86 results on '"Overton, L."'
Search Results
2. Pomalidomide, bortezomib, and dexamethasone for patients with relapsed or refractory multiple myeloma previously treated with lenalidomide (OPTIMISMM): a randomised, open-label, phase 3 trial
- Author
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Richardson, Paul G, Oriol, Albert, Beksac, Meral, Liberati, Anna Marina, Galli, Monica, Schjesvold, Fredrik, Lindsay, Jindriska, Weisel, Katja, White, Darrell, Facon, Thierry, San Miguel, Jesus, Sunami, Kazutaka, O'Gorman, Peter, Sonneveld, Pieter, Robak, Pawel, Semochkin, Sergey, Schey, Steve, Yu, Xin, Doerr, Thomas, Bensmaine, Amine, Biyukov, Tsvetan, Peluso, Teresa, Zaki, Mohamed, Anderson, Kenneth, Dimopoulos, Meletios, OPTIMISMM trial investigators, Abildgaard N, Adler H, Altuntas F, Akay OM, Amin B, Anagnostopoulos A, Anderson L, Anttila P, Araujo C, Arce-Lara C, Aydin Y, Basu S, Battini R, Beeker T, Benboubker L, Ben-Yehuda D, Bladé J, Blau IW, Boccia R, Burke L, Byeff P, Cascavilla N, Cavo M, Chantry A, Charles Y, Chaudhry A, Corso A, Coyne M, De Arriba F, Delimpasi S, Desjardins P, Dhakal B, Di Bartolomeo P, Di Raimondo F, Dürig J, Engelhardt M, Escoffre-Barbe M, Esteves G, Flogegard M, Gabrail N, Gamberi B, Garrison M, Gay J, Gisslinger H, Goldschmidt H, Goncalves C, Gressot L, Grosicki S, Hanna W, Hayden P, Henriques Bernardo MM, Hermann R, Holden V, Honkalehto K, Huben M, Huffman J, Hunter H, Hus M, Jagasia M, Jagganath S, Janakiram M, Jaiyesimi I, Jenner M, João C, Johnson P, Jurcyszyn A, Kalayoğlu Beşişik S, Kambhampati S, Kanate A, Karadoğan I, Khojasteh A, Kirkel D, Komarnicki M, Krauth MT, Kuriakose P, Larocca A, Lauri B, Leleu X, Lucio P, Luppi M, Mangiacavalli S, Mariette C, Matsue K, Mellqvist UH, Mendeleeva L, Meshad M, Miller C, Mohrbacher A, Moreau P, Morelli AM, Müldür E, Naassan A, Nahi H, Nair R, O'Dwyer M, Öngören Aydin S, Openshaw T, O'Rourke T, Osswald M, Overton L, Pati A, Pavic M, Pegourie B, Pehlivan M, Pierola AA, Plesner T, Pluta A, Rabin N, Ramasamy K, Rambaldi A, Rodriguez P, Röllig C, Rosenblatt J, Rosenbluth J, Salomo M, Samoylova O, Sastre Moral J, Sati H, Selleri C, Shafeek S, Shinagawa A, Sleckman B, Smith C, Sonmez M, Stone C, Streetly M, Suzuki K, Taetle R, Tafuri A, Takezako N, Teke HÜ, Vapaatalo M, Vassilopoulos G, Verma A, Vidito S, Viterbo L, Vural F, Wang XS, Yağci M, Yee A., Richardson, Paul G, Oriol, Albert, Beksac, Meral, Liberati, Anna Marina, Galli, Monica, Schjesvold, Fredrik, Lindsay, Jindriska, Weisel, Katja, White, Darrell, Facon, Thierry, San Miguel, Jesu, Sunami, Kazutaka, O'Gorman, Peter, Sonneveld, Pieter, Robak, Pawel, Semochkin, Sergey, Schey, Steve, Yu, Xin, Doerr, Thoma, Bensmaine, Amine, Biyukov, Tsvetan, Peluso, Teresa, Zaki, Mohamed, Anderson, Kenneth, Dimopoulos, Meletio, OPTIMISMM trial investigator, Abildgaard N, Adler H, Altuntas F, Akay OM, Amin B, Anagnostopoulos A, Anderson L, Anttila P, Araujo C, Arce-Lara C, Aydin Y, Basu S, Battini R, Beeker T, Benboubker L, Ben-Yehuda D, Bladé J, Blau IW, Boccia R, Burke L, Byeff P, Cascavilla N, Cavo M, Chantry A, Charles Y, Chaudhry A, Corso A, Coyne M, De Arriba F, Delimpasi S, Desjardins P, Dhakal B, Di Bartolomeo P, Di Raimondo F, Dürig J, Engelhardt M, Escoffre-Barbe M, Esteves G, Flogegard M, Gabrail N, Gamberi B, Garrison M, Gay J, Gisslinger H, Goldschmidt H, Goncalves C, Gressot L, Grosicki S, Hanna W, Hayden P, Henriques Bernardo MM, Hermann R, Holden V, Honkalehto K, Huben M, Huffman J, Hunter H, Hus M, Jagasia M, Jagganath S, Janakiram M, Jaiyesimi I, Jenner M, João C, Johnson P, Jurcyszyn A, Kalayoğlu Beşişik S, Kambhampati S, Kanate A, Karadoğan I, Khojasteh A, Kirkel D, Komarnicki M, Krauth MT, Kuriakose P, Larocca A, Lauri B, Leleu X, Lucio P, Luppi M, Mangiacavalli S, Mariette C, Matsue K, Mellqvist UH, Mendeleeva L, Meshad M, Miller C, Mohrbacher A, Moreau P, Morelli AM, Müldür E, Naassan A, Nahi H, Nair R, O'Dwyer M, Öngören Aydin S, Openshaw T, O'Rourke T, Osswald M, Overton L, Pati A, Pavic M, Pegourie B, Pehlivan M, Pierola AA, Plesner T, Pluta A, Rabin N, Ramasamy K, Rambaldi A, Rodriguez P, Röllig C, Rosenblatt J, Rosenbluth J, Salomo M, Samoylova O, Sastre Moral J, Sati H, Selleri C, Shafeek S, Shinagawa A, Sleckman B, Smith C, Sonmez M, Stone C, Streetly M, Suzuki K, Taetle R, Tafuri A, Takezako N, Teke HÜ, Vapaatalo M, Vassilopoulos G, Verma A, Vidito S, Viterbo L, Vural F, Wang XS, Yağci M, Yee A., and Hematology
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Oncology ,Adult ,Male ,medicine.medical_specialty ,Adolescent ,Population ,Dexamethasone ,Bortezomib ,03 medical and health sciences ,Young Adult ,0302 clinical medicine ,Internal medicine ,hemic and lymphatic diseases ,Antineoplastic Combined Chemotherapy Protocols ,medicine ,Humans ,education ,Survival rate ,Lenalidomide ,Multiple myeloma ,Aged ,Salvage Therapy ,education.field_of_study ,business.industry ,Pomalidomide, bortezomib, dexamethasone ,Middle Aged ,Pomalidomide ,medicine.disease ,Prognosis ,Thalidomide ,Survival Rate ,Regimen ,Drug Resistance, Neoplasm ,030220 oncology & carcinogenesis ,Female ,Neoplasm Recurrence, Local ,business ,Multiple Myeloma ,030215 immunology ,medicine.drug ,Follow-Up Studies - Abstract
Background As lenalidomide becomes increasingly established for upfront treatment of multiple myeloma, patients refractory to this drug represent a population with an unmet need. The combination of pomalidomide, bortezomib, and dexamethasone has shown promising results in phase 1/2 trials of patients with relapsed or refractory multiple myeloma. We aimed to assess the efficacy and safety of this triplet regimen in patients with relapsed or refractory multiple myeloma who previously received lenalidomide. Methods We did a randomised, open-label, phase 3 trial at 133 hospitals and research centres in 21 countries. We enrolled patients (aged >= 18 years) with a diagnosis of multiple myeloma and measurable disease, an Eastern Cooperative Oncology Group performance status of 0-2, who received one to three previous regimens, including a lenalidomide-containing regimen for at least two consecutive cycles. We randomly assigned patients (1:1) to bortezomib and dexamethasone with or without pomalidomide using a permutated blocked design in blocks of four, stratified according to age, number of previous regimens, and concentration of beta(2) microglobulin at screening. Bortezomib (1.3 mg/m(2)) was administered intravenously until protocol amendment 1 then either intravenously or subcutaneously on days 1,4, 8, and 11 for the first eight cycles and subsequently on days 1 and 8. Dexamethasone (20 mg [10 mg if age >75 years]) was administered orally on the same days as bortezomib and the day after. Patients allocated pomalidomide received 4 mg orally on days 1-14. Treatment cycles were every 21 days. The primary endpoint was progression-free survival in the intention-to-treat population, as assessed by an independent review committee. Safety was assessed in all patients who received at least one dose of study medication. This trial is registered at ClinicalTrials.gov, number NCT01734928; patients are no longer being enrolled. Findings Between Jan 7, 2013, and May 15,2017,559 patients were enrolled. 281 patients were assigned pomalidomide, bortezomib, and dexamethasone and 278 were allocated bortezomib and dexamethasone. Median follow-up was 15.9 months (IQR 9.9-21.7). Pomalidomide, bortezomib, and dexamethasone significantly improved progression-free survival compared with bortezomib and dexamethasone (median 11.20 months [95% CI 9.66-13-73] vs 7.10 months [5.88-8-48]; hazard ratio 0.61, 95% CI 0.49-0-77; p
- Published
- 2019
3. Pomalidomide, bortezomib, and dexamethasone for patients with relapsed or refractory multiple myeloma previously treated with lenalidomide (OPTIMISMM): a randomised, open-label, phase 3 trial
- Author
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Richardson, P.G. Oriol, A. Beksac, M. Liberati, A.M. Galli, M. Schjesvold, F. Lindsay, J. Weisel, K. White, D. Facon, T. San Miguel, J. Sunami, K. O'Gorman, P. Sonneveld, P. Robak, P. Semochkin, S. Schey, S. Yu, X. Doerr, T. Bensmaine, A. Biyukov, T. Peluso, T. Zaki, M. Anderson, K. Dimopoulos, M. Abildgaard, N. Adler, H. Altuntas, F. Akay, O.M. Amin, B. Anagnostopoulos, A. Anderson, L. Anttila, P. Araujo, C. Arce-Lara, C. Aydin, Y. Basu, S. Battini, R. Beeker, T. Benboubker, L. Ben-Yehuda, D. Bladé, J. Blau, I.W. Boccia, R. Burke, L. Byeff, P. Cascavilla, N. Cavo, M. Chantry, A. Charles, Y. Chaudhry, A. Corso, A. Coyne, M. De Arriba, F. Delimpasi, S. Desjardins, P. Dhakal, B. Di Bartolomeo, P. Di Raimondo, F. Dürig, J. Engelhardt, M. Escoffre-Barbe, M. Esteves, G. Flogegard, M. Gabrail, N. Gamberi, B. Garrison, M. Gay, J. Gisslinger, H. Goldschmidt, H. Goncalves, C. Gressot, L. Grosicki, S. Hanna, W. Hayden, P. Henriques Bernardo, M.M. Hermann, R. Holden, V. Honkalehto, K. Huben, M. Huffman, J. Hunter, H. Hus, M. Jagasia, M. Jagganath, S. Janakiram, M. Jaiyesimi, I. Jenner, M. João, C. Johnson, P. Jurcyszyn, A. Kalayoğlu Beşişik, S. Kambhampati, S. Kanate, A. Karadoğan, I. Khojasteh, A. Kirkel, D. Komarnicki, M. Krauth, M.-T. Kuriakose, P. Larocca, A. Lauri, B. Leleu, X. Lucio, P. Luppi, M. Mangiacavalli, S. Mariette, C. Matsue, K. Mellqvist, U.-H. Mendeleeva, L. Meshad, M. Miller, C. Mohrbacher, A. Moreau, P. Morelli, A.M. Müldür, E. Naassan, A. Nahi, H. Nair, R. O'Dwyer, M. Öngören Aydin, S. Openshaw, T. O'Rourke, T. Osswald, M. Overton, L. Pati, A. Pavic, M. Pegourie, B. Pehlivan, M. Pierola, A.A. Plesner, T. Pluta, A. Rabin, N. Ramasamy, K. Rambaldi, A. Rodriguez, P. Röllig, C. Rosenblatt, J. Rosenbluth, J. Salomo, M. Samoylova, O. Sastre Moral, J. Sati, H. Selleri, C. Shafeek, S. Shinagawa, A. Sleckman, B. Smith, C. Sonmez, M. Stone, C. Streetly, M. Suzuki, K. Taetle, R. Tafuri, A. Takezako, N. Teke, H.Ü. Vapaatalo, M. Vassilopoulos, G. Verma, A. Vidito, S. Viterbo, L. Vural, F. Wang, X.S. Yağci, M. Yee, A. OPTIMISMM trial investigators
- Subjects
hemic and lymphatic diseases - Abstract
Background: As lenalidomide becomes increasingly established for upfront treatment of multiple myeloma, patients refractory to this drug represent a population with an unmet need. The combination of pomalidomide, bortezomib, and dexamethasone has shown promising results in phase 1/2 trials of patients with relapsed or refractory multiple myeloma. We aimed to assess the efficacy and safety of this triplet regimen in patients with relapsed or refractory multiple myeloma who previously received lenalidomide. Methods: We did a randomised, open-label, phase 3 trial at 133 hospitals and research centres in 21 countries. We enrolled patients (aged ≥18 years)with a diagnosis of multiple myeloma and measurable disease, an Eastern Cooperative Oncology Group performance status of 0–2, who received one to three previous regimens, including a lenalidomide-containing regimen for at least two consecutive cycles. We randomly assigned patients (1:1)to bortezomib and dexamethasone with or without pomalidomide using a permutated blocked design in blocks of four, stratified according to age, number of previous regimens, and concentration of β2 microglobulin at screening. Bortezomib (1·3 mg/m2)was administered intravenously until protocol amendment 1 then either intravenously or subcutaneously on days 1, 4, 8, and 11 for the first eight cycles and subsequently on days 1 and 8. Dexamethasone (20 mg [10 mg if age >75 years])was administered orally on the same days as bortezomib and the day after. Patients allocated pomalidomide received 4 mg orally on days 1–14. Treatment cycles were every 21 days. The primary endpoint was progression-free survival in the intention-to-treat population, as assessed by an independent review committee. Safety was assessed in all patients who received at least one dose of study medication. This trial is registered at ClinicalTrials.gov, number NCT01734928; patients are no longer being enrolled. Findings: Between Jan 7, 2013, and May 15, 2017, 559 patients were enrolled. 281 patients were assigned pomalidomide, bortezomib, and dexamethasone and 278 were allocated bortezomib and dexamethasone. Median follow-up was 15·9 months (IQR 9·9–21·7). Pomalidomide, bortezomib, and dexamethasone significantly improved progression-free survival compared with bortezomib and dexamethasone (median 11·20 months [95% CI 9·66–13·73]vs 7·10 months [5·88–8·48]; hazard ratio 0·61, 95% CI 0·49–0·77; p
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- 2019
4. Recombinant protein expression in aDrosophila cell line: comparison with the baculovirus system
- Author
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Bernard, A. R., Kost, T. A., Overton, L., Cavegn, C., Young, J., Bertrand, M., Yahia-Cherif, Z., Chabert, C., and Mills, A.
