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The M. tuberculosis citrate synthase regulatory domain binds pyruvate to affect enzyme activity while cysteine oxidation in the same domain eliminates enzyme activity. This affords regulatory control at the protein level of entry to the TCA cycle.

Neurological emergencies carry significant morbidity and mortality, and it is necessary to have a multidisciplinary approach involving the emergency physician, the neurologist, the intensivist, and the critical care nursing staff. These disorders can be broadly divided into noninfectious and infectious etiologies. In this article, we review a few of the neurological emergencies that present to the neurological intensive unit, with emphasis on convulsive status epileptics, myasthenia gravis, Guillain-Barré syndrome, meningitis, encephalitis, and brain abscess.

Due to the problems associated with cracking, the tensile properties of bituminous mix are of great concern to pavement engineers. Consequently, the tensile strength of bituminous mix is crucial for applications involving pavement. The Indirect Tensile Strength (ITS) test is used to evaluate the tensile properties of the bituminous mix. Adding cement or lime to bituminous mixtures is known to impart anti-stripping properties. For adverse conditions, however, it is advisable to use antistripping chemicals. It is crucial to ensure that the addition of this compound will not have a negative impact on other properties. Numerous studies on ITS have been conducted separately; however, no effect to evaluate the effect of addition of both of them has been done. In this proposal, an effort has been made to investigate the addition techniques of Anti-Stripping and Cement to DBM, both individually and in combination with the appropriate proportion.

Tumor necrosis factor (TNF) α-induced nuclear translocation of the NF-κB subunit RELA has been implicated in several pathological conditions. Here we report the discovery of a spirocyclic dimer (SpiD7) that covalently modifies RELA to inhibit TNFα-induced nuclear translocation. This is a previously unexplored strategy to inhibit TNFα-induced NF-κB activation., Discovery of a spirocyclic dimer (SpiD7) that covalently modifies RELA to generate stable high molecular weight complexes. SpiD7 inhibits TNFα-induced nuclear translocation of RELA resulting in the blockade of NF-kB gene transcription, through a previously unexplored modality.

Summary Background There is an urgent unmet need for new therapies in amyotrophic lateral sclerosis. In a clinical study with healthy volunteers, levosimendan, a calcium sensitiser, was shown to improve neuromechanical efficiency and contractile function of the human diaphragm. We aimed to evaluate the safety and efficacy of oral levosimendan in people with amyotrophic lateral sclerosis, with a focus on respiratory function. Methods The REFALS study is a randomised, double-blind, placebo-controlled phase 3 trial at 99 amyotrophic lateral sclerosis specialist centres in 14 countries worldwide. People with amyotrophic lateral sclerosis were eligible for participation if they were at least 18 years of age and had a sitting slow vital capacity (SVC) of 60–90% predicted. Participants were randomly assigned (2:1) by interactive web-response system to receive either levosimendan or placebo. The capsules for oral administration were identical in appearance to maintain blinding of participants and investigators. The primary endpoint was the change from baseline in supine SVC at 12 weeks, assessed as the percentage of predicted normal sitting SVC. The key secondary endpoint was the combined assessment of function and survival (CAFS) up to 48 weeks. Analyses were done in the intention-to-treat population, comprising all participants who were randomly assigned. This trial is registered at ClinicalTrials.gov (NCT03505021) and has been completed. An extension study (REFALS-ES; NCT03948178 ) has also been completed, but will be reported separately. Findings Between June 21, 2018, and June 28, 2019, 871 people were screened for the study, of whom 496 were randomly assigned either levosimendan (n=329) or placebo (n=167). Participants were followed up between June 27, 2018 and June 26, 2020, for a median duration of 50·1 (IQR 37·5–51·1) weeks. The median duration of treatment was 47·9 (IQR 26·4–48·1) weeks. Change from baseline in supine SVC at 12 weeks was –6·73% with levosimendan and –6·99% with placebo, with no significant difference between the treatments (estimated treatment difference 0·26%, 95% CI –2·03 to 2·55, p=0·83). Similarly, at week 48, CAFS did not differ between treatment groups (least squares mean change from baseline 10·69, 95% CI –15·74 to 37·12; nominal p value=0·43). The most frequent adverse events were increased heart rate (106 [33%] of 326 receiving levosimendan vs 12 [7%] of 166 receiving placebo), fall (85 [26%] vs 48 [29%]), headache (93 [29%] vs 36 [22%]), and dyspnoea (59 [18%] vs 32 [19%]). 33 (10%) participants allocated levosimendan and 20 (12%) assigned placebo died during the trial, mainly due to respiratory failure or progression of amyotrophic lateral sclerosis. Interpretation Levosimendan was not superior to placebo in maintaining respiratory function in a broad population with amyotrophic lateral sclerosis. Although levosimendan was generally well tolerated, increased heart rate and headache occurred more frequently with levosimendan than with placebo. The possibility of a clinically relevant subgroup of responsive individuals requires further evaluation. Funding Orion Corporation.

