Immune Pressure Research Articles

Abstract LB005: High quality neoantigens are immunoedited in long term survivors of pancreatic cancer

Abstract Cancer immunoediting, a hallmark of cancer, predicts that lymphocytes recognize tumor-specific mutations (neoantigens) to kill immunogenic tumor cells and select less immunogenic clones to survive in immunocompetent hosts. Yet, though proven in mice, whether immunoediting governs how human tumors evolve naturally, and the principles of how the immune system edits tumors remain unclear. Here, we report that T cell rich tumors of rare long-term survivors of human pancreatic ductal adenocarcinoma longitudinally lose immunogenic features predicted by a fitness model based on immune recognition of high quality neoantigens. We find that compared to T cell poor tumors in short-term survivors, long-term survivors evolve less immunogenic recurrent tumors over 10 years with fewer clones, fewer neoantigens, and new neoantigens of markedly lower quality, to indicate that the immune system edits lower fitness clones with high-quality neoantigens. To explore if the immune system more broadly selects against non-self-like changes in host genomes, we define a minimal amino acid substitution distance sufficient for T cell receptors to discriminate self from non-self mutations. We find that both primary and recurrent tumors in long but not short-term survivors are enriched in self-like mutations, to suggest that the immune system depletes non-self-like mutations. Thus, we submit longitudinal evidence that the immune system edits neoantigens to sculpt recurrent human pancreatic ductal adenocarcinomas, a lethal tumor largely considered resistant to endogenous immune attack. Furthermore, we demonstrate that the neoantigen quality fitness model predicts how immune pressure changes tumor cell populations over time, with significant implications for cancer biology and immunotherapy. More broadly, our results argue that cancer immunoediting is a fundamental principle of how human cancers evolve in immunocompetent hosts. Citation Format: Marta Luksza, Zachary Sethna, Luis Rojas, Kevin Soares, Yuval Elhanati, Jayon Lihm, David Hoyos, Rajya Kappagantula, Alvin M. Moore, Erin Patterson, Christine Iacobuzio-Donahue, Benjamin Greenbaum, Vinod P. Balachandran. High quality neoantigens are immunoedited in long term survivors of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB005.

Abstract 3140: Tumor evolution selectively inactivates microRNA machinery for immune evasion

Abstract Cancer arises after somatic mutations override cellular and immunological control of cell proliferation1. Mutations in oncogenes and tumor suppressor genes endow cancer cells intrinsic growth advantage over neighboring cells. Mutations can also enable cancer cells to escape from immune surveillance during cancer evolution, but a targeted approach to uncover such mutations is lacking. We hypothesized that genetic heterogeneity intrinsic to cancer cells affects the cell fitness to T cell-mediated cytotoxicity. By deep sequencing of murine tumors grown under different conditions of immune editing, we identified 59 genetic mutations in clonal populations that were specifically enriched under host immune pressure. To differentiate driver from passenger mutations against immune selection, we performed in vivo and T cell co-culture CRISPR screens with a guide RNA library targeting the corresponding genes. We identified ANKRD52 as required for PD-1-independent T cell-mediated cytotoxicity, which enhanced the JAK-STAT-interferon-γ signaling and antigen presentation in cancer cells. ANKRD52, in complex with PPP6C, dephosphorylates AGO26 and promoted microRNA (miRNA)-targeted silencing of suppressor of cytokine signaling 1 (SOCS1), and deletion of SOCS1 reinstalled the sensitivity of the mutant cells to T cell killing. Introduction of patient hotspot ANKRD52 mutations, or deletion of AGO2, DICER1 or XPO5 in cancer cells compromised the interferon-γ and T cell response. Remarkably, expression of the core miRNA biogenesis and targeting machinery strongly correlates with intratumoral T cell abundance across nearly all human cancer types. Our data indicate that the evolutionarily conserved miRNA pathway is exploited by cancer cells for immune escape. Citation Format: Haixin Zhao, Tian-yu Song, Min Long, Hong-Jie Fan, Yang Liu, Chen-Lu Geng, Xiaoxiao Xie, Dawei Huang, Zhengang Peng, Bo Peng, Yong Cang. Tumor evolution selectively inactivates microRNA machinery for immune evasion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3140.

