crossmark LETTER TO THE EDITOR
Evaluation of the 2-Aminomethylphenol JPC-2997 in Aotus Monkeys Infected with Plasmodium falciparum Fiona McCallum,a Ivor Harris,a Karin van Breda,a Sai Lata De,b Danielle I. Stanisic,b Michael F. Good,b David P. Jacobus,c Michael D. Edsteina Department of Drug Evaluation, Australian Army Malaria Institute, Enoggera, Brisbane, Queensland, Australiaa; Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, Australiab; Jacobus Pharmaceutical Company, Princeton, New Jersey, USAc
P
reviously, we reported JPC-2997 [4-(tert-Butyl)-2-((tertbutylamino)methyl)-6-(6-(trifluoromethyl)pyridin-3-yl)phenol] to possess high in vitro antimalarial activity against Plasmodium falciparum lines, low cytotoxicity in mammalian cell lines, a long blood elimination half-life in Aotus monkeys (10.8 days), and potent in vivo efficacy against animal malaria models (1). Herein, as planned by Birrell et al. (1), we report on the in vivo efficacy of JPC-2997 against the chloroquine- and quinineresistant P. falciparum FVO strain in Aotus monkeys (2). As part of a malaria vaccine study of chemically attenuated parasites (CAP) in Aotus monkeys (3), various regimens of JPC2997 given alone (1- and 3-day treatments) and JPC-2997 coadministered with artesunate (1- and 3-day treatments) were used to treat monkeys that developed an FVO infection. Briefly, nonsplenectomized Aotus monkeys (n ⫽ 9; body weight range, 0.98 kg to 1.55 kg) were inoculated intravenously with 1 ⫻ 105 to 4 ⫻ 107 parasites of the P. falciparum FVO strain. Three to 13 days after the onset of patency, monkeys were treated by orogastric intubation with either JPC-2997 alone or JPC-2997 plus artesunate. After treatment of the monkeys, thick and thin blood smears were examined daily by counting parasitized erythrocytes against 200 leukocytes or 10,000 erythrocytes. When no parasites were detected in 100 microscopic high-power (⫻1,000) fields, follow-up blood thick smears were examined twice a week for 3 weeks after parasite inoculation and then approximately once a week up to at least day 60 after parasite inoculation. The study was approved by the AMI Animal Ethics Committee (protocol AEC 10-13).
Regimens evaluated and parasite responses after treatment with JPC-2997 alone and in combination with artesunate are summarized in Table 1. The 1-day JPC-2997 (20 mg/kg of body weight) treatment cleared parasites as rapidly as the two 3-day treatment courses of JPC-2997 (10 or 20 mg/kg/day), with ⬎99% reduction in parasitemia by day 3 after starting treatment. None of the 4 naive monkeys had a recrudescence after administration of the three JPC-2997 regimens. Previously, we reported that artesunate administered at 10 mg/kg daily for 3 days rapidly cleared parasites in Aotus monkeys infected with the P. falciparum FVO strain but that recrudescence occurred between days 9 and 20 (4). In the present study, a 1-day treatment of JPC-2997 (20 mg/kg) plus artesunate (10 mg/kg) administered to 4 monkeys (2 naive and 2 infected with CAP) with rising high parasitemia (range, 264.4 to 512.8 parasites ⫻ 103/l) cleared parasites by day 4 postdose, with no recrudescence observed over a 60-day follow-up period. Although the 1-day treat-
Accepted manuscript posted online 14 December 2015 Citation Mccallum F, Harris I, van Breda K, De SL, Stanisic DI, Good MF, Jacobus DP, Edstein MD. 2016. Evaluation of the 2-aminomethylphenol JPC-2997 in Aotus monkeys infected with Plasmodium falciparum. Antimicrob Agents Chemother 60:1948 –1949. doi:10.1128/AAC.02799-15. Address correspondence to Michael D. Edstein, [email protected]. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
TABLE 1 Parasitological responses of Aotus monkeys infected with the FVO strain of Plasmodium falciparum following 1-day and 3-day oral treatments with JPC-2997 or 1-day and 3-day treatments with JPC-2997 plus artesunatea Parasitemia (no. of parasites ⫻ 103/l) on day:
Regimen (1-day and 3-day treatments)
No. of monkeys
Immune status
0
1
2
3
4
7
JPC-2997
20 mg/kg on days 0, 1, and 2 10 mg/kg on days 0, 1, and 2 20 mg/kg on day 0
1 1* 1** 1
Naive Naive Naive Naive
110.9 68.0 59.5 6.3
83.1 3.1 2.1 0.5
1.7 1.6 1.4 NPS
0.7 0.5 0.2 NPS
ND ND ND NPS
NPS NPS NPS NPS
AS ⫹ JPC-2997
10 mg/kg AS on days 0 and 1, and 20 mg/kg JPC-2997 on day2 10 mg/kg AS ⫹ 20 mg/kg JPC-2997 on day 0
1
Naive
188.8
76.4
1.2
NPS
NPS
NPS
1 1 1 1 1* 1**
Naive Naive CAP vaccine CAP vaccine Semi-immne Semi-immne
512.8 418.4 264.4 309.8 16.0 3.6
219.9 29.3 10.6 17.7 1.0 0.1
26.0 4.5 0.4 0.3 0.1 NPS
2.7 1.2 NPS 0.2 NPS NPS
ND NPS NPS NPS NPS NPS
NPS NPS NPS NPS NPS NPS
Drug
a AS, artesunate; CAP, monkeys were exposed to chemically attenuated parasites 101 days before being challenged with the FVO strain; semi-immune, monkeys had been infected 101 days previously with the FVO strain and treated with JPC-2997 (10 mg/kg daily for 3 days or a single dose of 20 mg/kg); ND, not determined; NPS, no parasites seen. Day 0 is the day of treatment. Other days are the days after treatment was started. Nine Aotus monkeys were tested. Single and double asterisks indicate the same monkey. Parasite recrudescence was nil for all drugs. The follow-up period was at least 60 days after starting treatment.
