The Biology and Underlying Mechanisms of Cross-Presentation of Exogenous Antigens on MHC-I Molecules.

To monitor the health of cells, the immune system tasks antigen-presenting cells with gathering antigens from other cells and bringing them to CD8 T cells in the form of peptides bound to MHC-I molecules. Most cells would be unable to perform this function because they use their MHC-I molecules to exclusively present peptides derived from the cell's own proteins. However, the immune system evolved mechanisms for dendritic cells and some other phagocytes to sample and present antigens from the extracellular milieu on MHC-I through a process called cross-presentation. How this important task is accomplished, its role in health and disease, and its potential for exploitation are the subject of this review.

Manipulating the selection forces during affinity maturation to generate cross-reactive HIV antibodies.

Generation of potent antibodies by a mutation-selection process called affinity maturation is a key component of effective immune responses. Antibodies that protect against highly mutable pathogens must neutralize diverse strains. Developing effective immunization strategies to drive their evolution requires understanding how affinity maturation happens in an environment where variants of the same antigen are present. We present an in silico model of affinity maturation driven by antigen variants which reveals that induction of cross-reactive antibodies often occurs with low probability because conflicting selection forces, imposed by different antigen variants, can frustrate affinity maturation. We describe how variables such as temporal pattern of antigen administration influence the outcome of this frustrated evolutionary process. Our calculations predict, and experiments in mice with variant gp120 constructs of the HIV envelope protein confirm, that sequential immunization with antigen variants is preferred over a cocktail for induction of cross-reactive antibodies focused on the shared CD4 binding site epitope.

Frequent Loss of IRF2 in Cancers Leads to Immune Evasion through Decreased MHC Class I Antigen Presentation and Increased PD-L1 Expression.

To arise and progress, cancers need to evade immune elimination. Consequently, progressing tumors are often MHC class I (MHC-I) low and express immune inhibitory molecules, such as PD-L1, which allows them to avoid the main antitumor host defense, CD8+ T cells. The molecular mechanisms that led to these alterations were incompletely understood. In this study, we identify loss of the transcription factor IRF2 as a frequent underlying mechanism that leads to a tumor immune evasion phenotype in both humans and mice. We identified IRF2 in a CRISPR-based forward genetic screen for genes that controlled MHC-I Ag presentation in HeLa cells. We then found that many primary human cancers, including lung, colon, breast, prostate, and others, frequently downregulated IRF2. Although IRF2 is generally known as a transcriptional repressor, we found that it was a transcriptional activator of many key components of the MHC-I pathway, including immunoproteasomes, TAP, and ERAP1, whose transcriptional control was previously poorly understood. Upon loss of IRF2, cytosol-to-endoplasmic reticulum peptide transport and N-terminal peptide trimming become rate limiting for Ag presentation. In addition, we found that IRF2 is a repressor of PD-L1. Thus, by downregulating a single nonessential gene, tumors become harder to see (reduced Ag presentation), more inhibitory (increased checkpoint inhibitor), and less susceptible to being killed by CD8+ T cells. Importantly, we found that the loss of Ag presentation caused by IRF2 downregulation could be reversed by IFN-stimulated induction of the transcription factor IRF1. The implication of these findings for tumor progression and immunotherapy are discussed.

G-quadruplex-induced instability during leading-strand replication.

G-quadruplexes are four-stranded nucleic acid structures whose biological functions remain poorly understood. In the yeast S. cerevisiae, we report that G-quadruplexes form and, if not properly processed, pose a specific challenge to replication. We show that the G-quadruplex-prone CEB1 tandem array is tolerated when inserted near ARS305 replication origin in wild-type cells but is very frequently destabilized upon treatment with the potent Phen-DC(3) G-quadruplex ligand, or in the absence of the G-quadruplex-unwinding Pif1 helicase, only when the G-rich strand is the template of leading-strand replication. The orientation-dependent instability is associated with the formation of Rad51-Rad52-dependent X-shaped intermediates during replication detected by two-dimensional (2D) gels, and relies on the presence of intact G-quadruplex motifs in CEB1 and on the activity of ARS305. The asymmetrical behaviour of G-quadruplex prone sequences during replication has implications for their evolutionary dynamics within genomes, including the maintenance of G-rich telomeres.

Outcomes Among Patients With Mucosal Head and Neck Squamous Cell Carcinoma Treated With Checkpoint Inhibitors.

