Exploiting bacteria for cancer immunotherapy.

Immunotherapy has revolutionized the treatment of cancer but continues to be constrained by limited response rates, acquired resistance, toxicities and high costs, which necessitates the development of new, innovative strategies. The discovery of a connection between the human microbiota and cancer dates back 4,000 years, when local infection was observed to result in tumour eradication in some individuals. However, the true oncological relevance of the intratumoural microbiota was not recognized until the turn of the twentieth century. The intratumoural microbiota can have pivotal roles in both the pathogenesis and treatment of cancer. In particular, intratumoural bacteria can either promote or inhibit cancer growth via remodelling of the tumour microenvironment. Over the past two decades, remarkable progress has been made preclinically in engineering bacteria as agents for cancer immunotherapy; some of these bacterial products have successfully reached the clinical stages of development. In this Review, we discuss the characteristics of intratumoural bacteria and their intricate interactions with the tumour microenvironment. We also describe the many strategies used to engineer bacteria for use in the treatment of cancer, summarizing contemporary data from completed and ongoing clinical trials. The work described herein highlights the potential of bacteria to transform the landscape of cancer therapy, bridging ancient wisdom with modern scientific innovation.

Tumor Pre-Targeting System Using Streptavidin-Expressing Bacteria.

PURPOSE: A major obstacle to targeted cancer therapy is identifying suitable targets that are specifically and abundantly expressed by solid tumors. Certain bacterial strains selectively colonize solid tumors and can deliver genetically encoded cargo molecules to the tumor cells. Here, we engineered bacteria to express monomeric streptavidin (mSA) in tumors, and developed a novel tumor pre-targeting system by visualizing the presence of tumor-associated mSA using a biotinylated imaging probe. PROCEDURES: We constructed a plasmid expressing mSA fused to maltose-binding protein and optimized the ribosome binding site sequence to increase solubility and expression levels. E. coli MG1655 was transformed with the recombinant plasmid, expression of which is driven by the pBAD promotor. Expression of mSA was induced by L-arabinose 4 days after injection of bacteria into mice bearing CT26 mouse colon carcinoma cells. Selective accumulation of mSA in tumor tissues was visualized by optical imaging after administration of a biotinylated fluorescent dye. Counting of viable bacterial cells was also performed. RESULTS: Compared with a conventional system, the novel expression system resulted in significantly higher expression of mSA and sustained binding to biotin. Imaging signals in tumor tissues were significantly stronger in the mSA-expressing group than in non-expressing group (P = 0.0005). Furthermore, the fluorescent signal in tumor tissues became detectable again after multiple inductions with L-arabinose. The bacterial counts in tumor tissues showed no significant differences between conditions with and without L-arabinose (P = 0.45). Western blot analysis of tumor tissues confirmed expression and binding of mSA to biotin. CONCLUSIONS: We successfully engineered tumor-targeting bacteria carrying a recombinant plasmid expressing mSA, which was targeted to, and expressed in, tumor tissues. These data demonstrate the potential of this novel tumor pre-targeting system when combined with biotinylated imaging probes or therapeutic agents.

Engineering of an EPHA2-Targeted Monobody for the Detection of Colorectal Cancer.

BACKGROUND/AIM: Colorectal cancer (CRC) is the third most common cancer worldwide, and is second only to lung cancer with respect to cancer-related deaths. Noninvasive molecular imaging using established markers is a new emerging method to diagnose CRC. The human ephrin receptor family type-A 2 (hEPHA2) oncoprotein is overexpressed at the early, but not late, stages of CRC. Previously, we reported development of an E1 monobody that is specific for hEPHA2-expressing cancer cells both in vitro and in vivo. Herein, we investigated the ability of the E1 monobody to detect hEPHA2 expressing colorectal tumors in a mouse model, as well as in CRC tissue. MATERIALS AND METHODS: The expression of hEPHA2 on the surface of CRC cells was analyzed by western blotting and flow cytometry. The targeting efficacy of the E1 monobody for CRC cells was examined by flow cytometry, and immunofluorescence staining. E1 conjugated to the Renilla luciferase variant 8 (Rluc8) reporter protein was used for in vivo imaging in mice. Additionally, an enhanced green fluorescence protein (EGFP) conjugated E1 monobody was used to check the ability of the E1 monobody to target CRC tissue. RESULTS: The E1 monobody bound efficiently to hEPHA2-expressing CRC cell lines, and E1 conjugated to the Rluc8 reporter protein targeted tumor tissues in mice transplanted with HCT116 CRC tumor cells. Finally, E1-EGFP stained tumor tissues from human CRC patients, showing a pattern similar to that of an anti-hEPHA2 antibody. CONCLUSION: The E1 monobody has utility as an EPHA2 targeting agent for the detection of CRC.