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- 1994
- Full Text
- View/download PDF
5. Pembrolizumab plus lenalidomide and dexamethasone for patients with treatment-naive multiple myeloma (KEYNOTE-185): a randomised, open-label, phase 3 trial.
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Usmani S.Z., Oriol A., Karlin L., Cavo M., Rifkin R.M., Hayek F., Kirschner E., Bharany N., Overton L., Mannem S., Harroff A., Jain S., Roque T., McIntyre K., Yasencha C.K., Houck W., Schjesvold F., Yimer H.A., LeBlanc R., Takezako N., McCroskey R.D., Lim A.B.M., Suzuki K., Kosugi H., Grigoriadis G., Avivi I., Facon T., Jagannath S., Lonial S., Ghori R.U., Farooqui M.Z.H., Marinello P., San-Miguel J., Lim A., Walker T., Nicol A., Reece D., Elemary M., Boudreault Pedneault J.S., Attal M., Weisel K., Engelhardt M., Mackensen A., Quinn J., Cohen A., Magen-Nativ H., Benyamini N., Larocca A., Matsumoto M., Iida S., Ishikawa T., Kondo Y., Sunami K., Ando K., Teshima T., Chou T., Iwasaki H., Miki H., Matsumura I., Onishi Y., Izutsu K., Kizaki M., George A., Blacklock H., Simpson D., Waage A., Samoilova O., Nikitin E., Chagorova T., McDonald A., Patel M., Oriol Rocafiguera A., San Miguel Izquierdo J., Mateos M., Streetly M., Forsyth P., Jackson G., Jenkins S., Rifkin R., Yimer H., McCroskey R., Martincic D., Tarantolo S., Larson S., Faroun Y., Vaughn J., Baz R., Saylors G., Neppalli A., Raptis A., Fung H., Janosky M., Stevens D., Coleman M., Costa D., Cross S., Fanning S., Berges D.F., Harris T., Zackon I., Atanackovic D., Lee K., Oliff I., Lee W., Bensinger W., Lutzky J., Baron A., Usmani S.Z., Oriol A., Karlin L., Cavo M., Rifkin R.M., Hayek F., Kirschner E., Bharany N., Overton L., Mannem S., Harroff A., Jain S., Roque T., McIntyre K., Yasencha C.K., Houck W., Schjesvold F., Yimer H.A., LeBlanc R., Takezako N., McCroskey R.D., Lim A.B.M., Suzuki K., Kosugi H., Grigoriadis G., Avivi I., Facon T., Jagannath S., Lonial S., Ghori R.U., Farooqui M.Z.H., Marinello P., San-Miguel J., Lim A., Walker T., Nicol A., Reece D., Elemary M., Boudreault Pedneault J.S., Attal M., Weisel K., Engelhardt M., Mackensen A., Quinn J., Cohen A., Magen-Nativ H., Benyamini N., Larocca A., Matsumoto M., Iida S., Ishikawa T., Kondo Y., Sunami K., Ando K., Teshima T., Chou T., Iwasaki H., Miki H., Matsumura I., Onishi Y., Izutsu K., Kizaki M., George A., Blacklock H., Simpson D., Waage A., Samoilova O., Nikitin E., Chagorova T., McDonald A., Patel M., Oriol Rocafiguera A., San Miguel Izquierdo J., Mateos M., Streetly M., Forsyth P., Jackson G., Jenkins S., Rifkin R., Yimer H., McCroskey R., Martincic D., Tarantolo S., Larson S., Faroun Y., Vaughn J., Baz R., Saylors G., Neppalli A., Raptis A., Fung H., Janosky M., Stevens D., Coleman M., Costa D., Cross S., Fanning S., Berges D.F., Harris T., Zackon I., Atanackovic D., Lee K., Oliff I., Lee W., Bensinger W., Lutzky J., and Baron A.
- Abstract
Background: Lenalidomide and dexamethasone has been a standard of care in transplant-ineligible patients with newly diagnosed multiple myeloma. The addition of a third drug to the combination is likely to improve treatment efficacy. KEYNOTE-185 assessed the efficacy and safety of lenalidomide and dexamethasone with and without pembrolizumab in patients with previously untreated multiple myeloma. Here, we present the results of an unplanned interim analysis done to assess the benefit-risk of the combination at the request of the US Food and Drug Administration (FDA). Method(s): KEYNOTE-185 was a randomised, open-label, phase 3 trial done at 95 medical centres across 15 countries (Australia, Canada, France, Germany, Ireland, Israel, Italy, Japan, New Zealand, Norway, Russia, South Africa, Spain, UK, and USA). Transplantation-ineligible patients aged 18 years and older with newly diagnosed multiple myeloma, Eastern Cooperative Oncology Group performance status of 0 or 1, and who were treatment naive were enrolled, and randomly assigned 1:1 to receive either pembrolizumab plus lenalidomide and dexamethasone or lenalidomide and dexamethasone alone using an interactive voice or integrated web response system. Patients received oral lenalidomide 25 mg on days 1-21 and oral dexamethasone 40 mg on days 1, 8, 15, and 22 of repeated 28-day cycles, with or without intravenous pembrolizumab 200 mg every 3 weeks. The primary endpoint was progression-free survival, which was investigator-assessed because of early trial termination. Efficacy was analysed in all randomly assigned patients and safety was analysed in all patients who received at least one dose of study drug. This trial is registered at ClinicalTrials.gov, number NCT02579863, and it is closed for accrual. Finding(s): Between Jan 7, 2016, and June 9, 2017, 301 patients were randomly assigned to the pembrolizumab plus lenalidomide and dexamethasone group (n=151) or the lenalidomide and dexamethasone group (n=150). On Jul
- Published
- 2019
6. Recombinant protein expression in a Drosophila cell line: comparison with the baculovirus system
- Author
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Bernard, A. R., primary, Kost, T. A., additional, Overton, L., additional, Cavegn, C., additional, Young, J., additional, Bertrand, M., additional, Yahia-Cherif, Z., additional, Chabert, C., additional, and Mills, A., additional
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- 1994
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7. Funding retirement: The role of equity release (draft version)
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Overton, L. (author) and Overton, L. (author)
- Abstract
Across Europe there has been much debate about the potential of housing assets to ease the fiscal pressures inherent in current pension systems. However little is known about what people do with housing equity once it is released. Do they extract equity to supplement inadequate pension provision or do they use it for conspicuous consumption? And are they reluctant to use housing equity anyway because of a desire to leave it as a bequest? While the scale of equity release currently varies greatly across Europe, it has been largest by far in the UK. This paper presents the findings of a survey of over 500 equity release customers in the UK as a case study of the actual usage of equity release. It reveals that equity release products are not just used to supplement low pension income, but rather, a significant proportion are also using these products because they want a more enjoyable retirement while others appear to have early bequest motives. Consideration is given to what these findings might mean for the house as pension debate in other European countries.
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- 2010
8. Rosai Dorfman Disease presenting as unilateral chronic parotitis
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Byrd, J., primary, Overton, L., additional, Goldin, T., additional, and Lentsch, E., additional
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- 2012
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9. Inactivation of N-methyl-N′-nitro-N-nitrosoguanidine in the Ames Salmonella/microsome test
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Gentile, J. M., Overton, L. K., and Schubert, J.
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- 1978
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10. USE OF FERROELECTRICS FOR GAMMA-RAY DOSIMETRY
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Hester, D L, primary, Glower, D D, additional, and Overton, L J, additional
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- 1964
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11. High level expression of mammalian protein farnesyltransferase in a baculovirus system. The purified protein contains zinc
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Chen, W.J., primary, Moomaw, J.F., additional, Overton, L., additional, Kost, T.A., additional, and Casey, P.J., additional
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- 1993
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12. Human immunodeficiency virus infection and syncytium formation in HeLa cells expressing glycophospholipid-anchored CD4
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Kost, T A, primary, Kessler, J A, additional, Patel, I R, additional, Gray, J G, additional, Overton, L K, additional, and Carter, S G, additional
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- 1991
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13. Use of Ferroelectrics for Gamma-Ray Dosimetry.
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Hester, D. L., Glower, D. D., and Overton, L. J.
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- 1964
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14. Mouse BAC ends quality assessment and sequence analyses.
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Zhao, S, Shatsman, S, Ayodeji, B, Geer, K, Tsegaye, G, Krol, M, Gebregeorgis, E, Shvartsbeyn, A, Russell, D, Overton, L, Jiang, L, Dimitrov, G, Tran, K, Shetty, J, Malek, J A, Feldblyum, T, Nierman, W C, and Fraser, C M
- Abstract
A large-scale BAC end-sequencing project at The Institute for Genomic Research (TIGR) has generated one of the most extensive sets of sequence markers for the mouse genome to date. With a sequencing success rate of >80%, an average read length of 485 bp, and ABI3700 capillary sequencers, we have generated 449,234 nonredundant mouse BAC end sequences (mBESs) with 218 Mb total from 257,318 clones from libraries RPCI-23 and RPCI-24, representing 15x clone coverage, 7% sequence coverage, and a marker every 7 kb across the genome. A total of 191,916 BACs have sequences from both ends providing 12x genome coverage. The average Q20 length is 406 bp and 84% of the bases have phred quality scores > or = 20. RPCI-24 mBESs have more Q20 bases and longer reads on average than RPCI-23 sequences. ABI3700 sequencers and the sample tracking system ensure that > 95% of mBESs are associated with the right clone identifiers. We have found that a significant fraction of mBESs contains L1 repeats and approximately 48% of the clones have both ends with > or = 100 bp contiguous unique Q20 bases. About 3% mBESs match ESTs and > 70% of matches were conserved between the mouse and the human or the rat. Approximately 0.1% mBESs contain STSs. About 0.2% mBESs match human finished sequences and > 70% of these sequences have EST hits. The analyses indicate that our high-quality mouse BAC end sequences will be a valuable resource to the community.
