Immortality Research Articles

Disruption of the β1L Isoform of GABP Reverses Glioblastoma Replicative Immortality in a TERT Promoter Mutation-Dependent Manner

TERT promoter mutations reactivate telomerase, allowing for indefinite telomere maintenance and enabling cellular immortalization. These mutations specifically recruit the multimeric ETS factor GABP, which can form two functionally independent transcription factor species: a dimer or a tetramer. We show that genetic disruption of GABPβ1L (β1L), a tetramer-forming isoform of GABP that is dispensable for normal development, results in TERT silencing in a TERT promoter mutation-dependent manner. Reducing TERT expression by disrupting β1L culminates in telomere loss and cell death exclusively in TERT promoter mutant cells. Orthotopic xenografting of β1L-reduced, TERT promoter mutant glioblastoma cells rendered lower tumor burden and longer overall survival in mice. These results highlight the critical role of GABPβ1L in enabling immortality in TERT promoter mutant glioblastoma.

Precision Targeting of BFL-1/A1 and an ATM Co-dependency in Human Cancer

Cancer cells overexpress a diversity of anti-apoptotic BCL-2 family proteins, such as BCL-2, MCL-1, and BFL-1/A1, to enforce cellular immortality. Thus, intensive drug development efforts have focused on targeting this class of oncogenic proteins to overcome treatment resistance. Whereas a selective BCL-2 inhibitor has been FDA approved and several small molecule inhibitors of MCL-1 have recently entered phase I clinical testing, BFL-1/A1 remains undrugged. Here, we developed a series of stapled peptide design principles to engineer a functionally selective and cell-permeable BFL-1/A1 inhibitor that is specifically cytotoxic to BFL-1/A1-dependent human cancer cells. Because cancers harbor a diversity of resistance mechanisms and typically require multi-agent treatment, we further investigated BFL-1/A1 co-dependencies by mining a genome-scale CRISPR-Cas9 screen. We identified ataxia-telangiectasia-mutated (ATM) kinase as a BFL-1/A1 co-dependency in acute myeloid leukemia (AML), which informed the validation of BFL-1/A1 and ATM inhibitor co-treatment as a synergistic approach to subverting apoptotic resistance incancer.

Leibniz and the Problem of the Immortality of the Soul

Leibniz and the Problem of the Immortality of the Soul

The Responsibility of Immortality: Welcome to the New Transhumanism

The Responsibility of Immortality: Welcome to the New Transhumanism

Abstract 3078: The fer/FerT cell survival pathway of the ancestral sponge (Sycon raphanus) becomes a mitochondrial proto-oncogene in the human genome

Abstract The Fer/FerT (truncated) genes and their encoded 879 aa / 100 kDa proteins (fujinami poultry sarcoma; feline sarcoma-related kinases) were discovered in the sponge Sycon raphanus [1]. The recognition of other related genes and their protein products (ras/RAS; src/SRC; the human Bruton kinase-related Tec family proteins, etc) in the descendants of ancestral sponges (Suberites; Spongilla) rapidly followed [1]. Primordial cell survival pathways were essential for the life style of early diploblastic multicellular eukaryotes (ctenophores; cnidarians; ciona, etc) in the hostile environment of the ancient Earth [2]. These preserved ancient cell survival pathways evolved into proto-oncogenes in triploblastic hosts (Homo), in order to allow switching back to them under circumstances of life-threatening intrusions, and in chronic infections of the inflammasomes [2]. The Author refers to this inherent fundamental biological principle as “retrograde cellular immortalization” [2]. A common form of this process is recognized in medical clinics as 'cancer' [2]. Constitutively up-regulated Fer/FerT genes and proteins redirect sugar metabolism toward Warburg's aerobic glycolysis with reduced Oxphosph (for ATP production) and tricarboxylic acid cycles. Downregulation of Fer/FerT results in a non-functional ETC. Tumor cells with nonfunctional mitochondrial ETC undergo either mTOR-directed autophagy, and/or necrotic death promoted by poly ADP-ribose polymerase (PARP-1) [3]. The novel inhibitor E260 of enzyme-like activity to Fer/FerT-induced kinases, attacks selectively mitochondrial (or cytoplasmic) Fer/FerT proteins. E260-treated cancer cells lose their ATP content and increase their glycolytic activity (especially in high glucose concentration), thus delaying (but not canceling) their death. These cancer cells operate with low hexokinase II levels and deregulated PARP-1. ATP-deficient cells activate AMPK, de-activate mTOR, and fail to recover from the state of autophagy, resulting in necrosis. So far, all healthy cells remained intact upon exposure to E260 [4]. Recalled ancient cell survival pathways operate as "protooncogenes-to-oncoproteins" in extant triploblastic hosts [2].