The combination of calreticulin-targeting L-ASNase and anti-PD-L1 antibody modulates the tumor immune microenvironment to synergistically enhance the antitumor efficacy of radiotherapy.

Radiotherapy (RT) triggers immunogenic cell death (ICD). L-ASNase, which catalyzes the conversion of asparagine (Asn), thereby depleting it, is used in the treatment of blood cancers. In previous work, we showed that CRT3LP and CRT4LP, PASylated L-ASNases conjugated to the calreticulin (CRT)-specific monobodies CRT3 and CRT4, increase the efficacy of ICD-inducing chemotherapy. Here, we assessed their efficacy in tumor-bearing mice treated with RT. Methods: Monobody binding was evaluated by in silico molecular docking analysis. The expression and cellular localization of ecto-CRT were assessed by confocal imaging and flow cytometry. The antitumor effect and the roles of CRT3LP and CRT4LP in irradiation (IR)-induced ICD in tumors were analyzed by ELISA, immunohistochemistry, and immune analysis methods. Results: Molecular docking analysis showed that CRT3 and CRT4 monobodies were stably bound to CRT. Exposure to 10 Gy IR decreased the viability of CT-26 and MC-38 tumor cells in a time-dependent manner until 72 h, and increased the expression of the ICD marker ecto-CRT (CRT exposed on the cell surface) and the immune checkpoint marker PD-L1 until 48 h. IR enhanced the cytotoxicity of CRT3LP and CRT4LP in CT-26 and MC-38 tumor cells, and increased reactive oxygen species (ROS) levels. In mice bearing CT-26 and MC-38 subcutaneous tumors treated with 6 Gy IR, Rluc8-conjugated CRT-specific monobodies (CRT3-Rluc8 and CRT4-Rluc8) specifically targeted tumor tissues, and CRT3LP and CRT4LP increased total ROS levels in tumor tissues, thereby enhancing the antitumor efficacy of RT. Tumor tissues from these mice showed increased mature dendritic, CD4+ T, and CD8+ T cells and pro-inflammatory cytokines (IFNγ and TNFα) and decreased regulatory T cells, and the expression of tumor cell proliferation markers (Ki67 and CD31) was downregulated. These data indicate that the combination of IR and CRT-targeting L-ASNases activated and reprogramed the immune system of the tumor microenvironment. Consistent with these data, an immune checkpoint inhibitor (anti-PD-L1 antibody) markedly increased the therapeutic efficacy of combined IR and CRT-targeting L-ASNases. Conclusion: CRT-specific L-ASNases are useful as additive drug candidates in tumors treated with RT, and combination treatment with anti-PD-L1 antibody increases their therapeutic efficacy.

Prognostic Significance Of Sequential 18f-fdg Pet/Ct During Frontline Treatment Of Peripheral T Cell Lymphomas.