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- 2001
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15. Production of a urokinase plasminogen activator-IgG fusion protein (uPA-IgG) in the baculovirus expression system
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Kost, T. A., Ignar, D. M., Clay, W. C., Andrews, J., Leray, J. D., Overton, L., Hoffman, C. R., Kilpatrick, K. E., Ellis, B., and Emerson, D. L.
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- 1997
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16. In-depth view of structure, activity, and evolution of rice chromosome 10
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Yu, Ys, Rambo, T., Currie, J., Saski, C., Kim, Hr, Collura, K., Thompson, S., Simmons, J., Yang, Tj, Nah, G., Patel, Aj, Thurmond, S., Henry, D., Oates, R., Palmer, M., Pries, G., Gibson, J., Anderson, H., Paradkar, M., Crane, L., Dale, J., Carver, Mb, Wood, T., Frisch, D., Engler, F., Soderlund, C., Palmer, Le, Tetylman, L., Nascimento, L., La Bastide, M., Spiegel, L., Ware, D., O Shaughnessy, A., Dike, S., Dedhia, N., Preston, R., Huang, E., Ferraro, K., Kuit, K., Miller, B., Zutavern, T., Katzenberger, F., Muller, S., Balija, V., Martienssen, Ra, Stein, L., Minx, P., Johnson, D., Cordum, H., Mardis, E., Cheng, Zk, Jiang, Jm, Wilson, R., Mccombie, Wr, Wing, Ra, Yuan, Qp, Shu, Oy, Liu, J., Jones, Km, Gansberger, K., Moffat, K., Hill, J., Tsitrin, T., Overton, L., Bera, J., Kim, M., Jin, Sh, Tallon, L., Ciecko, A., Pai, G., Aken, S., Utterback, T., Reidmuller, S., Bormann, J., Feldblyum, T., Hsiao, J., Zismann, V., Blunt, S., Vazeilles, A., Shaffer, T., Koo, H., Suh, B., Yang, Q., Haas, B., Peterson, J., Pertea, M., Volfovsky, N., Wortman, J., White, O., Salzberg, Sl, Fraser, Cm, Buell, Cr, Messing, J., Rentao Song, Fuks, G., Llaca, V., Kovchak, S., Young, S., Bowers, Je, Paterson, Ah, Johns, Ma, Mao, L., Pan, Hq, Dean, Ra, and Rice Chromosome 10 Sequencing Cons
17. FERROELECTRIC TRANSITION FE1‐FE2 IN Pb(Zr0.65Ti0.35)O3 CONTAINING 1 wt % Nb2O5
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Glower, D. D., primary and Overton, L. J., additional
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- 1963
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18. THE LABORATORY.
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Overton, L. E.
- Abstract
The article discusses the factors in producing good optical prints for theater production. Flutter arising in printing is kept to a minimum by using short-pitched low shrinkage negative and correct tooth root diameter of the printer sprocket. Processing must be carefully controlled, and printing must be spread over a number of machines. The only way to optimum print density is to make a series of print tests at different densities and select one that gives satisfactory quality.
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- 1961
19. FERROELECTRIC TRANSITION FE1‐FE2IN Pb(Zr0.65Ti0.35)O3CONTAINING 1 wt % Nb2O5
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Glower, D. D. and Overton, L. J.
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- 1963
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20. Pembrolizumab plus lenalidomide and dexamethasone for patients with treatment-naive multiple myeloma (KEYNOTE-185): a randomised, open-label, phase 3 trial
- Author
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Saad Zafar Usmani, Fredrik Schjesvold, Albert Oriol, Lionel Karlin, Michele Cavo, Robert M Rifkin, Habte Aragaw Yimer, Richard LeBlanc, Naoki Takezako, Robert Donald McCroskey, Andrew Boon Ming Lim, Kenshi Suzuki, Hiroshi Kosugi, George Grigoriadis, Irit Avivi, Thierry Facon, Sundar Jagannath, Sagar Lonial, Razi Uddin Ghori, Mohammed Z H Farooqui, Patricia Marinello, Jesus San-Miguel, Andrew Lim, Trish Walker, Andrew Nicol, Donna Reece, Mohamed Elemary, Jean Samuel Boudreault Pedneault, Michel Attal, Katja Weisel, Monika Engelhardt, Andreas Mackensen, John Quinn, Amos Cohen, Hila Magen-Nativ, Noam Benyamini, Alessandra Larocca, Morio Matsumoto, Shinsuke Iida, Takayuki Ishikawa, Yukio Kondo, Kazutaka Sunami, Kiyoshi Ando, Takanori Teshima, Takaaki Chou, Hiromi Iwasaki, Hirokazu Miki, Itaru Matsumura, Yasushi Onishi, Koji Izutsu, Masahiro Kizaki, Anupkumar George, Hillary Blacklock, David Simpson, Anders Waage, Olga Samoilova, Evgeniy Nikitin, Tatiana Chagorova, Andrew McDonald, Moosa Patel, Albert Oriol Rocafiguera, Jesus San Miguel Izquierdo, Maria Mateos, Matthew Streetly, Peter Forsyth, Graham Jackson, Stephen Jenkins, Robert Rifkin, Habte Yimer, Robert McCroskey, Danko Martincic, Stefano Tarantolo, Sarah Larson, Yacoub Faroun, Jennifer Vaughn, Rachid Baz, Gene Saylors, Amarendra Neppalli, Anastasios Raptis, Henry Fung, Maxwell Janosky, Don Stevens, Morton Coleman, Dennis Costa, Scott Cross, Suzanne Fanning, Daniel Farray Berges, Thomas Harris, Ira Zackon, Djordje Atanackovic, Kelvin Lee, Ira Oliff, Wes Lee, William Bensinger, Jose Lutzky, Ari Baron, Fadi Hayek, Eli Kirschner, Neeraj Bharany, Lindsay Overton, Siva Mannem, Allyson Harroff, Sharad Jain, Tammy Roque, Kristi McIntyre, Christopher K Yasencha, William Houck, Usmani SZ1, Schjesvold F2, Oriol A3, Karlin L4, Cavo M5, Rifkin RM6, Yimer HA7, LeBlanc R8, Takezako N9, McCroskey RD10, Lim ABM11, Suzuki K12, Kosugi H13, Grigoriadis G14, Avivi I15, Facon T16, Jagannath S17, Lonial S18, Ghori RU19, Farooqui MZH19, Marinello P19, San-Miguel J20, and KEYNOTE-185 Investigators. Lim A, Grigoriadis G, Walker T, Nicol A, LeBlanc R, Reece D, Elemary M, Boudreault Pedneault JS, Karlin L, Facon T, Attal M, Weisel K, Engelhardt M, Mackensen A, Quinn J, Avivi I, Cohen A, Magen-Nativ H, Benyamini N, Cavo M, Larocca A, Takezako N, Suzuki K, Kosugi H, Matsumoto M, Iida S, Ishikawa T, Kondo Y, Sunami K, Ando K, Teshima T, Chou T, Iwasaki H, Miki H, Matsumura I, Onishi Y, Izutsu K, Kizaki M, George A, Blacklock H, Simpson D, Schjesvold F, Waage A, Samoilova O, Nikitin E, Chagorova T, McDonald A, Patel M, Oriol Rocafiguera A, San Miguel Izquierdo J, Mateos M, Streetly M, Forsyth P, Jackson G, Jenkins S, Rifkin R, Yimer H, McCroskey R, Martincic D, Tarantolo S, Larson S, Faroun Y, Vaughn J, Baz R, Saylors G, Neppalli A, Raptis A, Fung H, Janosky M, Stevens D, Coleman M, Costa D, Cross S, Fanning S, Berges DF, Harris T, Zackon I, Atanackovic D, Lee K, Oliff I, Lee W, Bensinger W, Lutzky J, Baron A, Hayek F, Kirschner E, Bharany N, Overton L, Mannem S, Harroff A, Jain S, Roque T, McIntyre K, Yasencha CK, Houck W.
- Subjects
Male ,medicine.medical_specialty ,Pembrolizumab ,Antibodies, Monoclonal, Humanized ,Dexamethasone ,law.invention ,03 medical and health sciences ,0302 clinical medicine ,Randomized controlled trial ,law ,Internal medicine ,Antineoplastic Combined Chemotherapy Protocols ,Humans ,Medicine ,Progression-free survival ,Lenalidomide ,Survival rate ,Multiple myeloma ,Aged ,Proportional Hazards Models ,Aged, 80 and over ,business.industry ,Pneumonia ,Hematology ,Interim analysis ,medicine.disease ,Progression-Free Survival ,Survival Rate ,Treatment Outcome ,030220 oncology & carcinogenesis ,Female ,Multiple Myeloma ,business ,Pembrolizumab, lenalidomide, dexamethasone ,030215 immunology ,medicine.drug - Abstract
Summary Background Lenalidomide and dexamethasone has been a standard of care in transplant-ineligible patients with newly diagnosed multiple myeloma. The addition of a third drug to the combination is likely to improve treatment efficacy. KEYNOTE-185 assessed the efficacy and safety of lenalidomide and dexamethasone with and without pembrolizumab in patients with previously untreated multiple myeloma. Here, we present the results of an unplanned interim analysis done to assess the benefit–risk of the combination at the request of the US Food and Drug Administration (FDA). Methods KEYNOTE-185 was a randomised, open-label, phase 3 trial done at 95 medical centres across 15 countries (Australia, Canada, France, Germany, Ireland, Israel, Italy, Japan, New Zealand, Norway, Russia, South Africa, Spain, UK, and USA). Transplantation-ineligible patients aged 18 years and older with newly diagnosed multiple myeloma, Eastern Cooperative Oncology Group performance status of 0 or 1, and who were treatment naive were enrolled, and randomly assigned 1:1 to receive either pembrolizumab plus lenalidomide and dexamethasone or lenalidomide and dexamethasone alone using an interactive voice or integrated web response system. Patients received oral lenalidomide 25 mg on days 1–21 and oral dexamethasone 40 mg on days 1, 8, 15, and 22 of repeated 28-day cycles, with or without intravenous pembrolizumab 200 mg every 3 weeks. The primary endpoint was progression-free survival, which was investigator-assessed because of early trial termination. Efficacy was analysed in all randomly assigned patients and safety was analysed in all patients who received at least one dose of study drug. This trial is registered at ClinicalTrials.gov , number NCT02579863 , and it is closed for accrual. Findings Between Jan 7, 2016, and June 9, 2017, 301 patients were randomly assigned to the pembrolizumab plus lenalidomide and dexamethasone group (n=151) or the lenalidomide and dexamethasone group (n=150). On July 3, 2017, the FDA decided to halt the study because of the imbalance in the proportion of death between groups. At database cutoff (June 2, 2017), with a median follow-up of 6·6 months (IQR 3·4–9·6), 149 patients in the pembrolizumab plus lenalidomide and dexamethasone group and 145 in the lenalidomide and dexamethasone group had received their assigned study drug. Median progression-free survival was not reached in either group; progression-free survival estimates at 6-months were 82·0% (95% CI 73·2–88·1) versus 85·0% (76·8–90·5; hazard ratio [HR] 1·22; 95% CI 0·67–2·22; p=0·75). Serious adverse events were reported in 81 (54%) patients in the pembrolizumab plus lenalidomide and dexamethasone group versus 57 (39%) patients in the lenalidomide and dexamethasone group; the most common serious adverse events were pneumonia (nine [6%]) and pyrexia (seven [5%]) in the pembrolizumab plus lenalidomide and dexamethasone group and pneumonia (eight [6%]) and sepsis (two [1%]) in the lenalidomide and dexamethasone group. Six (4%) treatment-related deaths occurred in the pembrolizumab plus lenalidomide and dexamethasone group (cardiac arrest, cardiac failure, myocarditis, large intestine perforation, pneumonia, and pulmonary embolism) and two (1%) in the lenalidomide and dexamethasone group (upper gastrointestinal haemorrhage and respiratory failure). Interpretation The results from this unplanned, FDA-requested, interim analysis showed that the benefit–risk profile of pembrolizumab plus lenalidomide and dexamethasone is unfavourable for patients with newly diagnosed, previously untreated multiple myeloma. Long-term safety and survival follow-up is ongoing. Funding Merck Sharp & Dohme, a subsidiary of Merck & Co, Inc (Kenilworth, NJ, USA).