Abstract 1905: In vivo CRISPR screen identifies tumor suppressors as drivers of tumor cell-intrinsic immune evasion

Abstract Cancer cell-intrinsic genetic alterations that allow cancer cells to evade destruction by the immune system remain poorly characterized. Here we performed a pooled CRISPR-Cas9-based in vivo genetic screen targeted to a pre-defined set of tumor suppressor genes to mimic loss-of-function mutations in syngeneic tumor models. CRISPR edited tumor cells were implanted into immune-deficient or immune-competent C57BL/6 mice, a subset of which were treated with anti-PD1 to simulate increased immune pressure. Tumor samples at the endpoint were subjected to next generation sequencing analysis to identify tumor suppressor genes driving immune evasion. The screen confirmed previously identified immunotherapy targets such as CD47 and Adar as well as known drivers of immune resistance in the interferon signaling and antigen presentation pathways. Importantly, we identified loss of STK11/LKB1 and KEAP1 as drivers of immune resistance in syngeneic models. TIL profiling analysis suggests that STK11/LKB1 knockout induces a “cold” tumor microenvironment. STK11/LKB1 and/or KEAP1 genomic alterations are found in ~25% of non-squamous non-small cell lung cancer and have been reported as a major predictor of primary resistance to PD-1 blockade. Together, these data demonstrate that high-throughput in vivo genetic screens can identify tumor cell-intrinsic drivers of immune evasion and establish murine system relevant to the study of primary resistance to PD-1 axis immunotherapy and more generally, tumor cell-intrinsic immune evasion. Citation Format: Chengyin Min, Ferdinandos Skoulidis, Wenrong Zhou, Jennifer Tsoi, Alan Huang, Matthew J. Goldstein. In vivo CRISPR screen identifies tumor suppressors as drivers of tumor cell-intrinsic immune evasion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1905.

Antigenic waves of virus–immune coevolution

The evolution of many microbes and pathogens, including circulating viruses such as seasonal influenza, is driven by immune pressure from the host population. In turn, the immune systems of infected populations get updated, chasing viruses even farther away. Quantitatively understanding how these dynamics result in observed patterns of rapid pathogen and immune adaptation is instrumental to epidemiological and evolutionary forecasting. Here we present a mathematical theory of coevolution between immune systems and viruses in a finite-dimensional antigenic space, which describes the cross-reactivity of viral strains and immune systems primed by previous infections. We show the emergence of an antigenic wave that is pushed forward and canalized by cross-reactivity. We obtain analytical results for shape, speed, and angular diffusion of the wave. In particular, we show that viral-immune coevolution generates an emergent timescale, the persistence time of the wave's direction in antigenic space, which can be much longer than the coalescence time of the viral population. We compare these dynamics to the observed antigenic turnover of influenza strains, and we discuss how the dimensionality of antigenic space impacts the predictability of the evolutionary dynamics. Our results provide a concrete and tractable framework to describe pathogen-host coevolution.