BACKGROUND/AIMS: The prognostic significance of 18F-fluorodeoxyglucose (FDG)-positron emission tomography-computed tomography (PET/CT) in peripheral T-cell lymphomas (PTCLs) are controversial. We explored the prognostic impact of sequential 18F-FDG PET/CT during frontline chemotherapy of patients with PTCLs. METHODS: In total, 143 patients with newly diagnosed PTCLs were included. Sequential 18F-FDG PET/CTs were performed at the time of diagnosis, during chemotherapy, and at the end of chemotherapy. The baseline total metabolic tumor volume (TMTV) was calculated using the the standard uptake value with a threshold method of 2.5. RESULTS: A baseline TMTV of 457.0 cm3 was used to categorize patients into high and low TMTV groups. Patients with a requirehigh TMTV had shorter progression-free survival (PFS) and overall survival (OS) than those with a low TMTV (PFS, 9.8 vs. 26.5 mo, p = 0.043; OS, 18.9 vs. 71.2 mo, p = 0.004). The interim 18F-FDG PET/CT response score was recorded as 1, 2-3, and 4-5 according to the Deauville criteria. The PFS and OS showed significant differences according to the interim 18F-FDG PET/CT response score (PFS, 120.7 vs. 34.1 vs. 5.1 mo, p < 0.001; OS, not reached vs. 61.1 mo vs. 12.1 mo, p < 0.001). CONCLUSION: The interim PET/CT response based on visual assessment predicts disease progression and survival outcome in PTCLs. A high baseline TMTV is associated with a poor response to anthracycline-based chemotherapy in PTCLs. However, TMTV was not an independent predictor for PFS in the multivariate analysis.

Mucosal TLR5 activation controls healthspan and longevity.

Addressing age-related immunological defects through therapeutic interventions is essential for healthy aging, as the immune system plays a crucial role in controlling infections, malignancies, and in supporting tissue homeostasis and repair. In our study, we show that stimulating toll-like receptor 5 (TLR5) via mucosal delivery of a flagellin-containing fusion protein effectively extends the lifespan and enhances the healthspan of mice of both sexes. This enhancement in healthspan is evidenced by diminished hair loss and ocular lens opacity, increased bone mineral density, improved stem cell activity, delayed thymic involution, heightened cognitive capacity, and the prevention of pulmonary lung fibrosis. Additionally, this fusion protein boosts intestinal mucosal integrity by augmenting the surface expression of TLR5 in a certain subset of dendritic cells and increasing interleukin-22 (IL-22) secretion. In this work, we present observations that underscore the benefits of TLR5-dependent stimulation in the mucosal compartment, suggesting a viable strategy for enhancing longevity and healthspan.

Reprogramming a Doxycycline-Inducible Gene Switch System for Bacteria-Mediated Cancer Therapy.

PURPOSE: Attenuated Salmonella typhimurium is a potential biotherapeutic antitumor agent because it can colonize tumors and inhibit their growth. The present study aimed to develop a doxycycline (Doxy)-inducible gene switch system in attenuated S. typhimurium and assess its therapeutic efficacy in various tumor-bearing mice models. PROCEDURES: A Doxy-inducible gene switch system comprising two plasmids was engineered to trigger the expression of cargo genes (Rluc8 and clyA). Attenuated S. typhimurium carrying Rluc8 were injected intravenously into BALB/c mice bearing CT26 tumors, and bioluminescence images were captured at specified intervals post-administration of doxycycline. The tumor-suppressive effects of bacteria carrying clyA were evaluated in BALB/c mice bearing CT26 tumors and in C57BL/6 mice bearing MC38 tumors. RESULTS: Expression of the fimE gene, induced only in the presence of Doxy, triggered a unidirectional switch of the POXB20 promoter to induce expression of the cargo genes. The switch event was maintained over a long period of bacterial culture. After intravenous injection of transformed Salmonella into mice bearing CT26 tumors, the bacteria transformed with the Doxy-inducible gene switch system for Rluc8 targeted only tumor tissues and expressed the payloads 2 days after Doxy treatment. Notably, bacteria carrying the Doxy-inducible gene switch system for clyA effectively suppressed tumor growth and prolonged survival, even after just one Doxy induction. CONCLUSIONS: These results suggest that attenuated S. typhimurium carrying this novel gene switch system elicited significant therapeutic effects through a single induction triggering and were a potential biotherapeutic agent for tumor therapy.

Synthesis and evaluation of 18F-labeled procainamide as a PET imaging agent for malignant melanoma.