BackgroundEndovascular thrombectomy (EVT) is efficacious for appropriately selected patients with large vessel occlusions (LVO) up to 24 hours from symptom onset. There is limited information on outcomes of nonagenarians, selected with computed tomography perfusion (CTP) imaging.MethodsWe retrospectively analyzed data from a large academic hospital between December 2017 and October 2019. Patients receiving EVT for anterior circulation LVO were stratified into nonagenarian (≥90 years) and younger (ResultsOverall, 214 consecutive patients (26 nonagenarians, 188 younger) underwent EVT. Nonagenarians were aged 92.8±2.9 years and younger patients were 74.5±13.5 years. Mortality rate was significantly greater in nonagenarians compared with younger patients (43.5% vs 10.4%, OR 9.33, 95% CI 2.88 to 47.97, PConclusionsNonagenarians were noted to have greater mortality and sICH rates following EVT compared with matched younger patients, which may be ameliorated by selecting patients with smaller CTP core volumes. Nonagenarians undergoing EVT had similar rates of successful reperfusion and functional independence compared with the younger cohort.

Background The rapid spread of the coronavirus disease 2019 (COVID-19) with high mortality rate necessitates disease characterization and accurate prognostication for prompt clinical decision-making. The aim of this study is to study clinical characteristics and predictors of mortality in hospitalized patients with COVID-19 in India. Methods Retrospective cohort study was conducted in a tertiary care hospital in northern India. All consecutive confirmed hospitalized COVID-19 cases aged 15 years and older from 13 Apr till 31 Aug 2020 are included. Primary end point was 30-day mortality. Results Of 1622 patients ,1536 cases were valid. Median age was 36 years, 88.3% were men and 58.1% were symptomatic. Fever (37.6%) was commonest presenting symptom. Dyspnea was reported by 15.4%. Primary hypertension (8.5%) was commonest comorbidity, followed by diabetes mellitus (6.7%). Mild, moderate, and severe hypoxemia were seen in 3.4%, 4.3%, and 0.8% respectively. Logistic regression showed greater odds of moderate/severe disease in patients with dyspnea, hypertension, Chronic Kidney Disease (CKD), and malignancy. Seventy six patients died (4.9%). In adjusted Cox proportional hazards model for mortality, patients with dyspnea (hazard ratio [HR]: 14.449 [5.043-41.402]), altered sensorium (HR: 2.762 [1.142-6.683]), Diabetes Mellitus (HR: 1.734 [1.001-3.009]), malignancy (HR:10.443 [4.396-24.805]) and Chronic Liver Disease (CLD) (HR: 14.432 [2.321-89.715]) had higher risk. Rising respiratory rate (HR: 1.098 [1.048-1.150]), falling oxygen saturation (HR: 1.057 per unit change 95% CI: 1.028-1.085) were significant predictors. Conclusion Analysis suggests that age, dyspnea, and malignancy were associated with both severe disease and mortality. Diabetes Mellitus and Chronic Liver Disease were associated with increased the risk of fatal outcome. Simple clinical parameters such as respiratory rate and oxygen saturation are strong predictors and with other risk factors at admission can be effectively used to triage patients.

Objective Indian data on cancer patients with coronavirus disease 2019 (COVID-19) infection and its outcome are limited. Infectivity and outcome among cancer patients staying in large congregations are not known. We conducted this study to address this lacuna in literature.Methods This was a retrospective–prospective, observational study of hospitalized cancer patients with proven COVID-19 infection, conducted at tertiary care hospital in New Delhi after ethical approval. We analyzed clinical, laboratorial parameters, and outcomes in these patients.Results All 32 admitted male patients became COVID-19 positive and 10 (31%) patients were symptomatic. Median age was 37.5 years (range: 16–64). Sixteen health-care workers (HCWs) were working in the cancer ward (paramedical staff: 4, nurses: 6, residents: 4, and consultants: 2). Among HCWs, two nursing staff and three paramedical staff contracted asymptomatic COVID-19. Eighteen (56%) and 14 (44%) patients were on curative and palliative treatment, respectively. Leukopenia (total leukocyte count Conclusion Seventy percent of cancer patients were asymptomatic. Cancer patients living in congregation areas are susceptible to COVID-19 with 3% mortality rate. Recent chemotherapy and associated cytopenias may not increase the risk in cancer patients with COVID-19 treated with curative intent. Palliative intended patients are at increased risk of death. N-95, personal protective equipment, and adherence to infection control measures should be encouraged.

