Discovery and Synthesis of Heterobifunctional Degraders of Rearranged during Transfection (RET) Kinase.

We describe the design, synthesis, and structure-activity relationship (SAR) of heterobifunctional RET ligand-directed degraders (LDDs) derived from three different second-generation RET inhibitors. These LDDs are composed of a target binding motif (TBM) that binds to the RET protein, a linker, and a cereblon binding motif (CBM) as the E3 ligase recognition unit. This led to the identification of a series of pyrazolopyridine-based heterobifunctional LDDs, as exemplified by compound 39. LDD 39 demonstrated high in vitro inhibitory and degradation potency against both RET wild-type and the two representative mutants, V804M and G810R. Importantly, in PK/PD studies, 39 exhibited a differentiated and favorable in vivo profile compared to the corresponding tyrosine kinase inhibitor (TKI), compound 3. Robust and sustained degradation of total-RET (tRET) protein and inhibition of phospho-RET (pRET) signaling were observed in TPC-1 xenograft tumors driven by RET and the RET/G810R mutant following a single dose of LDD 39 at 15 and 75 mg/kg, respectively.

Diagnostic incidence and pitfalls of rete testis hyperplasia and hyaline globules in a multi-institutional study of 348 testicular germ cell tumors.

OBJECTIVES: The concept of rete hyperplasia with hyaline globules simulating testicular yolk sac tumor was first reported in a mostly retrospective review over 30 years ago. Nonetheless, we continue to encounter examples where this scenario resulted in misdiagnosis. Herein, we sought to investigate the incidence of rete hyperplasia/hyaline globules in germ cell tumors and their associated subtypes and hypothesize an etiology. METHODS: A consecutive series of 348 germ cell tumor orchiectomies was evaluated for the presence of rete hyperplasia and hyaline globules, with clinicopathologic features recorded. RESULTS: The incidence of rete hyperplasia and/or hyaline globules in our cohort was 30%, with 56% of specimens with rete hyperplasia containing concomitant hyaline globules. Hyaline globules were more often identified in specimens with nonfocal rete hyperplasia (78%) vs focal rete hyperplasia (22%). Absence of a yolk sac tumor component was seen in over half (61%) of orchiectomies with concurrent rete hyperplasia/hyaline globules (n = 105), inclusive of tumors with "pure" subtypes (ie, pure seminoma, pure teratoma, or pure embryonal carcinoma). Of these 105 specimens, rete invasion was seen in only 48%; notably, Paneth cell-like metaplasia was identified in efferent ductules/epididymis in 13%. CONCLUSIONS: Rete hyperplasia and hyaline globules are not uncommon findings in the setting of germ cell tumors (including occurrences in various pure/mixed germ cell tumors) and can show striking overlap with yolk sac tumor. We hypothesize that these histologic pitfalls evolve secondary to testicular obstruction by the tumor mass. Recognition of and distinguishing this morphologic mimicry is fundamental to guide appropriate clinical management.

Hyperpolarized 129Xe MRI and spectroscopy of gas exchange abnormalities in chronic hypersensitivity pneumonitis.

PURPOSE: To evaluate 3D gas-exchange functional imaging characteristics using 129Xe MRI in a group of study participants with chronic hypersensitivity pneumonitis (CHP) as compared with healthy control participants. METHODS: In this prospective study, 11 participants with clinical and CT findings of CHP (4M 7F, mean age 67 ± 6.1 years) as well as 41 healthy subjects (25M 16F, mean age 44 ± 18 years) were enrolled between 2017 and 2022 and underwent 129Xe MRI. Three-dimensional images of ventilation, interstitial membrane uptake, and RBC transfer were rendered into quantitative 3D maps relative to a healthy reference cohort. In addition, 129Xe spectroscopy was used to assess the RBC:membrane ratio (RBC:M), the oxygen-dependent RBC chemical shift, and cardiogenically-driven RBC oscillation amplitude. Differences between the CHP participants and healthy subjects were assessed using the two-sample t-test or Wilcoxon rank-sum test as appropriate. RESULTS: CHP participants demonstrated significant differences in 6 parameters (p < 0.001) including regions of reduced ventilation, increased membrane uptake, and reduced RBC transfer as compared to healthy subjects. Gas exchange abnormalities measured on spectroscopy included a reduced RBC:M, reduced RBC chemical shift, and increased RBC oscillation amplitude. CONCLUSION: In participants with CHP, 129Xe MRI demonstrated gas exchange abnormalities common to other fibrotic lung diseases including increased membrane uptake, deficits in RBC transfer, and reduced RBC:M. However, CHP participants also exhibited prominent ventilation abnormalities, which may be reflective of the airway-centric nature of the disease. Further, the high variability observed in the membrane uptake could suggest varying degrees of disease progression or activity.