- Published
- 2019
21. FERROELECTRIC TRANSITION FE$sub 1$-FE$sub 2$ IN Pb(Zr$sub 0$.$sub 65$)O$sub 3$ CONTAINING 1 wt % Nb$sub 2$O$sub 5$
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Overton, L
- Published
- 1963
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22. Adeno-associated virus genome quantification with amplification-free CRISPR-Cas12a.
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Hetzler Z, Marinakos SM, Lott N, Mohammad N, Lass-Napiorkowska A, Kolbe J, Turrentine L, Fields D, Overton L, Marie H, Hucknall A, Rammo O, George H, and Wei Q
- Subjects
- Humans, Genetic Vectors genetics, HEK293 Cells, Dependovirus genetics, CRISPR-Cas Systems, Genome, Viral
- Abstract
Efficient manufacturing of recombinant Adeno-Associated Viral (rAAV) vectors to meet rising clinical demand remains a major hurdle. One of the most significant challenges is the generation of large amounts of empty capsids without the therapeutic genome. There is no standardized analytical method to accurately quantify the viral genes, and subsequently the empty-to-full ratio, making the manufacturing challenges even more complex. We propose the use of CRISPR diagnostics (CRISPR-Dx) as a robust and rapid approach to determine AAV genome titers. We designed and developed the CRISPR-AAV Evaluation (CRAAVE) assay to maximize sensitivity, minimize time-to-result, and provide a potentially universal design for quantifying multiple transgene constructs encapsidated within different AAV serotypes. We also demonstrate an on-chip CRAAVE assay with lyophilized reagents to minimize end user assay input. The CRAAVE assay was able to detect AAV titers as low as 7e7 vg/mL with high precision (<3% error) in quantifying unknown AAV titers when compared with conventional quantitative PCR (qPCR) method. The assay only requires 30 min of assay time, shortening the analytical workflow drastically. Our results suggest CRISPR-Dx could be a promising tool for efficient rAAV genome titer quantification and has the potential to revolutionize biomanufacturing process analytical technology (PAT)., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)
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- 2024
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23. Development and Delivery of a Hands-On Short Course in Adeno-Associated Virus Manufacturing to Support Growing Workforce Needs in Gene Therapy.
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Overton L, Boi C, Shastry S, Smith-Moore C, Balchunas J, Sambandan D, and Gilleskie G
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- Humans, Commerce, Genetic Vectors genetics, Dependovirus genetics, Genetic Therapy
- Abstract
The manufacturing of gene therapy products is a rapidly growing industry bolstered by the tremendous potential of these therapies to provide lifesaving treatment for rare and complex genetic diseases. The industry's steep rise has resulted in a high demand for skilled staff required to manufacture gene therapy products of the expected high quality. To address this skill shortage, more opportunities for education and training in all aspects of gene therapy manufacturing are needed. The Biomanufacturing Training and Education Center (BTEC) at the North Carolina State University (NC State) has developed and delivered (and continues to deliver) a 4-day, hands-on course titled Hands-on cGMP Biomanufacturing of Vectors for Gene Therapy. The course, which consists of 60% hands-on laboratory activities and 40% lectures, aims to provide a comprehensive understanding of the gene therapy production process, from vial thaw through the final formulation step, and analytical testing. This article discusses the design of the course, the backgrounds of the nearly 80 students who have participated in the seven offerings held since March 2019, and feedback from the course participants.
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- 2023
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24. Flexible sensor patch for continuous carbon dioxide monitoring.
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Hetzler Z, Wang Y, Krafft D, Jamalzadegan S, Overton L, Kudenov MW, Ligler FS, and Wei Q
- Abstract
Monitoring and measurement of carbon dioxide (CO
2 ) is critical for many fields. The gold standard CO2 sensor, the Severinghaus electrode, has remained unchanged for decades. In recent years, many other CO2 sensor formats, such as detection based upon pH-sensitive dyes, have been demonstrated, opening the door for relatively simple optical detection schemes. However, a majority of these optochemical sensors require complex sensor preparation steps and are difficult to control and repeatably execute. Here, we report a facile CO2 sensor generation method that suffers from none of the typical fabrication issues. The method described here utilizes polydimethylsiloxane (PDMS) as the flexible sensor matrix and 1-hydroxypyrene-3,6,8-trisulfonate (HPTS), a pH-sensitive dye, as the sensing material. HPTS, a base (NaOH), and glycerol are loaded as dense droplets into a thin PDMS layer which is subsequently cured around the droplet. The fabrication process does not require prior knowledge in chemistry or device fabrication and can be completed as quickly as PDMS cures (∼2 h). We demonstrate the application of this thin-patch sensor for in-line CO2 quantification in cell culture media. To this end, we optimized the sensing composition and quantified CO2 in the range of 0-20 kPa. A standard curve was generated with high fidelity ( R2 = 0.998) along with an analytical resolution of 0.5 kPa (3.7 mm Hg). Additionally, the sensor is fully autoclavable for applications requiring sterility and has a long working lifetime. This flexible, simple-to-manufacture sensor has a myriad of potential applications and represents a new, straightforward means for optical carbon dioxide measurement., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Hetzler, Wang, Krafft, Jamalzadegan, Overton, Kudenov, Ligler and Wei.)- Published
- 2022
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25. An Integrated Model to Accelerate Patient Satisfaction Improvement.
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Delisle DR, Dajani J, and Overton L
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- Hospitals, Community, Humans, Leadership, Patient Satisfaction
- Abstract
Patient satisfaction is gaining traction in the strategic direction and daily operations of hospital executives. The financial penalty/incentive tied to patient satisfaction scores creates a burning platform to accelerate progress. Previous studies have shown the effectiveness of various improvement strategies including leadership rounding and employee training, among others. There has not been a study utilizing an integrated model that incorporates known best practices into a holistic approach. The integrated model included service excellence training, nursing unit-specific action plans, and weekly leadership rounding. Implementation of the model led to significant and sustainable improvements in patient satisfaction in the community hospital setting. This approach can be leveraged and scaled in other organizations to accelerate the pace of change., (Copyright © 2021 The Authors. Published by Wolters Kluwer Health, Inc. All rights reserved.)
- Published
- 2021
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26. Immune checkpoint inhibitors in advanced nasopharyngeal carcinoma: Beyond an era of chemoradiation?
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Masterson L, Howard J, Gonzalez-Cruz J, Jackson C, Barnett C, Overton L, Liu H, Ladwa R, Simpson F, McGrath M, Wallwork B, Jones T, Ottensmeier C, Chua MLK, Perry C, Khanna R, Panizza B, Porceddu S, and Lechner M
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- Antibodies, Monoclonal, Humanized administration & dosage, Antineoplastic Combined Chemotherapy Protocols therapeutic use, B7-H1 Antigen immunology, CTLA-4 Antigen immunology, Chemoradiotherapy, Clinical Trials, Phase I as Topic, Clinical Trials, Phase II as Topic, Clinical Trials, Phase III as Topic, Humans, Nasopharyngeal Carcinoma immunology, Nasopharyngeal Neoplasms immunology, Nivolumab administration & dosage, Nivolumab therapeutic use, Programmed Cell Death 1 Receptor immunology, Antibodies, Monoclonal, Humanized therapeutic use, B7-H1 Antigen antagonists & inhibitors, CTLA-4 Antigen antagonists & inhibitors, Nasopharyngeal Carcinoma drug therapy, Nasopharyngeal Neoplasms drug therapy, Programmed Cell Death 1 Receptor antagonists & inhibitors
- Abstract
Now is an exciting era of development in immunotherapy checkpoint inhibitors and their effect on the treatment of NPC. While the general prognosis of R/M disease is poor, immunotherapy offers some promise in a malignancy associated with EBV and characterized by a peritumoural immune infiltrate. Our study aims to review past and on-going clinical trials of monoclonal antibody therapies against the checkpoint inhibitors (e.g. PD1 and CTLA-4), in R/M NPC. All randomized and nonrandomized controlled trials involving immune checkpoint inhibitor interventions for treatment of NPC were included in the study. We utilized a validated "risk of bias" tool to assess study quality. Four separate Phase I-II trials report the potential of PD1 inhibitor treatment for patients with NPC. Within the observed groups, camrelizumab combined with chemotherapy achieved an objective response in 91% of patients as first-line treatment for metastatic NPC (PFS 68% at 1-year) but this was associated with a high rate of grade >3 adverse events (87%; CTCAE version 4.03). The remaining three studies focused on recurrent NPC disease in patients who had received at least one line of prior chemotherapy. Within this group, camrelizumab monotherapy achieved an objective response in 34% of patients (PFS 27% at 1-year; range across all three studies 20.5-34%). No NPC trial has yet reported on specific outcomes for non-PD1 checkpoint inhibitors but 11 on-going studies include alternative targets (e.g. PD-L1/CTLA-4) as combination or monotherapy treatments. In considering checkpoint immunotherapies for NPC, initial results show promise for anti-PD1 interventions. Further phase I-III trials are in progress to clarify clinical outcomes, fully determine safety profiles, and optimize drug combinations and administration schedules., (© 2020 UICC.)
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- 2020
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27. Maintenance avelumab versus continuation of first-line chemotherapy in gastric cancer: JAVELIN Gastric 100 study design.
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Moehler M, Ryu MH, Dvorkin M, Lee KW, Coşkun HŞ, Wong R, Chung HC, Poltoratsky A, Tsuji A, Yen CJ, Muntean AS, Le Sourd S, Vaccaro GM, Overton L, Boku N, Wainberg ZA, Patel M, Sharma M, Xiong H, Conti I, Taieb J, and Bang YJ
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- Antibodies, Monoclonal administration & dosage, Antibodies, Monoclonal adverse effects, Antibodies, Monoclonal, Humanized, Antineoplastic Agents administration & dosage, Antineoplastic Agents adverse effects, Biomarkers, Tumor, Humans, Maintenance Chemotherapy, Molecular Targeted Therapy, Neoplasm Staging, Stomach Neoplasms pathology, Treatment Outcome, Antibodies, Monoclonal therapeutic use, Antineoplastic Agents therapeutic use, Stomach Neoplasms drug therapy
- Abstract
Avelumab is a human anti-PD-L1 IgG1 monoclonal antibody that has shown antitumor activity in early phase studies in advanced/metastatic gastric/gastroesophageal junction cancer, including as first-line maintenance therapy. Here, we describe the design of JAVELIN Gastric 100 (NCT02625610), an open-label, Phase III trial. A total of 499 patients with locally advanced/metastatic HER2- gastric/gastroesophageal junction cancer adenocarcinoma, who had achieved at least stable disease following 12 weeks of first-line oxaliplatin/fluoropyrimidine chemotherapy, have been randomized 1:1 to receive avelumab maintenance therapy or continue chemotherapy. The primary objective is to demonstrate superior overall survival in all randomized patients or in the PD-L1+ population. Secondary objectives are to demonstrate superiority for progression-free survival and objective response rate, compare quality of life measures, and determine safety.
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- 2019
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28. Trial Vocal Fold Injection Predicts Thyroplasty Outcomes in Nonparalytic Glottic Incompetence.
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Dumberger LD, Overton L, Buckmire RA, and Shah RN
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- Atrophy, Female, Humans, Laryngeal Diseases surgery, Male, Middle Aged, Polytetrafluoroethylene therapeutic use, Prognosis, Quality of Life, Retrospective Studies, Treatment Outcome, Voice, Carboxymethylcellulose Sodium therapeutic use, Cicatrix surgery, Collagen therapeutic use, Gelatin Sponge, Absorbable therapeutic use, Glottis, Laryngoplasty, Vocal Cord Paralysis surgery, Vocal Cords surgery
- Abstract
Objectives: Trial vocal fold injection (TVFI) may be used prior to permanent medialization when voice outcome is uncertain. We aimed to determine whether voice outcomes of TVFI are predictive of, or correlate with outcomes after type I Gore-Tex medialization thyroplasty (GMT) in patients with nonparalytic glottic incompetence (GI)., Methods: Thirty-five patients with nonparalytic GI who underwent TVFI followed by GMT were retrospectively reviewed. Change in voice-related quality of life (VRQOL) after TVFI was compared to change in VRQOL 3 to 9 months after GMT. Similar comparisons were made for change in glottal function index (GFI) and change in grade, roughness, breathiness, asthenia, and strain (GRBAS). Sample correlation coefficients were calculated., Results: Change in VRQOL after TVFI showed good correlation with change in VRQOL after GMT, r = 0.55. Change in GFI after TVFI showed strong correlation with change in GFI after GMT, r = 0.74. Change in GRBAS after TVFI showed excellent correlation with change in GRBAS after GMT, r = 0.90., Conclusion: The TVFI is a useful tool in nonparalytic GI when outcomes from glottic closure procedures are not clear. Voice outcome measures after TVFI strongly correlate with outcomes from GMT. These data may be used to more confidently counsel patients regarding their predicted outcomes of permanent medialization.