Paroxysmal Nocturnal Hemoglobinuria Results from Initial Clonal PIG-a Mosaicism

The pathogenesis of PNH involves alteration of at least one HSC clone, characterized by a somatic mutation in the PIG-A gene. The resulting phenotype appears to offer a growth advantage over other unaffected HSCs, yet the failure of PIG-A mutant clones to expand in animal models suggests that additional factors are necessary for the development of PNH. The association of PNH with aplastic anemia (AA) is evidence of selection pressure by immune attack, but secondary hits in additional genes may also be clonally acquired to render PNH clones competitive irrespective of immune pressure. With the advent of NGS, clonal architecture analysis has been revolutionized. Yet, despite increased resolution, NGS is hampered by the presence of normal/non-clonal cells at varying frequencies in the sequenced sample, often decreasing the detection threshold and precluding recognition of subclonal mosaicism.To study these issues in PNH, 133 patients with PNH (N=33), AA (N=33), and AA/PNH (N=67) were sequenced using deep targeted NGS covering all exons of the PIG-A gene. All patients had a detectable PNH clone by flow cytometry, with the mean WBC clone size of 0.22% in AA, 27.4% in AA/PNH, and 77.1% in PNH. 223 PIG-A mutations were identified in this cohort; 35% were frameshift mutations, 34% missense, 14% splice site, 13% nonsense, and 3% non-frameshift. In unpurified blood, PIG-A mutations were found in 27/33 (82%) of PNH patients, 36/67 (54%) of AA/PNH, and 2/33 (6%) of AA, with a clear association between PNH clone size and the ability to detect mutations by NGS (P<0.0001). In DNA extracted from cells flow sorted by FLAER negativity, detection rates were 26/29 (90%), with an average of nearly 3 mutations per patient, indicating that mechanisms such as microdeletions may affect as many as 10% of PNH or AA/PNH patients and that clonal mosaicism is common. Our results also show that indeed type III and II PNH cells are derived from 2 distinct clones, each with a distinct mutation.To examine disease pathogenesis, we studied 47 patients longitudinally using both PNH flow and deep sequencing. Serial testing illustrates possible modes of evolution: mutations can fluctuate in the contribution to clonal burden over time with the variant allelic frequency of some mutations increasing while others decrease. Linear regression of the number of mutations vs . time from diagnosis suggests that in AA/PNH (N=29), the number of mutant clones detected increases over time (P=.012) while in classic PNH (N=18) the trend is reversed, as the number of mutations detected decreases (P<.0001) and a dominant clone emerges to supply a majority of blood cells.Traditionally, PNH has been considered a monoclonal disease in which a PIG-A mutant HSC clone can outcompete normal hematopoiesis within the context of permissive conditions. Using deep NGS, our results clearly indicate that while a single clone ultimately may dominate, PNH begins as oligoclonal process with the eventual emergence of the most competitive HSC clone(s) populating hematopoietic space. Initially, an environment that “favors” a state of GPI-anchor deficiency thereby allows multiple clones to evolve. Over time, the most “fit” PNH HSC clones dominate, resulting in monoclonality. In addition, our results confirm the presence of distinct mutations with corresponding phenotype through sequencing of sorted type II and type III PNH cells. Clonal drift may also be observed, likely through a newly acquired advantage of one clone within the available pool of GPI-deficient PIGA mutant clones. Our previous work indicated that acquisition of additional enabling mutations results in a more complex clonal architecture comparable to that seen in MDS, an intrinsic mechanism of the selection process that involves mutations in genes altered in MDS. Additional intrinsic hits also explain patient-specific kinetics of PNH evolution and apparent independence of established PNH from the effect of immunosuppression. Thus, while immune selection may provide initial conditions favorable for oligoclonal PNH evolution, once pressure is relieved, intrinsic factors can lead to PNH monoclonality. Alternative mechanisms could include loss of dominance by exhaustion, senescence, or telomere shortening.In sum, our study clearly demonstrates initial polyclonality early in the pathogenesis of PNH with selection of dominant clones during disease duration. DisclosuresMakishima:Yasuda Medical Foundation: Research Funding.

HIV Compartmentalization in Male Genital Tract: Relevance for Viral Eradication

Abstract Human immunodeficiency virus (HIV) has been shown to evolve independently in different anatomical compartments. Characterizing HIV genetic evolution in different tissues and cells provides insights into the mechanisms that maintain the viral reservoir. HIV compartmentalization has been well documented in the semen but rarely in male genital tract (MGT) organs. The precise mechanisms that result in the development of HIV compartmentalization in multiple genitourinary sites have been poorly explored due to the difficulty in accessing these tissues. Based on evidence from lymph nodes and gut tissues, mechanisms that may influence compartmentalization include immune pressures, local concentrations of antiviral drugs, clonal expansion of different cell types and inflammation that alters the cellular microenvironment. We reviewed phylogenetic evidences supporting viral compartmentalization between the blood and multiple genitourinary sites in HIV-infected people. Characterizing distinct viral sub-populations enhances our overall understanding of the HIV reservoir in MGT and could ultimately lead to the development of novel therapies to eradicate the virus in tissues.