Activation of inhibitor of nuclear factor NF-κB kinase subunit-β (IKKβ), characterized by phosphorylation of activation loop serine residues 177 and 181, has been implicated in the early onset of cancer. On the other hand, tissue-specific IKKβ knockout in Kras mutation-driven mouse models stalled the disease in the precancerous stage. In this study, we used cell line models, tumor growth studies, and patient samples to assess the role of IKKβ and its activation in cancer. We also conducted a hit-to-lead optimization study that led to the identification of 39-100 as a selective mitogen-activated protein kinase kinase kinase (MAP3K) 1 inhibitor. We show that IKKβ is not required for growth of Kras mutant pancreatic cancer (PC) cells but is critical for PC tumor growth in mice. We also observed elevated basal levels of activated IKKβ in PC cell lines, PC patient-derived tumors, and liver metastases, implicating it in disease onset and progression. Optimization of an ATP noncompetitive IKKβ inhibitor resulted in the identification of 39-100, an orally bioavailable inhibitor with improved potency and pharmacokinetic properties. The compound 39-100 did not inhibit IKKβ but inhibited the IKKβ kinase MAP3K1 with low-micromolar potency. MAP3K1-mediated IKKβ phosphorylation was inhibited by 39-100, thus we termed it IKKβ activation modulator (IKAM) 1. In PC models, IKAM-1 reduced activated IKKβ levels, inhibited tumor growth, and reduced metastasis. Our findings suggests that MAP3K1-mediated IKKβ activation contributes to KRAS mutation-associated PC growth and IKAM-1 is a viable pretherapeutic lead that targets this pathway.

Introduction: Tenecteplase has been demonstrated to be an effective option for thrombolysis in patients with acute ischemic stroke (AIS) due to large vessel occlusion (LVO). Thrombolysis decision making is an important component of telestroke consultations. Data is scarce concerning tenecteplase usage in telestroke patients. We aimed to evaluate if tenecteplase was safe and feasible for patients with confirmed LVO in a large telestroke network. Methods: We conducted a retrospective analysis of AIS patients with LVO and treated with thrombolysis from May 2018 to April 2021. We compared outcomes in telestroke patients treated with IV alteplase (May 2018 - April 2020) to patients treated with IV tenecteplase (May 2020 - April 2021). We evaluated our primary efficacy and safety outcomes: 90 day functional outcome as measured by modified Rankin Scale (mRS) and complications related to thrombolysis. Secondary outcomes included door to needle time (DTN) and door in door out time (DIO). Ordinal regression assessed 90 day mRS and binomial logistic regression analysis evaluated complications between the groups. Quantile regression models assessed the median to compared groups for DTN and DIO. Results: There were 3747 telestroke consults during the study period and 537 (14.3%) were found to have an LVO, of which 109 (39 IV tenecteplase; 70 IV alteplase) were eligible, received thrombolysis and were included in this study. Patients treated with IV tenecteplase had significantly less disability at 90 days compared to patients treated with IV alteplase [1 (0 - 4.5) vs. 3 (1 - 6)], adjusted odds ratio (aOR) = 0.31 [95%CI, 0.14 - 0.72], p = 0.006, based on mRS ordinal shift analysis. Similar complication rates were reported between the thrombolysis groups, aOR = 3.73 [95%CI, 0.23 - 59.95], p = 0.35. An adjusted quantile regression model found IV tenecteplase was administered 9.40 minutes quicker, standard error (SE) = 3.67, [95%CI, 2.11 - 16.69] than IV alteplase, p = 0.01. There were no differences reported between thrombolysis groups and DIO, p = 0.63. Conclusion: Telestroke patients presenting with confirmed LVO and treated with IV tenecteplase were found to have better 90 day outcomes compared to patients treated with IV alteplase, without increased complication rates.