Malignant melanoma has an aggressive nature and a high metastatic propensity resulting in the highest mortality rate of any skin cancer. In this study, we synthesized 18F-labeled procainamide (PCA) for detection of melanoma using positron emission tomography (PET), and evaluated its biological characteristics. The non-decay-corrected radiochemical yield of 18F-PCA was 10-15% and its in vitro stability was over 98% for 2 h. At 1 h, cellular uptake of 18F-PCA was 3.8-fold higher in a group with the presence of l-tyrosine than in a non-l-tyrosine-treated group. Furthermore, 18F-PCA permitted visualization of B16F10 (mouse melanoma) xenografts on microPET after intravenous injection, and was retained in the tumor for 60 min, with a high tumor-to-liver uptake ratio. 18F-PCA showed specific melanoma uptake in primary lesions with a high melanin targeting ability in small animal models. 18F-PCA may have potential as a PET imaging agent for direct melanoma detection.

Enhanced resolution and sensitivity acoustic-resolution photoacoustic microscopy with semi/unsupervised GANs.

Acoustic-resolution photoacoustic microscopy (AR-PAM) enables visualization of biological tissues at depths of several millimeters with superior optical absorption contrast. However, the lateral resolution and sensitivity of AR-PAM are generally lower than those of optical-resolution PAM (OR-PAM) owing to the intrinsic physical acoustic focusing mechanism. Here, we demonstrate a computational strategy with two generative adversarial networks (GANs) to perform semi/unsupervised reconstruction with high resolution and sensitivity in AR-PAM by maintaining its imaging capability at enhanced depths. The b-scan PAM images were prepared as paired (for semi-supervised conditional GAN) and unpaired (for unsupervised CycleGAN) groups for label-free reconstructed AR-PAM b-scan image generation and training. The semi/unsupervised GANs successfully improved resolution and sensitivity in a phantom and in vivo mouse ear test with ground truth. We also confirmed that GANs could enhance resolution and sensitivity of deep tissues without the ground truth.

Synergistic Antitumor Effect of Combined Radiotherapy and Engineered Salmonella typhimurium in an Intracranial Sarcoma Mouse Model.

Intracranial sarcoma is an uncommon aggressive cancer with a poor prognosis and a high recurrence rate. Although postoperative adjuvant radiotherapy (RT) is the most recommended treatment strategy, it does not significantly improve survival rates. In this study, we used an attenuated Salmonella typhimurium strain engineered to secrete Vibrio vulnificus flagellin B (SLpFlaB) as an immunotherapy to assist with the antitumor effects of RT on intracranial sarcoma. In vitro, the expression of γH2AX and cleaved caspase-3 was analyzed by Western blot. In vivo detection of SLpFlaB colonization time in tumors was measured using an in vivo imaging system (IVIS). Tumor growth delay and elimination were demonstrated in an intracranial mouse model, and the distribution of macrophages, M1 macrophages, and CD8+ cells after treatment was measured using FACS analysis. Our findings in vitro suggest that combination therapy increases S-180 radiosensitivity, the expression of DNA double-strand breaks, and programmed cell death. In vivo, combination treatment causes intracranial sarcoma to be eliminated without tumor recurrence and redistribution of immune cells in the brain, with data showing the enhanced migration and infiltration of CD8+ T cells and macrophages, and an increased proportion of M1 macrophage polarization. Compared to RT alone, the combination therapy enhanced the radiosensitivity of S-180 cells, promoted the recruitment of immune cells at the tumor site, and prevented tumor recurrence. This combination therapy may provide a new strategy for treating intracranial sarcomas.

Synergistic cancer immunotherapy utilizing programmed Salmonella typhimurium secreting heterologous flagellin B conjugated to interleukin-15 proteins.

