the next frontier in medicine

PM003/2016publishersversionpublishe

Convergence of Logic of Cellular Regulation in Different Premalignant Cells by an Information Theoretic Approach

Abstract Background Surprisal analysis is a thermodynamic-like molecular level approach that identifies biological constraints that prevents the entropy from reaching its maximum. To examine the significance of altered gene expression levels in tumorigenesis we apply surprisal analysis to the WI-38 model through its precancerous states. The constraints identified by the analysis are transcription patterns underlying the process of transformation. Each pattern highlights the role of a group of genes that act coherently to define a transformed phenotype. Results We identify a major transcription pattern that represents a contraction of signaling networks accompanied by induction of cellular proliferation and protein metabolism, which is essential for full transformation. In addition, a more minor, "tumor signature" transcription pattern completes the transformation process. The variation with time of the importance of each transcription pattern is determined. Midway through the transformation, at the stage when cells switch from slow to fast growth rate, the major transcription pattern undergoes a total inversion of its weight while the more minor pattern does not contribute before that stage. Conclusions A similar network reorganization occurs in two very different cellular transformation models: WI-38 and the cervical cancer HF1 models. Our results suggest that despite differences in a list of transcripts expressed in different cancer models the rationale of the network reorganization remains essentially the same

A Thermodynamic-Based Interpretation of Protein Expression Heterogeneity in Different Glioblastoma Multiforme Tumors Identifies Tumor-Specific Unbalanced Processes

We describe a thermodynamic-motivated, information theoretic analysis of proteomic data collected from a series of 8 glioblastoma multiforme (GBM) tumors. GBMs are considered here as prototypes of heterogeneous cancers. That heterogeneity is viewed here as manifesting in different unbalanced biological processes that are associated with thermodynamic-like constraints. The analysis yields a molecular description of a stable steady state that is common across all tumors. It also resolves molecular descriptions of unbalanced processes that are shared by several tumors, such as hyperactivated phosphoprotein signaling networks. Further, it resolves unbalanced processes that provide unique classifiers of tumor subgroups. The results of the theoretical interpretation are compared against those of statistical multivariate methods and are shown to provide a superior level of resolution for identifying unbalanced processes in GBM tumors. The identification of specific constraints for each GBM tumor suggests tumor-specific combination therapies that may reverse this imbalance

Intercellular signaling through secreted proteins induces free-energy gradient-directed cell movement

Controlling cell migration is important in tissue engineering and medicine. Cell motility depends on factors such as nutrient concentration gradients and soluble factor signaling. In particular, cell–cell signaling can depend on cell–cell separation distance and can influence cellular arrangements in bulk cultures. Here, we seek a physical-based approach, which identifies a potential governed by cell–cell signaling that induces a directed cell–cell motion. A single-cell barcode chip (SCBC) was used to experimentally interrogate secreted proteins in hundreds of isolated glioblastoma brain cancer cell pairs and to monitor their relative motions over time. We used these trajectories to identify a range of cell–cell separation distances where the signaling was most stable. We then used a thermodynamics-motivated analysis of secreted protein levels to characterize free-energy changes for different cell–cell distances. We show that glioblastoma cell–cell movement can be described as Brownian motion biased by cell–cell potential. To demonstrate that the free-energy potential as determined by the signaling is the driver of motion, we inhibited two proteins most involved in maintaining the free-energy gradient. Following inhibition, cell pairs showed an essentially random Brownian motion, similar to the case for untreated, isolated single cells

Glioblastoma cellular architectures are predicted through the characterization of two-cell interactions

To understand how pairwise cellular interactions influence cellular architectures, we measured the levels of functional proteins associated with EGF receptor (EGFR) signaling in pairs of U87EGFR variant III oncogene receptor cells (U87EGFRvIII) at varying cell separations. Using a thermodynamics-derived approach we analyzed the cell-separation dependence of the signaling stability, and identified that the stable steady state of EGFR signaling exists when two U87EGFRvIII cells are separated by 80–100 μm. This distance range was verified as the characteristic intercellular separation within bulk cell cultures. EGFR protein network signaling coordination for the U87EGFRvIII system was lowest at the stable state and most similar to isolated cell signaling. Measurements of cultures of less tumorigenic U87PTEN cells were then used to correctly predict that stable EGFR signaling occurs for those cells at smaller cell–cell separations. The intimate relationship between functional protein levels and cellular architectures explains the scattered nature of U87EGFRvIII cells relative to U87PTEN cells in glioblastoma multiforme tumors

The immunogenic radiation and new players in immunotherapy and targeted therapy for head and neck cancer

Although treatment modalities for head and neck cancer have evolved considerably over the past decades, survival rates have plateaued. The treatment options remained limited to definitive surgery, surgery followed by fractionated radiotherapy with optional chemotherapy, and a definitive combination of fractionated radiotherapy and chemotherapy. Lately, immunotherapy has been introduced as the fourth modality of treatment, mainly administered as a single checkpoint inhibitor for recurrent or metastatic disease. While other regimens and combinations of immunotherapy and targeted therapy are being tested in clinical trials, adapting the appropriate regimens to patients and predicting their outcomes have yet to reach the clinical setting. Radiotherapy is mainly regarded as a means to target cancer cells while minimizing the unwanted peripheral effect. Radiotherapy regimens and fractionation are designed to serve this purpose, while the systemic effect of radiation on the immune response is rarely considered a factor while designing treatment. To bridge this gap, this review will highlight the effect of radiotherapy on the tumor microenvironment locally, and the immune response systemically. We will review the methodology to identify potential targets for therapy in the tumor microenvironment and the scientific basis for combining targeted therapy and radiotherapy. We will describe a current experience in preclinical models to test these combinations and propose how challenges in this realm may be faced. We will review new players in targeted therapy and their utilization to drive immunogenic response against head and neck cancer. We will outline the factors contributing to head and neck cancer heterogeneity and their effect on the response to radiotherapy. We will review in-silico methods to decipher intertumoral and intratumoral heterogeneity and how these algorithms can predict treatment outcomes. We propose that (a) the sequence of surgery, radiotherapy, chemotherapy, and targeted therapy should be designed not only to annul cancer directly, but to prime the immune response. (b) Fractionation of radiotherapy and the extent of the irradiated field should facilitate systemic immunity to develop. (c) New players in targeted therapy should be evaluated in translational studies toward clinical trials. (d) Head and neck cancer treatment should be personalized according to patients and tumor-specific factors