Simple Summary Cyclin-dependent kinases (CDKs) are rich and viable therapeutic targets for various cancers. The emergence of event-driven pharmacology as an alternative to occupancy-driven pharmacology has begun to address the challenges associated with selectively targeting CDKs. In this review article, we summarize the CDK inhibitors that are currently in clinical trials. In addition, we provide an overview of PROTAC- and molecular glue-based strategies to modulate CDK function. Abstract The cyclin-dependent kinase (CDK) family of proteins play prominent roles in transcription, mRNA processing, and cell cycle regulation, making them attractive cancer targets. Palbociclib was the first FDA-approved CDK inhibitor that non-selectively targets the ATP binding sites of CDK4 and CDK6. In this review, we will briefly inventory CDK inhibitors that are either part of over 30 active clinical trials or recruiting patients. The lack of selectivity among CDKs and dose-limiting toxicities are major challenges associated with the development of CDK inhibitors. Proteolysis Targeting Chimeras (PROTACs) and Molecular Glues have emerged as alternative therapeutic modalities to target proteins. PROTACs and Molecular glues utilize the cellular protein degradation machinery to destroy the target protein. PROTACs are heterobifunctional molecules that form a ternary complex with the target protein and E3-ligase by making two distinct small molecule–protein interactions. On the other hand, Molecular glues function by converting the target protein into a “neo-substrate” for an E3 ligase. Unlike small molecule inhibitors, preclinical studies with CDK targeted PROTACs have exhibited improved CDK selectivity. Moreover, the efficacy of PROTACs and molecular glues are not tied to the dose of these molecular entities but to the formation of the ternary complex. Here, we provide an overview of PROTACs and molecular glues that modulate CDK function as emerging therapeutic modalities.

We present the neutral hydrogen mass ($M_{\text{HI}}$) function (HIMF) and velocity width ($w_{50}$) function (HIWF) based on a sample of 7857 galaxies from the 40% data release of the ALFALFA survey ($\alpha.40$). The low mass (velocity width) end of the HIMF (HIWF) is dominated by the blue population of galaxies whereas the red population dominates the HIMF (HIWF) at the high mass (velocity width) end. We use a deconvolution method to estimate the HI rotational velocity ($V_{\text{rot}}$) functions (HIVF) from the HIWF for the total, red, and blue samples. The HIWF and HIVF for the red and blue samples are well separated at the knee of the function compared to their HIMFs. We then use recent stacking results from the ALFALFA survey to constrain the halo mass ($M_{\text{h}}$) function of HI-selected galaxies. This allows us to obtain various scaling relations between $M_{\text{HI}} - w_{50} - V_{\text{rot}} - M_{\text{h}}$, which we present. The $M_{\text{HI}} - M_{\text{h}}$ relation has a steep slope ~2.10 at small masses and flattens to ~0.34 at masses larger than a transition halo mass, $\log_{10}(M_{\text{ht}} h^2_{70}/M_{\odot})=10.62$. Our scaling relation is robust and consistent with a volume-limited sample of $\alpha.40$. The $M_{\text{HI}} - M_{\text{h}}$ relation is qualitatively similar to the $M_{\text{star}} - M_{\text{h}}$ relation but the transition halo mass is smaller by ~1.4 dex compared to that of the $M_{\text{star}} - M_{\text{h}}$ relation. Our results suggest that baryonic processes like heating and feedback in larger mass halos suppress HI gas on a shorter time scale compared to star-formation., Comment: 17 pages, 11 figures, 2 tables, Accepted for publication in MNRAS