Whole genome sequencing of human metapneumoviruses from clinical specimens using MinION nanopore technology

Human metapneumovirus (HMPV), a member of the Pneumoviridae family, is a causative agent of respiratory illness in young children, the elderly, and immunocompromised individuals. Globally, viruses belonging to two main genetic lineages circulate, A and B, which are further divided into four genetic sublineages (A1, A2, B1, B2). Classical genotyping of HMPV is based on the sequence of the fusion (F) and attachment (G) glycoprotein genes, which are under direct antibody-mediated immune pressure. Whole genome sequencing provides more information than sequencing of subgenomic fragments and is therefore a powerful tool for studying virus evolution and disease epidemiology and for identifying transmission events and nosocomial outbreaks. Here, we report a robust method to obtain whole genome sequences for HMPV using MinION Nanopore technology. This assay is able to generate HMPV whole genome sequences from clinical specimens with good coverage of the highly variable G gene and is equally sensitive for strains of all four genetic HMPV sublineages. This method can be used for studying HMPV genetics, epidemiology, and evolutionary dynamics.

Association of HLA loss of heterozygosity with allele-specific neoantigen expansion in response to immunotherapy.

e18030 Background: Human leukocyte antigen loss of heterozygosity (HLA LOH) restricts immune recognition of tumors by limiting the major histocompatibility complex (MHC) presentation of neoantigens to T cells and correlates with reduced response to immune checkpoint blockade therapy (ICB) in non-small cell lung cancer. To explore the mechanism behind the impairment of HLA LOH on ICB, we analyzed the relationship between the antigen presentation pathway, neoantigen presentation and response to ICB in a head and neck squamous cell carcinoma (HNSCC) cohort. Methods: Following baseline sample collection, a cohort of 14 HNSCC patients recieved a single dose of PD-1 inhibitor. The primary tumor mass was definitively resected approximately one month later. If resection was impractical, a second biopsy was taken. Response to therapy was evaluated using RECIST criteria. Each pre- and post-intervention tumor sample and normal PBMC sample were profiled using Personalis’ ImmunoID NeXT Platform, an HLA-enhanced exome/transcriptome platform. HLA LOH was detected using a digital PCR validated machine learning algorithm (DASH). Neoantigen presentation was computationally predicted using a machine learning algorithm (SHERPATM) trained on mono-allelic immunopeptidomics data. Results: We found that 50% of the HNSCC cohort had HLA LOH, a larger percentage than in a large pan-cancer cohort (23%, n=611) and a distinct HNSCC cohort (40%, n=20). Further, two patients had B2M LOH and one patient had a deleterious mutation in an HLA allele. Despite the high frequency of somatic alteration in the antigen presentation pathway, we did not find an association between HLA LOH and ICB response. However, if HLA LOH was still shaping tumor evolution in response to ICB, we would expect to see immune pressure against subclonal tumor populations with neoantigens presentable by the retained HLA alleles but not the deleted HLA alleles. Indeed, we found that significantly more novel post-treatment neoantigens were predicted to bind to deleted HLA alleles compared to their homologous alleles (p=0.045). Conclusions: Given the high prevalence of HLA LOH across tumor types, a greater understanding is needed regarding the impact of HLA LOH on tumor evolution during ICB treatment. Though HLA LOH does not correlate with response to ICB, the consistent shift in neoantigen composition suggests that it acts as an evolutionary force in resistance to response during immunotherapy.

Effect of Prorrenin/Renin Receptor Inhibition upon Immune Cells Signaling and Blood Pressure in An Experimental Model of Hypertension

Introduction Hypertension is a multifactorial disease. Several systems participate in its pathophysiology. The renin-angiotensin-aldosterone system (RAAS) and the immune system (IS) participate actively in maintaining inflammation and other alterations in the cardiovascular system. Interestingly, both systems, cardiovascular and the IS, share the expression and activity of prorenin/renin receptor (P)RR in different cells, as endothelial cells, cardiomyocytes, lymphocytes and NK cells, suggesting an orchestrated action during hypertension. Aim To evaluate the effect of prorrenin/renin receptor inhibition upon immune cells and blood pressure in 5/6 nephrectomized rats. Methods We used male Wistar rats of 250 ± 20 g submitted to a 5/6 nephrectomy. Control rats were submitted to sham operation. The animals were randomly distributed in 7 days and 15 days groups (n = 4, each group). HRP peptide was administered subcutaneously at a dose of 0.2 mg/kg, every 24 hours, during the last 4 days of each time period. Control animals received vehicle. We measured blood pressure and animals were sacrificed, the spleen was processed for flow cytometry to determine PRR and intracellular pH, and for immunoblot to determine PRR, DVL-1, PLZF using beta-actin as housekeeping protein. Procedures in animals complied with the requirements of the NOM – 062 – ZOO – 1999, and the NIH Guide for the Care and Use of Laboratory Animals. Results 5/6 nephrectomy groups had an increase in blood pressure compared to controls. HRP treatment decreased blood pressure significantly in nephrectomized groups. Seven days nephrectomy group increased PRR expression in lymphocytes and NK cells. HRP treatment reverted this effect; in the 15 days nephrectomized group there were no significant differences. DVL-1 and PLZF were measured in both groups. We found that, in the nephrectomy groups, these proteins had a higher expression compared to controls, and PRR blockade decreased this expression significantly. The intracellular pH in polymorphonuclear cells and lymphocytes was more acidic in the nephrectomy groups compared to the control group. This effect was reverted by treatment. Conclusion The PRR increased its expression in immune cells during hypertension. Treatment with HRP decreased this expression and reverted high blood pressure in early times. This effect seems to involve V-ATPase activity by affecting intracellular pH and the activation of the canonical WNT pathway

