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These are figures associated with a second manuscript that has been submitted for consideration for co-publication. It describes the platform and hopefully will facilitate the reviewers' assessment of this manuscript on XMT-1536. we are also happy to make this available as supplementary data for this publication if desired.

These are figures associated with a second manuscript that has been submitted for consideration for co-publication. It describes the platform and hopefully will facilitate the reviewers' assessment of this manuscript on XMT-1536. we are happy to make this available as supplementary data for publication in any format if desired.

Supplementary Figures

Introduction: In November 2016, surgical antibiotic prophylaxis (SAP) guidelines for cardiothoracic surgeries at the authors’ centre were updated. SAP was reduced from 48 to 24 hours, and dual cover with vancomycin and cefazolin instead of vancomycin monotherapy was recommended for patients colonised with methicillin-resistant Staphylococcus aureus. This study was conducted to review compliance to the updated guidelines, and compare the incidence of surgical site infections (SSI). Methods: A list of patients undergoing sternotomy in National Heart Centre, Singapore, from March 2016 to February 2019 was extracted from the hospital’s electronic database; every fourth patient was included in the analysis. The patients were divided into three groups: Group 1 (before guideline revision, March–October 2016), Group 2 (post-guideline implementation, July 2017–May 2018), and Group 3 (July 2018–February 2019). Compliance to guidelines, incidence, and epidemiology of SSIs within 90 days of surgery were evaluated. Results: 509 patients (Group 1: 149; Group 2: 184; Group 3: 176) were included. There was appropriate selection and timely administration of SAP across all three groups. Post-guideline implementation, the proportion of patients on SAP for >24 hours decreased from 149 (100%) in Group 1 to 55 (29.9%), and 67 (38.1%) in Group 2 and 3, respectively (p

Background Surgical site infection (SSI) rates post pancreaticoduodenectomy (PD) range from 20 to 40%. Biliary obstruction (± stent in situ) leads to colonization of the biliary tract with gut flora, and bile contamination during surgery increases SSI risk. Current guidelines recommend surgical prophylaxis (ppx) with 1st or 2nd generation cephalosporins. However, emerging data favor targeted ppx based on biliary cultures to reduce SSI. We reviewed the microbial profile of pathogens isolated from perioperative biliary stent and bile cultures, and evaluated the impact of ppx on culture positive SSIs within 30 days of PD. Methods This was a retrospective study conducted in Singapore General Hospital. Patients who had PD from 1/1/2013 to 31/12/2019 were included. The first positive cultures from perioperative biliary stent, bile, and SSI cultures obtained within 30 days of surgery were reviewed. Antibiotic use 30 days prior and during surgery was collected. Results A total of 341 patients with a mean age of 63.9 ± 11.3 years were included; 202 (59.2%) were male and 137 (40.2%) received antibiotics in the past 30 days. For surgical ppx, patients received either cephalosporins ± metronidazole (216 [63.3%] ceftriaxone, 25 [7.3%] cefazolin), piperacillin-tazobactam (TZP) (35 [10.3%)], ciprofloxacin ± metronidazole (16 [4.7%)] or other antibiotics (14 [4.1%]), while 35 had missing data. Of the positive cultures from 84 biliary stent, 86 bile and 79 SSI, the most common organisms isolated were Enteroccocus spp., E. coli, Klebsiella spp., Enterobacter spp. and Candida spp. Ceftriaxone susceptibility rates in E. coli were 50.0%, 54.8% and 22.2% in biliary stent, bile and SSI cultures respectively; similarly low rates were also seen in Klebsiella spp. (Tables 1-3). Antibiotic use 30-days prior was associated with higher ceftriaxone resistance rates in E. coli and Klebsiella spp. from bile cultures (70.8% vs 36.4%, p=0.019). In patients with biliary stents cultured, use of TZP ppx showed a trend towards lower rates of culture positive SSIs compared to non-TZP ppx (9.1% vs 29.4%, p=0.054). Conclusion Our local gram-negative bacteria susceptibility rates to ceftriaxone are low in biliary stent, bile fluid cultures, and even lower for SSI post PD. Antibiotic ppx for PD in high-risk patients may need to be broadened. Disclosures All Authors: No reported disclosures.

Objectives Voriconazole serum concentration, which is affected by several factors, is associated with treatment response and toxicity. There is paucity of data on voriconazole therapeutic drug monitoring (TDM) among Southeast Asians, who exhibit a higher prevalence of CYP2C19-poor metabolisers compared with Caucasians and East Asians. Hence, there are concerns for higher risk of voriconazole accumulation and toxicity. We aim to determine the utility of voriconazole TDM through establishing: (1) proportion of patients achieving therapeutic troughs without dose adjustments; (2) characterisation of patients with sub-therapeutic, therapeutic and supra-therapeutic levels; (3) appropriate dose titrations/dose required for therapeutic troughs; (4) correlation between troughs and adverse events, treatment response/fungal breakthrough. Patients and methods A single-centre retrospective analysis of data from adults (≥21 years old) with ≥1 voriconazole trough measured at Singapore General Hospital from 2015 to 2017 was performed. Results Thirty-two patients (45.7%) among 70 patients achieved therapeutic troughs (defined as 2.0–5.5 mg/L) without dose adjustments. Eleven patients (15.7%) experienced hepatotoxicity (troughs 0.5 to >7.5 mg/L). Neurotoxicity occurred in three patients (4.3%) (troughs ≥6.7 mg/L) and all patients had symptom resolution upon dose reduction. Treatment failure of invasive fungal infection appeared less in patients with therapeutic troughs compared with sub-therapeutic troughs (11.4% vs. 14.2%). Two patients experienced treatment failure despite supra-therapeutic voriconazole troughs. Conclusions TDM should be implemented due to significant unpredictability in dose exposure. TDM can reduce unnecessary switches to alternatives due to intolerability and rule in the possibility of resistant organisms in the event of treatment failure despite therapeutic troughs, alerting clinicians to switch to alternatives promptly.