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- 2017
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29. Longitudinal Voice Outcomes After Type I Gore-tex Thyroplasty for Nonparalytic Glottic Incompetence.
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Overton L, Adams K, Shah RN, and Buckmire RA
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- Female, Follow-Up Studies, Humans, Laryngoplasty methods, Male, Retrospective Studies, Voice Quality, Laryngeal Diseases surgery, Laryngoplasty instrumentation, Polytetrafluoroethylene, Prostheses and Implants
- Abstract
Objective: Type I Gore-tex thyroplasty (GTP) for nonparalytic glottic incompetence (GI) results in significantly improved subjective and perceptual voice outcomes. We endeavored to investigate the longitudinal course of voice outcomes measuring the same patients across time points stratified by diagnostic subgroup., Methods: Seventy-five patients with nonparalytic GI treated with GTP in the past 9 years were retrospectively reviewed and grouped according to their primary diagnoses (atrophy, scar, hypomobility, and paresis). Voice outcome measures, Voice-Related Quality of Life (VRQOL), Glottal Function Index (GFI), and GRBAS (grade, roughness, breathiness, asthenia, and strain) for each individual patient and diagnostic subgroup were grouped by time interval following surgery: 0 to 90 days, 3 to 9 months, 9 to 18 months, 18 to 36 months, 3 to 5 years, and 5 to 10 years., Results: Across all diagnoses, statistically significant improvement in VRQOL was maintained at 3 to 5 years (P = .03) and GFI at 5 to 10 years (P = .02). The GRBAS showed statistically significant improvements out to 18 to 36 months (P = .02). In the subgroup analysis, hypomobility/paresis patients maintained significant improvement voice measures longer than patients with other diagnoses. As a group, scar patients did not show statistically significant postoperative improvement in VRQOL or GFI at any of the tested time points., Conclusions: Gore-tex thyroplasty provides durable improvement in subjective and perceptual voice outcomes for patients with nonparalytic GI. Patients treated for hypomobility/paresis have the most durable vocal outcomes followed by atrophy and lastly, scar., (© The Author(s) 2016.)
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- 2017
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30. Trends in chronic rhinosinusitis research in the past three decades.
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Banglawala SM, Schlosser RJ, Wenztel J, Walsh T, Overton L, and Soler ZM
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- Bibliometrics, Biomedical Research methods, Chronic Disease, Humans, Outcome Assessment, Health Care methods, Outcome Assessment, Health Care trends, Research Design trends, Biomedical Research trends, Rhinitis diagnosis, Rhinitis etiology, Rhinitis physiopathology, Rhinitis therapy, Sinusitis diagnosis, Sinusitis etiology, Sinusitis physiopathology, Sinusitis therapy
- Abstract
Background: The objective of this work was to identify trends in chronic rhinosinusitis (CRS)-related publications for the past 3 decades., Methods: Literature review was conducted using multiple terms, including sinusitis, chronic rhinosinusitis, chronic sinus disease, nasal polyposis, ethmoid sinusitis, frontal sinusitis, and maxillary sinusitis. Abstracts were divided into 3 decades: 1983 through 1992, 1993 through 2002, and 2003 through 2012. For each decade, we compared the total number of publications and journals, study design, use of validated outcome measures, quality of evidence, number of authors, country of origin, and clinical vs basic science., Results: A total of 3406 abstracts were identified. There was a statistically significant increase in the number of publications with a 637% increase from 1983 through 1992 to 2003 through 2012 (p < 0.05). Likewise, the number of journals with CRS-related publications significantly increased during the study period (117 to 350; p < 0.05). Prospective studies increased (15.30% to 28.90%, p < 0.05) and retrospective studies decreased (33.00% to 17.36%, p < 0.05). Cohort studies were the most common type of design study (18.70% to 32.46%). In studies reporting outcome measures, the use of validated measures significantly increased over time (2.56% to 49.70%, p < 0.05). Although, the quality of evidence for most clinical publications for all 3 decades were grade C, the number and percentage of grade A and grade B publications increased significantly over time (0.99% to 7.23%, p < 0.05; 6.9% to 10.44%, p < 0.05; respectively)., Conclusion: CRS-related publication quantity and quality have increased over the last 3 decades., (© 2015 ARS-AAOA, LLC.)
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- 2016
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31. Search for varicella zoster virus and herpes simplex virus-1 in normal human cerebral arteries.
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Nagel MA, Choe A, Khmeleva N, Overton L, Rempel A, Wyborny A, Traktinskiy I, and Gilden D
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- Adolescent, Adult, Aged, Aged, 80 and over, Antigens, Viral genetics, Cerebral Arteries virology, DNA, Viral genetics, Female, Humans, Leukocyte Common Antigens genetics, Leukocytes cytology, Leukocytes metabolism, Male, Middle Aged, Antigens, Viral analysis, Cerebral Arteries chemistry, DNA, Viral analysis, Herpesvirus 1, Human genetics, Herpesvirus 3, Human genetics, Leukocyte Common Antigens analysis
- Abstract
Virological confirmation of varicella zoster virus (VZV) vasculopathy is provided by presence of virus in the cerebral arteries, frequently associated with inflammation. Yet, cerebral arteries from normal subjects have never been studied for VZV DNA or antigen. We analyzed 63 human cerebral arteries from 45 subjects for VZV DNA and antigen, control herpes simplex virus (HSV)-1 DNA and antigen, and leukocyte-specific CD45 antigen. No cerebral arteries contained VZV or HSV-1 DNA or antigen; eight arteries from seven subjects contained leukocytes expressing CD45. Thus, the presence of VZV antigen in cerebral arteries of patients with stroke is likely to be clinically significant.
- Published
- 2013
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32. Unseasonal transmission of H3N2 influenza A virus during the swine-origin H1N1 pandemic.
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Ghedin E, Wentworth DE, Halpin RA, Lin X, Bera J, DePasse J, Fitch A, Griesemer S, Hine E, Katzel DA, Overton L, Proudfoot K, Sitz J, Szczypinski B, StGeorge K, Spiro DJ, and Holmes EC
- Subjects
- Animals, Humans, Influenza A virus genetics, Influenza A virus isolation & purification, Influenza, Human epidemiology, Influenza, Human virology, New York epidemiology, Phylogeny, Seasons, Sequence Analysis, Swine, Disease Outbreaks, Influenza A Virus, H1N1 Subtype genetics, Influenza A Virus, H1N1 Subtype isolation & purification, Influenza A Virus, H3N2 Subtype genetics, Influenza A Virus, H3N2 Subtype isolation & purification, Influenza, Human transmission
- Abstract
The initial wave of swine-origin influenza A virus (pandemic H1N1/09) in the United States during the spring and summer of 2009 also resulted in an increased vigilance and sampling of seasonal influenza viruses (H1N1 and H3N2), even though they are normally characterized by very low incidence outside of the winter months. To explore the nature of virus evolution during this influenza "off-season," we conducted a phylogenetic analysis of H1N1 and H3N2 sequences sampled during April to June 2009 in New York State. Our analysis revealed that multiple lineages of both viruses were introduced and cocirculated during this time, as is typical of influenza virus during the winter. Strikingly, however, we also found strong evidence for the presence of a large transmission chain of H3N2 viruses centered on the south-east of New York State and which continued until at least 1 June 2009. These results suggest that the unseasonal transmission of influenza A viruses may be more widespread than is usually supposed.
- Published
- 2010
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33. Evolutionary dynamics of human rotaviruses: balancing reassortment with preferred genome constellations.
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McDonald SM, Matthijnssens J, McAllen JK, Hine E, Overton L, Wang S, Lemey P, Zeller M, Van Ranst M, Spiro DJ, and Patton JT
- Subjects
- Antigenic Variation genetics, Child, Child, Preschool, Gastroenteritis genetics, Gastroenteritis virology, Genotype, Humans, Infant, Models, Molecular, Molecular Sequence Data, Open Reading Frames genetics, Phylogeny, Rotavirus chemistry, Rotavirus Infections genetics, Rotavirus Infections virology, Sequence Analysis, DNA, Evolution, Molecular, Genome, Viral, Reassortant Viruses genetics, Rotavirus genetics
- Abstract
Group A human rotaviruses (RVs) are a major cause of severe gastroenteritis in infants and young children. Yet, aside from the genes encoding serotype antigens (VP7; G-type and VP4; P-type), little is known about the genetic make-up of emerging and endemic human RV strains. To gain insight into the diversity and evolution of RVs circulating at a single location over a period of time, we sequenced the eleven-segmented, double-stranded RNA genomes of fifty-one G3P[8] strains collected from 1974 to 1991 at Children's Hospital National Medical Center, Washington, D. C. During this period, G1P[8] strains typically dominated, comprising on average 56% of RV infections each year in hospitalized children. A notable exception was in the 1976 and 1991 winter seasons when the incidence of G1P[8] infections decreased dramatically, a trend that correlated with a significant increase in G3P[8] infections. Our sequence analysis indicates that the 1976 season was characterized by the presence of several genetically distinct, co-circulating clades of G3P[8] viruses, which contained minor but significant differences in their encoded proteins. These 1976 lineages did not readily exchange gene segments with each other, but instead remained stable over the course of the season. In contrast, the 1991 season contained a single major clade, whose genome constellation was similar to one of the 1976 clades. The 1991 clade may have gained a fitness advantage after reassorting with as of yet unidentified RV strain(s). This study reveals for the first time that genetically distinct RV clades of the same G/P-type can co-circulate and cause disease. The findings from this study also suggest that, although gene segment exchange occurs, most reassortant strains are replaced over time by lineages with preferred genome constellations. Elucidation of the selective pressures that favor maintenance of RVs with certain sets of genes may be necessary to anticipate future vaccine needs.
- Published
- 2009
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34. Sequence, annotation, and analysis of synteny between rice chromosome 3 and diverged grass species.
- Author
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Buell CR, Yuan Q, Ouyang S, Liu J, Zhu W, Wang A, Maiti R, Haas B, Wortman J, Pertea M, Jones KM, Kim M, Overton L, Tsitrin T, Fadrosh D, Bera J, Weaver B, Jin S, Johri S, Reardon M, Webb K, Hill J, Moffat K, Tallon L, Van Aken S, Lewis M, Utterback T, Feldblyum T, Zismann V, Iobst S, Hsiao J, de Vazeille AR, Salzberg SL, White O, Fraser C, Yu Y, Kim H, Rambo T, Currie J, Collura K, Kernodle-Thompson S, Wei F, Kudrna K, Ammiraju JS, Luo M, Goicoechea JL, Wing RA, Henry D, Oates R, Palmer M, Pries G, Saski C, Simmons J, Soderlund C, Nelson W, de la Bastide M, Spiegel L, Nascimento L, Huang E, Preston R, Zutavern T, Palmer L, O'Shaughnessy A, Dike S, McCombie WR, Minx P, Cordum H, Wilson R, Jin W, Lee HR, Jiang J, and Jackson S
- Subjects
- Arabidopsis genetics, Chromosome Mapping, Chromosomes, Artificial, Bacterial genetics, Genes, Plant, Minisatellite Repeats, Molecular Sequence Data, Oryza classification, Physical Chromosome Mapping, Poaceae classification, Proteome, Species Specificity, Zea mays classification, Zea mays genetics, Chromosomes, Plant genetics, Oryza genetics, Poaceae genetics
- Abstract
Rice (Oryza sativa L.) chromosome 3 is evolutionarily conserved across the cultivated cereals and shares large blocks of synteny with maize and sorghum, which diverged from rice more than 50 million years ago. To begin to completely understand this chromosome, we sequenced, finished, and annotated 36.1 Mb ( approximately 97%) from O. sativa subsp. japonica cv Nipponbare. Annotation features of the chromosome include 5915 genes, of which 913 are related to transposable elements. A putative function could be assigned to 3064 genes, with another 757 genes annotated as expressed, leaving 2094 that encode hypothetical proteins. Similarity searches against the proteome of Arabidopsis thaliana revealed putative homologs for 67% of the chromosome 3 proteins. Further searches of a nonredundant amino acid database, the Pfam domain database, plant Expressed Sequence Tags, and genomic assemblies from sorghum and maize revealed only 853 nontransposable element related proteins from chromosome 3 that lacked similarity to other known sequences. Interestingly, 426 of these have a paralog within the rice genome. A comparative physical map of the wild progenitor species, Oryza nivara, with japonica chromosome 3 revealed a high degree of sequence identity and synteny between these two species, which diverged approximately 10,000 years ago. Although no major rearrangements were detected, the deduced size of the O. nivara chromosome 3 was 21% smaller than that of japonica. Synteny between rice and other cereals using an integrated maize physical map and wheat genetic map was strikingly high, further supporting the use of rice and, in particular, chromosome 3, as a model for comparative studies among the cereals.
