Engineered T cell therapies.

Alongside advancements in gene therapy for inherited immune disorders, the need for effective alternative therapeutic options for other conditions has resulted in an expansion in the field of research for T cell gene therapy. T cells are easily obtained and can be induced to divide robustly ex vivo, a characteristic that allows them to be highly permissible to viral vector-mediated introduction of transgenes. Pioneering clinical trials targeting cancers and infectious diseases have provided safety and feasibility data and important information about persistence of engineered cells in vivo. Here, we review clinical experiences with γ-retroviral and lentiviral vectors and consider the potential of integrating transposon-based vectors as well as specific genome editing with designer nucleases in engineered T cell therapies

Genome-Edited T Cell Therapies

PURPOSE OF REVIEW: Alternative approaches to conventional drug-based cancer treatments have seen T cell therapies deployed more widely over the last decade. This is largely due to their ability to target and kill specific cell types based on receptor recognition. Introduction of recombinant T cell receptors (TCRs) using viral vectors and HLA-independent T cell therapies using chimeric antigen receptors (CARs) are discussed. This article reviews the tools used for genome editing, with particular emphasis on the applications of site-specific DNA nuclease mediated editing for T cell therapies. RECENT FINDINGS: Genetic engineering of T cells using TCRs and CARs with redirected antigen-targeting specificity has resulted in clinical success of several immunotherapies. In conjunction, the application of genome editing technologies has resulted in the generation of HLA-independent universal T cells for allogeneic transplantation, improved T cell sustainability through knockout of the checkpoint inhibitor, programmed cell death protein-1 (PD-1), and has shown efficacy as an antiviral therapy through direct targeting of viral genomic sequences and entry receptors. SUMMARY: The combined use of engineered antigen-targeting moieties and innovative genome editing technologies have recently shown success in a small number of clinical trials targeting HIV and hematological malignancies and are now being incorporated into existing strategies for other immunotherapies

Ex vivo gene modification therapy for genetic skin diseases—recent advances in gene modification technologies and delivery

Genetic skin diseases, also known as genodermatoses, are inherited disorders affecting skin and constitute a large and heterogeneous group of diseases. While genodermatoses are rare with the prevalence rate of less than 1 in 50,000 - 200,000, they frequently occur at birth or early in life and are generally chronic, severe, and could be life-threatening. The quality of life of patients and their families are severely compromised by the negative psychosocial impact of disease, physical manifestations, and the lack or loss of autonomy. Currently, there are no curative treatments for these conditions. Ex-vivo gene modification therapy that involves modification or correction of mutant genes in patients' cells in-vitro and then transplanted back to patients to restore functional gene expression has being developed for genodermatoses. In this review, the ex-vivo gene modification therapy strategies for genodermatoses are reviewed, focusing on current advances in gene modification and correction in patients' cells and delivery of genetically modified cells to patients with discussions on gene therapy trials which have been performed in this area

Association between polymorphism of the CRP gene rs1130864 and hypertension among Iraqi hypertensive patients

Background: C-reactive protein (CRP) a reactant of the acute phase and a measure of persistent low-grade inflammatory processes. Objectives of this paper to the role of rs1130864 polymorphism in CRP gene among Iraqi hypertensive patients.Methods: Samples of blood were collected from patients with hypertension and controls, their age (37 to 75). Hardy-Weinberg equilibrium (HWE) was used to calculate the risk score for the disease of hypertension, Moreover, PCR/SNP (specific primers) was used to do genotyping.Results: HWE analysis of hypertensive patients and controls showed that the rs1130864 genotypes were in agreement with the equilibrium, without differences clear between the observed and expected genotype frequencies (p >0.153, 0.238). The frequencies of genotype and alleles were compared in hypertensive patients with controls, with no significant differences. The common GG genotype of rs1130864 recorded a significant increase in hypertensive patients and controls and was regarded as a preventative fraction (RR = 0.76), whilst, GA , AA genotype were considered the etiological fraction (RR = 1.06, 2.59) and associated with hypertension. Compared to A allele, which might be the cause of the disease, G allele might be preventive.Conclusion: The findings showed that the Iraqi population's GA, AA genotype, and A allele are risk factors for hypertension. However, the need for additional findings utilizing bigger samples is needed are necessary to verify our findings.Keywords: Hypertension; C-reactive protein; Polymorphism, 1444G>A; rs1130864   

