Consequences of Zmat3 loss in c-MYC- and mutant KRAS-driven tumorigenesis.

TP53 is a critical tumor suppressor that is mutated in approximately 50% of human cancers. Unveiling the downstream target genes of TP53 that fulfill its tumor suppressor function is an area of intense investigation. Zmat3 (also known as Wig-1 or PAG608) is one such downstream target of p53, whose loss in hemopoietic stem cells lacking the apoptosis and cell cycle regulators, Puma and p21, respectively, promotes the development of leukemia. The function of Zmat3 in tumorigenesis however remains unclear. Here, to investigate which oncogenic drivers co-operate with Zmat3 loss to promote neoplastic transformation, we utilized Zmat3 knockout mice in models of c-MYC-driven lymphomagenesis and KrasG12D-driven lung adenocarcinoma development. Interestingly, unlike loss of p53, Zmat3 germline loss had little impact on the rate of tumor development or severity of malignant disease upon either the c-MYC or KrasG12D oncogenic activation. Furthermore, loss of Zmat3 failed to rescue KrasG12D primary lung tumor cells from oncogene-induced senescence. Taken together, we conclude that in the context of c-MYC-driven lymphomagenesis or mutant KrasG12D-driven lung adenocarcinoma development, additional co-occurring mutations are required to resolve Zmat3 tumor suppressive activity.

Distinct initiating events underpin the immune and metabolic heterogeneity of KRAS-mutant lung adenocarcinoma.

The KRAS oncoprotein, a critical driver in 33% of lung adenocarcinoma (LUAD), has remained an elusive clinical target due to its perceived undruggable nature. The identification of dependencies borne through common co-occurring mutations are sought to more effectively target KRAS-mutant lung cancer. Approximately 20% of KRAS-mutant LUAD carry loss-of-function mutations in KEAP1, a negative regulator of the antioxidant response transcription factor NFE2L2/NRF2. We demonstrate that Keap1-deficient KrasG12D lung tumors arise from a bronchiolar cell-of-origin, lacking pro-tumorigenic macrophages observed in tumors originating from alveolar cells. Keap1 loss activates the pentose phosphate pathway, inhibition of which, using 6-AN, abrogated tumor growth. These studies highlight alternative therapeutic approaches to specifically target this unique subset of KRAS-mutant LUAD cancers.

Heterogeneity of mesenchymal and pluripotent stem cell populations grown on nanogrooves and nanopillars.

Surface nanotopographies are an important way of mimicking the stem cell niche on biomaterial surfaces. Previous studies have focused on the differentiation of stem cells into a defined lineage using nanotopographies, but they have rarely considered the homogeneity of cell populations produced. We examined the impact of two types of substrates (i.e. nanogrooves and nanopillars made by soft lithography) on the surface-induced differentiation of human amniotic membrane-derived mesenchymal stem cells (hAM-MSCs) and mouse embryonic stem cells (mESCs) without the use of additional chemical induction medium components. Cell morphology and proliferation were analysed at day 1 and day 3. Gene expression was analysed at day 14 for hAM-MSCs and at day 7 for mESC-derived embryoid bodies (mEBs) using quantitative real-time polymerase chain reaction (qPCR). The substrates with nanogrooves had a noticeable effect on cell alignment in a depth dependent manner with both cell types showing strong alignment along the deep grooves. On the other hand, the nanopillar substrates showed inhibition of cell spreading for both cell types. The nanogrooves showed inhibition of hAM-MSC growth but enhanced mEB proliferation, especially on the deeper grooves. The nanopillars did not significantly affect hAM-MSC growth, but can modulate mEB growth depending on the pillar density, indicating that mEBs are more sensitive to nanotopographies in terms of proliferation, while hAM-MSCs are only sensitive to specific structures and sizes. Genes associated with bone, cartilage, and fat were investigated for hAM-MSCs, whereas genes of the endoderm, mesoderm, ectoderm, and pluripotency were investigated for mEBs. In general, gene expression for hAM-MSCs was not enhanced significantly by the nanotopographies compared to the flat control. On the other hand, genes of bone, cartilage, skeletal muscle, heart, and liver were up-regulated on both nanopillars and nanogrooves, especially OP65 (ordered pillars with 65% density) and SG40 (shallow grooves with 40 nm depth) in a feature size dependent manner. We found that a small portion of mEBs was composed of cardiac-like beating cells (i.e. GFP-NKX2.5 positive) and a bone cell marker (i.e. OCN) indicating a heterogeneous cell population being generated on those types of surfaces. This work highlights the importance of nanotopographies in stem cell differentiation and how studying multiple properties of the substrate and cells is needed as we strive to generate homogeneous and mature cell populations using biomaterials.

Modulation of human mesenchymal and pluripotent stem cell behavior using biophysical and biochemical cues: A review.

In vitro manipulation of human stem cells is a critical process in regenerative medicine and cellular therapies. Strategies and methods to maintain stem cells and direct them into specific lineages are ongoing challenges in these fields. To date, a number of studies have reported that besides biochemical stimulation, biophysical cues in the form of surface patterning and external stimulation also influence stem cell attachment, proliferation, and differentiation, and can be used in cell reprogramming and the maintenance of pluripotency. While biochemical cues are generally effective and easy to deliver, biophysical cues have many other advantages for scalability as they are cost efficient, have a longer lifetime, and can be easily defined. However, different protocols and cell sources utilized in a variety of studies have led to difficulties in obtaining clear conclusions about the effects of the biophysical environment on stem cells. In addition, the examination of different types of external stimulation is time consuming and limited by available fabrication techniques, resulting in a delay in commercialization and clinical applications. In this review, we aim to summarize the most important biophysical cues and methods for the culture of human stem cells, including mesenchymal and pluripotent stem cells, to facilitate their adoption in stem cell biology. The standard classical protocols of using biochemical cues will also be discussed for comparison. We believe that combining biochemical and biophysical stimulation has the greatest potential to generate functionally mature cells at a scalable and inexpensive rate for diverse applications in regenerative medicine and cell therapy. Biotechnol. Bioeng. 2017;114: 260-280. © 2016 Wiley Periodicals, Inc.