- Published
- 2005
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35. Molecular analysis of the bison phagocyte NADPH oxidase: cloning and sequencing of five NADPH oxidase cDNAs.
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Gauss KA, Bunger PL, Siemsen DW, Young CJ, Nelson-Overton L, Prigge JR, Swain SD, and Quinn MT
- Subjects
- Alternative Splicing, Amino Acid Sequence, Animals, Cloning, Molecular, Membrane Glycoproteins genetics, Molecular Sequence Data, NADPH Dehydrogenase genetics, NADPH Oxidase 2, NADPH Oxidases biosynthesis, Neutrophils enzymology, Phosphoproteins genetics, Sequence Alignment, Sequence Analysis, DNA, Bison genetics, DNA, Complementary chemistry, Membrane Transport Proteins, NADPH Oxidases genetics, Phagocytes enzymology
- Abstract
During the host defense process, neutrophils migrate into infected tissues where they become activated, resulting in the assembly of a superoxide anion-generating complex known as the NADPH oxidase. Despite the importance of this system in animal host defense, almost nothing is known about the NADPH oxidase in neutrophils from wild ruminant species. In the present studies, we provide a molecular analysis of the bison leukocyte NADPH oxidase. Using reverse transcriptase-polymerase chain reaction and rapid amplification of cDNA ends, we cloned and sequenced the full-length cDNAs for five bison NADPH oxidase components: p22(phox), p40(phox), p47(phox) and p67(phox), and gp91(phox). When compared to other species, the deduced amino acid sequences of the bison homologs were most similar to those of bovine. Interestingly, a bison p40(phox) alternative splice product was isolated, which was similar to that observed for human p40(phox) in that the cDNAs contained sequence from intron 8. Consistent with the high degree of similarity between bison and bovine amino acid sequences, immunoblot analysis showed that the bison homologs migrated similarly to their bovine counterparts. Overall, these studies show that the bison and bovine NADPH oxidase genes are highly conserved between these two species, despite their divergence from a common ancestor over 1 million years ago.
- Published
- 2002
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36. A physical map of the mouse genome.
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Gregory SG, Sekhon M, Schein J, Zhao S, Osoegawa K, Scott CE, Evans RS, Burridge PW, Cox TV, Fox CA, Hutton RD, Mullenger IR, Phillips KJ, Smith J, Stalker J, Threadgold GJ, Birney E, Wylie K, Chinwalla A, Wallis J, Hillier L, Carter J, Gaige T, Jaeger S, Kremitzki C, Layman D, Maas J, McGrane R, Mead K, Walker R, Jones S, Smith M, Asano J, Bosdet I, Chan S, Chittaranjan S, Chiu R, Fjell C, Fuhrmann D, Girn N, Gray C, Guin R, Hsiao L, Krzywinski M, Kutsche R, Lee SS, Mathewson C, McLeavy C, Messervier S, Ness S, Pandoh P, Prabhu AL, Saeedi P, Smailus D, Spence L, Stott J, Taylor S, Terpstra W, Tsai M, Vardy J, Wye N, Yang G, Shatsman S, Ayodeji B, Geer K, Tsegaye G, Shvartsbeyn A, Gebregeorgis E, Krol M, Russell D, Overton L, Malek JA, Holmes M, Heaney M, Shetty J, Feldblyum T, Nierman WC, Catanese JJ, Hubbard T, Waterston RH, Rogers J, de Jong PJ, Fraser CM, Marra M, McPherson JD, and Bentley DR
- Subjects
- Animals, Chromosomes genetics, Chromosomes, Human, Pair 6 genetics, Cloning, Molecular, Conserved Sequence genetics, Contig Mapping methods, Genome, Human, Humans, Molecular Sequence Data, Radiation Hybrid Mapping, Sequence Alignment, Sequence Homology, Nucleic Acid, Species Specificity, Synteny, Genome, Mice genetics, Physical Chromosome Mapping methods
- Abstract
A physical map of a genome is an essential guide for navigation, allowing the location of any gene or other landmark in the chromosomal DNA. We have constructed a physical map of the mouse genome that contains 296 contigs of overlapping bacterial clones and 16,992 unique markers. The mouse contigs were aligned to the human genome sequence on the basis of 51,486 homology matches, thus enabling use of the conserved synteny (correspondence between chromosome blocks) of the two genomes to accelerate construction of the mouse map. The map provides a framework for assembly of whole-genome shotgun sequence data, and a tile path of clones for generation of the reference sequence. Definition of the human-mouse alignment at this level of resolution enables identification of a mouse clone that corresponds to almost any position in the human genome. The human sequence may be used to facilitate construction of other mammalian genome maps using the same strategy.
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- 2002
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37. Pharmacokinetic parameters and mechanisms of inhibition of rat type 1 and 2 steroid 5alpha-reductases: determinants for different in vivo activities of GI198745 and finasteride in the rat.
- Author
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Stuart JD, Lee FW, Simpson Noel D, Kadwell SH, Overton LK, Hoffman CR, Kost TA, Tippin TK, Yeager RL, Batchelor KW, and Bramson HN
- Subjects
- 3-Oxo-5-alpha-Steroid 4-Dehydrogenase genetics, 3-Oxo-5-alpha-Steroid 4-Dehydrogenase metabolism, Animals, Azasteroids blood, Azasteroids pharmacology, Binding, Competitive, Cells, Cultured, Dutasteride, Enzyme Inhibitors blood, Enzyme Inhibitors pharmacology, Finasteride blood, Finasteride pharmacology, Insecta, Kinetics, Male, Rats, Rats, Sprague-Dawley, Testosterone metabolism, Time Factors, Transfection, 5-alpha Reductase Inhibitors, Azasteroids pharmacokinetics, Enzyme Inhibitors pharmacokinetics, Finasteride pharmacokinetics
- Abstract
The interaction of baculovirus expressed rat steroid 5alpha-reductase types 1 and 2 (r5AR1 and r5AR2) with 17beta-N-(2,5-bis(trifluoromethyl)phenyl)carbamoyl-4-aza-5alpha-androst-1-en-3-one (GI198745) was investigated at pH 7 and 37 degrees. This 5alpha-reductase inhibitor was found previously to be a time-dependent inhibitor of the two human 5alpha-reductase isozymes. In contrast, we demonstrate in the present study that although GI198745 is a potent time-dependent inhibitor of r5AR2, it is a classical rapid-equilibrium inhibitor of r5AR1. This type of behavior with human and rat 5alpha-reductases has been shown for the inhibitor 17beta-(N-tert-butylcarbamoyl)-4-aza-5alpha-androst-1-en-3-one (finasteride), a current therapy for benign prostatic hyperplasia. Inhibition of r5AR1 by GI198745 was competitive with testosterone and followed Michaelis-Menten kinetics with a K(i) value of 0.3 +/- 0.02 nM. Data for the inhibition of r5AR2 by GI198745 were consistent with a two-step mechanism, where K(i) is the dissociation constant for an initial enzyme-inhibitor complex and k(3) is the rate constant for the second slow step. The pseudo-bimolecular rate constant (k(3)/K(i)) for the association of GI198745 with r5AR2 was (2.0 +/- 0.4) x 10(7) M(-1) sec(-1). The high affinity of this inhibitor for r5AR2 was further demonstrated by the inability of the enzyme-inhibitor complex to dissociate after approximately 7 days of dialysis at 4 degrees. Both GI198745 and finasteride appear to inactivate r5AR2 by apparent irreversible modification, but are classical, reversible inhibitors of r5AR1. Therefore, we hypothesize that because of its pharmacokinetic parameters and increased potency against r5AR1, GI198745 is more effective than finasteride in preventing the growth of the rat prostate.
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- 2001
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38. Replication-associated activities of purified human papillomavirus type 11 E1 helicase.
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Rocque WJ, Porter DJ, Barnes JA, Dixon EP, Lobe DC, Su JL, Willard DH, Gaillard R, Condreay JP, Clay WC, Hoffman CR, Overton LK, Pahel G, Kost TA, and Phelps WC
- Subjects
- Acid Anhydride Hydrolases isolation & purification, Acid Anhydride Hydrolases metabolism, Animals, Antibodies, Monoclonal biosynthesis, Antigens, Polyomavirus Transforming metabolism, Baculoviridae genetics, Cells, Cultured, Circular Dichroism, DNA Helicases genetics, DNA Helicases isolation & purification, DNA Helicases metabolism, DNA-Binding Proteins genetics, DNA-Binding Proteins isolation & purification, DNA-Binding Proteins metabolism, Female, Humans, Insecta cytology, Insecta virology, Mice, Nucleoside-Triphosphatase, Point Mutation, Protein Structure, Secondary, Viral Proteins genetics, Viral Proteins isolation & purification, Viral Proteins metabolism, Acid Anhydride Hydrolases chemistry, DNA Helicases chemistry, DNA Replication, DNA-Binding Proteins chemistry, Papillomaviridae chemistry, Viral Proteins chemistry
- Abstract
Replication of human papillomavirus type11 (HPV11) requires both the E1 and the E2 proteins. E1 is structurally and functionally similar to SV40 large T-antigen and is a DNA helicase/NTPase that binds to the origin of replication and initiates viral DNA replication. The biochemical characterization of HPV E1 is incompletely documented in the literature in part because of difficulties in expressing and purifying the protein. Herein, we report a method for the overexpression of full-length, untagged E1 (73.5 kDa) in baculovirus-infected Trichoplusia ni insect cells and the purification to homogeneity using a two-step procedure. The purified protein is a nonspecific NTPase that hydrolyzes ATP, dATP, UTP, or GTP equally well. Point mutations were made in the putative NTPase domain to verify that the activities observed were encoded by E1. Purified mutant D523N had negligible ATPase and helicase activities but retained DNA-binding activity. Sedimentation equilibrium ultracentrifugation and glycerol gradient centrifugation demonstrated that the wild-type protein is primarily a hexamer in its purified form. Secondary structure determination by circular dichroism revealed a large percentage of alpha-helical structure consistent with secondary structure predictions. These data define a fundamental set of biochemical and kinetic parameters for HPV E1 which are a critical prerequisite to future mechanistic studies of the enzyme., (Copyright 2000 Academic Press.)
- Published
- 2000
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39. A scintillation proximity assay for the Raf/MEK/ERK kinase cascade: high-throughput screening and identification of selective enzyme inhibitors.
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McDonald OB, Chen WJ, Ellis B, Hoffman C, Overton L, Rink M, Smith A, Marshall CJ, and Wood ER
- Subjects
- Amino Acid Sequence, Animals, Baculoviridae genetics, Base Sequence, DNA Primers genetics, Enzyme Inhibitors pharmacology, Escherichia coli genetics, Humans, In Vitro Techniques, Kinetics, MAP Kinase Kinase 1, Mitogen-Activated Protein Kinase 1 antagonists & inhibitors, Mitogen-Activated Protein Kinase 1 genetics, Peptides chemistry, Peptides metabolism, Protein Serine-Threonine Kinases antagonists & inhibitors, Protein Serine-Threonine Kinases genetics, Protein-Tyrosine Kinases antagonists & inhibitors, Protein-Tyrosine Kinases genetics, Proto-Oncogene Proteins c-raf antagonists & inhibitors, Proto-Oncogene Proteins c-raf genetics, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Signal Transduction, Substrate Specificity, Mitogen-Activated Protein Kinase 1 metabolism, Mitogen-Activated Protein Kinase Kinases, Protein Serine-Threonine Kinases metabolism, Protein-Tyrosine Kinases metabolism, Proto-Oncogene Proteins c-raf metabolism, Scintillation Counting methods
- Abstract
We have developed a quantitative scintillation proximity assay (SPA) that reproduces the Raf/MEK/ERK signal transduction pathway. The components of this assay include human cRaf1, MEK1, and ERK2 and a biotinylated peptide substrate for ERK2. cRaf1 was expressed as a his-tagged protein in insect cells in an active form. MEK1 and ERK2 were expressed in Escherichia coli as glutathione S-transferase (GST)-fusion proteins in their inactive forms. ERK2 was removed from the GST portion of the fusion protein by cleavage with thrombin protease. When the purified components are incubated together, cRaf-1 phosphorylates and activates MEK1, MEK1 phosphorylates and activates ERK2, and ERK2 phosphorylates the peptide, biotin-AAATGPLSPGPFA. Phosphorylation of the peptide using [gamma-33P]ATP is detected following binding to streptavidin-coated SPA beads. The assay detects inhibitors of cRaf1, MEK1, or ERK2, and has been used to screen large numbers of compounds. The specific target of inhibition was subsequently identified with secondary assays described herein., (Copyright 1999 Academic Press.)