Web readability factors affecting users of all ages

With the increase in the growth of internet since 1990 WorldWide Web have gained a worldwide popularity.Web has become an ultimate source of information throughout the world. The web has been a source of information, knowledge, entertainment and carrying out different tasks such as online shopping, reservation etc. Every one finds ways to the web due to easiness of access and the bulk of information that has been available so ways to make web more attractive and easy to interact with has been an issue under discussion since its inception. In this paper we have focused how to make a webpage more usable for different age group in terms of readability. We have focused on eight eternal readability factors i.e. color contrast, white space, line spacing, font style, font size, text width, headings, graphics and animation. These eight factors are compared that how different age group people behave with the web applications by varying these eight factors. Based on literature review we have proposed an idea that how to make web more readable for people of different age groups e.g. children, teenagers and old /senior users

Process development of human mesenchymal stem cell microcarrier culture using an automated high-throughput microbioreactor

Improvements to process development technology will have a significant impact in reducing the overall costs associated with the manufacture and scale-up of human cell-based therapies. Small-scale models, including microbioreactors, play a critical role in this regard as they reduce reagent requirements and can facilitate high-throughput screening of process parameters and culture conditions. Here we have demonstrated, for the first time, the amenability of the automated ambr15 cell culture microbioreactor system (originally designed for free suspension culture) for adherent hMSC microcarrier culture. We also demonstrated that the ambr15 could be used for bioprocess development of a microcarrier process which was subsequently validated with larger-scale spinner flask studies. The results were achieved by a combination of strategies including adapting the free suspension design of the vessel to improve the suspension and mixing of the microcarriers. A more effective cell attachment method was also developed by using only 50% of the final working volume of medium for the first 24 h combined with an intermittent agitation strategy. These improvements led to a reduction in the initial lag phase which in turn resulted in \u3e 150 % increase in viable cell density after 24 h compared to the original process (no agitation for 24 h and 100 % working volume). Using the same methodology as in the ambr 15, similar improvements were obtained in larger scale spinner flask studies. Finally, this improved bioprocess methodology, which was developed for a serum-based medium process, was applied to a serum-free process in the ambr15; this resulted in \u3e 250% increase in yield compared to the ambr15 serum-based process. The use of the ambr15, with its improved control compared to the spinner flask, reduced the coefficient of variation on viable cell density in the serum containing medium from 7.65% to 4.08%, and the switch to the serum free medium further reduced these to 1.06% and 0.54% respectively. The combination of both serum-free and automated processing improved the consistency more than 10-fold compared to the initial manual, serum-based spinner flask work. The findings of this study demonstrate that the ambr15 microbioreactor is an effective tool for bioprocess development of hMSC microcarrier cultures and that a combination of serum-free medium and automation improves both process yield and consistency. Please click Additional Files below to see the full abstract

Progress and prospects for engineered T cell therapies

Proof-of-concept studies have demonstrated the therapeutic potential of engineered T cells. Transfer of recombinant antigen-specific T cell receptors (TCR) and chimaeric antigen receptors (CARs) against tumour and viral antigens are under investigation by multiple approaches, including viral- and nonviral-mediated gene transfer into both autologous and allogeneic T cell populations. There have been notable successes recently using viral vector-mediated transfer of CARs specific for B cell antigens, but also reports of anticipated and unanticipated complications in these and other studies. We review progress in this promising area of cellular therapy, and consider developments in antigen receptor therapies including the application of emerging gene-editing technologies

A potentially scalable method for the harvesting of hMSCs from microcarriers

The use of hMSCs for allogeneic therapies requiring lot sizes of billions of cells will necessitate large-scale culture techniques such as the expansion of cells on microcarriers in bioreactors. Whilst much research investigating hMSC culture on microcarriers has focused on growth, much less involves their harvesting for passaging or as a step towards cryopreservation and storage. A successful new harvesting method has recently been outlined for cells grown on SoloHill microcarriers in a 5L bioreactor [1]. Here, this new method is set out in detail, harvesting being defined as a two-step process involving cell 'detachment' from the microcarriers' surface followed by the 'separation' of the two entities. The new detachment method is based on theoretical concepts originally developed for secondary nucleation due to agitation. Based on this theory, it is suggested that a short period (here 7min) of intense agitation in the presence of a suitable enzyme should detach the cells from the relatively large microcarriers. In addition, once detached, the cells should not be damaged because they are smaller than the Kolmogorov microscale. Detachment was then successfully achieved for hMSCs from two different donors using microcarrier/cell suspensions up to 100mL in a spinner flask. In both cases, harvesting was completed by separating cells from microcarriers using a Steriflip® vacuum filter. The overall harvesting efficiency was >95% and after harvesting, the cells maintained all the attributes expected of hMSC cells. The underlying theoretical concepts suggest that the method is scalable and this aspect is discussed too