MCherry-expressing Mycobacterium tuberculosis gives insight into host cell tropism and immune pressure at the single bacterium level in macaques with tuberculosis

Abstract Tuberculosis is caused by Mycobacterium tuberculosis (Mtb) and is associated with granuloma formation in the lungs and other tissues. These lesions can limit bacterial replication but also contain host cells for Mtb and support bacterial persistence and dissemination in poorly controlled disease. Macrophages are often viewed as primary host cells for Mtb but work in mice demonstrates that other cell types host Mtb in acute infection. Less is known about host cell tropism in human TB, especially in the early stages of infection but these early interactions may have implications for later stages of disease. To identify which cells are infected in early and established TB, we infected two cynomolgus macaques with an mCherry-expressing Mtb strain and used flow cytometry and microscopy to identify and phenotype the infected cells at four- and ten-weeks post infection. This approach also allowed us to compare bacterial mCherry expression and size to determine if granulomas exert different levels of pressure on Mtb that vary by timepoint or lesion type. We found that granulomas contained numerous Mtb-infected neutrophils and macrophages at four weeks post infection but fewer infected neutrophils at ten weeks post infection. When the fluorescence and size of individual bacteria were compared across granulomas and timepoints, we found that bacteria in high Mtb burden granulomas were more fluorescent than bacteria in granulomas that contained fewer bacteria, and the bacteria in the four-week granulomas were longer, suggesting that early granulomas are better at supporting Mtb growth than granulomas in established disease. These results provide new insights into Mtb-host cell dynamics in a nonhuman primate model that replicates human TB.

Effects of aromatherapy on sleep disorders: A protocol for systematic review and meta-analysis.

Background:The progress of modern society and changes in lifestyle have not only increased the proportion of sub-healthy people, but also caused many people to suffer from sleep disorders and mental anxiety. Long-term lack of high-quality sleep will not only cause psychological problems such as anxiety and fatigue, but also cause physical abnormalities, such as abnormal hormone secretion, weakened immunity, neuroendocrine disorders, and high blood pressure. Therefore, the purpose of this study is to systematically evaluate the effectiveness of aromatherapy in improving sleep quality in people with sleep disorders.Methods:Computer search CNKI, SinoMed, Wanfang, PubMed, Web of science, and EMbase collect randomized controlled trials on aromatherapy to improve sleep quality of people with sleep disorders. The search time limit is to build the database until April 5, 2021. RevMan5.3 software is used for meta-analysis.Results:This systematic review will provide an assessment of the current state of sleep disorders, aiming to assess the efficacy of aromatherapy for patients with sleep disorders.Conclusion:This systematic review will provide a credible evidence-based for the clinical treatment of sleep disorders with aromatherapy.

Recipient factors in faecal microbiota transplantation: one stool does not fit all.