Provider Perceptions and Use of Audit-Feedback and Communication Strategies to Improve Human Papillomavirus Vaccine Uptake.

OBJECTIVE: Human papillomavirus (HPV) vaccine uptake remains suboptimal among US adolescents. A cluster randomized trial was conducted at six primary care practices in southeast Minnesota to assess the impact of parent reminder-recall letters and provider audit-feedback reports on 11-12-year-old HPV vaccine uptake. Audit-feedback reports included access to a web toolkit with instruction on two communication approaches. We evaluated the process of the audit-feedback report intervention to inform future adaptations. METHODS: We sent a survey to providers assigned to the intervention and asked about their use and perceptions of the reports, web toolkit, the communication approaches, and HPV vaccine recommendation. RESULTS: Surveys from 95 providers were analyzed. Most (97.9%) recalled receiving audit-feedback reports, with 92.4% finding them somewhat to very easy to understand, 86% somewhat to very familiar with their content and objectives, and 69.9% using them five or more times in the past year. Few respondents (11.6%) recalled receiving access to the web toolkit. Web analytics showed that the toolkit was rarely used. Most reported familiarity with communication approaches but less than half reported that these positively impacted the tone of the clinical encounter. Higher familiarity with audit-feedback reports (OR=2.58) and perceived peer approval about using presumptive language (the first of two communication approaches) to recommend HPV vaccination (OR=2.16) correlated with higher frequency of vaccine recommendation. CONCLUSIONS: Implementation of the audit-feedback reports showed good acceptability. Low utilization of the web toolkit suggests a need to further examine provider preferences on delivery and usability of training materials.

Accurate Co-Localization of Luciferase Expression and Fluorescent Anti-CEA Antibody Targeting of Liver Metastases in an Orthotopic Mouse Model of Colon Cancer.

BACKGROUND: The present study aimed to validate the accuracy of a tumor-specific antibody to target liver metastases of colorectal cancer. METHODS: A humanized anti-CEA antibody conjugated to a fluorescent dye (M5A-IR800) was tested for targeting human colorectal cancer liver metastases (CRLMs) expressing luciferase in an orthotopic mouse model. Orthotopic mouse models of CRLMs were established by implanting fragments of a luciferase-expressing human colorectal cancer cell line, LS174T, in the liver of nude mice. Mice received 50 µg M5A-IR800 72 h prior to imaging. To test co-localization, bioluminescence imaging was performed using D-luciferin, which was given via intraperitoneal injection just prior to imaging. RESULTS: Tumors were able to be visualized non-invasively through the skin with the luciferase-luciferin signal. Intra-abdominal imaging showed accurate labeling of CRLMs with M5A-IR800, which co-localized with the luciferase-luciferin signal. CONCLUSIONS: The present results validate the accuracy of a tumor-specific anti-CEA antibody in targeting liver metastases of colorectal cancer.

Post-traumatic reactions and quality of life after pelvic exenteration for gynecologic cancer: a retrospective cohort study.