Importance: Immune checkpoint inhibitors (CPIs) are now part of standard therapy for patients with recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) because of improved outcomes compared with chemotherapy in clinical trials. However, data on outcomes in patients with HNSCC in the general population who are treated with CPIs remain limited. Objective: To assess response rates, survival outcomes, and associations with key clinical covariates in a large, contemporary cohort of patients with recurrent or metastatic mucosal HNSCC who were treated with CPIs with or without chemotherapy. Design, Setting, and Participants: This retrospective cohort study included patients older than 18 years who received CPI-based therapy for recurrent or metastatic HNSCC at the University of Pennsylvania from January 1, 2015, through August 15, 2021. Clinical and survival data were abstracted through medical record review. Exposures: Treatment with CPIs with or without chemotherapy for a diagnosis of HNSCC. Main Outcomes and Measures: The main outcomes were overall survival, progression-free survival, and response rates. Overall survival and progression-free survival were estimated by Kaplan-Meier methods. Multivariable Cox proportional hazards regression was used to examine associations of key clinical variables with survival; a χ2 test and logistic regression were used to assess associations with response rate. Results: The study cohort consisted of 212 patients, of whom 165 (77.8%) were male, 148 (69.8%) were former or current smokers, and 66 (31.1%) had an Eastern Cooperative Oncology Group (ECOG) performance status of 2 or greater; median age was 63.2 years (IQR, 57.2-71.2 years). Primary tumor sites included the oropharynx (99 [46.7%]), oral cavity (61 [28.8%]), and larynx or hypopharynx (52 [24.5%]). Most (126 [59.4%]) received CPI as first-line systemic therapy, and 23 (10.8%) received combination CPI with chemotherapy. The overall response rate was 30.1%. Estimated 1-year overall survival was 51.8% (95% CI, 44.5%-58.8%), and estimated 1-year progression-free survival was 9.4% (95% CI, 5.0%-15.5%). Median overall survival was 12.9 months (IQR, 4.1-36.5 months), and median progression-free survival was 3.9 months (IQR, 1.9-17.8 months). Non-oral cavity primary site (vs oral cavity) was associated with improved overall survival (human papillomavirus-positive oropharynx: hazard ratio [HR], 0.567 [95% CI, 0.335-0.960]; all other sites: HR, 0.491 [95% CI, 0.298-0.810]), and T category of 4 at presentation (HR, 1.594; 95% CI, 1.062-2.394) and an ECOG performance status greater than 1 (HR, 2.720; 95% CI, 1.866-3.964) were associated with worse overall survival. Conclusions and Relevance: In this cohort study of patients with recurrent or metastatic HNSCC who received CPI therapy, the overall response rate was 30.1%. Patients with oral cavity cancer had worse overall survival compared with patients with HNSCC of other subsites. These findings support the use of CPI therapies for first- or second-line treatment of recurrent or metastatic HNSCC.

Women and Minorities in Otolaryngology: A Historical Perspective and Analysis of Current Representation.

Over the course of the last century, otolaryngology-head and neck surgery has made significant medical and surgical advancements. Several of these efforts are credited to women and minorities despite their having faced systemic barriers to entering medical schools and the medical professions. This article highlights some of these pioneering doctors and their contributions to the field. Additionally, the current representation of women and minorities in otolaryngology residency programs and the gender and racial disparities in academic positions are reviewed. The need for mentorship during undergraduate medical education to improve diversity and inclusion within this surgical subspecialty is reinforced.

Rapid conformational epitope mapping of anti-gp120 antibodies with a designed mutant panel displayed on yeast.

gp120 is a substrate for protein engineering both for human immunodeficiency virus (HIV) immunogen design and as a bait for isolating anti-HIV antibodies from patient samples. In this work, we describe the display of a stripped core gp120 on the yeast cell surface. Validation against a panel of neutralizing antibodies confirms that yeast-displayed gp120 presents the CD4 binding site in the correct conformation. We map the epitope of the broadly neutralizing anti-gp120 antibody VRC01 using both a random mutagenesis library and a defined mutant panel and find that the resultant epitope maps are consistent with one another and with the crystallographically identified contact residues. Mapping the VRC01-competitive antibodies b12 and b13 reveals energetic differences in their epitopes that are not obvious from existing crystal structures. These data suggest mutation sets that abrogate binding to broadly neutralizing antibodies with greater specificity than the canonical mutation D368R, useful in rapidly assessing the nature of a vaccine response.