To determine percentage of patients with sub-therapeutic beta-lactam exposure in our intensive care units (ICU) and to correlate target attainment with clinical outcomes.Multi-centre, prospective, observational study was conducted in ICUs from three hospitals in Singapore from July 2016 to May 2018. Adult patients (≥21 years) receiving meropenem or piperacillin-tazobactam were included. Four blood samples were obtained during a dosing interval to measure and determine attainment of therapeutic targets: unbound beta-lactam concentration above (i) minimum inhibitory concentration (MIC) at 40% (meropenem) or 50% (piperacillin) of dosing interval (40-50%fT MIC) and (ii) 5 × MIC at 100% of dosing interval (100%fT 5 × MIC). Correlation to clinical outcomes was evaluated using Cox regression.Beta-lactam levels were highly variable among 61 patients, with trough meropenem and piperacillin levels at 21.5 ± 16.8 mg/L and 101.6 ± 81.1 mg/L respectively. Among 85 sets of blood samples, current dosing practices were able to achieve 94% success for 40-50%fT MIC and 44% for 100%fT 5 × MIC. Failure to achieve 40-50%fT MIC within 48 h was significantly associated with all-cause mortality (HR: 9.0, 95% CI: 1.8-45.0), after adjustment for APACHE II score. Achievement of 100%fT 5 × MIC within 48 h was significantly associated with shorter length of hospital stay.Current dosing practices may be suboptimal for ICU patients. Beta-lactam TDM may be useful.

Antimicrobial stewardship programmes (ASPs) have often been recommended as a viable solution to minimise the incidence of Clostridium difficile infection (CDI), which can be life-threatening. This study aimed to evaluate whether ASP interventions have contributed to reducing CDI rates.A retrospective review of ASP interventions issued from January 2013 to April 2014 was performed using data from the ASP database of Singapore General Hospital, a 1600-bed tertiary-care hospital in Singapore. A total of 283 interventions satisfied the inclusion criteria, of which commonly audited antibiotics were piperacillin/tazobactam (41.3%) and carbapenems (54.8%). Comparisons were made at 30days post-intervention between those with accepted or rejected interventions. The primary outcome was CDI incidence; secondary outcomes included length of hospitalisation post-intervention, 30-day mortality and CDI recurrence rate.Whilst the median duration of antibiotic therapy was reduced by 2days (6days vs. 4 days; P0.001), acceptance of ASP interventions did not alter primary CDI incidence at 30days (P=0.644) post-intervention. However, reduced CDI recurrence rates were observed for patients positive for CDI in the accepted patient group compared with the rejected group (0% vs. 37.5%; P=0.03), with no difference in CDI 30-day mortality between the two groups.Intervention acceptance did not contribute to a significant reduction in CDI incidence but may be associated with lower recurrence rates, although further studies are required.

OBJECTIVES: To compare the effectiveness and tolerability of micafungin versus posaconazole during chemotherapy-induced neutropenia in acute leukemia (AL) and myelodysplastic syndrome (MDS). METHODS: Patients with AL or MDS undergoing chemotherapy were randomized to open-label micafungin 100 mg intravenously daily or posaconazole suspension 400 mg orally twice daily until neutrophil recovery, up to 28 days. Patients were followed for 12 weeks. The primary endpoint was prophylaxis failure (premature discontinuation due to infection, intolerance, adverse event, or death). Time to failure and survival were calculated by Kaplan-Meier analysis. RESULTS: From March 2011 to May 2016, 113 patients who received at least 2 doses of prophylaxis were analyzed (58 patients randomized to micafungin and 55 to posaconazole). Prophylaxis failure occurred in 34.5% and 52.7% of patients on micafungin and posaconazole, respectively (P = 0.0118). The median number of days on prophylaxis was 16 [interquartile range (IQR) 12-20] for micafungin and 13 [IQR 6-16] for posaconazole (P = 0.01). Micafungin failures were largely due to antifungal treatment; posaconazole failures were mostly due to gastrointestinal intolerance or adverse effects. IFI incidence and survival were similar between study arms. CONCLUSIONS: Our data support micafungin as alternative antifungal prophylaxis in patients with AL and MDS.