Developing small molecules that indirectly regulate Mcl-1 function has attracted a lot of attention in recent years. Here, we report the discovery of an aminopyrazole, 2-([1,1′-biphenyl]-4-yl)-N-(5-cyclobutyl-1H-pyrazol-3-yl)acetamide (analog 24), which selectively inhibited cyclin-dependent kinase (CDK) 5 over CDK2 in cancer cell lines. We also show that analog 24 reduced Mcl-1 levels in a concentration-dependent manner in cancer cell lines. Using a panel of doxycycline inducible cell lines, we show that CDK5 inhibitor 24 selectively modulates Mcl-1 function while the CDK4/6 inhibitor 6-acetyl-8-cyclopentyl-5-methyl-2-(5-(piperazin-1-yl)pyridin-2-ylamino)pyrido[2,3-day]pyrimidin-7(8H)-one does not. Previous studies using RNA interference and CRISPR showed that concurrent elimination of Bcl-xL and Mcl-1 resulted in induction of apoptosis. In pancreatic cancer cell lines, we show that either CDK5 knockdown or expression of a dominant negative CDK5 when combined with Bcl2 inhibitor results in synergistic induction of apoptosis. Moreover, concurrent pharmacological perturbation of Mcl-1 and Bcl-xL in pancreatic cancer cell lines using a CDK5 inhibitor analog 24 that reduced Mcl-1 levels and 4-(4-{[2-(4-chlorophenyl)-5,5-dimethyl-1-cyclohexen-1-yl]methyl}-1-piperazinyl)-N-[(4-{[(2R)-4-(4-morpholinyl)-1-(phenylsulfanyl)-2-butanyl]amino}-3-[(trifluoromethyl)sulfonyl]phenyl)sulfonyl] benzamide (navitoclax), a Bcl-2/Bcl-xL/Bcl-w inhibitor, resulted in synergistic inhibition of cell growth and induction of apoptosis. In conclusion, we demonstrate targeting CDK5 will sensitize pancreatic cancers to Bcl-2 inhibitors. SIGNIFICANCE STATEMENT Mcl-1 is stabilized by CDK5-mediated phosphorylation in pancreatic ductal adenocarcinoma, resulting in the deregulation of the apoptotic pathway. Thus, genetic or pharmacological targeting of CDK5 sensitizes pancreatic cancers to Bcl-2 inhibitors, such as navitoclax.

Development of selective kinase inhibitors that target the ATP binding site continues to be a challenge largely due to similar binding pockets. Palbociclib is a cyclin-dependent kinase inhibitor that targets the ATP binding site of CDK4 and CDK6 with similar potency. The enzymatic function associated with the kinase can be effectively probed using kinase inhibitors however the kinase-independent functions cannot. Herein, we report a palbociclib based PROTAC that selectively degrades CDK6 while sparing the homolog CDK4. We used competition studies to characterize the binding and mechanism of CDK6 degradation.

Genetic models validated Inhibitor of nuclear factor (NF) kappa B kinase beta (IKKβ) as a therapeutic target for KRAS mutation associated pancreatic cancer. Phosphorylation of the activation loop serine residue (S(177), S(181)) IKKβ is a key event that drives tumor necrosis factor (TNF) α induced NF-κB mediated gene expression. Here we conducted structure activity relationship (SAR) study to improve potency and oral bioavailability of a quinoxaline analog 13–197 that was previously reported as an NFκB inhibitor for pancreatic cancer therapy. The SAR led to the identification of a novel quinoxaline urea analog 84 that reduced the levels of p-IKKβ in dose- and time-dependent studies. When compared to 13–197, analog 84 was ~2.5-fold more potent in TNFα-induced NFκB inhibition and ~4-fold more potent in inhibiting pancreatic cancer cell growth. Analog 84 exhibited ~4.3-fold greater exposure (AUC(0−)∞) resulting in ~5.7-fold increase in oral bioavailability (%F) when compared to 13–197. Importantly, oral administration of 84 by itself and in combination of gemcitabine reduced p-IKKβ levels and inhibited pancreatic tumor growth in a xenograft model.

Inhibitor of nuclear factor kappa-B kinase subunit beta (IKKβ) is a key regulator of the cannonical NF-κB pathway. IKKβ has been validated as a drug target for pathological conditions, which include chronic inflammatory diseases and cancer. Pharmacological studies revealed that chronic administration of ATP-competitive IKKβ inhibitors resulted in unexpected toxicity. We previously reported the discovery of 13-197 as a non-toxic IKKβ inhibitor that reduced tumor growth. Here, we show that 13-197 inhibits IKKβ in a ATP non-competitive manner and an allosteric pocket at the interface of the kinase and ubiquitin like domains was identified as the potential binding site.

