inhibitor venetoclax induced greater apoptosis in ex vivo-cultured CLL cells obtained from patients on duvelisib compared with pre-treatment CLL cells from the same patients. In vitro combination of duvelisib and venetoclax resulted in enhanced apoptosis even in CLL cells cultured under conditions that simulate the tumor microenvironment. These data provide a mechanistic rationale for testing the combination of duvelisib and venetoclax in the clinic. Such combination regimen (NCT02640833) is being evaluated for patients with B-cell malignancies including CLL.

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Oncogenic mutations in PIK3CA, which encodes the phosphoinositide-3-kinase (PI3K) catalytic subunit p110α, occur in ∼25% of human breast cancers. In this study, we report the development of a knock-in mouse model for breast cancer where the endogenous Pik3ca allele was modified to allow tissue-specific conditional expression of a frequently found Pik3ca(H1047R) (Pik3ca(e20H1047R)) mutant allele. We found that activation of the latent Pik3ca(H1047R) allele resulted in breast tumors with multiple histological types. Whole-exome analysis of the Pik3ca(H1047R)-driven mammary tumors identified multiple mutations, including Trp53 mutations that appeared spontaneously during the development of adenocarinoma and spindle cell tumors. Further, we used this model to test the efficacy of GDC-0941, a PI3K inhibitor, in clinical development, and showed that the tumors respond to PI3K inhibition.

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BACKGROUND: Injury to reproductive organs including the uterus is a known complication of ionizing radiation, but the risks to the mother and fetus during subsequent pregnancies are not well defined. CASE: A young woman with a remote history of whole body irradiation for childhood leukemia had uterine rupture at 17 weeks' gestation. Pathologic examination of the supracervical hysterectomy specimen revealed a posterior-fundal placenta percreta with a diffusely thinned myometrium (1-6 mm). The clinicopathologic findings were consistent with prior radiation injury. CONCLUSION: Uterine irradiation may predispose to abnormal placentation and uterine rupture in a subsequent pregnancy.

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Development of the myotome within somites depends on unknown signals from the neural tube. The present study tested the ability of basic fibroblast growth factor (bFGF), transforming growth factor-beta1 (TGF-beta1) and dorsalin-1 (dsl-1) to promote myogenesis in stage 10-14 chick paraxial mesoderm utilizing 72 hour explant cultures. Each of these factors alone and the combination of bFGF with dsl-1 had limited to no myogenic-promoting activity, but the combination of bFGF with TGF-beta1 demonstrated a potent dose-dependent effect. In addition, bFGF enhanced the survival/proliferation of somite cells. 98% of stage 10-11 caudal segmental plate explants treated with bFGF plus TGF-beta1, exhibited myosin heavy chain (MHC)-positive cells (avg.=60 per explant), whereas only 15% of similarly treated somites responded with an average of 5 MHC-positive cells. Thus at stage 10-11, there are rostrocaudal differences in myogenic responsiveness with the caudal (more 'immature') paraxial mesoderm being more myogenically responsive to these factors than are somites. It was also discovered that 17% of stage 10-11 caudal segmental plate explants exhibited several MHC-positive cells even when cultured without added growth factors, further demonstrating a different myogenic potential of the caudal paraxial mesoderm. Stage 13-14 paraxial mesoderm also exhibited a myogenic response to bFGF/TGF-beta1 but, unlike stage 10-11 embryos, both somites and segmental plate exhibited a strong response. A two-step mechanism for the bFGF/TGF-beta1 effect is suggested by the finding that only TGF-beta1 was required during the first 12 hours of culture, whereas bFGF plus a TGF-beta-like factor were required for the remainder of the culture. The biological relevance of the findings with bFGF is underscored by the observation that a monoclonal antibody to bFGF inhibited myogenic signaling from the dorsal neural tube. However, a monoclonal antibody that can neutralize the three factors TGF-beta1, TGF-beta2 and TGF-beta3 did not block myogenic signals from the neural tube, raising the possibility that another TGF-beta family member may be involved in vivo.

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A model experimental system for investigating myogenic induction signals has been devised with mouse P19 embryonal carcinoma cells. When cocultured with pieces of chick neural tube, aggregated P19 cells are induced to become skeletal muscle. The most potent inducing activity is localized to the dorsal neural tube. Less activity was found in the ventral neural tube, notochord, ectoderm, and lateral plate mesoderm, and none was detected in the neural retina. These results suggest that P19 cells may be a useful model system for investigating the mechanisms underlying induction of somite myogenesis.

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Previous studies have demonstrated that the neural tube/notochord complex is required for skeletal muscle development within somites. In order to explore the localization of myogenic inducing signals within the neural tube, dorsal or ventral neural tube halves were cultured in contact with single somites or pieces of segmental plate mesoderm. Somites and segmental plates cultured with the dorsal half of the neural tube exhibited 70% and 85% myogenic response rates, as determined by immunostaining for myosin heavy chain. This response was slightly lower than the 100% response to whole neural tube/notochord, but was much greater than the 30% and 10% myogenic response to ventral neural tube with and without notochord. These results demonstrate that the dorsal neural tube emits a potent myogenic inducing signal which accounts for most of the inductive activity of whole neural tube/notochord. However, a role for ventral neural tube/notochord in somite myogenic induction was clearly evident from the larger number of myogenic cells induced when both dorsal neural tube and ventral neural tube/notochord were present. To address the role of a specific dorsal neural tube factor in somite myogenic induction, we tested the ability of Wnt-1-expressing fibroblasts to promote paraxial mesoderm myogenesis in vitro. We found that cells expressing Wnt-1 induced a small number of somite and segmental plate cells to undergo myogenesis. This finding is consistent with the localized dorsal neural tube inductive activity described above, but since the ventral neural tube/notochord also possesses myogenic inductive capacity yet does not express Wnt-1, additional inductive factors are likely involved.

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We have developed an in vitro assay for examining the neural tube/notochord influence on somite myogenesis. No cells committed to myogenesis (operationally defined as immunopositive staining for myosin heavy chain after a 48-hr culture period) are present in the most caudal somites (I-V) of stage 8-11 chicken embryos, whereas by stage 14-15, the most caudal somites occasionally produce differentiated muscle cells when cultured alone. To test for responsiveness to neural tube, one somite of each pair was cultured in contact with a piece of neural tube; as a control, the contralateral somite was cultured alone to test for prior myogenic commitment of somitic cells. Using this assay, we show that neural tube induces myogenesis in single somite explants, that close somite-neural tube proximity is required, and that the interaction can occur in defined medium. Neural tube induces myogenesis in the most recently formed somites and in segmental plate mesoderm, indicating that somite segmentation is not a prerequisite for neural tube mediated myogenesis in vitro. The assay was also used to determine whether continued exposure to neural tube is required for increasing myogenic cell numbers in somites which already contain cells committed to myogenesis. Since somites containing committed cells exhibit a fourfold increase in the number of myogenic cells when cocultured with neural tube, continued in vitro exposure to neural tube either recruits additional somite cells into the myogenic lineage, and/or it stimulates proliferation of existing myogenic cells. Consistent with this finding, the most rostral neural tube retains its inductive properties as evidenced by the induction of nascent myogenesis in the most caudal somites. Parallel studies of notochord mediated myogenesis indicate that the notochord promotes myogenesis within somites II-VIII, but not within somite I or the segmental plate. This observation suggests either a qualitative or quantitative difference in the mechanism by which the neural tube and notochord communicate with the youngest somite and presomitic mesoderm.

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