My clinical interests include trauma/critical care and acute care surgery. I am interested in how the application of innovative solutions (both device and informatics based) can improve patient care and outcomes in developed as well as developing world settings.
- A suite of phenotypic assays to ensure pipeline diversity when prioritizing drug-like Cryptosporidium growth inhibitors NATURE COMMUNICATIONS 2019; 10
A suite of phenotypic assays to ensure pipeline diversity when prioritizing drug-like Cryptosporidium growth inhibitors.
2019; 10 (1): 1862
Cryptosporidiosis is a leading cause of life-threatening diarrhea in children, and the only currently approved drug is ineffective in malnourished children and immunocompromised people. Large-scale phenotypic screens are ongoing to identify anticryptosporidial compounds, but optimal approaches to prioritize inhibitors and establish a mechanistically diverse drug development pipeline are unknown. Here, we present a panel of medium-throughput mode of action assays that enable testing of compounds in several stages of the Cryptosporidium life cycle. Phenotypic profiles are given for thirty-nine anticryptosporidials. Using a clustering algorithm, the compounds sort by phenotypic profile into distinct groups of inhibitors that are either chemical analogs (i.e. same molecular mechanism of action (MMOA)) or known to have similar MMOA. Furthermore, compounds belonging to multiple phenotypic clusters are efficacious in a chronic mouse model of cryptosporidiosis. This suite of phenotypic assays should ensure a drug development pipeline with diverse MMOA without the need to identify underlying mechanisms.
View details for PubMedID 31015448
A Novel Piperazine-Based Drug Lead for Cryptosporidiosis from the Medicines for Malaria Venture Open-Access Malaria Box.
Antimicrobial agents and chemotherapy
2018; 62 (4)
Cryptosporidiosis causes life-threatening diarrhea in children under the age of 5 years and prolonged diarrhea in immunodeficient people, especially AIDS patients. The standard of care, nitazoxanide, is modestly effective in children and ineffective in immunocompromised individuals. In addition to the need for new drugs, better knowledge of drug properties that drive in vivo efficacy is needed to facilitate drug development. We report the identification of a piperazine-based lead compound for Cryptosporidium drug development, MMV665917, and a new pharmacodynamic method used for its characterization. The identification of MMV665917 from the Medicines for Malaria Venture Malaria Box was followed by dose-response studies, in vitro toxicity studies, and structure-activity relationship studies using commercial analogues. The potency of this compound against Cryptosporidium parvum Iowa and field isolates was comparable to that against Cryptosporidium hominis Furthermore, unlike nitazoxanide, clofazimine, and paromomycin, MMV665917 appeared to be curative in a NOD SCID gamma mouse model of chronic cryptosporidiosis. MMV665917 was also efficacious in a gamma interferon knockout mouse model of acute cryptosporidiosis. To determine if efficacy in this mouse model of chronic infection might relate to whether compounds are parasiticidal or parasitistatic for C. parvum, we developed a novel in vitro parasite persistence assay. This assay suggested that MMV665917 was parasiticidal, unlike nitazoxanide, clofazimine, and paromomycin. The assay also enabled determination of the concentration of the compound required to maximize the rate of parasite elimination. This time-kill assay can be used to prioritize early-stage Cryptosporidium drug leads and may aid in planning in vivo efficacy experiments. Collectively, these results identify MMV665917 as a promising lead and establish a new method for characterizing potential anticryptosporidial agents.
View details for DOI 10.1128/AAC.01505-17
View details for PubMedID 29339392
View details for PubMedCentralID PMC5913971
Identification of Cryptosporidium parvum active chemical series by Repurposing the open access malaria box.
