Conditional Immune Escape during Chronic Simian Immunodeficiency Virus Infection.

UNLABELLED: Anti-HIV CD8 T cells included in therapeutic treatments will need to target epitopes that do not accumulate escape mutations. Identifying the epitopes that do not accumulate variants but retain immunogenicity depends on both host major histocompatibility complex (MHC) genetics and the likelihood for an epitope to tolerate variation. We previously found that immune escape during acute SIV infection is conditional; the accumulation of mutations in T cell epitopes is limited, and the rate of accumulation depends on the number of epitopes being targeted. We have now tested the hypothesis that conditional immune escape extends into chronic SIV infection and that epitopes with a preserved wild-type sequence have the potential to elicit epitope-specific CD8 T cells. We deep sequenced simian immunodeficiency virus (SIV) from Mauritian cynomolgus macaques (MCMs) that were homozygous and heterozygous for the M3 MHC haplotype and had been infected with SIV for about 1 year. When interrogating variation within individual epitopes restricted by M3 MHC alleles, we found three categories of epitopes, which we called categories A, B, and C. Category B epitopes readily accumulated variants in M3-homozygous MCMs, but this was less common in M3-heterozygous MCMs. We then determined that chronic CD8 T cells specific for these epitopes were more likely preserved in the M3-heterozygous MCMs than M3-homozygous MCMs. We provide evidence that epitopes known to escape from chronic CD8 T cell responses in animals that are homozygous for a set of MHC alleles are preserved and retain immunogenicity in a host that is heterozygous for the same MHC alleles. IMPORTANCE: Anti-HIV CD8 T cells that are part of therapeutic treatments will need to target epitopes that do not accumulate escape mutations. Defining these epitope sequences is a necessary precursor to designing approaches that enhance the functionality of CD8 T cells with the potential to control virus replication during chronic infection or after reactivation of latent virus. Using MHC-homozygous and -heterozygous Mauritian cynomolgus macaques, we have now obtained evidence that epitopes known to escape from chronic CD8 T cell responses in animals that are MHC homozygous are preserved and retain immunogenicity in a host that is heterozygous for the same MHC alleles. Importantly, our findings support the conditional immune escape hypothesis, such that the potential to present a greater number of CD8 T cell epitopes within a single animal can delay immune escape in targeted epitopes. As a result, certain epitope sequences can retain immunogenicity into chronic infection.

Comparative characterization of transfection- and infection-derived simian immunodeficiency virus challenge stocks for in vivo nonhuman primate studies.

Simian immunodeficiency virus (SIV) stocks for in vivo nonhuman primate models of AIDS are typically generated by transfection of 293T cells with molecularly cloned viral genomes or by expansion in productively infected T cells. Although titers of stocks are determined for infectivity in vitro prior to in vivo inoculation, virus production methods may differentially affect stock features that are not routinely analyzed but may impact in vivo infectivity, mucosal transmissibility, and early infection events. We performed a detailed analysis of nine SIV stocks, comprising five infection-derived SIVmac251 viral swarm stocks and paired infection- and transfected-293T-cell-derived stocks of both SIVmac239 and SIVmac766. Representative stocks were evaluated for (i) virus content, (ii) infectious titer, (iii) sequence diversity and polymorphism frequency by single-genome amplification and 454 pyrosequencing, (iv) virion-associated Env content, and (v) cytokine and chemokine content by 36-plex Luminex analysis. Regardless of production method, all stocks had comparable particle/infectivity ratios, with the transfected-293T stocks possessing the highest overall virus content and infectivity titers despite containing markedly lower levels of virion-associated Env than infection-derived viruses. Transfected-293T stocks also contained fewer and lower levels of cytokines and chemokines than infection-derived stocks, which had elevated levels of multiple analytes, with substantial variability among stocks. Sequencing of the infection-derived SIVmac251 stocks revealed variable levels of viral diversity between stocks, with evidence of stock-specific selection and expansion of unique viral lineages. These analyses suggest that there may be underappreciated features of SIV in vivo challenge stocks with the potential to impact early infection events, which may merit consideration when selecting virus stocks for in vivo studies.

Adoptive transfer of lymphocytes isolated from simian immunodeficiency virus SIVmac239Δnef-vaccinated macaques does not affect acute-phase viral loads but may reduce chronic-phase viral loads in major histocompatibility complex-matched recipients.

The live attenuated simian immunodeficiency virus (SIV) SIVmac239Δnef is the most effective SIV/human immunodeficiency virus (HIV) vaccine in preclinical testing. An understanding of the mechanisms responsible for protection may provide important insights for the development of HIV vaccines. Leveraging the uniquely restricted genetic diversity of Mauritian cynomolgus macaques, we performed adoptive transfers between major histocompatibility complex (MHC)-matched animals to assess the role of cellular immunity in SIVmac239Δnef protection. We vaccinated and mock vaccinated donor macaques and then harvested between 1.25 × 10(9) and 3.0 × 10(9) mononuclear cells from multiple tissues for transfer into 12 naive recipients, followed by challenge with pathogenic SIVmac239. Fluorescently labeled donor cells were detectable for at least 7 days posttransfer and trafficked to multiple tissues, including lung, lymph nodes, and other mucosal tissues. There was no difference between recipient macaques' peak or postpeak plasma viral loads. A very modest difference in viral loads during the chronic phase between vaccinated animal cell recipients and mock-vaccinated animal cell recipients did not reach significance (P = 0.12). Interestingly, the SIVmac239 challenge virus accumulated escape mutations more rapidly in animals that received cells from vaccinated donors. These results may suggest that adoptive transfers influenced the course of infection despite the lack of significant differences in the viral loads among animals that received cells from vaccinated and mock-vaccinated donor animals.