Background: Despite recent advances in therapies, resistance to chemotherapy remains a critical problem in the clinical management of colorectal cancer (CRC). Cancer stem cells (CSCs) play a central role in therapy resistance. Thus, elimination of CSCs is crucial for effective CRC therapy, however, such strategies are limited. Autophagy promotes resistance to cancer therapy, however, whether autophagy protects CSCs to promote resistance to CRC-therapy is not well understood. Moreover, specific and potent autophagy inhibitors are warranted as clinical trials with hydroxychloroquine have not been successful. Methods: Colon cancer cells and tumoroids were used. Fluorescent reporter-based analysis of autophagy flux, spheroid and side population (SP) culture, and qPCR were done. We synthesized 36-077, a potent inhibitor of PIK3C3/VPS34 kinase, to inhibit autophagy. Combination treatments were done using 5-fluorouracil (5-FU) and 36-077. Results: The 5-FU treatment induced autophagy only in a subset of the treated colon cancer. These autophagy-enriched cells also showed increased expression of CSC markers. Co-treatment with 36-077 significantly improved efficacy of the 5-FU treatment. Mechanistic studies revealed that combination therapy inhibited GSK-3β/Wnt/β-catenin signaling to inhibit CSC population. Conclusion: Autophagy promotes resistance to CRC-therapy by specifically promoting GSK-3β/Wnt/β-catenin signaling to promote CSC survival, and 36-077, a PIK3C3/VPS34 inhibitor, helps promote efficacy of CRC therapy.

Noninvasive ventilator support such as bi-level positive airway pressure (BiPAP) or continuous positive airway pressure (CPAP) is often used for patients with obstructive sleep apnea or neuromuscular disorders, such as amyotrophic lateral sclerosis (ALS), where respiratory muscles are weakened. Current commercially available masks for BiPAP and CPAP are often cited as being ill-fitting and leaky, leading to poor quality of sleep or reduced usage of therapy. This project seeks to minimize leaks and maximize comfort by developing custom-fit masks. Patient faces are imaged using an in-house camera system to obtain a three-dimensional (3D) facial contour. Custom interfaces are generated based on this contour using interactive computer software. Using 3D printing to enable rapid tooling, these interfaces are produced in a skin-safe silicone and attached to an off-the-shelf (OTS) mask to create a custom mask. The methodology has been initially tested on five healthy subjects who underwent a two-night sleep study, one night with an OTS mask and one night with a custom-fit mask, to evaluate the leakage and comfort of the custom-fit mask compared to the OTS version. Subjects filled out a questionnaire asking them about mask comfort, leakage, and quality of sleep along with open-ended questions. While the custom-fit mask did not reduce the average measured leakage for subjects, subjects reported experiencing less leakage. Overall, results suggest that the custom-fit masks are more comfortable and tolerable than the provided OTS option. Subject feedback will be implemented into future masks that will be used in a clinical study.

Introduction: Data remains scarce on which telestroke related sub-events (component parts/time intervals) are associated with delays in door-to-needle (DTN) time and goals for each telestroke sub-event. We aimed to assess the telestroke sub-events that contribute to DTN. After establishing set goals for each sub-event, we further evaluated the odds of DTN within 45 minutes if sub-event goals were achieved. Methods: We retrospectively analyzed prospectively collected data from a hub-and-spoke model telestroke network from January 2017 to September 2019. To determine which sub-events significantly contributed to DTN time, a sequential multiple regression analysis was performed. We entered covariates (age, sex, time of telestroke [day or night], NIHSS, average number of telestroke consults at a given site) in the first block followed by sub-events (door-to-telestroke request, door-to-CT, request-to-page, stroke physician response time, telestroke phone-to-video, video duration prior to needle and video completion-to-needle) in the second block. Logistic regression models were performed to estimate the odds of achieving a DTN within 45 minutes if sub-event goals were achieved. Results: During the study, 3361 telestrokes were completed and 306 (9.1%) patients received IV thrombolytics. After exclusions, 253 patients treated with IV thrombolytics were included. Five sub-events contributed to DTN time above and beyond the nuisance variables: door-to-telestroke request, stroke physician response time, telestroke phone-to-video, video duration prior to needle, and video completion-to-needle; each p Conclusions: Telestroke sub-events involving door-to-telestroke request, stroke physician response, telestroke phone-to-video, video duration prior to needle, and video completion-to-needle significantly contribute to DTN time. Successful achievement of sub-event goals was related to greater likelihood of administration of thrombolytic therapy within 45 minutes.

