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A patient with B-cell acute lymphoblastic leukemia (ALL) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) had persistent, progressive pneumonia with viremia after 5 months of infection despite monoclonal antibodies, intravenous (IV) remdesivir and prolonged oral steroids. Twenty days of nirmatrelvir/ritonavir and 10 days of IV remdesivir led to full recovery.

Background Nirmatrelvir with ritonavir (nirmatrelvir/r) is an oral antiviral COVID-19 treatment. We report its efficacy to shorten time to sustained alleviation and resolution of COVID-19 signs/symptoms in nonhospitalized adults with COVID-19 at high risk of severe disease as of primary completion data cut (11 Dec 2021). Methods In this phase 2/3 double-blind study, eligible adults with confirmed SARS-CoV-2 and ≤ 5 days (d) of symptoms were randomized 1:1 to nirmatrelvir/r 300 mg/100 mg or placebo (PBO) every 12 hrs for 5 d. Pts logged presence and severity (on 3- or 4-point scales) of prespecified COVID-19 signs/symptoms daily Day 1 (pre-dose) through 28. Times to sustained alleviation and resolution of all targeted signs/symptoms were assessed, summarized with Kaplan-Meier curves, and compared by treatment by log-rank test. Individual signs/symptoms were compared with descriptive analyses. Results From Jul–Dec 2021, 2246 pts enrolled; 2085 pts (nirmatrelvir/r, n=1039; PBO, n=1046) met criteria for the mITT1 population (≤ 5 d of symptom onset, did not/not expected to receive an mAb). More pts achieved sustained alleviation or sustained resolution with nirmatrelvir/r. Shorter median times to sustained alleviation/resolution were observed with nirmatrelvir/r (13/16 d) vs PBO (15/19 d; Fig 1 & 2). Also, a shorter median time to sustained alleviation was seen in pts treated ≤ 3 d of symptoms with nirmatrelvir/r (12 d) vs PBO (15 d). The most common symptoms were cough, muscle/body aches, and headache in both groups. The median time to sustained alleviation of cough and headache was 2 d less with nirmatrelvir/r vs PBO. The median time to sustained resolution of muscle aches and shortness of breath was 3 d and 4 d less with nirmatrelvir/r. The proportion of pts with severe signs/symptoms in the nirmatrelvir/r vs PBO group was significantly higher at baseline, but significantly lower after treatment, showing nirmatrelvir/r significantly reduced symptom severity through Day 28 (Fig 3). Pts who were seronegative vs seropositive or had high vs low viral load at baseline achieved faster times to sustained alleviation with nirmatrelvir/r vs PBO. Conclusion Nirmatrelvir/r treatment reduced duration and severity of COVID-19 symptoms vs PBO in pts at high risk of progressing to severe disease. NCT04960202. Disclosures Jennifer Hammond, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Heidi Leister-Tebbe, BSN, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Annie Gardner, MPH, MSPT, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Paula Abreu, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Weihang Bao, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wayne Wisemandle, MA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wajeeha Ansari, MPH, Pfizer Inc.: Stocks/Bonds Magdalena Alicja Harrington, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Jesus Abraham Simón Campos, MD, AstraZeneca: Board Member|AstraZeneca: Speaker|Eli Lilly: Board Member|Pfizer: Board Member|Roche: Board Member|Roche: Speaker Kara W. Chew, M.D., M.S., Merck Sharp & Dohme: Grant/Research Support Rienk Pypstra, MD, MBA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds James M Rusnak, MD, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds.

