Oncology

Jiayi He

Yujia Xia

Hongji Zhang

Dmitry Genkin

George Tetz

Methods: Subcutaneous MC38 (MSI-H adenocarcinoma of the colon) tumors were established in 6-week-old mice. Treatment was initiated six days after tumor inoculation with daily intraperitoneal (i.p) injections of DNase I followed by i.p anti-PD-1 treatment administered every 3 days. Animals were divided into treatment groups: control, DNase alone, anti-PD-1 alone and combination of DNase I with anti-PD-1 (n = 6 mice/ group).

Results: Tumors were successfully established after 6 days of inoculation. These tumors expressed high neutrophil infiltration and NETs. To determine the effects of combination therapy on tumor progression, we measured tumor volume and time to progression (TTP). We found that mice treated with combination DNase I and anti-PD-1 had significantly lower mean tumor volume compared to anti-PD-1 alone (1372.68mm³ vs 2193.20 mm³, p-value = 0.043). TTP was higher in the combination group compared to anti-PD-1 treatment alone (median time to progression 22 vs 17 days, p-value = 0.004). Next we sought to determine the mechanism behind this effect by investigating if NETs blockade modulated the tumor microenvironment. We found that treatment with DNase I decreased exhaustion of CD8+ T-cells in the tumor compared to no treatment.

Conclusions: Targeting NETs in combination with immune checkpoint inhibition may be an option to improve response rate to immune checkpoint inhibitors through decreasing exhausted CD8+ T-cells.

Methods

Paenibacillus sp. strain VT-400 was isolated from the saliva of four children with acute lymphoblastic leukemia. The genome was annotated using RAST and the NCBI Prokaryotic Genome Annotation Pipeline to characterize features of antibiotic resistance and virulence factors. Susceptibility to antibiotics was determined by the Kirby–Bauer disc diffusion method. We used a mouse model of pneumonia to study virulence in vivo. Mice were challenged with 7.5 log10–9.5 log10 CFU, and survival was monitored over 7 days. Bacterial load was measured in the lungs and spleen of surviving mice 48 h post-infection to reveal bacterial invasion and dissemination.

Results

Whole-genome sequencing revealed a large number of virulence factors such as hemolysin D and CD4+ T cell-stimulating antigen. Furthermore, the strain harbors numerous antibiotic resistance genes, including small multidrug resistance proteins, which have never been previously found in the Paenibacillus genus. We then compared the presence of antibiotic resistance genes against results from antibiotic susceptibility testing. Paenibacillus sp. strain VT-400 was found to be resistant to macrolides such as erythromycin and azithromycin, as well as to chloramphenicol and trimethoprim–sulphamethoxazole. Finally, the isolate caused mortality in mice infected with ≥8.5 log10 CFU.

Conclusions

Based on our results and on the available literature, there is yet no strong evidence that shows Paenibacillus species as an opportunistic pathogen in immunocompromised patients. However, the presence of spore-forming bacteria with virulence and antibiotic resistance genes in such patients warrants special attention because infections caused by spore-forming bacteria are poorly treatable.