Abstract 1901: Structural basis for selective therapeutic targeting of BFL-1 in cancer by a covalent stapled peptide inhibitor

Abstract BCL-2 family proteins are master regulators of the mitochondrial apoptotic pathway and dictate whether a cell will live or die in response to stress. Cancer cells hijack the anti-apoptotic arm of this signaling network, overexpressing BCL-2, MCL-1, and BFL-1, to enforce cellular immortality. Whereas BCL-2 has been drugged by the selective small-molecule inhibitor, venetoclax, and new MCL-1 inhibitor molecules have just entered phase I clinical testing, anti-apoptotic BFL-1 remains undrugged. Importantly, the expression of BFL-1 can cause resistance to these molecules and has been independently linked to the pathogenesis and chemoresistance of human cancers, including melanomas and hematologic malignancies. Given the emergence of BCL-2 family proteins as high-priority cancer targets, solving their structures, alone and in complex with their natural BH3 domain ligands, yields important blueprints for drug design. Whereas the structures of numerous “BH3-in-groove” complexes have been solved, the corresponding panel of apo structures remains incomplete. Here, we report the crystal structure of apo BFL-1 at 1.69 Å resolution, revealing similarities and key differences among unliganded anti-apoptotic targets. Unlike all other BCL-2 proteins, apo BFL-1 contains a surface-accessible cysteine within its BH3-binding groove, allowing for selective covalent targeting by BH3-based stapled peptide inhibitors. The crystal structure of a complex between BFL-1 and a cysteine-reactive and stapled NOXA BH3 α-helix demonstrated the sulfhydryl bond and fortuitous interactions between the acrylamide-bearing moiety and a newly formed hydrophobic cavity. Comparison of the apo and BH3-liganded structures further revealed an induced conformational change. The two BFL-1 structures expand our understanding of the surface landscapes available for selective targeting of BFL-1, and inform a new therapeutic strategy for overcoming BFL-1-dependent apoptotic blockades in human cancer. Citation Format: Edward P. Harvey, Hyuk-Soo Seo, Rachel M. Guerra, Gregory H. Bird, Sirano Dhe-Paganon, Loren D. Walensky. Structural basis for selective therapeutic targeting of BFL-1 in cancer by a covalent stapled peptide inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1901.

PO-389 Epigenomic and mutation determinants of carcinogen-driven primary epithelial cell immortalization