Background Nirmatrelvir coadministered with ritonavir (nirmatrelvir/r) is a COVID-19 treatment. This study evaluated nirmatrelvir/r in nonhospitalized, symptomatic adults with COVID-19 at high risk of progressing to severe disease. We report secondary efficacy endpoints associated with COVID-19─related medical visits, including hospitalization details and oxygen support, as of the primary completion data cutoff (Dec 11, 2021). Methods In this phase 2/3 double-blind, interventional study, adults with confirmed SARS-CoV-2 and symptom onset ≤ 5 days (d) were randomized 1:1 to receive nirmatrelvir/r 300 mg/100 mg or placebo (PBO) orally every 12 hours for 5 d. COVID-19─related medical visits were collected through Day 28. Oxygen support for COVID-19 and details of COVID-19─related hospitalization, including duration, intensive care unit (ICU) status, and mechanical ventilation, were assessed. Results Of the 2246 patients (pts) enrolled globally from Jul to Dec2021, 2085 (nirmatrelvir/r, n=1039; PBO, n=1046) started treatment and met criteria for the modified intent-to-treat population (mITT1; ≤ 5 d of symptom onset, did not/not expected to receive a mAb). Fewer overall COVID-19-related medical visits were reported with nirmatrelvir/r vs PBO (Table 1). In addition to fewer hospitalizations being reported with nirmatrelvir/r (n=8 [0.8%]) vs PBO (n=65 [6.2%]), pts receiving nirmatrelvir/r had fewer hospitalized d (Table 2), with mean durations of 9.6 (range, 5.0, 16.0) d with nirmatrelvir/r and 11.2 (range, 2.0, 57.0) d with PBO in hospitalized pts. No pts in the nirmatrelvir/r group and 9 pts (0.9%) in PBO group were admitted to the ICU. No pts in the nirmatrelvir/r group received mechanical ventilation vs 3 pts in the PBO group. Fewer other COVID-19─related nonhospital medical visits were reported with nirmatrelvir/r vs PBO (Table 3). In the full analysis set, fewer pts required oxygen therapy for COVID-19 with nirmatrelvir/r (n=9/1120 [0.8%]) vs PBO (n=54/1126 [4.8%]). Conclusion High-risk adults with symptomatic COVID-19 treated with nirmatrelvir/r within 5 d of symptom onset had fewer COVID-19─related medical visits and reduced healthcare utilization (no ICU visits, no mechanical ventilation, fewer days in hospital) vs pts receiving PBO. Clinical Trial: NCT04960202. Disclosures Jennifer Hammond, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Heidi Leister-Tebbe, BSN, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Annie Gardner, MPH, MSPT, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Paula Abreu, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Weihang Bao, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wayne Wisemandle, MA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds Wajeeha Ansari, MPH, Pfizer Inc.: Stocks/Bonds Rienk Pypstra, MD, MBA, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds James M Rusnak, MD, PhD, Pfizer Inc: Employee|Pfizer Inc: Stocks/Bonds.

A patient with B-cell acute lymphoblastic leukemia (ALL) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) had persistent, progressive pneumonia with viremia after 5 months of infection despite monoclonal antibodies, IV remdesivir and prolonged oral steroids. Twenty days of nirmatrelvir/ritonavir and 10 days of IV remdesivir led to full recovery.

Nirmatrelvir is an orally administered severe acute respiratory syndrome coronavirus 2 main protease (MWe conducted a phase 2-3 double-blind, randomized, controlled trial in which symptomatic, unvaccinated, nonhospitalized adults at high risk for progression to severe coronavirus disease 2019 (Covid-19) were assigned in a 1:1 ratio to receive either 300 mg of nirmatrelvir plus 100 mg of ritonavir (a pharmacokinetic enhancer) or placebo every 12 hours for 5 days. Covid-19-related hospitalization or death from any cause through day 28, viral load, and safety were evaluated.A total of 2246 patients underwent randomization; 1120 patients received nirmatrelvir plus ritonavir (nirmatrelvir group) and 1126 received placebo (placebo group). In the planned interim analysis of patients treated within 3 days after symptom onset (modified intention-to treat population, comprising 774 of the 1361 patients in the full analysis population), the incidence of Covid-19-related hospitalization or death by day 28 was lower in the nirmatrelvir group than in the placebo group by 6.32 percentage points (95% confidence interval [CI], -9.04 to -3.59; P0.001; relative risk reduction, 89.1%); the incidence was 0.77% (3 of 389 patients) in the nirmatrelvir group, with 0 deaths, as compared with 7.01% (27 of 385 patients) in the placebo group, with 7 deaths. Efficacy was maintained in the final analysis involving the 1379 patients in the modified intention-to-treat population, with a difference of -5.81 percentage points (95% CI, -7.78 to -3.84; P0.001; relative risk reduction, 88.9%). All 13 deaths occurred in the placebo group. The viral load was lower with nirmatrelvir plus ritonavir than with placebo at day 5 of treatment, with an adjusted mean difference of -0.868 logTreatment of symptomatic Covid-19 with nirmatrelvir plus ritonavir resulted in a risk of progression to severe Covid-19 that was 89% lower than the risk with placebo, without evident safety concerns. (Supported by Pfizer; ClinicalTrials.gov number, NCT04960202.).