- Published
- 1999
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40. Optimization of transient gene expression in mammalian cells and potential for scale-up using flow electroporation.
- Author
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Parham JH, Iannone MA, Overton LK, and Hutchins JT
- Abstract
The goals of this study were to identify mammalian cell lines which could be efficiently transiently-transfected and scaled-up for protein production. The transfection efficiencies of eight cell lines (NSO, NSO-TAg, CV-1, COS-7, CHO, CHO-TAg, HEK 293, and 293-EBNA) were measured using electroporation for DNA delivery and green fluorescent protein (Evans, 1996) as the reporter gene. In addition, we have evaluated the effects of stable expression of viral proteins, cell cycle manipulation, and butyrate post-treatment in small scale experiments. The cell lines varied widely in their GFP transfection efficiencies. Stable expression of simian virus 40 large T-antigen or Epstein Barr nuclear antigen failed to significantly increase transfection efficiency above that seen in the parental lines. Aphidicolin (a DNA polymerase inhibitor), which blocked cells from S or G2/M, brought about an increase in transfection efficiency in two cell lines. The primary effect of butyrate (a histone deacetylase inhibitor) post-treatment was an increased intensity of the fluorescent signal of green fluorescent protein, as measured by flow cytometry (1.0 to 4.2-fold, depending on the cell line). The combined use of aphidicolin pretreatment followed by butyrate treatment post- electroporation yielded increases in fluorescence intensities ranging from 0.9 to 6.8-fold. Based on their high transfection efficiencies in small scale experiments, rapid growth, and ability to grow in suspension culture, CHO, CHO-TAg, and 293-EBNA were selected to assess the feasibility of using flow electroporation for large-scale transfections. Using secreted placental alkaline phosphatase as a reporter, 293-EBNA cells produced the highest protein levels in both the presence and absence of butyrate. These data indicate that flow electroporation provides an efficient method of DNA delivery into large numbers of cells for mammalian protein production.
- Published
- 1998
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41. Detailed characterization of a purified type 4 phosphodiesterase, HSPDE4B2B: differentiation of high- and low-affinity (R)-rolipram binding.
- Author
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Rocque WJ, Holmes WD, Patel IR, Dougherty RW, Ittoop O, Overton L, Hoffman CR, Wisely GB, Willard DH, and Luther MA
- Subjects
- 3',5'-Cyclic-AMP Phosphodiesterases genetics, 3',5'-Cyclic-AMP Phosphodiesterases isolation & purification, Baculoviridae genetics, Binding Sites, Cloning, Molecular, Genetic Vectors chemistry, Genetic Vectors genetics, Genetic Vectors isolation & purification, Humans, Kinetics, Molecular Weight, Peptide Fragments biosynthesis, Peptide Fragments genetics, Peptide Fragments isolation & purification, Protein Structure, Secondary, Protein Structure, Tertiary, Pyrrolidinones chemistry, Recombinant Proteins, Rolipram, 3',5'-Cyclic-AMP Phosphodiesterases chemistry, 3',5'-Cyclic-AMP Phosphodiesterases metabolism, Pyrrolidinones metabolism
- Abstract
We have overexpressed in a baculovirus expression system, and purified to > 95% homogeneity, milligram quantities of a human recombinant rolipram-sensitive cAMP phosphodiesterase, HSPDE4B2B (amino acid residues 81-564). The protein expression levels were approximately 8 mg of HSPDE4B2B (81-564) per liter of Sf9 cells. The Km of the purified enzyme for cAMP was 4 microM and the Ki for the Type 4 phosphodiesterase-specific inhibitor (R)-rolipram was 0.6 microM. The specific activity of the purified protein was 40 mumol/min/mg protein. A nonequilibrium filter binding assay revealed a high-affinity (R)-rolipram binding site on the purified enzyme with a Kd of 1.5 nM and a stoichiometry of 0.05-0.3 mol of (R)-rolipram per mol of HSPDE4B2B (81-564). Equilibrium dialysis experiments revealed a single binding constant of 140 nM with a stoichiometry of 0.75 mol of (R)-rolipram per mol of HSPDE4B2B (81-564). Size exclusion chromatography and analytical ultracentrifugation experiments suggest that the protein exists in multiple association states larger than a monomer. Proteolysis experiments revealed a 43-kDa fragment that contained catalytic and rolipram-inhibitable activities, but the fragment showed no high-affinity (R)-rolipram binding. Based on the proteolytic cleavage studies a 43-kDa protein was constructed, expressed, and purified. This protein, HSPDE4B2B (152-528), had Km and Vmax similar to those of the HSPDE4B2B (81-564) protein, but did not exhibit high-affinity (R)-rolipram binding. The protein did show low-affinity (R)-rolipram binding using the equilibrium binding assay. These results show that a low-affinity binding site for (R)-rolipram is solely contained within the catalytic domain of HSPDE4B2B, whereas high-affinity (R)-rolipram binding requires residues within the catalytic domain and residues flanking N- and/or C-terminal to the catalytic region.
- Published
- 1997
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42. Cloning of a disintegrin metalloproteinase that processes precursor tumour-necrosis factor-alpha.
- Author
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Moss ML, Jin SL, Milla ME, Bickett DM, Burkhart W, Carter HL, Chen WJ, Clay WC, Didsbury JR, Hassler D, Hoffman CR, Kost TA, Lambert MH, Leesnitzer MA, McCauley P, McGeehan G, Mitchell J, Moyer M, Pahel G, Rocque W, Overton LK, Schoenen F, Seaton T, Su JL, and Becherer JD
- Subjects
- ADAM Proteins, ADAM17 Protein, Amino Acid Sequence, Animals, Binding Sites, Cell Line, Cloning, Molecular, Conserved Sequence, Disintegrins isolation & purification, Disintegrins metabolism, Humans, Membrane Proteins genetics, Membrane Proteins isolation & purification, Membrane Proteins metabolism, Metalloendopeptidases isolation & purification, Metalloendopeptidases metabolism, Molecular Sequence Data, Recombinant Proteins genetics, Recombinant Proteins metabolism, Sequence Homology, Amino Acid, Swine, Disintegrins genetics, Metalloendopeptidases genetics, Protein Precursors metabolism, Tumor Necrosis Factor-alpha metabolism
- Abstract
Tumour-necrosis factor-alpha (TNF-alpha) is a cytokine that contributes to a variety of inflammatory disease states. The protein exists as a membrane-bound precursor of relative molecular mass 26K which can be processed by a TNF-alpha-converting enzyme (TACE), to generate secreted 17K mature TNF-alpha. We have purified TACE and cloned its complementary DNA. TACE is a membrane-bound disintegrin metalloproteinase. Structural comparisons with other disintegrin-containing enzymes indicate that TACE is unique, with noteable sequence identity to MADM, an enzyme implicated in myelin degradation, and to KUZ, a Drosophila homologue of MADM important for neuronal development. The expression of recombinant TACE (rTACE) results in the production of functional enzyme that correctly processes precursor TNF-alpha to the mature form. The rTACE provides a readily available source of enzyme to help in the search for new anti-inflammatory agents that target the final processing stage of TNF-alpha production.
- Published
- 1997
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43. Comparative analyses of relative ERCC3 and ERCC6 mRNA levels in gliomas and adjacent non-neoplastic brain.
- Author
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Dabholkar MD, Berger MS, Vionnet JA, Overton L, Thompson C, Bostick-Bruton F, Yu JJ, Silber JR, and Reed E
- Subjects
- Astrocytoma genetics, Astrocytoma therapy, Brain Neoplasms therapy, DNA Repair Enzymes, Glioblastoma genetics, Glioblastoma therapy, Glioma therapy, Humans, Oligodendroglioma genetics, Oligodendroglioma therapy, Poly-ADP-Ribose Binding Proteins, RNA, Messenger genetics, RNA, Neoplasm genetics, Regression Analysis, Brain physiology, Brain Neoplasms genetics, DNA Helicases genetics, DNA Repair, DNA-Binding Proteins genetics, Drosophila Proteins, Gene Expression Regulation, Neoplastic, Glioma genetics
- Abstract
Nucleotide excision repair (NER) is an ordered process in nonmalignant cells, in both human and nonhuman systems. We previously reported that in human brain there is discordant mRNA expression of excision repair cross-complementing (ERCC) 1 and ERCC2 in malignant tissues, concurrent with excellent concordance of these genes in nonmalignant tissues from the same patients. Here we have extended these studies to compare low-grade tumors to high-grade tumors and to include ERCC3 (which links DNA repair with DNA transcription) and ERCC6 (which is essential for gene-specific repair). Glial tumor and adjacent normal brain specimens from 19 individuals were studied. Paired malignant and nonmalignant tissues were obtained from 12 of these patients. For ERCC3, there was excellent concordance of mRNA expression between malignant and nonmalignant tissues from the same individuals (P = 0.003). For ERCC6, no concordance was observed (P = 0.314). Tumor tissue from patients with high-grade gliomas exhibited marked discordance of mRNA expression patterns in situations in which good concordance was observed in tumor tissue from low-grade gliomas. We previously established that malignant brain tumors show increased disorder of genes in the NER process, as compared with nonmalignant tissues. These data suggest that increasing disorder in the NER process may occur as cells move from low-grade to high-grade malignancy.
- Published
- 1996
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44. Inhibition of human steroid 5alpha reductases type I and II by 6-aza-steroids: structural determinants of one-step vs two-step mechanism.
- Author
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Moss ML, Kuzmic P, Stuart JD, Tian G, Peranteau AG, Frye SV, Kadwell SH, Kost TA, Overton LK, and Patel IR
- Subjects
- Humans, Isoenzymes antagonists & inhibitors, Kinetics, Structure-Activity Relationship, 5-alpha Reductase Inhibitors, Azasteroids chemistry, Enzyme Inhibitors chemistry, Finasteride chemistry
- Abstract
We have discovered that 17beta-[N,N-(diethyl)carbamoyl]-6-azaandrost-4-en-3-one is a time-dependent inhibitor of type II 5alpha-reductase, as is the drug finasteride. Unlike finasteride, the 6-aza-steroid is not a time-dependent inhibitor of type I 5 alpha-reductase. Finasteride inhibition of type II enzyme proceeds in a two-step mechanism. At pH 6 and 37 degrees C, an initial finasteride-reductase complex is formed with a K(i)(app) of 11.9 +/- 4.1 nM. In a second step, an irreversible complex is formed with a rate constant of inactivation of 0.09 +/- 0.01 s(-1). In contrast, the 6-aza-steroid is a reversible inhibitor. From the results of a simplified mathematical analysis, based on the rapid equilibrium approximation, the inhibitor and the enzyme form an initial complex with a K(i) of 6.8 +/- 0.2 nM. The reversible formation of a final complex, with an overall K(i) of 0.07 +/- 0.02 nM, is characterized by a first-order isomerization rate constant 0.0035 +/- 0.0001 s(-1) for the forward step and 0.00025 +/- 0.00006 s(-1) for the backward step. All rate constants for the two-step mechanism were obtained by using a general numerical integration method. The best fit values for the association and dissociation rate constants were 5.0 microM(-1) s(-1) and 0.033 +/- 0.008 s(-1), respectively, and the isomerization rate constants were 0.0035 +/- 0.007 s(-1) and 0.000076 +/- 0.000019 s(-1). These values correspond to an initial K(i) of 6.5 nM and an overall dissociation constant of 0.14 nM. The data presented here show that both finasteride and the 6-aza-steroid analogs are potent against type II 5alpha-reductase, although their mechanisms of inhibition are different.
- Published
- 1996
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45. Expression of tissue inhibitor of metalloproteinases by recombinant baculovirus-infected insect cells cultured in an airlift fermentor.