Introduction: Prior studies have demonstrated the importance of measuring and monitoring telestroke sub-events of door-to-needle time (DTN) such as door-to-telestroke request, but there is limited data on educational efforts to reduce sub-events contributing to DTN. We educated spoke coordinators, nurses and physicians at our telestroke sites on tips to reduce sub-events of DTN, developed a reference pocket card and reviewed sub-event metrics during monthly telestroke quality meetings with spoke sites. We aimed to evaluate if our educational activities and monthly data review affected door-to-telestroke request within 10 minutes of arrival, completion-of-video to needle within 1 minute and DTN within 60 minutes. Methods: Prospectively collected data was analyzed retrospectively from a hub-and-spoke model telestroke network. Education of the spokes was completed between January 2019 and April 2019. We compared data for one year prior to education (January 2018 - December 2018) and one year after education (May 2019 - April 2020). Logistic regression analyses were performed to determine the odds of achieving a door-to-telestroke request within 10 minutes, DTN within 60 minutes and completion-of-video to needle within 1 minute in treated patients. We entered possible confounding variables (EMS arrival; NIHSS; posterior symptoms) in the first block followed by the pre/post education groups in the second block. Results: Overall, telestroke was requested 2574 times during the study (1338 pre-education; 1236 post-education). A Chi square test suggested a trend towards more thrombolytic and thrombectomy treatments occurred after education compared to before education (14.6% vs. 12%), OR = 1.25 (95%CI 0.99 - 1.56), p=0.06. Door-to-telestroke request Conclusions: Education and regular review of sub-events data reduced door-to-telestroke request and door-to-needle time in our telestroke network.

The IKK-NFκB complex is a key signaling node that facilitates activation of gene expression in response to extracellular signals. The kinase IKKβ and the transcription factor RELA have been targeted by covalent modifiers that bind to surface exposed cysteine residues. A common feature in well characterized covalent modifiers of RELA and IKKβ is the Michael acceptor containing α-methylene-γ-butyrolactone functionality. Through synthesis and evaluation of a focused set of α-methylene-γ-butyrolactone containing spirocyclic dimers (SpiDs) we identified SpiD3 as an anticancer agent with low nanomolar potency. Using cell-free and cell-based studies we show that SpiD3 is a covalent modifier that generates stable RELA containing high molecular weight complexes. SpiD3 inhibits TNFα-induced IκBα phosphorylation resulting in the blockade of RELA nuclear translocation. SpiD3 induces apoptosis, inhibits colony formation and migration of cancer cells. The NCI-60 cell line screen revealed that SpiD3 potently inhibits growth of leukemia cell lines, making it a suitable pre-therapeutic lead for hematological malignancies.

The unfolded protein response (UPR) is an adaptation mechanism activated to resolve transient accumulation of unfolded/misfolded proteins in the endoplasmic reticulum. Failure to resolve the transient accumulation of such proteins results in UPR-mediated programmed cell death. Loss of tumor suppressor gene or oncogene addiction in cancer cells can result in sustained higher basal UPR levels; however, it is not clear if these higher basal UPR levels in cancer cells can be exploited as a therapeutic strategy. We hypothesized that covalent modification of surface-exposed cysteine (SEC) residues could simulate unfolded/misfolded proteins to activate the UPR, and that higher basal UPR levels in cancer cells would provide the necessary therapeutic window. To test this hypothesis, here we synthesized analogs that can covalently modify multiple SEC residues and evaluated them as UPR activators. We identified a spirocyclic dimer, SpiD7, and evaluated its effects on UPR activation signals, that is, XBP1 splicing, phosphorylation of eIF2α, and a decrease in ATF 6 levels, in normal and cancer cells, which were further confirmed by RNA-Seq analyses. We found that SpiD7 selectively induced caspase-mediated apoptosis in cancer cells, whereas normal cells exhibited robust XBP1 splicing, indicating adaptation to stress. Furthermore, SpiD7 inhibited the growth of high-grade serous carcinoma cell lines ~3-15-fold more potently than immortalized fallopian tube epithelial (paired normal control) cells and reduced clonogenic growth of high-grade serous carcinoma cell lines. Our results suggest that induction of the UPR by covalent modification of SEC residues represents a cancer cell vulnerability and can be exploited to discover novel therapeutics.