The use of appropriately designed immunotherapeutic bacteria is an appealing approach to tumor therapy because the bacteria specifically target tumor tissue and deliver therapeutic payloads. The present study describes the engineering of an attenuated strain of Salmonella typhimurium deficient in ppGpp biosynthesis (SAM) that could secrete Vibrio vulnificus flagellin B (FlaB) conjugated to human (hIL15/FlaB) and mouse (mIL15/FlaB) interleukin-15 proteins in the presence of L-arabinose (L-ara). These strains, named SAMphIF and SAMpmIF, respectively, secreted fusion proteins that retained bioactivity of both FlaB and IL15. SAMphIF and SAMpmIF inhibited the growth of MC38 and CT26 subcutaneous (sc) tumors in mice and increased mouse survival rate more efficiently than SAM expressing FlaB alone (SAMpFlaB) or IL15 alone (SAMpmIL15 and SAMphIL15), although SAMpmIF had slightly greater antitumor activity than SAMphIF. The mice treated with these bacteria showed enhanced macrophage phenotype shift, from M2-like to M1-like, as well as greater proliferation and activation of CD4+ T, CD8+ T, NK, and NKT cells in tumor tissues. After tumor eradication by these bacteria, ≥50% of the mice show no evidence of tumor recurrence upon rechallenge with the same tumor cells, indicating that they had acquired long-term immune memory. Treatment of mice of 4T1 and B16F10 highly malignant sc tumors with a combination of these bacteria and an immune checkpoint inhibitor, anti-PD-L1 antibody, significantly suppressed tumor metastasis and increased mouse survival rate. Taken together, these findings suggest that SAM secreting IL15/FlaB is a novel therapeutic candidate for bacterial-mediated cancer immunotherapy and that its antitumor activity is enhanced by combination with anti-PD-L1 antibody.

The anticancer effect of PASylated calreticulin-targeting L-ASNase in solid tumor bearing mice with immunogenic cell death-inducing chemotherapy.

L-Asparaginase (L-ASNase), a bacterial enzyme that degrades asparagine, has been commonly used in combination with several chemical drugs to treat malignant hematopoietic cancers such as acute lymphoblastic leukemia (ALL). In contrast, the enzyme was known to inhibit the growth of solid tumor cells in vitro, but not to be effective in vivo. We previously reported that two novel monobodies (CRT3 and CRT4) bound specifically with calreticulin (CRT) exposed on tumor cells and tissues during immunogenic cell death (ICD). Here, we engineered L-ASNases conjugated with monobodies at the N-termini and PAS200 tags at the C-termini (CRT3LP and CRT4LP). These proteins were expected to possess four monobody and PAS200 tag moieties, which did not disrupt the L-ASNase conformation. These proteins were expressed 3.8-fold more highly in E. coli than those without PASylation. The purified proteins were highly soluble, with much greater apparent molecular weights than expected ones. Their affinity (Kd) against CRT was about 2 nM, 4-fold higher than that of monobodies. Their enzyme activity (∼6.5 IU/nmol) was similar to that of L-ASNase (∼7.2 IU/nmol), and their thermal stability was significantly increased at 55 °C. Their half-life times were > 9 h in mouse sera, about 5-fold longer than that of L-ASNase (∼1.8 h). Moreover, CRT3LP and CRT4LP bound specifically with CRT exposed on tumor cells in vitro, and additively suppressed the tumor growth in CT-26 and MC-38 tumor-bearing mice treated with ICD-inducing drugs (doxorubicin and mitoxantrone) but not with a non-ICD-inducing drug (gemcitabine). All data indicated that PASylated CRT-targeted L-ASNases enhanced the anticancer efficacy of ICD-inducing chemotherapy. Taken together, L-ASNase would be a potential anticancer drug for treating solid tumors.

Genetic Programming by Nitric Oxide-Sensing Gene Switch System in Tumor-Targeting Bacteria.

Recent progress in synthetic biology has enabled bacteria to respond to specific disease signals to perform diagnostic and/or therapeutic tasks. Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) colonization of tumors results in increases in nitric oxide (NO) levels, suggesting that NO may act as a candidate inducer of tumor-specific gene expression. The present study describes a NO-sensing gene switch system for triggering tumor-specific gene expression in an attenuated strain of S. Typhimurium. The genetic circuit was designed to sense NO via NorR, thus initiating the expression of FimE DNA recombinase. This was found to lead sequentially to the unidirectional inversion of a promoter region (fimS), which induced the expression of target genes. Target gene expression in bacteria transformed with the NO-sensing switch system was triggered in the presence of a chemical source of NO, diethylenetriamine/nitric oxide (DETA/NO) in vitro. In vivo results revealed that the gene expression is tumor-targeted, and specific to NO generated by inducible nitric oxide synthase (iNOS) after S. Typhimurium colonization. These results showed that NO was a promising inducer to finely tune the expression of target genes carried by tumor-targeting bacteria.

