Mutations can accumulate in the protease and gag genes of human immunodeficiency virus in patients who fail therapy with protease inhibitor drugs. Mutations within protease, the drug target, have been extensively studied. Mutations in gag have been less well studied, mostly concentrating on cleavage sites. A retroviral vector system has been adapted to study full-length gag, protease, and reverse transcriptase genes from patient-derived viruses. Patient plasma-derived mutant full-length gag, protease, and gag-protease from a multidrug-resistant virus were studied. Mutant protease alone led to a 95% drop in replication capacity that was completely rescued by coexpressing the full-length coevolved mutant gag gene. Cleavage site mutations have been shown to improve the replication capacity of mutated protease. Strikingly, in this study, the matrix region and part of the capsid region from the coevolved mutant gag gene were sufficient to achieve full recovery of replication capacity due to the mutant protease, without cleavage site mutations. The same region of gag from a second, unrelated, multidrug-resistant clinical isolate also rescued the replication capacity of the original mutant protease, suggesting a common mechanism that evolves with resistance to protease inhibitors. Mutant gag alone conferred reduced susceptibility to all protease inhibitors and acted synergistically when linked to mutant protease. The matrix region and partial capsid region of gag sufficient to rescue replication capacity also conferred resistance to protease inhibitors. Thus, the amino terminus of Gag has a previously unidentified and important function in protease inhibitor susceptibility and replication capacity.The development of antiretroviral drugs and their use in highly active antiretroviral therapy have led to the ability to effectively control human immunodeficiency virus (HIV) replication in infected patients. The emergence of resistance to these drugs, while reduced when the drugs are used in combination, remains a problem. A recent study has estimated that by 10 years of treatment, nearly 10% of patients will have experienced resistance to the three main classes of drugs that form the basis of highly active antiretroviral therapy. Further, the risk of death within 5 years after extensive triple-class failure is 10% (31). Resistance develops due to several factors, including the high error rate of reverse transcriptase (RT), viral recombination, high viral turnover, and suboptimal drug levels in patients. Mutations emerging in viruses can lead to cross-class resistance.Resistance to protease inhibitors (PI) occurs by the stepwise accumulation of mutations in protease itself, many of which lead to a reduced replication capacity (RC) of the virus (5-7, 22, 23, 37). Primary, or major, mutations mostly lead to a reduced affinity for the inhibitor through alterations in the enzyme active site. Further secondary, or minor, mutations also arise in protease that have little effect on inhibitor binding and therefore do not convey res...

These findings further support the concept of retroviral involvement in amyotrophic lateral sclerosis (ALS) and demonstrate that serum is more suitable than CSF for assay of reverse transcriptase (RT) activity in this disease. The levels of serum RT activity detected are comparable to those found in HIV infection. XMRV is not detectable in the blood of ALS patients, and the agent responsible for ALS-associated RT activity therefore remains unidentified.

Our findings demonstrate the importance of considering protease inhibitor susceptibility in the context of full-length gag, particularly with respect to the range of HIV-1 subtypes circulating worldwide.

SUMMARYThere is great potential for novel vaccines based on recombinant proteins and synthetic peptides. Unfortunately these antigens often lack the immunogenicity of whole, killed pathogens used in traditional vaccines. Thus there is strong interest in the identi®cation of immunological adjuvants with low reactogenicity, but high potency, to enhance immune responses and realize the potential of these new vaccine strategies. CD40 antibodies have been shown to have adjuvant effects when administered at very high doses. These large doses are impractical and induce a cascade of cytokine release giving rise to septic shock-like symptoms, as well as splenomegaly and polyclonal antibody production. We show here that a very small amount of CD40 antibody can exhibit potent adjuvant effects when attached to soluble antigen. The lack of detectable systemic effects indicates that this method may be a powerful and practical means of enhancing the ef®cacy of recombinant vaccines.

Over the past two decades a number of studies have demonstrated activity of the retroviral enzyme reverse transcriptase in the serum of patients with sporadic amyotrophic lateral sclerosis (ALS). Known human exogenous retroviruses such as HIV-1 have been eliminated as possible sources of this activity and investigators have therefore considered the possibility that human endogenous retroviruses (HERVs) might be involved. HERV-K (HML-2) is the most recent retroviral candidate to be proposed following the observation of elevated HERV-K expression in cortical and spinal neurons of ALS patients and the demonstration of HERV-K envelope protein neurotoxicity in vitro and in transgenic mice. This retroviral hypothesis is an attractive one, not least because it raises the possibility that ALS might become treatable using antiretroviral drugs. In the present study we have attempted independent confirmation of the observation that HERV-K RNA levels are elevated in ALS brain. Total RNA was extracted from the postmortem premotor cortex of 34 patients with ALS and 23 controls. Quantitative real-time reverse transcription PCR (RT-qPCR) was performed according to the MIQE guidelines using HERV-K gag, pol and env primer sets. Data was analysed by the 2-∆∆Ct method with normalisation against two reference genes, GAPDH and XPNPEP1. Geometric mean HERV-K RNA expression levels in the premotor cortex of ALS patients were not found to be different from the expression levels in non-ALS controls. Our findings do not confirm the recently reported association between elevated cortical HERV-K RNA levels and ALS, and thus raise doubts about the role of this endogenous retrovirus in ALS pathogenesis. The results of this study may have implications for ongoing clinical trials aiming to suppress HERV-K activity with antiretroviral drugs.Electronic supplementary materialThe online version of this article (10.1186/s40478-019-0698-2) contains supplementary material, which is available to authorized users.