KDM5B predicts temozolomide-resistant subclones in glioblastoma.

Adaptive plasticity to the standard chemotherapeutic temozolomide (TMZ) leads to glioblastoma progression. Here, we examine early stages of this process in patient-derived cellular models, exposing the human lysine-specific demethylase 5B (KDM5B) as a prospective indicator for subclonal expansion. By integration of a reporter, we show its preferential activity in rare, stem-like ALDH1A1+ cells, immediately increasing expression upon TMZ exposure. Naive, genetically unmodified KDM5Bhigh cells phosphorylate AKT (pAKT) and act as slow-cycling persisters under TMZ. Knockdown of KDM5B reverses pAKT levels, simultaneously increasing PTEN expression and TMZ sensitivity. Pharmacological inhibition of PTEN rescues the effect. Interference with KDM5B subsequent to TMZ decreases cellular vitality, and clonal tracing with DNA barcoding demonstrates high individual levels of KDM5B to predict subclonal expansion already before TMZ exposure. Thus, KDM5Bhigh treatment-naive cells preferentially contribute to the dynamics of drug resistance under TMZ. These findings may serve as a cornerstone for future biomarker-assisted clinical trials.

Cross-sectional study: prevalence of oedema disease Escherichia coli (EDEC) in weaned piglets in Germany at pen and farm levels.

BACKGROUND: Escherichia coli bacteria capable of producing the toxin Stx2e and possessing F18-fimbriae (edema disease E. coli, EDEC) are considered causative agents of porcine oedema disease. This disease, which usually occurs in piglets shortly after weaning, has a high lethality in affected animals and can lead to high economic losses in piglet rearing. The aim of this cross-sectional field study was to determine the prevalence of EDEC in weaned piglets in Germany at pen and farm levels. RESULTS: Ninety-nine farms with unknown history of infections with shigatoxin-producing E. coli (STEC) and oedema disease were sampled. On each farm, up to five pens were selected for sampling (n = 481). The piglets in these pens were at an age 1-3 weeks after weaning. Single faecal samples (n = 2405) and boot swabs (n = 479) were collected from the floor. On 50 farms, cotton ropes were additionally used to collect oral fluid samples (n = 185) and rope wash out samples (n = 231) from the selected pens. All samples were analyzed by bacterial culture combined with a duplex PCR for the presence of the corresponding genes stx2e and fedA (major subunit protein of F18 fimbriae). In addition, whole DNA specimens extracted from boot swabs, oral fluid samples, and rope wash out samples were directly examined by duplex PCR for DNA of stx2e and fedA. A pen was classified as positive if at least one of the samples, regardless of the technique, yielded a positive result in the PCR, and farms were considered positive if at least one pen was classified as positive. Overall, genes stx2e and fedA were found simultaneously in 24.9% (95% CI 22.1-29.1%) of sampled pens and in 37.4% (95% CI 27.9-47.7%) of sampled farms. Regardless of the presence of F18-fimbriae, Escherichia coli encoding for Stx2e (STEC-2e) were found in 35.1% (95% CI 31.0-39.1%) of the pens and 53.5% (95% CI 44.4-63.6%) of the farms sampled. CONCLUSIONS: Escherichia coli strains considered capable to cause oedema disease in swine (EDEC) are highly prevalent in the surveyed pig producing farms in Germany. Due to intermittent shedding of EDEC and a potentially low within-farm prevalence, we recommend a combination of different sampling techniques for EDEC monitoring at pen and farm levels. Further studies are needed to understand which STEC-2e strains really pose the risk of causing severe porcine disease.

A Sequential Targeting Strategy Interrupts AKT-Driven Subclone-Mediated Progression in Glioblastoma.

PURPOSE: Therapy resistance and fatal disease progression in glioblastoma are thought to result from the dynamics of intra-tumor heterogeneity. This study aimed at identifying and molecularly targeting tumor cells that can survive, adapt, and subclonally expand under primary therapy. EXPERIMENTAL DESIGN: To identify candidate markers and to experimentally access dynamics of subclonal progression in glioblastoma, we established a discovery cohort of paired vital cell samples obtained before and after primary therapy. We further used two independent validation cohorts of paired clinical tissues to test our findings. Follow-up preclinical treatment strategies were evaluated in patient-derived xenografts. RESULTS: We describe, in clinical samples, an archetype of rare ALDH1A1+ tumor cells that enrich and acquire AKT-mediated drug resistance in response to standard-of-care temozolomide (TMZ). Importantly, we observe that drug resistance of ALDH1A1+ cells is not intrinsic, but rather an adaptive mechanism emerging exclusively after TMZ treatment. In patient cells and xenograft models of disease, we recapitulate the enrichment of ALDH1A1+ cells under the influence of TMZ. We demonstrate that their subclonal progression is AKT-driven and can be interfered with by well-timed sequential rather than simultaneous antitumor combination strategy. CONCLUSIONS: Drug-resistant ALDH1A1+/pAKT+ subclones accumulate in patient tissues upon adaptation to TMZ therapy. These subclones may therefore represent a dynamic target in glioblastoma. Our study proposes the combination of TMZ and AKT inhibitors in a sequential treatment schedule as a rationale for future clinical investigation.