Background In early months of COVID-19 pandemic, SGH recorded a year-on-year increase in antibiotic (ABx) use for community acquired acute respiratory infection (CA ARI) from Feb-Apr 2019 (48.7 defined daily doses (DDD)/100 bed-days) to 2020 (50.8 DDD/100 bed-days). To address concerns of misuse, the antibiotic stewardship unit (ASU) expanded prospective audit feedback (PAF) to CA ARI patients admitted to ARI wards, with low procalcitonin (PCT). PAF was conducted on day 2-3 of ABx, on weekdays. Doctors received feedback to stop/modify when ABx was deemed inappropriate. Here, we describe the impact of ASU’s adaptive approach to curb rising ABx use in patients admitted for ARI during COVID-19 pandemic. Methods A Pre- & Post-intervention study was conducted. All patients started on ABx (ceftriaxone/co-amoxiclav/piptazo/carbapenems/levofloxacin) for CA ARI & PCT < 0.5µg/L were analysed. Those who died ≤48h of admission; admitted to intensive care; required ABx escalation; >1 infective sites; complex lung infection were excluded. Primary objective was to compare the proportion of ABx stopped ≤4 days (time to final infection diagnosis) Pre (22/3-18/4/20) & Post (21/4-13/7/20). Results 184 (Pre) & 528 (Post) ABx courses were analysed. ASU audited 51 (Pre) & 380 (Post) courses with the rest discontinued/discharged before review. Patients were largely similar in both periods; a third had low likelihood of bacterial infection (C reactive protein < 30mg/L). In Post, 73 feedback was given to stop ABx (often because symptoms suggested viral/fluid overload) & 18 to switch to oral ABx. 82 (90%) feedback was accepted. No ABx was restarted ≤48h or deaths ≤30 days due to ARI. 1 patient had C. difficile diarrhoea a day after ABx cessation as per ASU feedback. Proportion of all ABx stopped ≤4 days was higher in Post than Pre [27/184 (15%) vs 152/528 (29%), p< 0.01]. Median duration of therapy of IV ABx was reduced (6.5 vs 3 days, p< 0.01), with corresponding shorter median length of stay (10.5 vs 6 days, p< 0.01). Conclusion PAF directly and indirectly reduced ABx duration in patients treated for CA ARI as prescribers become more conscious about stopping ABx when investigations show low likelihood of bacterial infection. ASU must remain agile during pandemics to detect emerging problems and adapt processes to counter early. Disclosures All Authors: No reported disclosures

Background Antimicrobial stewardship programs (ASP) in hospitals improve antibiotic prescribing, slow antimicrobial resistance, reduce hospitalisation duration, mortality and readmission rates, and save costs. However, the strategy of prospective audit and feedback is laborious. In Singapore General Hospital (SGH), 10 reviews are required to identify 2 inappropriate cases. Limited manpower constraints ASP audits to only about 30% of antibiotics prescribed. This proof-of-concept study explored the feasibility of developing a predictive model to prioritise inappropriate antibiotic prescriptions for ASP review. Methods ASP-audited adult pneumonia patients from January 2016 to December 2018 in SGH were included. Patient data e.g., demographics, allergies, past medical history, and relevant laboratory investigations at each antibiotic use episode were extracted from electronic medical records and re-assembled through linking for analysis. Ground truth for model training was based on ASP-defined appropriateness for each encounter. The dataset was split into 80% and 20% for training and testing respectively. Three modelling techniques, XGBoost, decision tree and logistic regression, were assessed for their relative performance in terms of precision, sensitivity and specificity. Results There were 12471 unique patient encounters. Training was done on 10459 encounters and 39 data elements were included. When tested on 2012 encounters, the logistic regression model performed the best (86.7% sensitivity, 71.4% specificity). The model correctly classified 1377 out of 1388 (99.2%) encounters as “appropriate” (do not require ASP intervention). 624 antibiotic use encounters were classified as “inappropriate”, of which only 72 were truly inappropriate (positive predictive value for ASP intervention, PPV 11.5%). The low PPV was likely due to inadequate representation of “inappropriate” cases in the training dataset (4.1%). Applying this model would prioritise the number of immediate ASP reviews needed to identify cases for intervention by two-thirds, from 2012 to 624 (Figure 1). Figure 1. Illustration of AI benefits in ASP Conclusion ASPs can leverage on machine learning capabilities to improve audit efficiency. This can increase ASP’s productivity and staff’s job satisfaction as they are freed up to perform other work. Disclosures All Authors: No reported disclosures