Synthetic lethality is a successful strategy employed to develop selective chemotherapeutics against cancer cells. Inactivation of RAD52 is synthetically lethal to homologous recombination (HR) deficient cancer cell lines. Replication protein A (RPA) recruits RAD52 to repair sites, and the formation of this protein-protein complex is critical for RAD52 activity. To discover small molecules that inhibit the RPA:RAD52 protein-protein interaction (PPI), we screened chemical libraries with our newly developed Fluorescence-based protein-protein Interaction Assay (FluorIA). Eleven compounds were identified, including FDA-approved drugs (quinacrine, mitoxantrone, and doxorubicin). The FluorIA was used to rank the compounds by their ability to inhibit the RPA:RAD52 PPI and showed mitoxantrone and doxorubicin to be the most effective. Initial studies using the three FDA-approved drugs showed selective killing of BRCA1-mutated breast cancer cells (HCC1937), BRCA2-mutated ovarian cancer cells (PE01), and BRCA1-mutated ovarian cancer cells (UWB1.289). It was noteworthy that selective killing was seen in cells known to be resistant to PARP inhibitors (HCC1937 and UWB1 SYr13). A cell-based double-strand break (DSB) repair assay indicated that mitoxantrone significantly suppressed RAD52-dependent single-strand annealing (SSA) and mitoxantrone treatment disrupted the RPA:RAD52 PPI in cells. Furthermore, mitoxantrone reduced radiation-induced foci-formation of RAD52 with no significant activity against RAD51 foci formation. The results indicate that the RPA:RAD52 PPI could be a therapeutic target for HR-deficient cancers. These data also suggest that RAD52 is one of the targets of mitoxantrone and related compounds.

Effective and sustained inhibition of non-enzymatic oncogenic driver proteins represents a major pharmacologic challenge. The clinical success of thalidomide analogs demonstrates the therapeutic efficacy of drug-induced degradation of transcription factors and other cancer targets1–3, but a significant subset of proteins are recalcitrant to targeted protein degradation using current approaches4,5. Here we report an alternative mechanism, whereby a small molecule induces highly specific, reversible polymerization, sequestration into cellular foci, and subsequent degradation of a target protein. BI-3802 is a small molecule that binds the BTB domain of the oncogenic transcription factor BCL6 and results in proteasomal degradation6. We used cryo-EM to reveal how the solvent-exposed moiety of a BCL6 inhibitor contributes to a composite ligand/protein surface that engages BCL6 homodimers to form a supramolecular structure. Drug-induced formation of BCL6 filaments facilitates ubiquitination by the SIAH1 E3 ubiquitin ligase. Our findings demonstrate that a small molecule can induce polymerization coupled to highly specific protein degradation, which in the case of BCL6 leads to superior pharmacological activity. These findings create new avenues for the development of therapeutics and synthetic biology.

As per the Japanese or SUPAC guidance to maintain formulation composition similarity across tablet strengths, the coating should be applied based on the core tablet surface area or weight, respectively. These two coating approaches were compared by evaluating protective effects of coating on the light stability of three model compounds. Core tablets of three light sensitive drugs, nifedipine, rosuvastatin calcium, and montelukast sodium were coated either with PVA-based Opadry® II white or Opadry® II beige. The coated tablets were exposed to light up to three ICH cycles. For Opadry® II white, the surface area based coating provided consistent light protection across tablet strengths when the coating amount was more than 0.1 mg/mm2 compared to that based on core tablet weights. For Opadry® II beige, both approaches gave comparable and better light protection due to presence of iron oxides. The light protection by Opadry® II white could be because of physical barrier of coating, which was uniform across the strengths when it was based on core tablet surface area. For a routine tablet formulation development with Opadry color coating, it does not matter whether the coating is applied based on the core tablet surface area or weight.

The release of an active drug from the prodrug generates a pro-fragment that typically has no biological activity and could result in adverse effects. By combining two drugs, wherein each drug acts as a pro-fragment of the other drug will eliminate the pro-fragment in the prodrug. As they are prodrugs of each other and are symbiotic, we termed these as symbiotic prodrugs (SymProDs). To test this idea, we generated SymProDs using NFκB inhibitors that contain the reactive α-methylene-γ-butyrolactone moiety and CDK inhibitors with solvent exposed secondary nitrogen atoms. We show that secondary amine prodrugs of α-methylene-γ-butyrolactone containing NFκB inhibitors undergo slow release over a 72 hr period. Using an alkyne-tagged secondary amine prodrug of α-methylene-γ-butyrolactone containing NFκB inhibitor, we demonstrate target engagement. The NFκB-CDK SymProDs were ~20- to 200-fold less active against the corresponding CDK inhibitors in in vitro CDK kinase assays. Growth inhibition studies in a panel of ovarian cancer cell lines revealed potency trends of the SymProDs mirrored those of the single treatments suggesting their dissociation in cells. In conclusion, our results suggest that SymProDs offer a productive path forward for advancing compounds with reactive functionality and can be used as dual targeting agents.