IntroductionBased on cancer genome sequencing, almost 600 genes have been implicated in cancer development to date, and recently, chromatin structural changes have emerged as a key mechanism for translating exogenous cues, such as carcinogen exposures, to the deregulation of biological processes implicated in tumorigenesis. However, the functional relationship between genetic and epigenetic alterations in the context of early events of carcinogen-driven cell transformation is not well understood.Material and methodsWe employ an in vitro carcinogen exposure system using primary human mammary epithelial cells (HMEC), which utilises a biological barrier bypass to mimic early steps of cell transformation, to introduce genetic and epigenetic alterations and to investigate their functional interplay during cell immortalization. A temporal, integrated (epi)genomic analysis is applied to benzo[a]pyrene (B[a]P)-exposed, immortalised HMEC, involving whole genome sequencing (WGS), analyses of chromatin structure (ChIP-seq, ATAC-seq), and patterns of gene regulation (DNA methylome 850K array, RNA-seq).Results and discussionsWGS of sequentially derived post-stasis and immortalised clones, as well as tumorigenic clones (generated by TP53 insertional mutagenesis of post-stasis lines), identified a prominent B[a]P mutational signature across all clones. While all sequenced clones had comparable SBS counts, we observed a gradual increase of structural variants, most notably copy number variations, from post-stasis to immortal and tumorigenic clones. Reproducibly distinct methylome patterns also correlated closely with the individual phenotypic stages of cell transformation. Chromatin accessibility analysis further showed changes characteristic to individual as well as multiple exposed clones, such as in the breast cancer tumour suppressor gene BRCA2 or the promoter region of PCDH18, a gene frequently inactivated by promoter methylation in colorectal cancer. Cell type-matched histone modification and RNA expression patterns are being established to enable a thorough integrated analysis of genetic and epigenetic determinants of carcinogen-driven HMEC transformation.ConclusionWe propose that in-depth, integrated (epi)genomic analysis of this innovative model system can reveal novel molecular determinants of carcinogen-driven cellular immortalization, thus improving our understanding of cancer causes, development and progression.Funding INCa-INSERM Plan Cancer 2015 Grant.

Susceptibility of epithelial cells cultured from different regions of human cervix to HPV16-induced immortalization.

Persistent infection with high-risk human papillomavirus (HPV) is a major risk factor for cervical cancer. Greater than 90% of these cancers originate in the cervical transformation zone (TZ), a narrow region of metaplastic squamous epithelium that develops at the squamocolumnar junction between the ectocervix and endocervix. It is unclear why the TZ has high susceptibility to malignant transformation and few studies have specifically examined cells from this region. We hypothesized that cells cultured from TZ are more susceptible to cellular immortalization, an alteration that contributes to malignant development. We cultured primary epithelial cells from each region of human cervix (ectocervix, endocervix and TZ) and measured susceptibility to immortalization after transfection with the complete HPV-16 genome or infection of HPV16 E6/E7 retroviruses. Cells cultured from each cervical region expressed keratin markers (keratin 14 and 18) that confirmed their region of origin. In contrast to our prediction, cells from TZ were equally susceptible to immortalization as cells from ectocervix or endocervix. Thus, increased susceptibility of the TZ to cervical carcinogenesis is not due to increased frequency of immortalization by HPV-16. We developed a series of HPV16-immortalized cell lines from ectocervix, endocervix and TZ that will enable comparisons of how these cells respond to factors that promote cervical carcinogenesis.

Survival studies: competing risks, immortality and censoring.

Survival studies: competing risks, immortality and censoring.

Administration of a Nucleoside Analog Promotes Cancer Cell Death in a Telomerase-Dependent Manner

Telomerase, the end-replication enzyme, is reactivated in malignant cancers to drive cellular immortality. While this distinction makes telomerase an attractive target for anti-cancer therapies, most approaches for inhibiting its activity have been clinically ineffective. As opposed to inhibiting telomerase, we use its activity to selectively promote cytotoxicity incancer cells. We show that several nucleotide analogs, including 5-fluoro-2'-deoxyuridine (5-FdU) triphosphate, are effectively incorporated by telomerase into a telomere DNA product. Administration of 5-FdU results in an increased number of telomere-induced foci, impedes binding of telomere proteins, activates the ATR-related DNA-damage response, and promotes cell death in a telomerase-dependent manner. Collectively, our data indicate that telomerase activity can be exploited as a putative anti-cancer strategy.