Background International guidelines recommend up to 24 hours of perioperative antibiotic prophylaxis (AP) in joint replacement procedures. However, some observational studies support AP beyond 24 hours for the prevention of surgical site infections (SSI) and prosthetic joint infections (PJI), and this practice is also observed in our institution. This study aims to evaluate the incidence of SSI and PJI in patients receiving short- vs extended-course AP after unilateral primary total knee arthroplasty (TKA) at our center. Methods This was a retrospective cohort study of patients who underwent elective unilateral primary TKA from October to December 2019 at Singapore General Hospital. Patients were excluded if they received antibiotics for reasons other than post-operative AP or underwent other procedures in addition to unilateral primary TKA. Data was collected from electronic medical records and patients who received a short-course of AP (≤24 hours) were compared with patients who received an extended-course of AP. Primary outcomes were 30-day SSI and 30-day PJI rates. Secondary outcome was the impact of duration of AP on length of stay. Statistical analysis was performed using SPSS software version 20. Results There were 394 patients included in the study. 247 received short-course and 147 received extended-course AP. There were no differences in demographics (Table 1). Amongst those who received extended-course AP, median duration was 7 (IQR 4, 8) days, during which 119 (81.0%) patients switched from intravenous to oral route of antibiotics. Between the short- and extended-course arms, there were no differences observed in 30-day SSI (6.9% vs 6.1%, p=0.769) or PJI rates (0.4% vs 0.7%, p=0.999). However, extended-course AP was associated with a longer median length of stay (4 [IQR 3, 6] vs 5 [IQR 4, 7] days, p=0.001). In a subgroup analysis of 106 diabetic patients, there were no differences in 30-day SSI rates (12.3% vs 9.8%, p=0.763) and 30-day PJI rates (0% vs 2.4%, p=0.387) between both groups. Conclusion In this single center study, short-course AP in elective TKA was safe and effective. Extending AP did not reduce SSI or PJI rates, even in diabetic patients. In addition, extending AP was associated with increased length of stay, which translates to higher healthcare costs. Disclosures All Authors: No reported disclosures

BACKGROUND: Adopting the family-centered care (FCC) approach in the neonatal care has been shown to improve breastfeeding rate and parental satisfaction. To minimize the transmission of COVID-19, family visit in neonatal intensive care unit (NICU) was suspended in China. In order to maintain the benefits of FCC, the Hong Kong University-Shenzhen Hospital NICU modified FCC strategies. We evaluated the effects of new strategies and aimed to share our results and experience with other NICUs during the COVID-19 pandemic. METHODS: Using prospectively collected hospital databases, we retrospectively compared the demographic and clinical data of neonates, rates of breastfeeding at discharge, nosocomial infection and parental satisfaction one month before (open group) and after (closed group) the implementation of alternative FCC management strategies when family visit was suspended during COVID-19 pandemic. RESULTS: During the COVID-19 pandemic, we organized a multidisciplinary task force and adopted strategies of triage and screening, management of suspected infants, and breastfeeding promotion with effective communication. The nosocomial infection rate and parental satisfaction for open and closed groups (144 and 108 term and near-term neonates with brief hospitalization, respectively) were not different (1% vs. 0%, p = 1.00; 98.6 vs. 98.8, p = .80; respectively). Breastfeeding rate at discharge decreased but the difference was not significant (74% vs. 80%, p = .29). CONCLUSIONS: In our experience, in term and near-term neonates with brief hospitalization, the alternative FCC strategies maintained high parental satisfaction without increased nosocomial infection rate, but strong support for breastfeeding was needed. Through multidisciplinary collaboration, the continuation of "modified" FCC in a level III NICU is feasible in the context of COVID-19 pandemic with reduced family visitation and participation in the care.

Background STING pathway agonism has emerged as a potential therapeutic mechanism to stimulate an innate anti-tumor immune response. While in principle systemic administration of a STING agonist would have many therapeutic benefits, including the delivery of STING to all tumor lesions, such an approach may be limited by toxicity. Antibody-drug conjugates (ADCs) constitute a proven therapeutic modality that is ideally suited to allow systemic administration while stimulating the innate immunity in a targeted manner. We have previously demonstrated that targeted delivery of a STING agonist with an ADC induces robust anti-tumor immune responses. Methods Herein we investigated the mechanism of action of tumor cell-targeted STING agonist ADCs. We evaluated STING pathway activation and anti-tumor activity elicited by ADCs harboring either wild type (wt) or mutant Fc deficient in Fcγ receptor (FcγR) binding in wt or STING knockout (ko) cancer cell mono-cultures, immune cell co-cultures, and in in vivo tumor models. Results Consistent with previous reports, the majority of cancer cell lines tested failed to induce STING pathway following STING agonist payload treatment in mono-cultures. In cancer cell:THP1 monocytic cell co-cultures, tumor-targeted STING agonist ADCs with wt Fc exhibited robust STING activation, whereas Fc-mutant ADCs or non-targeted control ADCs had minimal activity. Similar results were obtained when THP1 cells were treated in plates coated with target antigen without cancer cells, demonstrating STING activation in THP1 cells following FcγR-mediated uptake of antigen-bound ADCs. Tumor-targeted Fc-wt ADCs led to marked induction of STING pathway and cancer cell-killing in cancer cell:PBMC or primary monocyte co-cultures, and complete tumor regressions in in vivo tumors. Surprisingly, while at reduced levels relative to the Fc-wt ADCs, Fc-mutant ADCs exhibited significant activity in these in vitro and in vivo models, suggesting that tumor cell-intrinsic STING pathway may be activated in the presence of cues from immune cells. Consistently, STING agonist payload treatment in the presence of conditioned media from PBMC and primary monocyte but not from THP1 cultures, led to STING activation in cancer cell mono-cultures. Moreover, Fc-mutant ADCs had diminished activity in STING ko cancer cell:PBMC or primary monocyte co-cultures, demonstrating the contribution of tumor cell-intrinsic STING activation to the anti-tumor activity elicited by tumor cell-targeted STING agonist ADCs. Conclusions In conclusion, we demonstrated that tumor cell-targeted STING agonist ADCs induce robust anti-tumor activity through mechanisms involving both FcγR and tumor antigen-mediated ADC internalization and subsequent induction of STING pathway in immune cells and tumor cells.