Faecal microbiota transplantation (FMT) is a promising therapy for chronic diseases associated with gut microbiota alterations. FMT cures 90% of recurrent Clostridioides difficile infections. However, in complex diseases, such as inflammatory bowel disease, irritable bowel syndrome and metabolic syndrome, its efficacy remains variable. It is accepted that donor selection and sample administration are key determinants of FMT success, yet little is known about the recipient factors that affect it. In this Perspective, we discuss the effects of recipient parameters, such as genetics, immunity, microbiota and lifestyle, on donor microbiota engraftment and clinical efficacy. Emerging evidence supports the possibility that controlling inflammation in the recipient intestine might facilitate engraftment by reducing host immune system pressure on the newly transferred microbiota. Deciphering FMT engraftment rules and developing novel therapeutic strategies are priorities to alleviate the burden of chronic diseases associated with an altered gut microbiota such as inflammatory bowel disease.

MrHAMER yields highly accurate single molecule viral sequences enabling analysis of intra-host evolution.

Technical challenges remain in the sequencing of RNA viruses due to their high intra-host diversity. This bottleneck is particularly pronounced when interrogating long-range co-evolved genetic interactions given the read-length limitations of next-generation sequencing platforms. This has hampered the direct observation of these genetic interactions that code for protein-protein interfaces with relevance in both drug and vaccine development. Here we overcome these technical limitations by developing a nanopore-based long-range viral sequencing pipeline that yields accurate single molecule sequences of circulating virions from clinical samples. We demonstrate its utility in observing the evolution of individual HIV Gag-Pol genomes in response to antiviral pressure. Our pipeline, called Multi-read Hairpin Mediated Error-correction Reaction (MrHAMER), yields >1000s of viral genomes per sample at 99.9% accuracy, maintains the original proportion of sequenced virions present in a complex mixture, and allows the detection of rare viral genomes with their associated mutations present at <1% frequency. This method facilitates scalable investigation of genetic correlates of resistance to both antiviral therapy and immune pressure and enables the identification of novel host-viral and viral-viral interfaces that can be modulated for therapeutic benefit.

UNCOVERING ASSOCIATIONS BETWEEN BLOOD PRESSURE PHENOTYPES AND IMMUNE TRAITS IN TWINSUK

Objective: The primary aim of this work was to define the contribution of both the innate and adaptive immune system arms to blood pressure changes. Design and method: In order to define the inter-relationships between immune cell subtypes, blood pressure (BP) and susceptibility to hypertension (HT), we carried out association analyses between 23,394 immune phenotypes and blood pressure measures [systolic blood pressure (SBP), diastolic blood pressure (DBP), pulse pressure (PP)] and HT. After calculating the mean SBP and DBP values from two BP measurements, we adjusted for medication use by adding 15- and 10-mm Hg to SBP and DBP respectively for individuals reported to be on BP-lowering treatment. PP was calculated as SBP minus DBP according to medication adjusted traits. HT was defined as (i) SBP&gt;=140 mm Hg, (ii) DBP&gt;=90 mm Hg or taking BP-lowering medication. We undertook a comprehensive and high-resolution deep immunophenotyping flow-cytometry approach, to examine immune traits in 502 female twins from TwinsUK. The platform used covered all major lymphocyte and myeloid subsets in peripheral blood. A linear/logistic mixed model regression analysis adjusted for family structure, age and BMI was used to examine the associations between immune traits and blood pressure phenotypes and HT status respectively. Results: We observed statistically significant associations (Bonferroni P-value &lt; 1.71x10–4) between blood pressure phenotypes and immune cell subtypes. In particular, DBP showed 47 significant associations with distinct myeloid cell types of CD8 differentiation T cells. SBP showed 59 significant associations with both CD8 differentiation T cells and specific myeloid subpopulations such as monocytes. PP showed 4 significant associations with monocyte subpopulations. Analysis of HT revealed 7 associations with monocytes, CD8 T cells and dendritic cells. Conclusions: The results reported in this study provide preliminary evidence of an inter-relationship between blood pressure and human immune cell profile associations. We have shown for the first time that change in the proportions of immune cells correlates with blood pressure measurements.