Flagellin synergistically enhances anti-tumor effect of EGFRvIII peptide in a glioblastoma-bearing mouse brain tumor model.

BACKGROUND: Glioblastoma (GBM) is the most aggressive type of brain tumor with heterogeneity and strong invasive ability. Treatment of GBM has not improved significantly despite the progress of immunotherapy and classical therapy. Epidermal growth factor receptor variant III (EGFRvIII), one of GBM-associated mutants, is regarded as an ideal therapeutic target in EGFRvIII-expressed GBM patients because it is a tumor-specific receptor expressed only in tumors. Flagellin B (FlaB) originated from Vibrio vulnificus, is known as a strong adjuvant that enhances innate and adaptive immunity in various vaccine models. This study investigated whether FlaB synergistically could enhance the anti-tumor effect of EGFRvIII peptide (PEGFRvIII). METHODS: EGFRvIII-GL261/Fluc cells were used for glioblastoma-bearing mouse brain model. Cell-bearing mice were inoculated with PBS, FlaB alone, PEGFRvIII alone, and PEGFRvIII plus FlaB. Tumor growth based on MRI and the survival rate was investigated. T cell population was examined by flow cytometry analysis. Both cleaved caspase-3 and CD8 + lymphocytes were shown by immunohistochemistry (IHC) staining. RESULTS: The PEGFRvIII plus FlaB group showed delayed tumor growth and increased survival rate when compared to other treatment groups. As evidence of apoptosis, cleaved caspase-3 expression and DNA disruption were more increased in the PEGFRvIII plus FlaB group than in other groups. In addition, the PEGFRvIII plus FlaB group showed more increased CD8 + T cells and decreased Treg cells than other treatment groups in the brain. CONCLUSIONS: FlaB can enhance the anti-tumor effect of PEGFRvIII by increasing CD8 + T cell response in a mouse brain GBM model.

Multistage adaptive noise reduction technique for optical resolution photoacoustic microscopy.

Photoacoustic microscopy has received great attention due to the benefits of the optical resolution contrast as well as its superior spatial resolution and relatively deep depth. Like other imaging modalities, photoacoustic images suffer from noise, and filtering techniques are required to remove them. To overcome the noise, we proposed a combination of filters, including an adaptive median filter, an effective filter for impulsive noise, and a nonlocal means filter, an effective filter for background noise, for noise removal and image quality enhancement. Our proposed method enhanced the signal-to-noise ratio by 16 dB in an in vivo study compared to the traditional image reconstruction approach and preserved the image detail with minimal blurring, which usually occurs when filtering. These experimental results verified that the proposed adaptive multistage denoising techniques could effectively improve image quality under noisy data acquisition conditions, providing a strong foundation for photoacoustic microscopy with limited laser power.

Comparison of Anticancer Activities and Biosafety Between Salmonella enterica Serovar Typhimurium ΔppGpp and VNP20009 in a Murine Cancer Model.

Salmonella Typhimurium defective in guanosine 5'-diphosphate-3'-diphosphate (ppGpp) synthesis (ΔppGpp) is an attenuated strain with good biosafety and excellent anticancer efficacy. It has been widely applied in preclinical studies of anticancer therapy for various types of solid cancer. VNP20009 is another genetically modified auxotrophic strain with 108-kb deletion, purI- , msbB- , and many single nucleotide polymorphisms (SNPs); it has shown promising therapeutic efficacy in various preclinical tumor models and entered phase I clinical trials. Here, the invasion activities and virulence of ΔppGpp were obviously lower than those of the VNP20009 strain when tested with cancer cells in vitro. In addition, the MC38 tumor-bearing mice showed comparable cancer suppression when treated with ΔppGpp or VNP20009 intravenously. However, the ΔppGpp-treated mice showed 16.7% of complete cancer eradication and prolonged survival in mice, whereas VNP20009 showed higher toxicity to animals, even with equal tumor size individually. Moreover, we found decreased levels of inflammatory cytokines in circulation but strengthened immune boost in tumor microenvironments of ΔppGpp-treated mice. Therefore, the engineered ΔppGpp has high potential for cancer therapeutics, and it is a promising option for future clinical cancer therapy.