Recent studies in our laboratory have shown that UCN-01 (7-hydroxystaurosporine), which is a derivative of the non-selective protein kinase inhibitor staurosporine that exhibits relative selectivity for protein kinase C (PKC), is a potent inhibitor of glioma growth in in vitro and in vivo models. This agent exhibits both cytotoxic and cytostatic effects, depending on the time period of drug exposure. In the present study, we examined whether UCN-01-induced cytotoxicity correlated with the induction of apoptosis, and characterized further the time course of this process as a prelude to application of UCN-01 in clinical trials. We first demonstrated that the cytotoxic effects of UCN-01 were associated with the induction of morphological features of apoptosis. Secondly, we identified electrophoretic features of apoptosis semiquantitatively at a series of time points using field inversion gel electrophoresis. These studies showed a peak in the induction of high-molecular-weight DNA fragmentation after 3-6 days of drug treatment. Thirdly, we measured the percentage of cells undergoing apoptosis at various time points using a terminal transferase-catalyzed in situ end-labeling technique, which confirmed a time- and concentration-dependent increase in apoptotic cell numbers. This correlated with a progressive decrease in the percentage of cells that were viable as assessed by trypan blue exclusion. Cell killing peaked within 2-4 days after beginning UCN-01 treatment, but continued at a lower level in the ensuing days. Taken together, these studies demonstrated that extended periods of exposure to UCN-01 are needed for optimal manifestation of cytotoxic effects against glioma cells, a factor that must be taken into consideration in the design of future clinical trials with this agent for malignant gliomas.

Tyrosine phosphatases (PTPases) dephosphorylate phosphotyrosines while dual-specificity phosphatases (DSPases) dephosphorylate contiguous and semicontiguous phosphothreonine and phosphotyrosine on cyclin dependent kinases and mitogen-activated protein kinases. Consequently, PTPases and DSPases have a central role controlling signal transduction and cell cycle progression. Currently, there are few readily available potent inhibitors of PTPases or DSPases other than vanadate. Using a pharmacophore modeled on natural product inhibitors of phosphothreonine phosphatases, we generated a refined library of novel, phosphate-free, small-molecule compounds synthesized by a parallel, solid-phase combinatorial-based approach. Among the initial 18 members of this targeted diversity library, we identified several inhibitors of DSPases: Cdc25A, -B, and -C and the PTPase PTP1B. These compounds at 100 microM did not significantly inhibit the protein serine/threonine phosphatases PP1 and PP2A. Kinetic studies with two members of this library indicated competitive inhibition for Cdc25 DSPases and noncompetitive inhibition for PTP1B. Compound AC-alphaalpha69 had a Ki of approximately 10 microM for recombinant human Cdc25A, -B, and -C, and a Ki of 0.85 microM for the PTP1B. The marked differences in Cdc25 inhibition as compared to PTP1B inhibition seen with relatively modest chemical modifications in the modular side chains demonstrate the structurally demanding nature of the DSPase catalytic site distinct from the PTPase catalytic site. These results represent the first fundamental advance toward a readily modifiable pharmacophore for synthetic PTPase and DSPase inhibitors and illustrate the significant potential of a combinatorial-based strategy that supplements the rational design of a core structure by a randomized variation of peripheral substituents.

SUMMARY Metallothioneins (MTs) are major intracellular, zinc-binding proteins with antioxidant properties. Mouse embryonic cells null for MT due to loss of functional MT I and II genes (MT2/2) were more susceptible to apoptotic death after exposure to tert-butyl hydroperoxide or the anti-cancer agents cytosine arabinoside, bleomycin, melphalan, and cis-dichlorodiammineplatinum(II) compared with wild-type mouse embryonic cells (MT1/1). We measured basal levels of the tumor suppressor protein p53 and the death effector protein Bax and found the basal levels of both proteins were higher in MT null cells compared with MT1/1 cells. After treatment with the DNA-damaging agent cis-dichlorodiammineplatinum(II), p53 protein levels were induced in both MT1/1 and MT2/2 cells with MT null cells always maintaining the highest p53 levels. The elevated sensitivity to apoptosis was not restricted to embryonic cells. Primary pulmonary fibroblasts were isolated from distinct litters of MT null, heterozygous, and wild-type mice, and all had undetectable basal MT levels. Zinc exposure increased MT levels in the wild-type and heterozygous fibroblasts but not in the MT null fibroblasts. Consistent with the induced MT levels, we found MT1/1 and MT1/2 embryonic cells were less sensitive to cis-dichlorodiammineplatinum(II)-induced apoptosis compared with MT2/2 cells. Our results implicate MT as a stressresponsive factor that can regulate apoptotic engagement.