The Covid Calamity, the Human Life, and the Surviving Hope

Introduction - Ongoing Covid-19 Calamity: As a disease, COVID-19 is still in the pandemic phase because infections continue to increasingly occur world-wide and various population groups are still susceptible. It is likely that the SARS-CoV-2 will not be eradicated but become endemic and continue to circulate and cause infections in pockets of the global populations for years to come. It may evolve into more transmissible and virulent forms with novel mutations and variants, and associated factors may worsen the overall scenario with involvement of newer population groups and world regions. Mutations, Variants and Immune Escape: The unabated prevalence increases risk of mutations, as the virus has more chances to mutate. Further, in areas where the incidence rates are high, selection pressures favour the emergence of variants that evade neutralising antibodies. Furthermore, as population groups receive vaccination, immune pressure is conjectured to facilitate and speed up the emergence of such variants by selecting for escape mutants. In due course, these selected variants would replace previous versions of the virus propelling the pandemic or the endemic disease later on. Associated Uncertainties with SARS-CoV-2: There are various associated uncertainties with the SARS-CoV-2 virus and the disease it causes. Due to evolving genomic changes, the virus elicits erratic and labile immune response. Simultaneously, the host factors are highly variable and largely uncontrollable. Further, the control measures and available vaccines for COVID-19 may not reduce the prevalence of infections drastically for multiple reasons. These epidemiological drivers would lead to persistence of the virus and endemicity of the disease interspersed by periodic outbreaks and re-emergence. The Human Life during Covid-19 Pandemic: With COVID-19 becoming an endemic disease, the SARS-CoV-2 virus would be first encountered during childhood, typically causing mild manifestations or none. The population groups will develop some immunity through natural infection or vaccination and may not suffer with severe illness, except in those with comorbid conditions or immune-compromised states, and the disease course would depend on evolving variants, efficacy of vaccines, and nature of immunity to the virus. The herd immunity against SARS-CoV-2 may remain a myth and with individual immunity being labile and waning after 6-8 months, booster doses of updated vaccine will be required at regular intervals. Future Scenario and Search for Solutions: To mitigate the spread of SARS-CoV-2 virus, various countries have implemented a wide range of control measures from time to time and likely to resort to, in future as well. There is need for genotyping and genomic sequencing capability for quick and effective utilization of epidemiological data. Simultaneously, the large deployment of COVID-19 vaccines under way needs a rapid and effective global effort. The next-generation vaccines may stimulate T cells effectively, apart from generating antibodies against the virus, and there is possibility of designing a universal coronavirus vaccine or pan-virus vaccine for immunisation against multiple variants and strains. On the therapeutic side, use of probiotics as adjuvant therapy may Improve the prognosis and clinical outcomes in COVID-19.

Tracking Cancer Evolution through the Disease Course.

During cancer evolution, constituent tumor cells compete under dynamic selection pressures. Phenotypic variation can be observed as intratumor heterogeneity, which is propagated by genome instability leading to mutations, somatic copy-number alterations, and epigenomic changes. TRACERx was set up in 2014 to observe the relationship between intratumor heterogeneity and patient outcome. By integrating multiregion sequencing of primary tumors with longitudinal sampling of a prospectively recruited patient cohort, cancer evolution can be tracked from early- to late-stage disease and through therapy. Here we review some of the key features of the studies and look to the future of the field. SIGNIFICANCE: Cancers evolve and adapt to environmental challenges such as immune surveillance and treatment pressures. The TRACERx studies track cancer evolution in a clinical setting, through primary disease to recurrence. Through multiregion and longitudinal sampling, evolutionary processes have been detailed in the tumor and the immune microenvironment in non-small cell lung cancer and clear-cell renal cell carcinoma. TRACERx has revealed the potential therapeutic utility of targeting clonal neoantigens and ctDNA detection in the adjuvant setting as a minimal residual disease detection tool primed for translation into clinical trials.

Virus Evolution and Neutralization Sensitivity in an HIV-1 Subtype B' Infected Plasma Donor with Broadly Neutralizing Activity.

We sought to analyze the evolutionary characteristics and neutralization sensitivity of viruses in a human immunodeficiency virus type 1 (HIV-1) subtype B′ infected plasma donor with broadly neutralizing activity, which may provide information for new broadly neutralizing antibodies (bNAbs) isolation and immunogen design. A total of 83 full-length envelope genes were obtained by single-genome amplification (SGA) from the patient’s plasma at three consecutive time points (2005, 2006, and 2008) spanning four years. In addition, 28 Env-pseudotyped viruses were constructed and their neutralization sensitivity to autologous plasma and several representative bNAbs were measured. Phylogenetic analysis showed that these env sequences formed two evolutionary clusters (Cluster I and II). Cluster I viruses vanished in 2006 and then appeared as recombinants two years later. In Cluster II viruses, the V1 length and N-glycosylation sites increased over the four years of the study period. Most viruses were sensitive to concurrent and subsequent autologous plasma, and to bNAbs, including 10E8, PGT121, VRC01, and 12A21, but all viruses were resistant to PGT135. Overall, 90% of Cluster I viruses were resistant to 2G12, while 94% of Cluster II viruses were sensitive to 2G12. We confirmed that HIV-1 continued to evolve even in the presence of bNAbs, and two virus clusters in this donor adopted different escape mechanisms under the same humoral immune pressure.

