DSP-0509, a systemically available TLR7 agonist, exhibits combination effect with immune checkpoint blockade by activating anti-tumor immune effects.

TLR7 is an innate immune receptor that recognizes single-stranded RNAs, and its activation leads to anti-tumor immune effects. Although it is the only approved TLR7 agonist in cancer therapy, imiquimod is allowed to be administered with topical formulation. Thus, systemic administrative TLR7 agonist is expected in terms of expanding applicable cancer types. Here, we demonstrated the identification and characterization of DSP-0509 as a novel small-molecule TLR7 agonist. DSP-0509 is designed to have unique physicochemical features that could be administered systemically with a short half-life. DSP-0509 activated bone marrow-derived dendritic cells (BMDCs) and induced inflammatory cytokines including type I interferons. In the LM8 tumor-bearing mouse model, DSP-0509 reduced tumor growth not only in subcutaneous primary lesions but also in lung metastatic lesions. DSP-0509 inhibited tumor growth in several syngeneic tumor-bearing mouse models. We found that the CD8+ T cell infiltration of tumor before treatment tended to be positively correlated with anti-tumor efficacy in several mouse tumor models. The combination of DSP-0509 with anti-PD-1 antibody significantly enhanced the tumor growth inhibition compared to each monotherapy in CT26 model mice. In addition, the effector memory T cells were expanded in both the peripheral blood and tumor, and rejection of tumor re-challenge occurred in the combination group. Moreover, synergistic anti-tumor efficacy and effector memory T cell upregulation were also observed for the combination with anti-CTLA-4 antibody. The analysis of the tumor-immune microenvironment by using the nCounter assay revealed that the combination of DSP-0509 with anti-PD-1 antibody enhanced infiltration by multiple immune cells including cytotoxic T cells. In addition, the T cell function pathway and antigen presentation pathway were activated in the combination group. We confirmed that DSP-0509 enhanced the anti-tumor immune effects of anti-PD-1 antibody by inducing type I interferons via activation of dendritic cells and even CTLs. In conclusion, we expect that DSP-0509, a new TLR7 agonist that synergistically induces anti-tumor effector memory T cells with immune checkpoint blockers (ICBs) and can be administered systemically, will be used in the treatment of multiple cancers.

Intravenous administration of the selective toll-like receptor 7 agonist DSR-29133 leads to anti-tumor efficacy in murine solid tumor models which can be potentiated by combination with fractionated radiotherapy.

Strategies to augment anti-cancer immune responses have recently demonstrated therapeutic utility. To date clinical success has been achieved through targeting co-inhibitory checkpoints such as CTLA-4, PD-1, and PD-L1. However, approaches that target co-activatory pathways are also being actively being developed. Here we report that the novel TLR7-selective agonist DSR-29133 is well tolerated in mice and leads to acute immune activation. Administration of DSR-29133 leads to the induction of IFNα/γ, IP-10, TNFα, IL-1Ra and IL-12p70, and to a reduction in tumor burden in syngeneic models of renal cancer (Renca), metastatic osteosarcoma (LM8) and colorectal cancer (CT26). Moreover, we show that the efficacy of DSR-29133 was significantly improved when administered in combination with low-dose fractionated radiotherapy (RT). Effective combination therapy required weekly administration of DSR-29133 commencing on day 1 of a fractionated RT treatment cycle, whereas no enhancement of radiation response was observed when DSR-29133 was administered at the end of the fractionated RT cycle. Combined therapy resulted in curative responses in a high proportion of mice bearing established CT26 tumors which was dependent on the activity of CD8+ T-cells but independent of CD4+ T-cells and NK/NKT cells. Moreover, long-term surviving mice originally treated with DSR-29133 and RT were protected by a tumor-specific memory immune response which could prevent tumor growth upon rechallenge. These results demonstrate that DSR-29133 is a potent selective TLR7 agonist that when administered intravenously can induce anti-tumor immune responses that can be further enhanced through combination with low-dose fractionated RT.

Novel carbapenem antibiotics for parenteral and oral applications: in vitro and in vivo activities of 2-aryl carbapenems and their pharmacokinetics in laboratory animals.