Verdad, Vida y Camino del bodisatva: lectura inter-confesional del Sutra del Loto

Tres ideas básicas del Sutra del Loto son: el Vehículo Único hacia la Verdad Última; la Vida Eterna del Buda Originario; y el Camino de la Compasión Universal de los Bodisatvas. Dejando para los estudios históricos la tarea de interpretación filológica y doctrinal de este texto clásico religioso, aquí intentaremos presentar la posibilidad de una lectura que lo descontextualice y recontextualice para destacarlo que nos hace pensar al leerlo en el marco de un diálogo intercultural e interconfesional. Tanto una filosofía o espiritualidad no teista como otra teista, sin renunciar cada una a su identidad, podrían encontrarse al enfocar algunos aspectos principales del Sutra del Loto como, por ejemplo, la verdad unificadora del universo y el auténtico aspecto de todas las realidades; la vida eterna del Buda Shakamuni y su presencia en la interioridad humana como naturaleza búdica o capacidad de alcanzar la iluminación; y en la acción compasiva universal de los bodisatvas.

Teaching as an Immortality Project: Positing Weakness in Response to Terror

Teaching as an Immortality Project: Positing Weakness in Response to Terror

A VHCF life prediction method based on surface crack density for FRP

In order to solve the difficulty of predicting very high cycle fatigue (VHCF) life of fiber reinforced plastic (FRP), a prediction method based on surface crack density is presented in this paper. According to the general rule of FRP fatigue damage evolution, the crack density is used to characterize the damage state of FRP in this method and further, the crack density-life evolution model of matrix can be established. The VHCF life prediction can be accomplished after obtaining the parameters in the evolution model through multi-load fatigue tests. The threshold of FRP fatigue strain is also assumed to exist in this paper. If the service strain level is lower than this threshold value, no new damage will occur after the matrix crack density reaches the characteristic damage state (CDS), which implies that FPR bearing below-threshold loads possesses an infinite life expectancy. In this paper, the very high cycle fatigue test of single-layer orthotropic woven CFRP has been carried out. The matrix crack density-life curve and the strain threshold of fatigue have been obtained by providing the rationality of this method.

Survival of a Nation and Duality in Narrative - Focusing on 2009 of the Series Guide of Immortality

Survival of a Nation and Duality in Narrative - Focusing on 2009 of the Series Guide of Immortality

Lightweight design of gears in the wheel-side reducer based on Shanghai road driving cycle

The development of wheel-side reducer with proper durability is important for electric vehicles equipped with a wheel-side reducer system. Based on the Shanghai road driving cycle, a novel conversion method is proposed from vehicle speed to load, which provides a theoretical foundation for lightweight design and a durability analysis of the key components of the wheel-side reducer system. With the gears of the wheel-side reducer as an example, an electric drive system durability test specification is also developed on the basis of the Miner linear cumulative damage theory considering the material strengthening under low-amplitude loads. The durability test shows that the second round of the prototype design no longer has an infinite life compared with the first round of the prototype design. The method based on active load spectrum is more effective than the traditional design method.

Fatigue life prediction and stress distribution of straight and angled abutments using numerical techniques

The durability of an implant-bone structure depends mainly on the capability of the implants and bone-implant interface to support masticatory forces. The mechanical behavior of both bone and implant varies from one configuration to another and thus, it is still an attractive topic for many researchers in dental biomechanics. This paper investigated stress distribution in four different applications/configurations of commercial implants by employing three dimensional finite element analyses. Fatigue analysis was also conducted to estimate the assembly’s life-cycles. The results indicated that the application of a straight abutment is more suitable for a situation where the implant is placed in an ideal position, perpendicular to the occlusal plane. Whereas an angled abutment may be implanted for conditions in which the implant is not placed in a perpendicular position to the occlusal plane. The results were found to be in agreement with the experimental data available in the literature. Although different clinical conditions, using angled abutments, may lead to different severities and regions of crack initiation, approximately infinite life was predicted for all analyzed cases. The abutment geometry whether straight or angled and the implant position, do not have a significant effect on the assembly’s life.