OBJECTIVE: We examined post-traumatic reactions and quality of life in women with recurrent gynecologic cancer who underwent a pelvic exenteration (PE), a potentially life-saving radical surgery associated with life-altering sequelae. METHODS: Twenty-one women who had completed PE at least 6 months prior completed the Impact of Event Scale-Revised, a measure of post-traumatic stress, the Post-Traumatic Growth Inventory, a measure of post-traumatic growth, the Center for Epidemiologic Studies-Depression Scale, and the European Organization for Research and Treatment of Cancer 30-item core Quality of Life Questionnaire. We examined the associations between these outcome variables, and quality of life scores were compared to normative values for the general and gynecologic cancer populations. RESULTS: Thirty percent of women reported clinically significant post-traumatic stress symptoms and 71% endorsed clinically significant depressive symptoms. More post-traumatic stress was associated with less post-traumatic growth, more depressive symptoms, and worse quality of life. In general, women's quality of life was worse than the general population but comparable to women with stage III-IV ovarian cancer and women with cervical cancer. Social functioning was markedly lower in our sample and women reported more pain, diarrhea, and financial difficulties post-PE compared to published norms for the general population and women with ovarian or cervical cancer. There were no differences in quality of life based on age, type of PE, type of urinary diversion, or cancer type. CONCLUSIONS: Findings support long-term continued symptom management and the ongoing rehabilitation of patients to optimize physical, psychological, and social well-being in PE survivorship.

Robotic manipulation of cardiomyocytes to identify gap junction modifiers for arrhythmogenic cardiomyopathy.

Arrhythmogenic cardiomyopathy (ACM) is a leading cause of sudden cardiac death among young adults. Aberrant gap junction remodeling has been linked to disease-causative mutations in plakophilin-2 (PKP2). Although gap junctions are a key therapeutic target, measurement of gap junction function in preclinical disease models is technically challenging. To quantify gap junction function with high precision and high consistency, we developed a robotic cell manipulation system with visual feedback from digital holographic microscopy for three-dimensional and label-free imaging of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). The robotic system can accurately determine the dynamic height changes in the cells' contraction and resting phases, microinject drug-treated healthy and diseased iPSC-CMs in their resting phase with constant injection depth across all cells, and deposit a membrane-impermeable dye that solely diffuses between cells through gap junctions for measuring the gap junction diffusion function. The robotic system was applied toward a targeted drug screen to identify gap junction modulators and potential therapeutics for ACM. Five compounds were found to dose-dependently enhance gap junction permeability in cardiomyocytes with PKP2 knockdown. In addition, PCO 400 (pinacidil) reduced beating irregularity in a mouse model of ACM expressing mutant PKP2 (R735X). These results highlight the utility of the robotic cell manipulation system to efficiently assess gap junction function in a relevant preclinical disease model, thus providing a technique to advance drug discovery for ACM and other gap junction-mediated diseases.

Comparing stand-alone endovascular embolization versus stereotactic radiosurgery in the treatment of arteriovenous malformations with Spetzler-Martin grades I-III: a propensity score matched study.

BACKGROUND: Arteriovenous malformations (AVMs) are uncommon cerebral lesions that can cause significant neurological complications. Surgical resection is the gold standard for treatment, but endovascular embolization and stereotactic radiosurgery (SRS) are viable alternatives. OBJECTIVE: To compare the outcomes of endovascular embolization versus SRS in the treatment of AVMs with Spetzler-Martin grades I-III. METHODS: This study combined retrospective data from 10 academic institutions in North America and Europe. Patients aged 1 to 90 years who underwent endovascular embolization or SRS for AVMs with Spetzler-Martin grades I-III between January 2010 and December 2023 were included. RESULTS: The study included 244 patients, including 84 who had endovascular embolization and 160 who had SRS. Before propensity score matching (PSM), complete obliteration at the last follow-up was achieved in 74.5% of the SRS group compared with 57.8% of the embolization group (OR=0.47; 95% CI 0.26 to 0.48; P=0.01). After propensity score matching, SRS still achieved significantly higher occlusion rates at last follow-up (78.9% vs 55.3%; OR=0.32; 95% CI 0.12 to 0.90; P=0.03).Hemorrhagic complications were higher in the embolization group than in the SRS group, although this difference did not reach statistical significance after PSM (13.2% vs 2.6%; OR=5.6; 95% CI 0.62 to 50.47; P=0.12). Similarly, re-treatment rate was higher in the embolization group (10.5% vs 5.3%; OR=2.11; 95% CI 0.36 to 12.31; P=0.40) compared with the SRS group. CONCLUSION: Our findings indicate that SRS has a significantly higher obliteration rate at last follow-up compared with endovascular embolization. Also, SRS has a higher tendency for fewer hemorrhagic complications and lower re-treatment rate. Further prospective studies are needed.

