Prostate cancer immunotherapy: Improving clinical outcomes with a multi-pronged approach.

Cancer immunotherapy has gained traction in recent years owing to remarkable tumor clearance in some patients. Despite the notable success of immune checkpoint blockade (ICB) in multiple malignancies, engagement of the immune system for targeted prostate cancer (PCa) therapy is still in its infancy. Multiple factors contribute to limited response, including the heterogeneity of PCa, the cold tumor microenvironment, and a low number of neoantigens. Significant effort is being invested in improving immune-based PCa therapies. This review is a summary of the status of immunotherapy in treating PCa, with a discussion of multiple immune modalities, including vaccines, adoptively transferred T cells, and bispecific T cell engagers, some of which are undergoing clinical trials. In addition, this review also focuses on emerging mechanism-based small-molecule tyrosine kinase inhibitors with immune modulatory properties that, either as single agents or in combination with other immunotherapies, have the potential to improve clinical outcomes.

Identification of a T-bethi Quiescent Exhausted CD8 T Cell Subpopulation That Can Differentiate into TIM3+CX3CR1+ Effectors and Memory-like Cells.

Persistent Ag induces a dysfunctional CD8 T cell state known as "exhaustion" characterized by PD-1 expression. Nevertheless, exhausted CD8 T cells retain functionality through continued differentiation of progenitor into effector cells. However, it remains ill-defined how CD8 T cell effector responses are sustained in situ. In this study, we show using the mouse chronic lymphocytic choriomeningitis virus infection model that CX3CR1+ CD8 T cells contain a T-bet-dependent TIM3-PD-1lo subpopulation that is distinct from the TIM3+CX3CR1+PD-1+ proliferative effector subset. The TIM3-CX3CR1+ cells are quiescent and express a low but significant level of the transcription factor TCF-1, demonstrating similarity to TCF-1hi progenitor CD8 T cells. Furthermore, following the resolution of lymphocytic choriomeningitis virus viremia, a substantial proportion of TCF-1+ memory-like CD8 T cells show evidence of CX3CR1 expression during the chronic phase of the infection. Our results suggest a subset of the CX3CR1+ exhausted population demonstrates progenitor-like features that support the generation of the CX3CR1+ effector pool from the TCF-1hi progenitors and contribute to the memory-like pool following the resolution of viremia.

Prevention of mitochondrial genomic instability in yeast by the mitochondrial recombinase Mhr1.

Mitochondrial (mt) DNA encodes factors essential for cellular respiration, therefore its level and integrity are crucial. ABF2 encodes a mitochondrial DNA-binding protein and its null mutation (Δabf2) induces mtDNA instability in Saccharomyces cerevisiae. Mhr1 is a mitochondrial recombinase that mediates the predominant form of mtDNA replication and acts in mtDNA segregation and the repair of mtDNA double-stranded breaks (DSBs). However, the involvement of Mhr1 in prevention of mtDNA deletion mutagenesis is unknown. In this study we used Δabf2 mhr1-1 double-mutant cells, which lose mitochondrial function in media containing fermentable carbon sources, to investigate whether Mhr1 is a suppressor of mtDNA deletion mutagenesis. We used a suppresivity assay and Southern blot analysis to reveal that the Δabf2 mutation causes mtDNA deletions rather than an mtDNA-lacking (ρ0) phenotype, and observed that mtDNA deletions are exacerbated by an additional mhr1-1 mutation. Loss of respiratory function due to mtDNA fragmentation occurred in ∆mhr1 and ∆abf2 mhr1-1 cells. However, exogenous introduction of Mhr1 into Δabf2 mhr1-1 cells significantly rescued respiratory growth, suggesting that Mhr1-driven homologous mtDNA recombination prevents mtDNA instability.

Regulation of Small Mitochondrial DNA Replicative Advantage by Ribonucleotide Reductase in Saccharomyces cerevisiae.

Small mitochondrial genomes can behave as selfish elements by displacing wild-type genomes regardless of their detriment to the host organism. In the budding yeast Saccharomyces cerevisiae, small hypersuppressive mtDNA transiently coexist with wild-type in a state of heteroplasmy, wherein the replicative advantage of the small mtDNA outcompetes wild-type and produces offspring without respiratory capacity in >95% of colonies. The cytosolic enzyme ribonucleotide reductase (RNR) catalyzes the rate-limiting step in dNTP synthesis and its inhibition has been correlated with increased petite colony formation, reflecting loss of respiratory function. Here, we used heteroplasmic diploids containing wild-type (rho+) and suppressive (rho-) or hypersuppressive (HS rho-) mitochondrial genomes to explore the effects of RNR activity on mtDNA heteroplasmy in offspring. We found that the proportion of rho+ offspring was significantly increased by RNR overexpression or deletion of its inhibitor, SML1, while reducing RNR activity via SML1 overexpression produced the opposite effects. In addition, using Ex Taq and KOD Dash polymerases, we observed a replicative advantage for small over large template DNA in vitro, but only at low dNTP concentrations. These results suggest that dNTP insufficiency contributes to the replicative advantage of small mtDNA over wild-type and cytosolic dNTP synthesis by RNR is an important regulator of heteroplasmy involving small mtDNA molecules in yeast.

Eukaryotic translation initiation factor 5A (eIF5A) is essential for HIF-1α activation in hypoxia.

The eukaryotic initiation factor 5A (eIF5A) is an essential protein involved in translation elongation and cell proliferation. eIF5A undergoes several post-translational modifications including hypusination and acetylation. Hypusination is indispensable for the function of eIF5A. On the other hand, the precise function of acetylation remains unknown, but it may render the protein inactive since hypusination blocks acetylation. Here, we report that acetylation of eIF5A increases under hypoxia. During extended hypoxic periods an increase in the level of eIF5A acetylation correlated with a decrease in HIF-1α, suggesting involvement of eIF5A activity in HIF-1α expression under hypoxia. Indeed, suppression of eIF5A by siRNA oligo-mediated knockdown or treatment with GC7, a deoxyhypusine synthase inhibitor, led to significant reduction of HIF-1α activity. Furthermore, knockdown of eIF5A or GC7 treatment reduced tumor spheroid formation with a concomitant decrease in HIF-1α expression. Our results suggest that functional, hypusinated eIF5A is necessary for HIF-1α expression during hypoxia and that eIF5A is an attractive target for cancer therapy.

Mitochondrial fission proteins Fis1 and Mdv1, but not Dnm1, play a role in maintenance of heteroplasmy in budding yeast.

In budding yeast, the mitochondrial DNA (mtDNA) replication pathway involving the homologous DNA pairing protein Mhr1 promotes mitochondrial allele segregation. Mitochondrial fusion facilitates the recombination-mediated replication pathway; however, the role of fission remains largely unknown. By monitoring mitochondrial allele segregation during zygotic division, we found that the absence of fission proteins Fis1 or Mdv1, but not Dnm1, resulted in increased initial homoplasmy levels and decreased mtDNA copy number. However, decreases in mtDNA copy number alone were not sufficient for rapid establishment of homoplasmy, suggesting that inhibiting the activities of certain fission proteins promotes homoplasmy by reducing the number of mtDNA segregation units.