Survivin Expression in Luminal Breast Cancer and Adjacent Normal Tissue for Immuno-Oncology Applications.

Survivin (BIRC5) is a tumor-associated antigen (TAA) overexpressed in various tumors but present at low to undetectable levels in normal tissue. Survivin is known to have a high expression in breast cancer (e.g., Ductal Carcinoma in situ (DCIS) and triple negative breast cancer). Previous studies have not compared survivin expression levels in DCIS tumor samples to levels in adjacent, normal breast tissue from the same patient. To ensure the effective use of survivin as a target for T cell immunotherapy of breast cancer, it is essential to ascertain the varying levels of survivin expression between DCIS tumor tissue samples and the adjacent normal breast tissue taken from the same patient simultaneously. Next-generation sequencing of RNA (RNA-seq) in normal breast tissue and tumor breast tissue from five women presenting with DCIS for lumpectomy was used to identify sequence variation and expression levels of survivin. The identity of both tumor and adjacent normal tissue samples were corroborated by histopathology. Survivin was overexpressed in human breast tissue tumor samples relative to the corresponding adjacent human normal breast tissue. Wild-type survivin transcripts were the predominant species identified in all tumor tissue sequenced. This study demonstrates upregulated expression of wild type survivin in DCIS tumor tissue versus normal breast tissue taken from the same patient at the same time, and provides evidence that developing selective cytotoxic T lymphocyte (CTL) immunotherapy for DCIS targeting survivin warrants further study.

Survivin (BIRC5) Peptide Vaccine in the 4T1 Murine Mammary Tumor Model: A Potential Neoadjuvant T Cell Immunotherapy for Triple Negative Breast Cancer: A Preliminary Study.

A triple negative breast cancer model using the murine 4T1 tumor cell line was used to explore the efficacy of an adjuvanted survivin peptide microparticle vaccine using tumor growth as the outcome metric. We first performed tumor cell dose titration studies to determine a tumor cell dose that resulted in sufficient tumor takes but allowed multiple serial measurements of tumor volumes, yet with minimal morbidity/mortality within the study period. Later, in a second cohort of mice, the survivin peptide microparticle vaccine was administered via intraperitoneal injection at the study start with a second dose given 14 days later. An orthotopic injection of 4T1 cells into the mammary tissue was performed on the same day as the administration of the second vaccine dose. The mice were followed for up to 41 days with subcutaneous measurements of tumor volume made every 3-4 days. Vaccination with survivin peptides was associated with a peptide antigen-specific gamma interferon enzyme-linked immunosorbent spot response in the murine splenocyte population but was absent from the control microparticle group. At the end of the study, we found that vaccination with adjuvanted survivin peptide microparticles resulted in statistically significant slower primary tumor growth rates in BALB/c mice challenged with 4T1 cells relative to the control peptideless vaccination group. These studies suggest that T cell immunotherapy specifically targeting survivin might be an applicable neoadjuvant immunotherapy therapy for triple negative breast cancer. More preclinical studies and clinical trials are needed to explore this concept further.

Analysis of well-annotated next-generation sequencing data reveals increasing cases of SARS-CoV-2 reinfection with Omicron.

SARS-CoV-2 has extensively mutated creating variants of concern (VOC) resulting in global infection surges. The Omicron VOC reinfects individuals exposed to earlier variants of SARS-CoV-2 at a higher frequency than previously seen for non-Omicron VOC. An analysis of the sub-lineages associated with an Omicron primary infection and Omicron reinfection reveals that the incidence of Omicron-Omicron reinfections is occurring over a shorter time interval than seen after a primary infection with a non-Omicron VOC. Our analysis suggests that a single infection from SARS-CoV-2 may not generate the protective immunity required to defend against reinfections from emerging Omicron lineages. This analysis was made possible by Next-generation sequencing (NGS) of a Danish cohort with clinical metadata on both infections occurring in the same individual. We suggest that the continuation of COVID-19 NGS and inclusion of clinical metadata is necessary to ensure effective surveillance of SARS-CoV-2 genomics, assist in treatment and vaccine development, and guide public health recommendations.