The increase of carbapenem-resistant Enterobacterales (CRE) and lack of therapeutic options due to the scarcity of new antibiotics has sparked interest toward the use of intravenous fosfomycin against systemic CRE infections. We aimed to investigate the in vitro pharmacodynamics of fosfomycin against carbapenem-resistant Enterobacter cloacae and Klebsiella aerogenes. Time-kill studies and population analysis profiles were performed with eight clinical CRE isolates, which were exposed to fosfomycin concentrations ranging from 0.25 to 2,048 mg/liter. The 24-h mean killing effect was characterized by an inhibitory sigmoid maximum effect (E(max)) model. Whole-genome sequencing was performed to elucidate known fosfomycin resistance mechanisms. Fosfomycin MICs ranged from 0.5 to 64 mg/liter. The isolates harbored a variety of carbapenemase genes including bla(IMP), bla(KPC), and bla(NDM). Five out of eight isolates harbored the fosA gene, while none harbored the recently discovered fosL-like gene. Heteroresistant subpopulations were detected in all isolates, with two out of eight isolates harboring heteroresistant subpopulations at up to 2,048 mg/liter. In time-kill studies, fosfomycin exhibited bactericidal activity at 2 to 4 h at several fosfomycin concentrations (one isolate at ≥16 mg/liter, two at ≥32 mg/liter, two at ≥64 mg/liter, two at ≥128 mg/liter, and one at ≥512 mg/liter). At 24 h, bactericidal activity was only observed in two isolates (MICs, 0.5 and 4 mg/liter) at 2,048 mg/liter. From the E(max) model, no significant bacterial killing was observed beyond 500 mg/liter. Our findings suggest that the use of fosfomycin monotherapy may be limited against CRE due to heteroresistance and rapid bacterial regrowth. Further optimization of intravenous fosfomycin dosing regimens is required to increase efficacy against such infections.

Population pharmacokinetic studies have suggested that high polymyxin B (PMB) doses (&ge, 30,000 IU/kg/day) can improve bacterial kill in carbapenem-resistant Gram-negative bacteria (CR-GNB). We aim to describe the efficacy and nephrotoxicity of patients with CR-GNB infections prescribed high-dose PMB. A single-centre cohort study was conducted from 2013 to 2016 on septic patients with CR-GNB infection and prescribed high-dose PMB (~30,000 IU/kg/day) for &ge, 72 h. Study outcomes included 30-day mortality and acute kidney injury (AKI) development. Factors associated with AKI were identified using multivariable regression. Forty-three patients with 58 CR-GNB received high-dose PMB, 57/58 (98.3%) CR-GNB were susceptible to PMB. The median daily dose and duration of high-dose PMB were 32,051 IU/kg/day (IQR, 29,340&ndash, 34,884 IU/kg/day) and 14 days (IQR, 7&ndash, 28 days), respectively. Thirty-day mortality was observed in 7 (16.3%) patients. AKI was observed in 25 (58.1%) patients with a median onset of 8 days (IQR, 6&ndash, 13 days). Higher daily PMB dose (aOR,1.01, 95% CI, 1.00&ndash, 1.02) and higher number of concurrent nephrotoxins (aOR, 2.14, 95% CI, 1.03&ndash, 4.45) were independently associated with AKI. We observed that a sizable proportion developed AKI in CR-GNB patients described high-dose PMB, hence, the potential benefits must be weighed against increased AKI risk. Concurrent nephrotoxins should be avoided to reduce nephrotoxicity.

Target selection for antibody–drug conjugates (ADC) frequently focuses on identifying antigens with differential expression in tumor and normal tissue, to mitigate the risk of on-target toxicity. However, this strategy restricts the possible target space. SLC34A2/NaPi2b is a sodium phosphate transporter expressed in a variety of human tumors including lung and ovarian carcinoma, as well as the normal tissues from which these tumors arise. Previous clinical trials with a NaPi2b targeting MMAE-ADCs have shown objective durable responses. However, the protein-based biomarker assay developed for use in that study was unable to discern a statistically significant relationship between NaPi2b protein expression and the probability of response. XMT-1536 is a NaPi2b targeting ADC comprised of a unique humanized antibody conjugated with 10–15 auristatin F- hydroxypropylamide (AF-HPA) payload molecules via the Dolaflexin platform. AF-HPA is a cell-permeable, antimitotic compound that is slowly metabolized intratumorally to an active, very low-permeable metabolite, auristatin F (AF), resulting in controlled bystander killing. We describe the preclinical in vitro and in vivo antitumor effects of XMT-1536 in models of ovarian and lung adenocarcinoma. Pharmacokinetic analysis showed approximately proportional increases in exposure in rat and monkey. Systemic free AF-HPA and AF concentrations were observed to be low in all animal species. Finally, we describe a unique IHC reagent, generated from a chimeric construct of the therapeutic antibody, that was used to derive a target expression and efficacy relationship in a series of ovarian primary xenograft cancer models.