Paclitaxel chemotherapy disrupts microbiota-enterohepatic bile acid metabolism in mice.

Balanced interactions between the enteric microbiota and enterohepatic organs are essential to bile acid homeostasis, and thus normal gastrointestinal function. Disruption of these interactions by cancer treatment instigates bile acid malabsorption, leading to treatment delays, malnutrition, and decreased quality of life. However, the nature of chemotherapy-induced bile acid malabsorption remains poorly characterized with limited treatment options. Therefore, this study sought to characterize changes in hepatic, enteric, and microbial bile acid metabolism in a mouse model of chemotherapy-induced toxicity. Consistent with clinical bile acid malabsorption, chemotherapy increased fecal excretion of primary bile acids and water, while diminishing microbiome diversity, secondary bile acid formation, and small intestinal bile acid signaling. We identified new contributors to pathology of bile acid malabsorption in the forms of lipopolysaccharide-induced cholestasis and colonic crypt hyperplasia from reduced secondary bile acid signaling. Chemotherapy reduced markers of hepatic bile flow and bile acid synthesis, elevated markers of fibrosis and endotoxemia, and altered transcription of genes at all stages of bile acid metabolism. Primary hepatocytes exposed to lipopolysaccharide (but not chemotherapy) replicated chemotherapy-induced transcriptional differences, while gut microbial transplant into germ-free mice replicated very few differences. In the colon, chemotherapy-altered bile acid profiles (particularly higher tauromuricholic acid and lower hyodeoxycholic acid) coincided with crypt hyperplasia. Exposing primary colonoids to hyodeoxycholic acid reduced proliferation, while gut microbiota transplant enhanced proliferation. Together, these investigations reveal complex involvement of the entire microbiota-enterohepatic axis in chemotherapy-induced bile acid malabsorption. Interventions to reduce hepatic lipopolysaccharide exposure and enhance microbial bile acid metabolism represent promising co-therapies to cancer treatment.

The Impact of Spliceosome Inhibition in SF3B1-Mutated Uveal Melanoma.

Purpose: Unfortunately, treatment of patients with uveal melanoma (UM) with metastatic disease is limited. Twenty percent of patients with UM harbor a mutation in the splicing factor gene SF3B1, suggesting that aberrant spliceosome function plays a vital role in tumorigenesis. Splicing inhibitors exploit the preferential sensitivity of spliceosome-compromised leukemic cells to these compounds. Methods: We studied the effect of the splicing inhibitor E7107 using two UM cell lines and ex vivo cultured SF3B1- and BAP1-mutated primary UM tumor slices. These UM cell lines and ex vivo tumor slices were exposed for 24 hours to different concentrations of E7107. Tumor slices were stained with hematoxylin and eosin (H&E) and incubated with BAP1, MelanA, MIB-1, and caspase-3 antisera. Results: The E7107-exposed UM cell lines exhibited decreased cell viability and increased apoptosis, with the greatest effect on SF3B1-mutated UM cells. A similar effect on UM tumor slices was observed upon exposure to E7107. Additionally, RNA was isolated for differential isoform expression analysis. No significant difference in isoform usage was found genome-wide. However, specific genes were differentially expressed after E7107 treatment in the SF3B1-mutated samples. Moreover, E7107 had the greatest effect on intron retention. Conclusions: This study indicates/suggests that mutated SF3B1 UM cells are more sensitive to the splicing inhibitor E7107 than wild-type SF3B1 UM cells.

Timely Hip Surgery Access in Children With Cerebral Palsy: Unaffected by Social Disadvantage at a Large Urban Safety Net Hospital.