SM-295291 and SM-369926 are new parenteral 2-aryl carbapenems with strong activity against major causative pathogens of community-acquired infections such as methicillin-susceptible Staphylococcus aureus, Streptococcus pneumoniae (including penicillin-resistant strains), Streptococcus pyogenes, Enterococcus faecalis, Klebsiella pneumoniae, Moraxella catarrhalis, Haemophilus influenzae (including ß-lactamase-negative ampicillin-resistant strains), and Neisseria gonorrhoeae (including ciprofloxacin-resistant strains), with MIC(90)s of ≤ 1 µg/ml. Unlike tebipenem (MIC(50), 8 µg/ml), SM-295291 and SM-369926 had no activity against hospital pathogens such as Pseudomonas aeruginosa (MIC(50), ≥ 128 µg/ml). The bactericidal activities of SM-295291 and SM-369926 against penicillin-resistant S. pneumoniae and ß-lactamase-negative ampicillin-resistant H. influenzae were equal or superior to that of tebipenem and greater than that of cefditoren. The therapeutic efficacies of intravenous administrations of SM-295291 and SM-369926 against experimentally induced infections in mice caused by penicillin-resistant S. pneumoniae and ß-lactamase-negative ampicillin-resistant H. influenzae were equal or superior to that of tebipenem and greater than that of cefditoren, respectively, reflecting their in vitro activities. SM-295291 and SM-369926 showed intravenous pharmacokinetics similar to those of meropenem in terms of half-life in monkeys (0.4 h) and were stable against human dehydropeptidase I. SM-368589 and SM-375769, which are medoxomil esters of SM-295291 and SM-369926, respectively, showed good oral bioavailability in rats, dogs, and monkeys (4.2 to 62.3%). Thus, 2-aryl carbapenems are promising candidates that show an ideal broad spectrum for the treatment of community-acquired infections, including infections caused by penicillin-resistant S. pneumoniae and ß-lactamase-negative ampicillin-resistant H. influenzae, have low selective pressure on antipseudomonal carbapenem-resistant nosocomial pathogens, and allow parenteral, oral, and switch therapies.

Experimental verification of the efficacy of optimized two-step infusion therapy with meropenem using an in vitro pharmacodynamic model and Monte Carlo simulation.

In this study we compared the efficacy of a theoretically optimized two-step infusion therapy (OTIT; rapid first-step infusion and slow second-step infusion) to the efficacies of prolonged infusion therapy (PIT) and traditional 0.5 h infusion therapy (TIT) with meropenem against Pseudomonas aeruginosa using an in vitro pharmacodynamic model and a Monte Carlo simulation. In the in vitro pharmacodynamic model, the bactericidal effect against P. aeruginosa was evaluated for 8 h, which is the usual dosing interval of meropenem. It was confirmed that the durability of the bactericidal effect of OTIT (0.25-1 g/0.5 h + 0.25-1 g/4 h t.i.d.) was almost equal to that of PIT and superior to that of TIT (0.5-2 g/0.5-4 h t.i.d.). In addition, the initial bactericidal effects of OTIT were superior to those of the prolonged 4 h infusion. In the Monte Carlo simulation study, the probability of target attainments (PTAs) of all dosing regimens of OTIT at MICs of 2-8 µg/ml were apparently superior to those of TIT and 4- and 6 h-PIT, when the target therapeutic condition for serious life-threatening infections was the achievement of both the percentage of the dosing interval at which the drug concentration exceeds the MIC (%T(>MIC)) ≥ 50% and the peak level divided by the MIC (Cmax/MIC) ≥ 4. Especially, the PTAs of the dosing regimens of 0.25-1 g/0.5 h + 0.25-1 g/4-6 h were excellent at MICs of 2-8 µg/ml. Against recent clinical isolates of P. aeruginosa in Japan, the dosing regimens of OTIT provided higher PTAs compared with those of TIT and 4- and 6 h-PIT. These results suggested that OTIT with sufficient pharmacokinetic conditions could be useful for enhancing the therapeutic efficacy of meropenem against serious life-threatening infections.

Pharmacodynamics of SMP-601 (PTZ601) against vancomycin-resistant Enterococcus faecium and methicillin-resistant Staphylococcus aureus in neutropenic murine thigh infection models.

SMP-601 (also known as PTZ601, PZ-601, or SM-216601) is a novel parenteral carbapenem with potent activity against multidrug-resistant gram-positive pathogens, including vancomycin-resistant Enterococcus faecium (VREF) and methicillin-resistant Staphylococcus aureus (MRSA). The pharmacodynamics of SMP-601 against VREF and MRSA were investigated in neutropenic murine thigh infection models. The percentage of the dosing interval that the unbound SMP-601 concentration exceeded the MIC (f%T>MIC) was the pharmacokinetic-pharmacodynamic parameter that correlated most closely with efficacy with R(2) values of 0.81 to 0.84 for two strains of VREF and 0.92 to 0.93 for two strains of MRSA, whereas the R(2) values for the area under the concentration-time curve from 0 to 24 h divided by the MIC were 0.12 to 0.89, and the R(2) values for the peak level divided by the MIC were 0 to 0.22. The f%T>MIC levels required for static or killing efficacy against two strains of VREF (9 to 19%) apparently were lower than those against two strains of MRSA (23 to 37%). These results suggested that SMP-601 showed time-dependent in vivo efficacy against VREF and MRSA, and SMP-601 had a sufficient therapeutic effect against VREF infections at lower exposure conditions compared to those for with MRSA infections.

The mode of action of 2-(thiazol-2-ylthio)-1beta-methylcarbapenems against Pseudomonas aeruginosa: the impact of outer membrane permeability and the contribution of MexAB-OprM efflux system.