Highlights • During the COVID-19 outbreak, antibiotic use has increased, especially respiratory antibiotics. • Healthcare resources are diverted to containment and management of COVID-19 cases, but the efforts of antibiotic stewardship programmes (ASPs) must run in parallel to mitigate antibiotic misuse and antimicrobial resistance. • ASPs must be operationally adaptable, and possess robust surveillance systems that detect subtle changes in prescribing and resistance trends. • This is an opportunity to execute a syndromic approach to guide antibiotic prescribing. • Continued engagement and education of prescribers to ensure appropriate prescription of antibiotics is important., Healthcare resources are being diverted for the containment and control of coronavirus disease 2019 (COVID-19). During this outbreak, it is cautioned that antibiotic misuse may be increased, especially for respiratory tract infections. With stewardship interventions, the duration of antibiotic therapy and length of stay of hospitalized patients can be reduced significantly. Antibiotic stewardship programmes should continually engage and educate prescribers to mitigate antibiotic misuse during the COVID-19 pandemic.

Background: Recent neonatal resuscitation guidelines suggest to perform chest compression (CC) at over-the-head (OTH) position instead of lateral position when further interventions including umbilical venous access are needed. Little information is available regarding the quality of cardiopulmonary resuscitation at different positions. Our study compared the quality of CC and ventilation at OTH position vs. lateral position in simulated neonatal resuscitation. Methods: Thirty-nine neonatal practitioners who attended the NRP®-based Provider renewal course workshop participated this study. Laerdal QCPR infant model were used to collect the data (2-miutes continuous recording) on quality of CC and ventilation of all participants at OTH position and lateral position in randomized order, both coordinated with mask ventilation or endotracheal ventilation through a Neopuff© T-piece system. The quality of CC and ventilation were compared. Participants also reported their demographics and opinions in anonymous questionnaires after the session. Results: The quality of CC and ventilation was not different when CPR was performed at OTH position and lateral position, in both mask and endotracheal ventilation. When CPR was performed with endotracheal ventilation, there were small faster frequencies of CC and ventilation at OTH position, compared with those at lateral position (p = 0.004). Most participants (87%) liked the CC performed at OTH position and had no adverse feedback. Conclusions: Performing CC at OTH position was generally well-received in simulated resuscitation; the quality of CC and ventilation at OTH position was not significantly different from that at lateral position, irrespective of mask or endotracheal ventilation.

Tze-Peng Lim,1,2 Reyna Wang,1 Gang Quan Poh,3 Tse-Hsien Koh,4 Thean-Yen Tan,5 Winnie Lee,1 Jocelyn Qi-Min Teo,1 Yiying Cai,1 Thuan-Tong Tan,6 Pui Lai Rachel Ee,3 Andrea L Kwa1,3,7 1Department of Pharmacy, Singapore General Hospital, Singapore, Singapore; 2SingHealth Duke-NUS Medicine Academic Clinical Programme, Singapore, Singapore; 3Department of Pharmacy, Faculty of Science, National University of Singapore, Singapore, Singapore; 4Department of Microbiology, Singapore General Hospital, Singapore, Singapore; 5Department of Laboratory Medicine, Changi General Hospital, Singapore, Singapore; 6Department of Infectious Diseases, Singapore General Hospital, Singapore, Singapore; 7Emerging Infectious Diseases, Duke-NUS Medical School, Singapore, Singapore Objectives: Treatment for nosocomial bloodstream infections (BSI) caused by multidrug-resistant (MDR) Gram-negative bacteria (GNB) is challenging. Rising antimicrobial resistance, especially in extended spectrum beta-lactamase production, inadvertently increases empiric carbapenem consumption. Three antipseudomonal carbapenems (imipenem, meropenem [MER], and doripenem [DOR]) are available commercially against MDR GNB in Singapore. The study aims to determine the most optimal empiric carbapenem dosing regimens (CDR) and evaluate their cost-effectiveness for GNB-BSI in the face of increasing MDR GNB. Methods: Carbapenem minimum inhibitory concentrations (MICs) were generated for non-repeat GNB-BSI obtained in 2013–2014 from two hospitals. Monte Carlo simulations were used to assess the cumulative fraction of response (CFR) of various CDRs using the percentage of time above MIC for 40% (%T>MIC of 40%) as the pharmacokinetic (PK)–pharmacodynamic (PD) parameter for efficacy. Carbapenem costs were based on patient antibiotic costs. Antibiotic cost-effectiveness was calculated as total daily drug cost/CFR. Results: A total of 1,140 bloodstream isolates were collected. They comprised 116 Acinetobacter baumannii, 237 Pseudomonas aeruginosa, and 787 Enterobacteriaceae. All CDRs achieved ~40, ~80, and ≥90% CFRs against A. baumannii, P. aeruginosa, and Enterobacteriaceae, respectively. Against P. aeruginosa, MER 2g every 8h infused over 3h and DOR 1g every 8h infused over 4h achieved CFRs 84 and 81%, respectively. Against Enterobacteriaceae, the cost of MER 2g every 8h infused over 3h was the lowest among the three carbapenems at $0.40/percentage of CFR. Conclusion: This study demonstrates the utility of PK–PD modeling to formulate the optimal selection of a cost-effective empiric CDR in antibiotics guidelines and formulary inclusion. The findings support the selection of high MER doses of prolonged infusions as empiric coverage for GNB-BSI in our institutions. Keywords: empiric carbapenem regimens, multidrug resistant, Gram-negative bacteria, bloodstream infections