Molecular imaging approaches to facilitate bacteria-mediated cancer therapy.

Bacteria-mediated cancer therapy is a potential therapeutic strategy for cancer that has unique properties, including broad tumor-targeting ability, various administration routes, the flexibility of delivery, and facilitating the host's immune responses. The molecular imaging of bacteria-mediated cancer therapy allows the therapeutically injected bacteria to be visualized and confirms the accurate delivery of the therapeutic bacteria to the target lesion. Several hurdles make bacteria-specific imaging challenging, including the need to discriminate therapeutic bacterial infection from inflammation or other pathologic lesions. To realize the full potential of bacteria-specific imaging, it is necessary to develop bacteria-specific targets that can be associated with an imaging assay. This review describes the current status of bacterial imaging techniques together with the advantages and disadvantages of several imaging modalities. Also, we describe potential targets for bacterial-specific imaging and related applications.

Pattern of F-18 FDG Uptake in Colon Cancer after Bacterial Cancer Therapy Using Engineered Salmonella Typhimurium: A Preliminary In Vivo Study.

Purpose: Bacterial cancer therapy (BCT) research using engineered Salmonella typhimurium has increased in recent years. 2-Deoxy-2[18F] fluoro-D-glucose positron emission tomography (FDG PET) is widely used in cancer patients to detect cancer, monitor treatment responses, and predict prognoses. The aim of this pilot study was to investigate FDG uptake patterns in a mouse tumor model after BCT. Procedures. BCT was performed via the intravenous injection of attenuated S. typhimurium (SLΔppGpp/lux) into female mice bearing a tumor (derived from CT26 murine colon cancer cells) in the right thigh. 18F-FDG PET images acquired before BCT and at different time points after BCT. In vivo bioluminescence imaging confirmed bacterial presence in the tumor. The tumor volume, standardized uptake value (SUV) of FDG (SUVmax and SUVmean), early SUV reduction%, and normalized tumor volume change were analyzed. Results: Early after BCT (1 or 2 days post-injection (dpi)), FDG tumor uptake decreased in 10 out of 11 mice and then increased at later stages. FDG uptake before BCT was correlated with normalized tumor volume change after BCT. Early FDG reduction% after BCT was correlated with normalized volume change after BCT. Conclusions: Early after BCT, FDG tumor uptake decreased and then increased at later stages. The higher the FDG tumor uptake before BCT, the better the BCT response. FDG uptake patterns were related to tumor volume change after BCT. Therefore, FDG uptake was a good candidate for evaluating BCT.

In vivo imaging of invasive aspergillosis with 18F-fluorodeoxysorbitol positron emission tomography.

Invasive aspergillosis is a critical complication in immunocompromised patients with hematologic malignancies or with viral pneumonia caused by influenza virus or SARS­CoV­2. Although early and accurate diagnosis of invasive aspergillosis can maximize clinical outcomes, current diagnostic methods are time-consuming and poorly sensitive. Here, we assess the ability of 2-deoxy-2-18F-fluorosorbitol (18F-FDS) positron emission tomography (PET) to specifically and noninvasively detect Aspergillus infections. We show that 18F-FDS PET can be used to visualize Aspergillus fumigatus infection of the lungs, brain, and muscles in mouse models. In particular, 18F-FDS can distinguish pulmonary aspergillosis from Staphylococcus aureus infection, both of which induce pulmonary infiltrates in immunocompromised patients. Thus, our results indicate that the combination of 18F-FDS PET and appropriate clinical information may be useful in the differential diagnosis and localization of invasive aspergillosis.