A Synthetic Peptide CTL Vaccine Targeting Nucleocapsid Confers Protection from SARS-CoV-2 Challenge in Rhesus Macaques.

BACKGROUND: Persistent transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has given rise to a COVID-19 pandemic. Several vaccines, conceived in 2020, that evoke protective spike antibody responses are being deployed in mass public health vaccination programs. Recent data suggests, however, that as sequence variation in the spike genome accumulates, some vaccines may lose efficacy. METHODS: Using a macaque model of SARS-CoV-2 infection, we tested the efficacy of a peptide-based vaccine targeting MHC class I epitopes on the SARS-CoV-2 nucleocapsid protein. We administered biodegradable microspheres with synthetic peptides and adjuvants to rhesus macaques. Unvaccinated control and vaccinated macaques were challenged with 1 × 108 TCID50 units of SARS-CoV-2, followed by assessment of clinical symptoms and viral load, chest radiographs, and sampling of peripheral blood and bronchoalveolar lavage (BAL) fluid for downstream analysis. RESULTS: Vaccinated animals were free of pneumonia-like infiltrates characteristic of SARS-CoV-2 infection and presented with lower viral loads relative to controls. Gene expression in cells collected from BAL samples of vaccinated macaques revealed a unique signature associated with enhanced development of adaptive immune responses relative to control macaques. CONCLUSIONS: We demonstrate that a room temperature stable peptide vaccine based on known immunogenic HLA class I bound CTL epitopes from the nucleocapsid protein can provide protection against SARS-CoV-2 infection in nonhuman primates.

Paired SARS-CoV-2 spike protein mutations observed during ongoing SARS-CoV-2 viral transfer from humans to minks and back to humans.

A mutation analysis of SARS-CoV-2 genomes collected around the world sorted by sequence, date, geographic location, and species has revealed a large number of variants from the initial reference sequence in Wuhan. This analysis also reveals that humans infected with SARS-CoV-2 have infected mink populations in the Netherlands, Denmark, United States, and Canada. In these animals, a small set of mutations in the spike protein receptor binding domain (RBD), often occurring in specific combinations, has transferred back into humans. The viral genomic mutations in minks observed in the Netherlands and Denmark show the potential for new mutations on the SARS-CoV-2 spike protein RBD to be introduced into humans by zoonotic transfer. Our data suggests that close attention to viral transfer from humans to farm animals and pets will be required to prevent build-up of a viral reservoir for potential future zoonotic transfer.

Anti-IL-6 Versus Anti-IL-6R Blocking Antibodies to Treat Acute Ebola Infection in BALB/c Mice: Potential Implications for Treating Cytokine Release Syndrome.

Cytokine release syndrome (CRS) is known to be a factor in morbidity and mortality associated with acute viral infections including those caused by filoviruses and coronaviruses. IL-6 has been implicated as a cytokine negatively associated with survival after filovirus and coronavirus infection. However, IL-6 has also been shown to be an important mediator of innate immunity and important for the host response to an acute viral infection. Clinical studies are now being conducted by various researchers to evaluate the possible role of IL-6 blockers to improve outcomes in critically ill patients with CRS. Most of these studies involve the use of anti-IL-6R monoclonal antibodies (α-IL-6R mAbs). We present data showing that direct neutralization of IL-6 with an α-IL-6 mAb in a BALB/c Ebolavirus (EBOV) challenge model produced a statistically significant improvement in outcome compared with controls when administered within the first 24 h of challenge and repeated every 72 h. A similar effect was seen in mice treated with the same dose of α-IL-6R mAb when the treatment was delayed 48 h post-challenge. These data suggest that direct neutralization of IL-6, early during the course of infection, may provide additional clinical benefits to IL-6 receptor blockade alone during treatment of patients with virus-induced CRS.