STING pathway plays a critical role in inducing anti-tumor immunity by upregulating Type 1 Interferon (IFN) and IFN-stimulated genes within the tumor microenvironment in response to cytosolic nucleic acid ligands. Therefore, the STING pathway agonism has emerged as a potential therapeutic mechanism to stimulate an anti-tumor innate immune response. Intratumorally injected free STING-agonists that are currently being evaluated in the clinic by others have shown limited effects in non-injected lesions. Antibody-drug conjugates (ADCs) constitute a proven therapeutic modality that enables tumor-targeted drug delivery with systemic administration. We have previously demonstrated that the tumor cell-intrinsic STING pathway is activated in the presence of cues from immune cells and contributes to the anti-tumor activity of tumor cell-targeted Immunosynthen STING-agonist ADCs, in which a STING-agonist payload is conjugated to a tumor cell-targeting antibody. Here we investigated the nature of the STING pathway activation in tumor cells and its contribution to the anti-tumor activity elicited by STING agonism. Leveraging ADCs with a wild type (wt) or mutant Fc (deficient in Fcγ receptor -FcγR- binding), we delivered a STING-agonist simultaneously to tumor-resident immune and cancer cells or only to cancer cells through FcγR-mediated and/or tumor antigen-mediated ADC internalization. We utilized these ADCs in in vivo human tumor xenograft models and STING wt or knock out (ko) cancer cell:immune cell co-cultures and evaluated gene expression, cytokine production, and anti-tumor activities induced by STING-agonist ADCs. Surprisingly, Nanostring analysis of the human tumor xenografts from mice treated with tumor cell-targeted STING-agonist ADCs revealed human tumor cell-specific activation of Type III IFNs. In human cancer cell:immune cell co-cultures, treatment with tumor cell-targeted STING-agonist ADCs also led to marked upregulation of Type III IFNs, which was significantly reduced in STING ko cancer cell:immune cell co-cultures, suggesting that the cancer cells may contribute majority of the Type III IFNs downstream of STING pathway activation. Blocking Type III IFNs with neutralizing antibodies in cancer cell:immune cell co-cultures inhibited the production of key cytokines, including Type I IFN, and nearly abolished tumor cell-killing in response to STING-agonist ADC treatment, indicating that the Type III IFNs play an important role in the anti-tumor activity induced by STING activation. These studies reveal a previously underappreciated mechanism of STING agonist anti-tumor activity. The ability of tumor cell-targeted STING-agonist ADCs to activate STING in both tumor cells and in tumor-resident immune cells may represent a significant therapeutic advantage of an Immunosynthen ADC approach to STING agonism. Citation Format: Naniye Malli Cetinbas, Travis Monnell, Kalli Catcott, Winnie Lee, Pamela Shaw, Kelly Slocum, Kenneth Avocetien, Keith Bentley, Susan Clardy, Brian Jones, Eoin Kelleher, Rebecca Mosher, Joshua D. Thomas, Dorin Toader, Jeremy Duvall, Raghida A. Bukhalid, Marc Damelin, Timothy B. Lowinger. Tumor cell-intrinsic STING pathway activation leads to robust induction of Type III Interferons and contributes to the anti-tumor activity elicited by STING agonism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1773.

STING pathway agonism has emerged as a potential therapeutic mechanism to stimulate an innate anti-tumor immune response. However, the systemic administration of a free STING agonist may be limited by toxicity, and broad biodistribution may not be ideal. Antibody-drug conjugates (ADCs) constitute a proven therapeutic modality that enables tumor-targeted delivery and thus is ideally suited to systemic administration with reduced toxicity. To develop an optimized STING-agonist ADC platform, we designed a novel STING-agonist specifically tailored for use in an ADC. Determination of the co-crystal structure confirmed that the agonist binds to the closed, or ‘active', conformation of the STING homodimer. The resulting Immunosynthen platform, which was developed specifically for the selected STING agonist payload, was used to generate XMT-2056, a tumor antigen-targeted STING-agonist ADC with excellent drug-like properties and >100-fold increased potency as compared to the free STING-agonist payload. In mice, XMT-2056 induced robust anti-tumor immune activity, with only minimal increases in systemic cytokine levels, and exhibited significant benefit over the benchmark free STING-agonist payload in both regards. Additionally, in vitro and in vivo studies demonstrate that XMT-2056 is able to activate the STING pathway in both tumor-resident immune cells and tumor cells, offering a potential advantage over other innate immune activating pathways. XMT-2056 was well-tolerated in non-human primates at significantly higher exposure levels than those required for anti-tumor activity, and the ADC exhibited favorable pharmacokinetics after repeat doses. Together these data support the clinical development of XMT-2056. Citation Format: Jeremy R. Duvall, Raghida A. Bukhalid, Naniye M. Cetinbas, Kalli C. Catcott, Kelly Slocum, Kenneth Avocetien, Keith W. Bentley, Stephen Bradley, Susan Clardy, Scott D. Collins, Elizabeth Ditty, Timothy Eitas, Brian D. Jones, Eugene W. Kelleher, Winnie Lee, Travis Monnell, Rebecca Mosher, Marina Protopopova, LiuLiang Qin, Pamela Shaw, Elena Ter-Ovanesyan, Joshua D. Thomas, Phonphimon Wongthida, Ling Xu, Liping Yang, Jeffrey Zurita, Dorin Toader, Marc Damelin, Timothy B. Lowinger. XMT-2056, a well-tolerated, Immunosynthen-based STING-agonist antibody-drug conjugate which induces anti-tumor immune activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1738.