BACKGROUND: Socioeconomic disadvantage has been shown to limit timely access to pediatric orthopaedic care and can result in poor surgical outcomes. Insurance coverage has often served as a proxy for socioeconomic status; however, area deprivation index (ADI) and child opportunity index (COI) are more comprehensive measures of social determinants of health (SDOH). The treatment of hip displacement in children with cerebral palsy (CP) requires early radiographic identification and continuous surveillance, which may be impacted by SDOH. This study seeks to evaluate the influence of insurance, ADI, and COI on preoperative Reimer migration percentage and need for pelvic osteotomy during varus derotation osteotomy (VDRO) in children with CP. METHODS: This retrospective cohort study examined 219 patients with CP who underwent VDRO surgery for hip subluxation or dislocation at a tertiary referral center (135 male, mean age 7.9 y, SD: 2.9, range: 2.4 to 17.2; 17 GMFCS II, 21 GMFCS III, 89 GMFCS IV, 92 GMFCS V) from 2004 to 2022. Imaging and clinical documentation for patients with CP and hip displacement, age <18 years with ≥1 year of follow-up, treated with VDRO were reviewed. GMFCS level, preoperative Reimer migration percentages (MP), surgical details, and demographic and socioeconomic data were collected, and addresses were used to determine ADI (2018 version) and COI (2.0 database). The relationship of ADI, COI, and insurance type to preoperative Reimer MP of the more displaced hip and the need for pelvic osteotomy were analyzed with linear regressions and logistic regressions. RESULTS: The mean preoperative Reimer MP was 64.4% (SD: 25.0, range: 0 to 100). As expected, patients functioning at higher GMFCS levels presented with greater Reimer MPs. The average Reimer MP was 34.0 for GMFCS II, 44.2 for GMFCS III, 64.6 for GMFCS IV, and 74.5 for GMFCS V (P<0.01). The mean ADI state decile (1 to 10 scale) and COI (1 to 100 scale) for the cohort were 5.6 (SD: 2.2, range: 1 to 10) and 37.2 (SD: 28.1, range: 4 to 100), respectively. ADI (P=0.77), COI (P=0.30), and insurance type (P=0.78) were not related to preoperative Reimer MP. However, patients with lower ADIs (OR 0.83, 95% CI [0.70, 0.99], P=0.04) and higher COIs (OR 1.01, 95% CI [1.00, 1.03], P=0.03) underwent pelvic osteotomies at a higher rate. CONCLUSIONS: ADI, COI, and insurance type were not related to preoperative Reimer MP. Interestingly, greater social disadvantage was associated with a lower frequency of pelvic osteotomy at the time of VDRO. Our data demonstrate that at our institution, greater social disadvantage does not result in limited access to timely orthopaedic care for children with CP. This is likely due to adequate governmental insurance coverage for children with neuromuscular disorders in this state and the active involvement of pediatric orthopaedic surgeons in government-sponsored clinics, including ongoing hip screening programs for children with CP. These results provide hope that healthcare disparities can potentially be mitigated.

Missed Adjuvant Therapy in Human Papillomavirus Positive Oropharyngeal Squamous Cell Carcinoma.

OBJECTIVE: Investigate missed adjuvant therapy and associated disparities in overall survival (OS) for human papillomavirus-associated (HPV+) oropharyngeal squamous cell carcinoma (OPSCC). METHODS: The 2010-2017 National Cancer Database was queried for patients with surgically resected HPV+ OPSCC. Indications for adjuvant radiotherapy (aRT) included pT3-4 classification, pN2-3 classification, lymphovascular invasion, pathologic extranodal extension (pENE), and/or positive surgical margins (PSM). Indication(s) for adjuvant chemoradiotherapy (aCRT) included pENE and/or PSM. Multivariable logistic and Cox regression models were implemented. RESULTS: Of 5297 patients satisfying inclusion criteria, 4288 had indication(s) for aRT; 775 did not receive any adjuvant therapy and were considered as missing aRT. A total of 2234 patients had indication(s) for aCRT. Of these, 1383 (61.9%) received aCRT, 555 (24.8%) patients received aRT alone and were considered as having missed aCRT, and 296 (13.2%) did not receive any adjuvant therapy. Missed aRT and missed aCRT were each associated with age, treatment facility type, pN classification, and surgical margin status (p < 0.015). Among patients with indication(s) for aRT alone, OS of those receiving no adjuvant therapy, aRT alone, and aCRT was 90.0%, 94.8%, and 93.4%, respectively (p = 0.080). Among patients with indication(s) for aCRT, those receiving aRT alone and aCRT had similar OS (89.0% vs. 86.6%, p = 0.357) which was superior to receiving no adjuvant therapy (74.9%, p < 0.001). These patterns in OS persisted on multivariable Cox regression. CONCLUSION: Among patients with HPV+ OPSCC and indication(s) for aRT, missed aRT was not associated with worse OS. For patients with indication(s) for aCRT, aRT alone was associated with similar OS as aCRT. LEVEL OF EVIDENCE: 3 Laryngoscope, 2024.

