English
Abstract
Extensive studies about the human immunodeficiency virus type 1 (HIV-1) have allowed the generation of lentiviral vectors as gene delivery vehicles with enhanced safety and efficacy features. In this review, several strategies for controlling the molecular mechanisms occurring during the lentiviral vector manufacturing process are presented. Specifically, modifications focused on LVV manufacturing components, such as plasmids or the producer cell line, that enable increased safety, integrity, and potency of the produced LVV, as well as manufacturing efficiency. Considering the stochasticity of the LVV manufacturing process from plasmid transfection until the budding of the virus from the target cell, minimal modifications might have a huge impact on the final LVV yield. Indeed, the extent of a potential impact may vary depending on the specificities of each LVV regarding the particular genetic payload or the envelope protein. Thus, the feasibility of each of the optimizations described herein requires thorough evaluation. The second part of the review examines the potential multi-purpose nature of the LVV. Growing research in the field has enabled the development of new engineered modalities of LVV, expanding their application scope beyond the traditional ex vivo DNA delivery approach. LVVs are becoming a versatile tool for the packaging or delivery of cargo in the form of DNA, RNA, or protein, allowing their use for in vivo approaches, vaccinology, or gene editing, among others.
Keywords:
lentiviral vectors; HIV-1; LVV manufacturing; CAR-T; in vivo gene therapy; ex vivo gene therapy; RNA delivery; vaccinology
1. Introduction
1.1. Overview of Lentiviruses Origin
Lentiviruses, a genus within the Retroviridae family, include human immunodeficiency virus type 1 (HIV-1) as their most well-characterized member. Current lentiviral vectors (LVVs) have primarily been derived from HIV-1 through extensive research and molecular engineering for maximized safety without compromising functionality [1]. HIV-1 is characterized by its genome (Figure 1), which comprises two copies of single-stranded, positive-sense RNA [2]. Each strand is flanked by long terminal repeats (LTRs) at both ends, which are essential for transcription, reverse transcription, and integration of the viral genome. Each LTR contains three elements: U3, R, and U5. The U3 element acts as a promoter/enhancer in the 3′LTR, while the R element serves as a polyadenylation signal in the 5′LTR. Thus, provirus mRNA is devoid of 5′ U3 and 3′ U5, resulting in a viral RNA flanked by R elements [3]. In addition, the U5 element in the 5′ end is part of a complex secondary structure that modulates key steps like packaging or reverse transcription [4]. The genome encodes nine proteins through nine Open Reading Frames (ORFs), some of which are proteolytically processed to generate additional proteins. Among these, the two primary proteins are the Gag/Pol polyprotein and the Env glycoprotein. Gag expresses the viral core proteins, which are matrix protein (MA or p17), capsid (CA or p24), nucleocapsid (NC or p7), and smaller core proteins p6, p1, and p2. Pol translates into the viral enzymes protease (PR) for proteolytic processing of Gag and Gag-Pol polyproteins, the reverse transcriptase (RT), which has both DNA polymerase and RNase H activity, and the integrase (IN) for viral genome integration into the host cell. In addition, the Env gene encodes for the viral surface glycoprotein (SU or gp120) as well as the transmembrane glycoprotein (TM or gp41). Additionally, the viral genome encodes the regulatory protein Tat (trans-activator of transcription) for transcription activation and Rev (Regulator of Expression of Virion proteins) for splicing control and nuclear export. The remaining genes are responsible for encoding the accessory proteins Vif (Virion Infectivity Factor), Vpr (Viral Protein R), Vpu (Viral Protein U), and Nef (Negative Factor) [3,5].
The article is reprinted from MDPI, original link:https://www.mdpi.com/1422-0067/26/17/8497
The FAI climbed 5.9 percent year-on-year in the first 11 months of 2018, quickening from the 5.7-percent growth in Jan-Oct, the National Bureau of Statistics (NBS) said Friday in an online statement.
The key indicator of investment, dubbed a major growth driver, hit the bottom in August and has since started to rebound steadily.
In the face of emerging economic challenges home and abroad, China has stepped up efforts to stabilize investment, in particular rolling out measures to motivate private investors and channel funds into infrastructure.
Friday's data showed private investment, accounting for more than 60 percent of the total FAI, expanded by a brisk 8.7 percent.
NBS spokesperson Mao Shengyong said funds into weak economic links registered rapid increases as investment in environmental protection and agriculture jumped 42 percent and 12.5 percent respectively, much faster than the average.
In breakdown, investment in high-tech and equipment manufacturing remained vigorous with 16.1-percent and 11.6-percent increases respectively in the first 11 months. Infrastructure investment gained 3.7 percent, staying flat. Investment in property development rose 9.7 percent, also unchanged.