Polymyxin B-based combinations are increasingly prescribed as a last-line option against extensively drug-resistant (XDR) Acinetobacter baumannii. It is unknown if such combinations can result in the development of nondividing persister cells in XDR A. baumannii. We investigated persister development upon exposure of XDR A. baumannii to polymyxin B-based antibiotic combinations using flow cytometry. Time-kill studies (TKSs) were conducted in three nonclonal XDR A. baumannii strains with 5 log(10) CFU/ml bacteria against polymyxin B alone and polymyxin B-based two-drug combinations over 24 h. At different time points, samples were obtained and enumerated by viable plating and flow cytometry. Propidium iodide and carboxyfluorescein succinimidyl ester dyes were used to differentiate between live and dead cells and between dividing and nondividing cells, respectively, at the single-cell level, and nondividing live cells were resuscitated and characterized phenotypically. Our results from viable plating showed that polymyxin B plus meropenem and polymyxin B plus rifampin were each bactericidal (>99.9% kill compared to the initial inoculum) against 2/3 XDR A. baumannii strains at 24 h. By flow cytometry, however, none of the combinations were bactericidal against XDR A. baumannii at 24 h. Further analysis using cellular dyes in flow cytometry revealed that upon exposure to polymyxin B-based combinations, XDR A. baumannii entered a viable but nondividing persister state. These bacterial cells reinitiated division upon the removal of antibiotic pressure and did not have a growth deficit compared to the parent strain. We conclude that persister cells develop in XDR A. baumannii upon exposure to polymyxin B-based combinations and that nonplating methods appear to complement viable-plating methods in describing the killing activity of polymyxin B-based combinations against XDR A. baumannii.

In the last decade, considerable advancements have been made to identify the pharmacokinetic/pharmacodynamic (PK/PD) index that defines the antimicrobial activity of polymyxins. Dose-fractionation studies performed in hollow-fiber models found that altering the dosing schedule had little impact on the killing or suppression of resistance emergence, alluding to AUC/MIC as the pharmacodynamic index that best describes polymyxin's activity. For in vivo efficacy, the PK/PD index that was the most predictive of the antibacterial effect of colistin against P. aeruginosa and A. baumannii was ƒAUC/MIC.

Salmonellosis in children from Ethiopia is caused mainly by Salmonella Concord, which are highly invasive, multi-drug resistant and extended-spectrum β-lactamase (ESBL) producers. S. Concord infections have been observed in children adopted from Ethiopia, now living in Europe and United States. S. Concord infections are present in parents of these adopted children, posing a significant dilemma for treatment. Data from Salmonella isolates are stored in Public Health England’s in-house Gastro Data Warehouse (GDW) database. Resistance profiles for 37 pure S. Concord isolates were determined using agar incorporation methods using EUCAST guidelines. The minimum inhibitory concentration (MIC) was determined for 18 antimicrobial agents and ESBL resistance was confirmed using Double Disk Synergy and ESBL E-test methods. ESBL resistance was present in 8 isolates with resistance most commonly seen against penicillins and cephalosporins. Isolates 408 and 537, and isolate 527 displayed resistance to 78 % and 56 % of the antibiotics respectively. Drug resistant regions in isolates 408, 527 and 537 that have been previously sequenced by an Illumina HiSeq 2500 were characterised. ESBL genes blaCTXM-15 was present in isolate 527 and 408 and blaSHV-12 was found in isolate 537. Replicons from plasmids, InCI2, InCFIB, InCF2 and IncH2 were located in the assembled sequences of the three previously sequenced isolates. In conclusion, the possible mechanisms causing spread of ESBL resistance in S. Concord is most likely due to acquisition of plasmids through horizontal gene transfer from various Enterobacteriaceae, however further research must be conducted to confirm this to advance antimicrobial research in this area.

Rapid and accurate differentiation of Salmonella spp. causing enteric fever from nontyphoidal Salmonella is essential for clinical management of cases, laboratory risk management, and implementation of public health measures. Current methods used for confirmation of identification, including biochemistry and serotyping as well as whole-genome sequencing analyses, take several days. Here we report the development and evaluation of a real-time PCR assay that can be performed directly on crude DNA extracts from bacterial colonies for the rapid identification of typhoidal and nontyphoidal Salmonella.