Mesenchymal plasticity of devil facial tumour cells during in vivo vaccine and immunotherapy trials.

Immune evasion is critical to the growth and survival of cancer cells. This is especially pertinent to transmissible cancers, which evade immune detection across genetically diverse hosts. The Tasmanian devil (Sarcophilus harrisii) is threatened by the emergence of Devil Facial Tumour Disease (DFTD), comprising two transmissible cancers (DFT1 and DFT2). The development of effective prophylactic vaccines and therapies against DFTD has been restricted by an incomplete understanding of how allogeneic DFT1 and DFT2 cells maintain immune evasion upon activation of tumour-specific immune responses. In this study, we used RNA sequencing to examine tumours from three experimental DFT1 cases. Two devils received a vaccine prior to inoculation with live DFT1 cells, providing an opportunity to explore changes to DFT1 cancers under immune pressure. Analysis of DFT1 in the non-immunised devil revealed a 'myelinating Schwann cell' phenotype, reflecting both natural DFT1 cancers and the DFT1 cell line used for the experimental challenge. Comparatively, immunised devils exhibited a 'dedifferentiated mesenchymal' DFT1 phenotype. A third 'immune-enriched' phenotype, characterised by increased PDL1 and CTLA-4 expression, was detected in a DFT1 tumour that arose after immunotherapy. In response to immune pressure, mesenchymal plasticity and upregulation of immune checkpoint molecules are used by human cancers to evade immune responses. Similar mechanisms are associated with immune evasion by DFTD cancers, providing novel insights that will inform modification of DFTD vaccines. As DFT1 and DFT2 are clonal cancers transmitted across genetically distinct hosts, the Tasmanian devil provides a 'natural' disease model for more broadly exploring these immune evasion mechanisms in cancer.

Antigenic cartography reveals complexities of genetic determinants that lead to antigenic differences among pandemic GII.4 noroviruses

Significance The GII.4 genotype of human noroviruses accounts for the majority of gastroenteritis outbreaks worldwide. This predominance is characterized by the chronological emergence of new variants in response to immune pressure. We performed a comprehensive analysis on a large panel of GII.4 viruses to investigate the genetic determinants of antigenic diversification. Using immunoassays and neutralization data, we observed major changes to antigenic properties over the course of GII.4 evolution and increased intervariant cross-reactivity across contemporary viruses. Antigenic cartography and sequence analyses indicated a minimum number of coevolving amino acid changes on the structural protein necessary for the emergence of antigenically distinct variants. These insights could facilitate the monitoring and characterization of emerging GII.4 noroviruses and the development of cross-protective vaccines.

The nucleocapsid protein of rice stripe virus in cell nuclei of vector insect regulates viral replication

Rice stripe virus (RSV) transmitted by the small brown planthopper causes severe rice yield losses in Asian countries. Although viral nuclear entry promotes viral replication in host cells, whether this phenomenon occurs in vector cells remains unknown. Therefore, in this study, we systematically evaluated the presence and roles of RSV in the nuclei of vector insect cells. We observed that the nucleocapsid protein (NP) and viral genomic RNAs were partially transported into vector cell nuclei by utilizing the importin α nuclear transport system. When blocking NP nuclear localization, cytoplasmic RSV accumulation significantly increased. In the vector cell nuclei, NP bound the transcription factor YY1 and affected its positive regulation to FAIM. Subsequently, decreased FAIM expression triggered an antiviral caspase-dependent apoptotic reaction. Our results reveal that viral nuclear entry induces completely different immune effects in vector and host cells, providing new insights into the balance between viral load and the immunity pressure in vector insects.