On notch and crack size effects in fatigue, Paris’ law and implications for Wöhler curves

As often done in design practice, the Wohler curve of a specimen, in the absence of more direct information, can be crudely retrieved by interpolating with a power-law curve between static strength at a given conventional low number of cycles N0 (of the order of 10-103), and the fatigue limit at a “infinite life”, also conventional, typically N∞=2·106 or N∞=107 cycles. These assumptions introduce some uncertainty, but otherwise both the static regime and the infinite life are relatively well known. Specifically, by elaborating on recent unified treatments of notch and crack effects on infinite life, and using similar concepts to the static failure cases, an interpolation procedure is suggested for the finite life region. Considering two ratios, i.e. toughness to fatigue threshold FK=KIc/DKth, and static strength to endurance limit, FR =sR /Ds0, qualitative trends are obtained for the finite life region. Paris’ and Wohler’s coefficients fundamentally depend on these two ratios, which can be also defined “sensitivities” of materials to fatigue when cracked and uncracked, respectively: higher sensitivity means stringent need for design for fatigue. A generalized Wohler coefficient, k’, is found as a function of the intrinsic Wohler coefficient k of the material and the size of the crack or notch. We find that for a notched structure, k<k’<m, as a function of size of the notch: in particular, k’=k holds for small notches, then k’ decreases up to a limiting value (which depends upon Kt for mildly notched structures, or on FK and FR only for severe notch or crack). A perhaps surprising return to the original slope k is found for very large blunt notches. Finally, Paris’ law should hold for a distinctly cracked structure, i.e. having a long-crack; indeed, Paris’ coefficient m is coincident with the limiting value of k’lim. The scope of this note is only qualitative and aims at a discussion over unified treatments in fatigue.

An Improved Method for Valuing Mature Companies and Estimating Terminal Value

The theory underlying Discounted cash flow (DCF) models is uncontroversial in academia, the economic intuition behind them is straightforward, and the mathematics reassuringly simple. Nevertheless, in practice, they are applied inconsistently with very different valuation results. Because of the assumed infinite life (“going concern”) of a business enterprise, DCF models implicitly assume the ability to forecast future cash flows forever. This forces analysts to make assumptions about the terminal period using simplistic metrics such as P/E or EV/EBITDA to estimate terminal values or to embed a perpetual stream of excess profitability and value creation in the terminal period.The author offers an uncomplicated alternative to these unrealistic assumptions. The first step is to introduce an adjustable fade rate called f. A fade rate of 100% brings about immediate convergence, and a fade rate of 0% specifies no fade and perpetual excess profitability. The notion that excess profits get competed away over time can be modeled by assuming that the spread of (ROIC ‐ r) fades to zero and that economic profit dissipates.Intrinsic value is very sensitive to the fade rate assumption and this helps explain the risk premium for quality stocks. The risk of owning quality stocks is that they lose their economic moat and competitive advantage. An adjustable fade rate provides an excellent means to value the effect of profitability attenuation in a DCF model.

A novel window opened: EBV-driven enhancer-promoter loops in lymphocytic immortalization

Epstein-Barr virus (EBV) is one of the human herpesvirus, which infects more than 90% of the global population (1). EBV infection causes the disorder of the immune system and has an essential role in the development of cancers (2).

Telomeres and Telomerase: The Switch of Tumor Genesis in Vivo

A line of evidence shows that telomere extension by telomerase leads to cellular immortalization that is characteristic of cancer cells. Recent findings revealed that oncogenes are activated through the inhibition of tumor suppressors and the activation of hTERT promoter following telomere extension. It is thus critical to prevent the activation of telomerase and telomere extension for the suppression of tumorigenesis. This review summarizes the structure of telomere and the mechanism of telomere extension and discusses the molecular targets for the development of novel cancer therapy.