Pancreatic cancer (PC) is a highly aggressive malignancy characterized by limited opportunities for early diagnosis and poor clinical outcomes, underscoring the need for minimally invasive biomarkers to improve detection and patient stratification. Given emerging evidence that mitochondrial DNA (mtDNA) alterations reflect cancer-related biological processes, this study investigated whether blood-derived mtDNA profiles could provide clinically relevant information in PC. In this exploratory study, whole-blood mtDNA from 33 PC patients and 10 healthy individuals were analyzed using next-generation sequencing to assess single-nucleotide variants (SNVs), allele frequencies, and mtDNA copy number. A total of 252 unique mtDNA SNVs were identified, including variants exclusive to PC patients, variants unique to controls, and variants shared between groups. While the overall SNV burden did not differ significantly between groups, PC patients showed distinct mutation distributions and allele frequency patterns, with cancer-exclusive variants occurring predominantly at low allele frequencies. Mutation hotspots were observed in the ND5, COI, and D-loop regions, implicating genes involved in oxidative phosphorylation and mtDNA maintenance. Although mtDNA copy number did not differ significantly between groups, greater variability was observed among PC patients and was associated with differences in survival outcomes. Overall, these findings indicate that blood-based mtDNA profiling captures biologically relevant variation associated with PC and supports further development of integrated mtDNA-based approaches for improved risk assessment and patient stratification.

Background: Systematic studies providing differentiated insight into the contribution of immunity directed against conserved and non-conserved epitopes of SARS-CoV-2 Spike on long-term protection are rare and insufficiently guide future pan-variant vaccine research. The present observational cohort study aimed to evaluate the correlation of neutralizing antibody levels and cellular immunity against the Spike protein with symptomatic Omicron breakthrough infection. Methods: Neutralizing antibody levels against multiple (sub)variants were analyzed 6 months following the second wild-type mRNA vaccination and 6 months after booster in 107 subjects using a multiplex surrogate virus neutralization assay. To assess cellular immunity, cytokine mRNA expression levels were determined after peptide pool stimulation in whole blood samples of a study subgroup. Results: Neutralizing antibody titers were found to serve as a reasonably reliable correlate of protection prior to booster immunization. However, the predictive power of neutralizing antibody titers was diminished after boosting. This loss appears to be due to a critical remodeling of the antibody repertoire—a process that was dose-dependent on pre-boost humoral immunity. Vaccination against Omicron infection was most effective when a balanced immune response to both conserved and non-conserved epitopes of the viral Spike protein was induced. While neutralizing antibodies against receptor-binding domain epitopes affected by mutations were specifically associated with protection from symptomatic variant infection, cellular immunity was most effective when targeting conserved Spike epitopes. Conclusions: Optimal long-term protection against Omicron infection requires balanced immunity to both conserved and non-conserved epitopes of the viral Spike protein. The limited availability of cross-neutralizing antibodies targeting non-conserved epitopes and their inherently lower efficacy renders them a limiting factor as humoral immunity wanes over time. Future pan-SARS-CoV-2 variant vaccines that primarily target conserved epitopes may therefore provide less effective long-term protection against symptomatic variant infection than vaccines targeting a broader epitope spectrum including both conserved and non-conserved epitopes.