Targeting Human Pancreatic Cancer with a Fluorophore-Conjugated Mucin 4 (MUC4) Antibody: Initial Characterization in Orthotopic Cell Line Mouse Models.

Background/Objectives: Pancreatic cancer is the third leading cause of death related to cancer. The only possible cure presently is complete surgical resection; however, this is limited by difficulty in clearly defining tumor margins. Enhancement of the visualization of pancreatic ductal adenocarcinoma (PDAC) tumor margins using near-infrared dye-conjugated tumor-specific antibodies was pioneered by using anti-CEA, anti-CA19.9, and anti-MUC5AC in orthotopic mouse models of pancreatic cancer. Recently, an antibody to Mucin 4 (MUC4) conjugated to a fluorescent probe has shown promise in targeting colon tumors in orthotopic mouse models. Methods: In the present study, we targeted pancreatic cancer using an anti-MUC4 antibody conjugated to IRDye800 (anti-MUC4-IR800) in orthotopic mouse models. Two pancreatic cancer human cell lines were used, SW1990 and CD18/HPAF. Results: Anti-MUC4-IR800 targeted the two pancreatic cancer cell line tumors in orthotopic mouse models with high tumor-to-pancreas ratios and high tumor-to-liver ratios, with greater targeting seen in SW1990. Conclusions: The present results suggest anti-MUC4-IR800's potential to be used in fluorescence-guided surgical resection of pancreatic cancer.

Quinoline-based compounds can inhibit diverse enzymes that act on DNA.

DNA methylation, as exemplified by cytosine-C5 methylation in mammals and adenine-N6 methylation in bacteria, is a key epigenetic process. Developing non-nucleoside inhibitors to cause DNA hypomethylation is crucial for treating various conditions without the toxicities associated with existing cytidine-based hypomethylating agents. This study characterized fifteen quinoline-based analogs, particularly compounds with additions like a methylamine (9) or methylpiperazine (11), which demonstrate similar low micromolar inhibitory potency against human DNMT1 and Clostridioides difficile CamA. These compounds (9 and 11) intercalate into CamA-bound DNA via the minor groove, causing a conformational shift that moves the catalytic domain away from the DNA. This study adds to the limited examples of DNA methyltransferases being inhibited by non-nucleotide compounds through DNA intercalation. Additionally, some quinoline-based analogs inhibit other DNA-interacting enzymes, such as polymerases and base excision repair glycosylases. Finally, compound 11 elicits DNA damage response via p53 activation in cancer cells.

A review of OSHA-permissible exposure limits for occupational carcinogens in relation to quantitative risk assessments based on epidemiological findings.

A very small proportion of all chemicals in commerce have occupational exposure limits (OELs) based on quantitative risk assessments which require estimates of exposure-response relationships (XRs). For only 18 of the 94 chemicals declared by NIOSH to be carcinogens were human XRs reported in or calculable from published reports. For the 18 carcinogens, 96 such XRs could be derived (corresponding to chemicals with multiple associated cancer end-points and/or multiple source studies). Twenty-four of 96 XR estimates came directly from reported statistical models (on continuous cumulative exposure), 45 were derived from summary study-population attributes, and 27 came from categorical analyses. Using the 96 XRs, OEL conferring one-per-thousand excess lifetime risk were calculated. OSHA's OEL, permissible exposure limits (PEL) were then compared to OEL derived from the 96 XRs. For 88 of the 96 calculated OELs (for which a corresponding PEL exists) all but 10 fell below the current PEL. Thirty-four OEL estimates were 10- to 100-fold below the PEL and 21 were greater than 100-fold below the PEL. This same pattern was observed using the different methods for deriving XRs. These findings can guide priorities in setting standards and the method is not limited to carcinogens.