Pharmacogenomics is a term recently coined to embody the concept of individualized and rational drug selection based on the genotype of a particular patient. Customization of drug therapy offers the potential for optimal safety and efficacy in an individual patient. Such a process contrasts current prescribing practices, which use medications shown to be safe and effective in patient populations or based on anecdotal experiences. Within patient populations, medications vary in their efficacy among individual patients. More importantly, a medication that is safe and effective in one patient may be ineffective or even harmful in another. Underlying many of these phenotypic differences are genotypic variants (polymorphisms) of key enzymes and proteins that affect the safety and efficacy of a drug in an individual patient. An understanding of these polymorphisms has the potential to enhance patient care by allowing physicians to customize the selection of medication to meet individual patient needs. Pharmacogenomics may also lead to improved compliance and shorter time to optimal disease management, thereby reducing morbidity and mortality. Significant cost savings could result from reductions in polypharmacy as well as from fewer physician encounters and hospitalizations for exacerbations of underlying illness and because of adverse drug reactions.

N 4 -Dipeptidyl slow-release forms of the anticancer drug ara -C were prepared by acylation of the lithiated nucleotide with 4,4-dialkyloxazolinones. An azapeptide prodrug of ara -C was obtained by condensation of an amino acid hydrazide with an activated nucleotide urea. The use of unnatural amino acid residues at N 4 prevented nonspecific proteolytic cleavage in biological medium. ara -C prodrugs 10, 15, 17 , and 19 released active drug with half-lives from a few minutes to several days, respectively. Activation via intramolecular N 4 -deacylation did not require enzymatic intervention but was strictly dependent on the structure of the peptide chain. The prodrugs 10, 15 , and 17 produced similar growth inhibition as ara -C in cultured murine leukemia cells while the azapeptide prodrug 19 was less potent reflecting the slow release of active drug with this compound. All four prodrugs retained the ability to induce apoptosis in human HL-60 leukemia cells with kinetics dictated by the rate of intramolecular N 4 -deacylation. This the first demonstration for the control of apoptotic cell death by the modulation of drug release from prodrugs.

Protein kinase C (PKC) has been implicated in propagating signals for apoptosis. We have investigated the effect of pharmacological modulation of PKC activity in DU-145 human androgen-independent prostatic carcinoma cells. The apoptotic death of these cells is characterized by the acquisition of classical apoptotic morphology and generation ofor = 1-mbp and 450- to 600-kbp DNA fragments in attached preapoptotic cell populations prior to cellular detachment and accrual of 30- to 50-kbp DNA fragments. We found that induction of apoptosis was arrested by downregulation of PKC activity and not by transient activation or inhibition of the enzyme. Concentrations and durations of exposure to phorbol esters that downregulated PKC activity correlated with inhibition of VP-16 or melphalan-induced morphological apoptosis and generation of the 30-to-50-kbp DNA fragments. Chronic exposure to phorbol-12,13-dibutyrate (PDBu) did not, however, suppress production of theor = 1-mbp and 450- to 600-kbp DNA fragments found in preapoptotic cell populations, suggesting that PKC downregulation may interfere with the transition between a preapoptotic cell and an apoptotic cell. PKC isozyme analysis revealed that chronic PDBu treatment caused downregulation of PKC-alpha and -epsilon in DU-145 cells. Using concentrations of the PKC inhibitor UCN-01 that were consistent with PKC-alpha inhibition (but not PKC-epsilon inhibition), however, did not mimic the effects of chronic PDBu treatment, implying that downregulation of PKC-epsilon may be of particular importance. Together, these findings suggest that phorbol esters may act as tumor promoters by suppressing apoptosis.

A two step synthesis of the title compound from the rubber serum by-product, quebrachitol, is reported.

A synthesis of 1L-1-deoxy-1-fluoro-myo-inositol in optically pure form starting from myo-inositol is reported.