Optimization of Energy-Consuming Pathways towards Rapid Growth in HPV-Transformed Cells

Cancer is a complex, multi-step process characterized by misregulated signal transduction and altered metabolism. Cancer cells divide faster than normal cells and their growth rates have been reported to correlate with increased metabolic flux during cell transformation. Here we report on progressive changes in essential elements of the biochemical network, in an in vitro model of transformation, consisting of primary human keratinocytes, human keratinocytes immortalized by human papillomavirus 16 (HPV16) and passaged repeatedly in vitro, and the extensively-passaged cells subsequently treated with the carcinogen benzo[a]pyrene. We monitored changes in cell growth, cell size and energy metabolism. The more transformed cells were smaller and divided faster, but the cellular energy flux was unchanged. During cell transformation the protein synthesis network contracted, as shown by the reduction in key cap-dependent translation factors. Moreover, there was a progressive shift towards internal ribosome entry site (IRES)-dependent translation. The switch from cap to IRES-dependent translation correlated with progressive activation of c-Src, an activator of AMP-activated protein kinase (AMPK), which controls energy-consuming processes, including protein translation. As cellular protein synthesis is a major energy-consuming process, we propose that the reduction in cell size and protein amount provide energy required for cell survival and proliferation. The cap to IRES-dependent switch seems to be part of a gradual optimization of energy-consuming mechanisms that redirects cellular processes to enhance cell growth, in the course of transformation

Anomalous Features of EMT during Keratinocyte Transformation

During the evolution of epithelial cancers, cells often lose their characteristic features and acquire a mesenchymal phenotype, in a process known as epithelial-mesenchymal transition (EMT). In the present study we followed early stages of keratinocyte transformation by HPV16, and observed diverse cellular changes, associated with EMT. We compared primary keratinocytes with early and late passages of HF1 cells, a cell line of HPV16-transformed keratinocytes. We have previously shown that during the progression from the normal cells to early HF1 cells, immortalization is acquired, while in the progression to late HF1, cells become anchorage independent. We show here that during the transition from the normal state to late HF1 cells, there is a progressive reduction in cytokeratin expression, desmosome formation, adherens junctions and focal adhesions, ultimately leading to poorly adhesive phenotype, which is associated with anchorage-independence. Surprisingly, unlike “conventional EMT”, these changes are associated with reduced Rac1-dependent cell migration. We monitored reduced Rac1-dependent migration also in the cervical cancer cell line SiHa. Therefore we can conclude that up to the stage of tumor formation migratory activity is eliminated

Understanding Glioblastoma Signaling, Heterogeneity, Invasiveness, and Drug Delivery Barriers

The most prevalent and aggressive type of brain cancer, namely, glioblastoma (GBM), is characterized by intra- and inter-tumor heterogeneity and strong spreading capacity, which makes treatment ineffective. A true therapeutic answer is still in its infancy despite various studies that have made significant progress toward understanding the mechanisms behind GBM recurrence and its resistance. The primary causes of GBM recurrence are attributed to the heterogeneity and diffusive nature; therefore, monitoring the tumor’s heterogeneity and spreading may offer a set of therapeutic targets that could improve the clinical management of GBM and prevent tumor relapse. Additionally, the blood–brain barrier (BBB)-related poor drug delivery that prevents effective drug concentrations within the tumor is discussed. With a primary emphasis on signaling heterogeneity, tumor infiltration, and computational modeling of GBM, this review covers typical therapeutic difficulties and factors contributing to drug resistance development and discusses potential therapeutic approaches

Information-theoretic analysis of phenotype changes in early stages of carcinogenesis

Cancer is a multistep process characterized by altered signal transduction, cell growth, and metabolism. To identify such processes in early carcinogenesis we use an information theoretic approach to characterize gene expression quantified as mRNA levels in primary keratinocytes (K) and human papillomavirus 16 (HPV16)-transformed keratinocytes (HF1 cells) from early (E) and late (L) passages and from benzo(a)pyrene-treated (BP) L cells. Our starting point is that biological signaling processes are subjected to the same quantitative laws as inanimate, nonequilibrium chemical systems. Environmental and genomic constraints thereby limit the maximal thermodynamic entropy that the biological system can reach. The procedure uncovers the changes in gene expression patterns in different networks and defines the significance of each altered network in the establishment of a particular phenotype. The development of transformed HF1 cells is shown to be represented by one major transcription pattern that is important at all times. Two minor transcription patterns are also identified, one that contributes at early times and a distinguishably different pattern that contributes at later times. All three transcription patterns defined by our analysis were validated by gene expression values and biochemical means. The major transcription pattern includes reduced transcripts participating in the apoptotic network and enhanced transcripts participating in cell cycle, glycolysis, and oxidative phosphorylation. The two minor patterns identify genes that are mainly involved in lipid or carbohydrate metabolism