T cell response specificity and magnitude against SIVmac239 are not concordant in major histocompatibility complex-matched animals.

BACKGROUND: CD8+ T cell responses, restricted by major histocompatibility complex (MHC) class I molecules, are critical to controlling human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) replication. Previous studies have used MHC-matched siblings and monozygotic twins to evaluate genetic and stochastic influences on HIV-specific T cell responses and viral evolution. Here we used a genetically restricted population of Mauritian cynomolgus macaques (MCM) to characterize T cell responses within nine pairs of MHC-matched animals. FINDINGS: In MHC-matched animals, there was considerable heterogeneity in the specificity and magnitude of T cell responses detected via individual peptide gamma interferon (IFN-γ) enzyme-linked immunospot (ELISPOT) assays. These findings were further supported by full proteome pooled peptide matrix ELISPOT data collected from this cohort at 52 weeks post-infection. Interestingly, peptide regions that elicited dominant T cell responses were more commonly shared between MHC-matched MCM than peptide regions that elicited non-dominant T cell responses. CONCLUSIONS: Our findings suggest that, while some T cell responses mounted during chronic infection by MHC-matched MCM are similar, the majority of responses are highly variable. Shared responses detected in this study between MHC-matched MCM were directed against epitopes that had previously elicited relatively dominant responses in MCM with the same MHC class I haplotype, suggesting that the factors that influence dominance may influence the reproducibility of responses as well. This may be an important consideration for future T cell-based vaccines aiming to consistently and reproducibly elicit protective T cell responses.

Specific CD8+ T cell responses correlate with control of simian immunodeficiency virus replication in Mauritian cynomolgus macaques.

Specific major histocompatibility complex (MHC) class I alleles are associated with an increased frequency of spontaneous control of human and simian immunodeficiency viruses (HIV and SIV). The mechanism of control is thought to involve MHC class I-restricted CD8(+) T cells, but it is not clear whether particular CD8(+) T cell responses or a broad repertoire of epitope-specific CD8(+) T cell populations (termed T cell breadth) are principally responsible for mediating immunologic control. To test the hypothesis that heterozygous macaques control SIV replication as a function of superior T cell breadth, we infected MHC-homozygous and MHC-heterozygous cynomolgus macaques with the pathogenic virus SIVmac239. As measured by a gamma interferon enzyme-linked immunosorbent spot assay (IFN-γ ELISPOT) using blood, T cell breadth did not differ significantly between homozygotes and heterozygotes. Surprisingly, macaques that controlled SIV replication, regardless of their MHC zygosity, shared durable T cell responses against similar regions of Nef. While the limited genetic variability in these animals prevents us from making generalizations about the importance of Nef-specific T cell responses in controlling HIV, these results suggest that the T cell-mediated control of virus replication that we observed is more likely the consequence of targeting specificity rather than T cell breadth.

Case Report: New Onset Lymphadenopathy After Immune Checkpoint Inhibitor Therapy Presents a Clinicopathological and Radiological Challenge.

The use of immune-checkpoint inhibitor (ICI) therapy has significantly improved patient outcomes in a wide variety of cancers and has become a cornerstone in the treatment of renal cell carcinoma. However, ICI treatment has the potential to cause a variety of immune-related adverse events (irAEs) that can affect any tissue or organ. This report describes the diagnostic dilemma of a patient with both RCC and diffuse large B-cell lymphoma who developed acute onset of fever and diffuse lymphadenopathy following treatment with combined ipilimumab and nivolumab. While diagnostic considerations included worsening lymphoma, hyperprogression of RCC, sarcoid-like reaction from immunotherapy, and fungal infection, his lymphadenopathy eventually resolved with treatment for histoplasmosis and discontinuation of immunotherapy. Despite only receiving two doses of immunotherapy, he has not required additional systemic therapy for RCC. This case demonstrates both the effectiveness of ICI therapy and the need for multidisciplinary approach to potential irAEs.

Liquid Biopsy in Solid Malignancy.

The clinical utility of tissue biopsies in cancer management will continue to expand, especially with the evolving role of targeted therapies. "Liquid biopsy" refers to testing a patient's biofluid samples such as blood or urine to detect tumor-derived molecules and cells that can be used diagnostically and prognostically in the assessment of cancer. Many proof-of-concept and pilot studies have shown the clinical potential of liquid biopsies as diagnostic and prognostic markers which would provide a surrogate for the conventional "solid biopsy". In this review, we focus on three methods of liquid biopsy-circulating tumor cells, extracellular vesicles, and circulating tumor DNA-to provide a landscape view of their clinical applicability in cancer management and research.