- Author
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Overton LK, Patel I, Becherer JD, Chandra G, and Kost TA
- Subjects
- Animals, Blotting, Western, Cell Division, Cell Line, Culture Techniques methods, Glycoproteins genetics, Glycoproteins isolation & purification, Glycoproteins pharmacology, Matrix Metalloproteinase Inhibitors, Nucleopolyhedroviruses growth & development, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins isolation & purification, Spodoptera cytology, Tissue Inhibitor of Metalloproteinases, Virus Cultivation methods, Cloning, Molecular methods, Culture Techniques instrumentation, Glycoproteins biosynthesis, Nucleopolyhedroviruses genetics, Recombinant Fusion Proteins biosynthesis, Virus Cultivation instrumentation
- Published
- 1995
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46. Agouti protein is an antagonist of the melanocyte-stimulating-hormone receptor.
- Author
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Lu D, Willard D, Patel IR, Kadwell S, Overton L, Kost T, Luther M, Chen W, Woychik RP, and Wilkison WO
- Subjects
- Adenylyl Cyclases metabolism, Adrenocorticotropic Hormone metabolism, Agouti Signaling Protein, Animals, Baculoviridae, Cattle, Cell Line, Enzyme Activation, Hair metabolism, Humans, Mice, Obesity etiology, Recombinant Proteins, Tumor Cells, Cultured, Intercellular Signaling Peptides and Proteins, Proteins physiology, Receptors, Pituitary Hormone antagonists & inhibitors
- Abstract
The genetic loci agouti and extension control the relative amounts of eumelanin (brown-black) and phaeomelanin (yellow-red) pigments in mammals: extension encodes the receptor for melanocyte-stimulating hormone (MSH) and agouti encodes a novel 131-amino-acid protein containing a signal sequence. Agouti, which is produced in the hair follicle, acts on follicular melanocytes to inhibit alpha-MSH-induced eumelanin production, resulting in the subterminal band of phaeomelanin often visible in mammalian fur. Here we use partially purified agouti protein to demonstrate that agouti is a high-affinity antagonist of the MSH receptor and blocks alpha-MSH stimulation of adenylyl cyclase, the effector through which alpha-MSH induces eumelanin synthesis. Agouti was also found to be an antagonist of the melanocortin-4 receptor, a related MSH-binding receptor. Consequently, the obesity caused by ectopic expression of agouti in the lethal yellow (Ay) mouse may be due to the inhibition of melanocortin receptor(s) outside the hair follicle.
- Published
- 1994
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47. 17 beta-(N-tert-butylcarbamoyl)-4-aza-5 alpha-androstan-1-en-3-one is an active site-directed slow time-dependent inhibitor of human steroid 5 alpha-reductase 1.
- Author
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Tian G, Stuart JD, Moss ML, Domanico PL, Bramson HN, Patel IR, Kadwell SH, Overton LK, Kost TA, and Mook RA Jr
- Subjects
- Binding Sites, Humans, Kinetics, 5-alpha Reductase Inhibitors, Finasteride pharmacology
- Abstract
17 beta-(N-tert-butylcarbamoyl)-4-aza-5 alpha-androstan-1-en-3-one (finasteride), which has been approved for treatment of benign prostatic hyperplasia, is shown here to be a slow time-dependent inhibitor of human steroid 5 alpha-reductase isozyme 1. This inhibition is characterized by an initial, fast step where the inhibitor binds to the enzyme followed by a slow step that leads to a final enzyme-inhibitor complex (EI*). No recovery of activity from this EI* complex was observed after dialysis for 3 days. The formation of EI* is diminished in the presence of a competitive, reversible inhibitor, indicating that the inhibition is active site-directed. At 37 degrees C and pH 7.0, the rate constant for the second, slow inhibition step, k3, is (1.40 +/- 0.04) x 10(-3) s-1 and the pseudo-bimolecular rate constant, k3/Ki, is (4.0 +/- 0.3) x 10(3) M-1 s-1. This latter rate constant is less than the value of 2.7 x 10(5) M-1 s-1 determined for the inhibition of 5 alpha-reductase 2 by finasteride [Faller, B., Farley, D., & Nick, H. (1993) Biochemistry 32, 5705-5710].(ABSTRACT TRUNCATED AT 250 WORDS)
- Published
- 1994
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48. X-ray-induced specific-locus mutations in the ad-3 region of two-component heterokaryons of Neurospora crassa. XI. Heterozygous effects of gene/point mutations of genotype ad-3A or ad-3B.
- Author
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de Serres FJ, Overton LK, and Sadler BM
- Subjects
- Dose-Response Relationship, Radiation, Genes, Lethal genetics, Genes, Recessive genetics, Neurospora crassa genetics, X-Rays, Adenine, Heterozygote, Mutation, Neurospora crassa radiation effects
- Abstract
Previous studies on X-ray-induced irreparable adenine-3 mutants (designated ad-3IR), induced in heterokaryon 12 of Neurospora crassa, showed that they were not recessive, and that they demonstrated heterozygous effects in terms of markedly reduced linear growth rates as compared with a wild-type control (de Serres, 1965, 1988). Homology tests on X-ray-induced irreparable mutants showed that they map, in the main part, as a series of overlapping multilocus deletions that extend both proximally and distally into the immediately adjacent genetic regions, as well as into the 'X' region (a region of unknown, but essential, function) between ad-3A and ad-3B (de Serres, 1969, 1989a). Studies on a larger sample of X-ray-induced multilocus deletion mutations of genotype (ad-3A)IR or (ad-3B)IR (de Serres et al., 1992) demonstrated that heterozygous effects are allele specific and that there was no correlation with genotype, radiation dose or complementation map position. Furthermore, the heterozygous effects of multilocus deletions in the ad-3 region can be modified genetically and biochemically (de Serres and Miller, 1988). In the present paper, the heterozygous effects of X-ray-induced gene/point mutations of genotype ad-3AR or ad-3BR, induced in heterokaryon 12 (Webber and de Serres, 1965; de Serres, 1988, 1989a), were determined. The studies presented in this paper show that 8.1% (3/37) of X-ray-induced ad-3AR mutations exhibit heterozygous effects in terms of reduced linear growth rates in forced dikaryons with a gene/point mutant at the ad-3B locus, and 10.8% (4/37) in forced dikaryons with a multilocus deletion mutation covering the ad-3B locus. In addition, 24.3% (9/37) of ad-3AR mutations exhibit heterozygous effects in terms of enhanced linear growth rates in forced dikaryons with a gene/point mutant at the ad-3B locus. Similar studies with X-ray-induced ad-3BR mutations showed that 54.9% (28/51) exhibit heterozygous effects in terms of reduced growth rates in forced dikaryons with a gene/point mutant at the ad-3A locus and 100.0% (48/48) in forced dikaryons with a multilocus deletion covering the ad-3A locus. These studies have also shown that about a 13-fold higher percentage of X-ray-induced multiple-locus mutations of genotype ad-3AR + RLCL have heterozygous effects resulting in reduced growth rates than X-ray-induced single-locus mutations of genotype ad-3AR. The overall data base on X-ray-induced ad-3 gene/point mutations in the present studies demonstrates that heterozygous effects are allele specific, genotype specific, and locus specific.(ABSTRACT TRUNCATED AT 400 WORDS)
- Published
- 1992
- Full Text
- View/download PDF
49. X-ray-induced specific-locus mutations in the ad-3 region of two-component heterokaryons of Neurospora crassa. X. Heterozygous effects of multilocus deletion mutations of genotype ad-3A or ad-3B.
- Author
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de Serres FJ, Overton LK, and Sadler BM
- Subjects
- Adenine, Animals, Chromosome Deletion, Dose-Response Relationship, Radiation, Drosophila melanogaster genetics, Genetic Complementation Test, Heterozygote, Mice genetics, Mutagenesis, Neurospora crassa genetics, Neurospora crassa growth & development, X-Rays, Genes, Fungal, Neurospora crassa radiation effects
- Abstract
Previous studies on X-ray-induced irreparable adenine-3 mutations (designated [ad-3]IR), induced in heterokaryon 12 of Neurospora crassa, demonstrated that they were not recessive and exhibited heterozygous effects in terms of markedly reduced linear growth rates (de Serres, 1965). Complementation tests with a series of tester strains carrying multilocus deletion mutations in the ad-3 and immediately adjacent genetic regions demonstrated that X-ray-induced irreparable mutations map, in the main part, as a series of overlapping multilocus deletion mutations that extend both proximally and distally into the immediately adjacent genetic regions, as well as into the 'X' region (a region of unknown, but essential function) between ad-3A and ad-3B (de Serres, 1968, 1989). Further studies (de Serres and Miller, 1988) have shown that the heterozygous effects of multilocus deletion mutations in the ad-3 region can be modified genetically and biochemically. In the present paper, the heterozygous effects of X-ray-induced multilocus deletion mutations of genotype ad-3A or ad-3B, induced in heterokaryon 12 (Webber and de Serres, 1965; de Serres, 1988, 1989), have been determined. These data show that 57.7% (15/26) of X-ray-induced multilocus deletion mutations covering the ad-3A locus have heterozygous effects, in terms of reduced linear growth rates, in forced dikaryons with a gene/point mutant at the ad-3B locus and 80.0% (20/25) in forced dikaryons with a multilocus deletion mutation covering the ad-3B locus. In addition, 35.1% (20/57) of X-ray-induced multilocus deletion mutations covering the ad-3B locus have heterozygous effects in forced dikaryons with a gene/point mutant at the ad-3A locus, and 100.0% (35/35) in forced dikaryons with a multilocus deletion mutation covering the ad-3A locus. These results demonstrate that the dominant or recessive characteristics of X-ray-induced specific-locus mutations resulting from multilocus deletion mutations are allele specific.
- Published
- 1992
- Full Text
- View/download PDF
50. 2-Amino-N6-hydroxyadenine induces gene/point mutations and multiple-locus mutations, but not multilocus deletion mutations, in the ad-3 region of a two-component heterokaryon of Neurospora crassa.
- Author
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de Serres FJ, Brockman HE, and Overton LK
- Subjects
- 2-Aminopurine toxicity, Adenine toxicity, Alleles, Chromosome Deletion, DNA Damage, Genetic Complementation Test, Genotype, Neurospora crassa drug effects, Adenine analogs & derivatives, Mutagens, Neurospora crassa genetics
- Abstract
The mutagenicity of 2-amino-N6-hydroxyadenine (AHA) has been studied in Neurospora crassa by treating a two-component heterokaryon (H-12) and recovering specific-locus mutations induced in the ad-3 region. This assay system permits the identification of ad-3A and/or ad-3B mutants resulting from gene/point mutations, multilocus deletion mutations, and multiple-locus mutations of various genotypes, involving one or both loci. Genetic characterization of the ad-3 mutants recovered from experiments with AHA in H-12 shows that 98.9% (270/273) of the ad-3 mutants are gene/point mutations (ad-3R), 1.1% (3/270) are unknowns, and none is a multilocus deletion mutation (ad-3IR). Among the gene/point mutations, 3.3% (9/273) are multiple-locus mutations (gene/point mutations with a closely-linked recessive lethal mutation [ad-3R + RLCL]). Another 25.3% (69/273) are multiple-locus mutations with a recessive lethal mutation located elsewhere in the genome [ad-3R + RL]. Heterokaryon tests for allelic complementation among the ad-3BR mutants showed that 90.8% (139/153) of the mutants were complementing, and 20.3% (31/153) were leaky. In addition, 32.5% (38/117) of the ad-3AR mutants were leaky. These data are consistent with the hypothesis that AHA produces specific-locus mutations in the ad-3 region of N. crassa by base-pair substitution. The data from the present experiments are compared with the data for 2-aminopurine (2AP)-induced ad-3 mutants in H-12 (de Serres and Brockman, 1991). Whereas, 2AP is a weak mutagen in H-12, AHA is extremely potent (Brockman et al., 1987). In contrast with 2AP, AHA induces ad-3 mutants exclusively by gene/point mutation in H-12. We conclude that whereas AHA induces ad-3 mutants predominantly by AT to GC base-pair transitions, 2AP induces ad-3 mutants by a wide variety of mechanisms including: (1) AT to GC and GC to AT base-pair transitions, (2) frameshift mutations, (3) other, as yet unidentified, intragenic alterations, (4) small multilocus deletion mutations, and (5) multiple-locus ad-3R mutations with closely linked recessive lethal mutations.
- Published
- 1991
- Full Text
- View/download PDF
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