Recombinant Methioninase Synergistically Reverses High-docetaxel Resistance Developed in Osteosarcoma Cells.

BACKGROUND/AIM: Docetaxel combined with gemcitabine is a second-line therapy for osteosarcoma, but its efficacy is limited by the development of docetaxel resistance. The aim of the present study was to determine whether recombinant methioninase (rMETase) could reverse docetaxel resistance developed in osteosarcoma cells. MATERIALS AND METHODS: Docetaxel-resistant 143B (DTR-143B) osteosarcoma cells were established by treating the parental 143B cells to increasing docetaxel concentrations (0.14-24 nM) over 5 months. The 50% inhibitory concentration (IC50) of docetaxel and rMETase as well as their combination on human osteosarcoma cells 143B and DTR-143B were determined. Four groups were analysed in vitro: untreated control; docetaxel; rMETase; docetaxel plus rMETase. RESULTS: The IC50 value of docetaxel for DTR-143B cells was 31.1 nM, compared to 4.38 nM for the parental 143B cells, a 7-fold increase. The combination of rMETase (0.53 U/ml) and docetaxel (4.38 nM) sensitized DTR-143B cells to docetaxel resulting in an inhibition of 73.7% compared to docetaxel alone (7.3%) or rMETase alone (54.6%) (p<0.05). rMETase thus increased the efficacy of docetaxel 10-fold on docetaxel-resistant osteosarcoma cells. CONCLUSION: rMETase reversed docetaxel resistance of DTR-143B in vitro. The present results indicate the clinical potential of rMETase to overcome docetaxel resistance in osteosarcoma patients.

Accurate and Safe Tumor Targeting of Orally-administered Salmonella typhimurium A1-R Leads to Regression of an Aggressive Fibrosarcoma in Nude Mice.

BACKGROUND/AIM: Salmonella typhimurium A1-R has been shown to target and inhibit many types of cancers in mouse models without continuous infection of normal tissue. The objective of the present study was to determine the effective dose of orally-administered Salmonella typhimurium A1-R, expressing-green fluorescent protein (GFP), on an HT1080 human-fibrosarcoma nude-mouse model. MATERIALS AND METHODS: The HT1080-human- fibrosarcoma nude-mouse models were randomized into the following three groups: G1: untreated control; G2: Oral Salmonella typhimurium A1-R (5×107 colony forming units [CFU]/body, twice a week, 2 weeks); G3: Oral Salmonella typhimurium A1-R (3.3×108 CFU/body, twice a week, 2 weeks). Each group comprised five mice. Body weight and tumor volume were measured twice a week. The number of colonies of Salmonella typhimurium A1-R-GFP in excised tumors and excised livers in groups G2 and G3 were determined on day 3, day 7 and 14 by growth on agar plates. Tukey-Kramer analysis was used to examine the relationships between variables. Statistically-significant results are defined as those with p≤0.05. RESULTS: Salmonella typhimurium A1-R was administered orally at a dose of 3.3×108 CFU, which successfully regressed the HT1080 tumor in nude mice. However, this effect was not observed at a lower dose of 5×107 CFU. After administering Salmonella typhimurium A1-R at 3.3×108 CFU, tumors and liver tissues were harvested, homogenized, and cultured on days 3, 7 and 14. Resulting GFP-expressing Salmonella typhimurium A1-R colonies were then counted. The number of GFP-bacterial colonies derived from excised tumors at intervals of 3, 7, and 14 days increased over time post-administration of oral GFP-Salmonella typhimurium. Conversely, the number of GFP-Salmonella typhimurium A1-R colonies that could be grown from excised livers decreased over time, following oral administration of GFP-Salmonella typhimurium. Additionally, the GFP-bacterial colonies grown from the excised tumors were significantly larger than those grown from the excised livers. CONCLUSION: The present study showed that an aggressive fibrosarcoma could be regressed by orally-administered Salmonella typhimurium A1-R which accurately targeted tumors without continuous growth in normal organs. The present results suggested the potential of orally-administered Salmonella typhimurium A1-R as a probiotic to treat aggressive soft-tissue sarcoma.