The mode of action of a series of 2-(4-dihydropyrrolylthiazol-2-ylthio) and 2-(4-tetrahydropyridinylthiazol-2-ylthio)-1beta-methylcarbapenem analogues against Pseudomonas aeruginosa was investigated with regard to contributions of the affinity for penicillin binding proteins (PBPs), the outer membrane permeability, and the effect of the MexAB-OprM efflux system. In this series of carbapenems, the introduction of a substituent in C-2 side chain with a change in physicochemical properties affected the antipseudomonal activity depending on the molecular weight. However, these structural modifications did not affect the affinity for pseudomonal PBPs significantly. It was confirmed that the affinity for PBPs was not an important determinant of the antipseudomonal activity of this series of carbapenems. OprD porin-deficiency did not affect antipseudomonal activity either. On the other hand, the MIC of these carbapenems against P. aeruginosa significantly decreased in the presence of outer membrane permeabilizer. This result strongly suggests that the cause of the relatively low antipseudomonal activity of these carbapanems is their low permeability through the outer membrane of P. aeruginosa. And also, in the presence of outer membrane permeabilizer, the MICs against MexAB-OprM deficient mutants remarkably decreased and were very close to the value of the IC(50) for pseudomonal PBPs. From this result, it was clear that the effect of the MexAB-OprM efflux system was also an important determinant of antipseudomonal activity of these carbapenems. In conclusion, the major determinants of the antipseudomonal activity of the 2-(thiazol-2-ylthio)-1beta-methylcarbapenems are the outer membrane permeability and the effect of the MexAB-OprM efflux system, not the affinity for pseudomonal PBPs.

Synthesis and properties of 2'-O,4'-C-methyleneoxymethylene bridged nucleic acid.

A novel bridged nucleic acid (BNA) analogue, 2'-O,4'-C-methyleneoxymethylene bridged nucleic acid (2',4'-BNA(COC)), was synthesized and incorporated into oligonucleotides. The 2',4'-BNA(COC) modified oligonucleotides showed high binding affinity with an RNA complement and significant enzymatic stability against snake venom phosphodiesterase.

In vitro and in vivo antibacterial activities of SM-216601, a new broad-spectrum parenteral carbapenem.

SM-216601 is a novel parenteral 1beta-methylcarbapenem. In agar dilution susceptibility testing, the MIC of SM-216601 for 90% of the methicillin-resistant Staphylococcus aureus (MRSA) strains tested (MIC(90)) was 2 microg/ml, which was comparable to those of vancomycin and linezolid. SM-216601 was also very potent against Enterococcus faecium, including vancomycin-resistant strains (MIC(90) = 8 microg/ml). SM-216601 exhibited potent activity against penicillin-resistant Streptococcus pneumoniae, ampicillin-resistant Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis, with MIC(90)s of less than 0.5 microg/ml, and intermediate activity against Citrobacter freundii, Enterobacter cloacae, Serratia marcescens, and Pseudomonas aeruginosa. The therapeutic efficacy of SM-216601 against experimentally induced infections in mice caused by S. aureus, E. faecium, E. coli, and P. aeruginosa reflected its in vitro activity and plasma level. Thus, SM-216601 is a promising candidate for nosocomial bacterial infections caused by a wide range of gram-positive and gram-negative bacteria, including multiresistant pathogens.

[Transitional cell carcinoma of the ureter with inverted proliferation: a difficult case to make a differential diagnosis with ureteral polyp].

A 59-year-old man was admitted to our hospital in June 2001 for evaluation of an asymptomatic microscopic hematuria. One year prior to presentation, he had a spontaneous discharge of a left ureteral stone. Excretory urography and retrograde pyelography showed a filling defect in the middle portion of the left ureter. Cystoscopic examination did not reveal any abnormality, and urinary cytology was class I. Cold cup biopsy was performed under ureteroscopy, and pathology revealed inflammatory fibrovascular tissue but with no malignancy. Selective washing cytology was class III, whereas selective washing cytology done at the referring hospital was reported to be class V. Under a preoperative diagnosis of a left fibroepithelial ureteral polyp or a transitional cell carcinoma, left segmental ureterectomy was performed. The tumor was 5 x 5 x 5 mm in size, pedunculated, and smooth-surfaced. Intraoperative pathological examination of a frozen section showed an inverted type transitional cell carcinoma. Therefore, a left nephroureterectomy was performed, and the final histopathological examination confirmed an inverted type transitional cell carcinoma of grade 2. The patient is healthy and free of disease 15 months after operation. We also reviewed the current literature relating to transitional cell carcinomas of the ureter with inverted proliferation.

Synthesis and properties of oligonucleotides containing novel 2',4'-BNA analogues (2',4'-BNACOC).

Preorganization of the nucleoside into proper conformation is one of the most promising approaches to develop the oligonucleotides strongly interacting with nucleic acid targets in a sequence-specific manner. We designed and synthesized the 2',4'-BNACOC monomer as a novel bridged nucleic acid analogue possessing a fixed N-type sugar conformation, and also successfully achieved its incorporation into oligonucleotides. The 2',4'-BNACOC modified oligonucleotides were found to have selective and strong binding-affinity for complementary RNA rather than DNA, and to show an excellent nuclease resistance ability.