Antimicrobial agents and chemotherapy
2014; 58 (5): 2731–39
The apicomplexan parasites Cryptosporidium parvum and Cryptosporidium hominis are major etiologic agents of human cryptosporidiosis. The infection is typically self-limited in immunocompetent adults, but it can cause chronic fulminant diarrhea in immunocompromised patients and malnutrition and stunting in children. Nitazoxanide, the current standard of care for cryptosporidiosis, is only partially efficacious for children and is no more effective than a placebo for AIDS patients. Unfortunately, financial obstacles to drug discovery for diseases that disproportionately affect low-income countries and technical limitations associated with studies of Cryptosporidium biology impede the development of better drugs for treating cryptosporidiosis. Using a cell-based high-throughput screen, we queried the Medicines for Malaria Venture (MMV) Open Access Malaria Box for activity against C. parvum. We identified 3 novel chemical series derived from the quinolin-8-ol, allopurinol-based, and 2,4-diamino-quinazoline chemical scaffolds that exhibited submicromolar potency against C. parvum. Potency was conserved in a subset of compounds from each scaffold with varied physicochemical properties, and two of the scaffolds identified exhibit more rapid inhibition of C. parvum growth than nitazoxanide, making them excellent candidates for further development. The 2,4-diamino-quinazoline and allopurinol-based compounds were also potent growth inhibitors of the related apicomplexan parasite Toxoplasma gondii, and a good correlation was observed in the relative activities of the compounds in the allopurinol-based series against T. gondii and C. parvum. Taken together, these data illustrate the utility of the Open Access Malaria Box as a source of both potential leads for drug development and chemical probes to elucidate basic biological processes in C. parvum and other apicomplexan parasites.
View details for DOI 10.1128/AAC.02641-13
View details for PubMedID 24566188
View details for PubMedCentralID PMC3993250
Drug repurposing screen reveals FDA-approved inhibitors of human HMG-CoA reductase and isoprenoid synthesis that block Cryptosporidium parvum growth.
Antimicrobial agents and chemotherapy
2013; 57 (4): 1804–14
Cryptosporidiosis, a diarrheal disease usually caused by Cryptosporidium parvum or Cryptosporidium hominis in humans, can result in fulminant diarrhea and death in AIDS patients and chronic infection and stunting in children. Nitazoxanide, the current standard of care, has limited efficacy in children and is no more effective than placebo in patients with advanced AIDS. Unfortunately, the lack of financial incentives and the technical difficulties associated with working with Cryptosporidium parasites have crippled efforts to develop effective treatments. In order to address these obstacles, we developed and validated (Z' score = 0.21 to 0.47) a cell-based high-throughput assay and screened a library of drug repurposing candidates (the NIH Clinical Collections), with the hopes of identifying safe, FDA-approved drugs to treat cryptosporidiosis. Our screen yielded 21 compounds with confirmed activity against C. parvum growth at concentrations of <10 μM, many of which had well-defined mechanisms of action, making them useful tools to study basic biology in addition to being potential therapeutics. Additional work, including structure-activity relationship studies, identified the human 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor itavastatin as a potent inhibitor of C. parvum growth (50% inhibitory concentration [IC(50)] = 0.62 μM). Bioinformatic analysis of the Cryptosporidium genomes indicated that the parasites lack all known enzymes required for the synthesis of isoprenoid precursors. Additionally, itavastatin-induced growth inhibition of C. parvum was partially reversed by the addition of exogenous isopentenyl pyrophosphate, suggesting that itavastatin reduces Cryptosporidium growth via on-target inhibition of host HMG-CoA reductase and that the parasite is dependent on the host cell for synthesis of isoprenoid precursors.
View details for DOI 10.1128/AAC.02460-12
View details for PubMedID 23380723
View details for PubMedCentralID PMC3623326
Drug repurposing: mining protozoan proteomes for targets of known bioactive compounds.
Journal of the American Medical Informatics Association : JAMIA
2013; 21 (2): 238–44
To identify potential opportunities for drug repurposing by developing an automated approach to pre-screen the predicted proteomes of any organism against databases of known drug targets using only freely available resources.We employed a combination of Ruby scripts that leverage data from the DrugBank and ChEMBL databases, MySQL, and BLAST to predict potential drugs and their targets from 13 published genomes. Results from a previous cell-based screen to identify inhibitors of Cryptosporidium parvum growth were used to validate our in-silico prediction method.In-vitro validation of these results, using a cell-based C parvum growth assay, showed that the predicted inhibitors were significantly more likely than expected by chance to have confirmed activity, with 8.9-15.6% of predicted inhibitors confirmed depending on the drug target database used. This method was then used to predict inhibitors for the following 13 disease-causing protozoan parasites, including: C parvum, Entamoeba histolytica, Giardia intestinalis, Leishmania braziliensis, Leishmania donovani, Leishmania major, Naegleria gruberi (in proxy of Naegleria fowleri), Plasmodium falciparum, Plasmodium vivax, Toxoplasma gondii, Trichomonas vaginalis, Trypanosoma brucei and Trypanosoma cruzi.Although proteome-wide screens for drug targets have disadvantages, in-silico methods can be developed that are fast, broad, inexpensive, and effective. In-vitro validation of our results for C parvum indicate that the method presented here can be used to construct a library for more directed small molecule screening, or pipelined into structural modeling and docking programs to facilitate target-based drug development.
View details for DOI 10.1136/amiajnl-2013-001700
View details for PubMedID 23757409
View details for PubMedCentralID PMC3932453
Avian hosts of West Nile virus in Puerto Rico.
Vector borne and zoonotic diseases (Larchmont, N.Y.)
2012; 12 (1): 47–54
West Nile virus (WNV) ecology in neotropical ecosystems is poorly understood, and vertebrate hosts responsible for infecting mosquitoes remain unidentified throughout the Caribbean Basin. After a period of intense WNV transmission among sentinel chickens near Ceiba, Puerto Rico, we measured abundance of resident birds and species-specific prevalence of WNV infection. Taking the product of these measures indicates the relative number of WNV infections by species. Greater Antillean grackle (Quiscalus niger) accounted for the most WNV infections among birds in our 100-km(2) study site. In urban habitats, the house sparrow (Passer domesticus) was frequently infected. Immature birds less than one year of age were more likely to have detectable WNV-reactive antibodies than older birds of the same species.
View details for DOI 10.1089/vbz.2011.0609
View details for PubMedID 21923260
Dengue virus seroprevalence among febrile patients in Bamako, Mali: results of a 2006 surveillance study.
Vector borne and zoonotic diseases (Larchmont, N.Y.)
2011; 11 (11): 1479–85
Dengue viruses (DENV) are endemic in over 100 countries worldwide, and annually 50 to 100 million people are infected by one of the four DENV serotypes, whereas over 2.5 billion people are at risk for infection. West African countries lack the surveillance to determine the true incidence of dengue; hence, this disease is likely significantly underestimated. In Mali, ?14 million people are potentially at risk of acquiring a dengue infection.A serosurvey for DENV was conducted on 95 human serum samples obtained from the Institute National de Recherche en Sante Publique in 2006. DENV-specific IgM and IgG enzyme-linked immunosorbent assays were performed on all samples, and a subset was tested using the plaque-reduction neutralization test against the DENV and yellow fever virus (YFV). Samples collected during the acute infection (0-5 days postonset of symptoms) were tested for dengue NS1 antigen and reverse-transcriptase polymerase chain reaction for Flaviviruses, Alphaviruses, and Bunyaviruses RNA. A total of 87 (93%) of samples were positive for anti-DENV IgG antibodies. Of a subset of 13 IgG positive samples, 2 samples neutralized monotypically against DENV-1 and -2, whereas 3 others neutralized broadly against YFV and multiple DENV. Although no polymerase chain reaction positives were found, DENV NS1 was detected in 1 of the 20 acute samples tested.Of the 93 human serum samples tested, the dengue prevalence based on dengue IgG enzyme-linked immunosorbent assay results was 93%. Three DENV specific positive samples and two YFV positives were identified by plaque-reduction neutralization test. Finally, one sample tested positive for dengue NS1, thus suggestive of an acute infection within 14 days of obtaining the sample from the patient. Based on these serological data from this study, YFV and DENV appear to be co-circulating in Mali.
View details for DOI 10.1089/vbz.2011.0622
View details for PubMedID 21767159
Utility of a commercial nonstructural protein 1 antigen capture kit as a dengue virus diagnostic tool.
Clinical and vaccine immunology : CVI
2010; 17 (6): 949–53
Annually, over 2.5 billion people are at risk for infection with dengue virus (DENV), while between 50 and 100 million people contract the infection. There is an urgent need for alternative diagnostic tools that can detect DENV during acute infection. Recent studies have shown that DENV nonstructural protein 1 (NS1) is detectable in the blood as early as the onset of symptoms and persists well into the convalescent phase of the infection. We evaluated the utility of the Bio-Rad Platelia DENV NS1 antigen capture kit in combination with real-time reverse transcriptase PCR (RT-PCR) and an IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) for refining a new algorithm for the diagnosis of acute- or convalescent-phase DENV infection with a single clinical sample. We tested the Bio-Rad kit with three panels of sera. These panels were designed to evaluate the sensitivities of the NS1 kit for (i) early-convalescent-phase samples, (ii) acute-phase samples with false-negative PCR results, and (iii) IgM-negative convalescent-phase samples from patients with confirmed secondary DENV infections. Results show that NS1 can be detected in 22% of serum samples collected more than 10 days after the onset of illness and in 22% of samples that did not elicit an IgM response. Additionally, NS1 was detected in 37% of the tested acute-phase samples with false-negative PCR results, suggesting that NS1 detection may be valuable in increasing the sensitivity of current acute-phase diagnostics. These results will improve diagnosis with a single acute-phase or early-convalescent-phase sample for disease surveillance and clinical diagnosis.
View details for DOI 10.1128/CVI.00041-10
View details for PubMedID 20410325
View details for PubMedCentralID PMC2884419
Evaluation of commercially available anti-dengue virus immunoglobulin M tests.
Emerging infectious diseases
2009; 15 (3): 436–40
Anti-dengue virus immunoglobulin M kits were evaluated. Test sensitivities were 21%-99% and specificities were 77%-98% compared with reference ELISAs. False-positive results were found for patients with malaria or past dengue infections. Three ELISAs showing strong agreement with reference ELISAs will be included in the World Health Organization Bulk Procurement Scheme.
View details for DOI 10.3201/eid1503.080923
View details for PubMedID 19239758
View details for PubMedCentralID PMC2681117
West Nile virus from blood donors, vertebrates, and mosquitoes, Puerto Rico, 2007.
Emerging infectious diseases
2009; 15 (8): 1298–1300
West Nile virus (WNV) was isolated from a human blood donor, a dead falcon, and mosquitoes in Puerto Rico in 2007. Phylogenetic analysis of the 4 isolates suggests a recent introduction of lineage I WNV that is closely related to WNV currently circulating in North America.
View details for DOI 10.3201/eid1508.090333
View details for PubMedID 19751597
View details for PubMedCentralID PMC2815984
First isolation of West Nile virus in the Caribbean.
The American journal of tropical medicine and hygiene
2008; 78 (4): 666–68
A sentinel chicken program for West Nile virus (WNV) surveillance was initiated in July 2006 in eastern Puerto Rico, yielding the first seroconversions on June 4, 2007. WNV was isolated from sentinel chicken serum and mosquito pools (Culex nigripalpus, Culex bahamensis) for the first time in Tropical America. Preliminary sequence analysis of the prM and E genes revealed a 1-amino acid difference (V159A) between the Puerto Rican 2007 and the NY99. This mutation has been observed in the current dominant clade circulating in the United States. Sentinel chicken surveillance was a useful tool for the detection of West Nile virus in the tropics.
View details for PubMedID 18385366
Comparison of two commercially available dengue virus (DENV) NS1 capture enzyme-linked immunosorbent assays using a single clinical sample for diagnosis of acute DENV infection.
Clinical and vaccine immunology : CVI
2008; 15 (10): 1513–18
Dengue virus (DENV) nonstructural protein 1 (NS1) has shown promise as a novel diagnostic marker of acute DENV infection. Current techniques used to diagnose acute DENV infection, including virus isolation and reverse transcription-PCR (RT-PCR), are costly and difficult to perform, while traditional serological assays have low sensitivities during the acute stage of infection. Two commercially available NS1 antigen capture enzyme-linked immunosorbent assays (ELISAs), the Platelia dengue NS1Ag test (Bio-Rad Laboratories, Marnes La Coquette, France) and the Pan-E dengue early ELISA test (Panbio Diagnostics, Brisbane, Australia), were evaluated against a well-characterized panel of 208 real-time RT-PCR- and virus isolation-positive sera, as well as 45 real-time RT-PCR- and serologically negative sera from patients with other acute febrile illnesses. The overall sensitivities were 64.9% (95% confidence interval [CI(95)], 58.2 to 71.1%) for the Panbio test and 83.2% (CI(95), 77.5 to 87.7%) for the Bio-Rad test, with interserotype variation, especially for DENV serotype 4. Predictive models were constructed to identify factors that had a significant influence on a test's outcome with respect to this panel of samples in order to identify the conditions in which the test will be most effective as a diagnostic tool. The immunoglobulin G titer was found to be the only covariate that significantly influenced results in the Bio-Rad test, while serotype and the day postonset were found to significantly influence results in the Panbio test. We concluded that the NS1 capture ELISA is a useful tool that can improve testing algorithms to diagnose DENV infection in single samples from acute and early convalescent cases.
View details for DOI 10.1128/CVI.00140-08
View details for PubMedID 18685015
View details for PubMedCentralID PMC2565928