1948
aac.asm.org
Antimicrobial Agents and Chemotherapy
March 2016 Volume 60 Number 3
Letter to the Editor
ment with JPC-2997 plus artesunate cured the FVO infections, parasite clearance was more rapid when a 3-day course of artesunate at 10 mg/kg daily was used, with no parasites seen on day 3 after starting treatment of an initial parasite load of 145 to 515 parasites ⫻ 103/l (4). The lack of recrudescence reported in this study following treatment of FVO infections with either JPC-2997 alone or JPC2997 in combination with artesunate provides further evidence of the in vivo potency and long prophylactic activity of JPC-2997. These findings suggest that JPC-2997 is a promising long-acting candidate antimalarial drug. ACKNOWLEDGMENTS We thank Joanne Beckett and John Hunter for veterinary support. We also acknowledge the excellent technical support of Stephen McLeodRobertson and Nerissa Walpole for animal husbandry and handling during drug administration and blood collections, Kerryn Rowcliffe for preparation of parasite inoculums, and Thomas Travers and Donna MacKenzie for microscopy analysis. The opinions expressed are those of the authors and do not necessarily reflect those of the Australian Defense Organization or any extant policy. The study was supported and funded by the National Health and Medical Research Council (Australia Fellowship and Program Grant), the Australian Defense Organization, and the Jacobus Pharmaceutical Company. With the exception of D.P.J., who is the Director of Jacobus Pharmaceutical Company, we have no conflicts of interest to declare.
March 2016 Volume 60 Number 3
FUNDING INFORMATION Australian Defence Organisation provided funding to Fiona McCallum, Ivor Harris, and Michael D. Edstein. Jacobus Pharmaceutical Company provided funding to Karin van Breda and David P. Jacobus. Department of Health | National Health and Medical Research Council (NHMRC) provided funding to Sai Lata De, Danielle I. Stanisic, and Michael F. Good under grant number AAP1037304.
REFERENCES 1. Birrell GW, Chavchich M, Ager AL, Shieh HM, Heffernan GD, Zhao W, Krasucki PE, Saionz KW, Terpinski J, Schiehser GA, Jacobus LR, Shanks GD, Jacobus DP, Edstein MD. 2015. JPC-2997, a new aminomethylphenol with high in vitro and in vivo antimalarial activities against blood stages of Plasmodium. Antimicrob Agents Chemother 59:170 –177. http://dx.doi .org/10.1128/AAC.03762-14. 2. Schmidt LH. 1978. Plasmodium falciparum and Plasmodium vivax infections in the owl monkey (Aotus trivirgatus). II. Responses to chloroquine, quinine, and pyrimethamine. Am J Trop Med Hyg 27:703–717. 3. De SL, Stanisic DI, van Breda K, Bellete B, Harris I, McCallum F, Edstein MD, Good MF. 2015. Persistence and immunogenicity of chemically attenuated blood stage Plasmodium falciparum in Aotus monkeys, poster no. LB5144 for poster session 79. 64th Annual Meeting of the American Society of Tropical Medicine and Hygiene (ASTMH) (25 to 29 October 2015). ASTMH, Philadelphia, PA. 4. Haynes RK, Fugmann B, Stetter J, Rieckmann K, Heilmann HD, Chan HW, Cheung MK, Lam WL, Wong HN, Croft SL, Vivas L, Rattray L, Stewart L, Peters W, Robinson BL, Edstein MD, Kotecka B, Kyle DE, Beckermann B, Gerisch M, Radtke M, Schmuck G, Steinke W, Wollborn U, Schmeer K, Romer A. 2006. Artemisone—a highly active antimalarial drug of the artemisinin class. Angew Chem Int Ed Engl 45:2082–2088. http: //dx.doi.org/10.1002/anie.200503071.
Antimicrobial Agents and Chemotherapy
aac.asm.org
1949
Evaluation of the 2-Aminomethylphenol JPC-2997 in Aotus Monkeys Infected with Plasmodium falciparum Fiona McCallum,a Ivor Harris,a Karin van Breda,a Sai Lata De,b Danielle I. Stanisic,b Michael F. Good,b David P. Jacobus,c Michael D. Edsteina Department of Drug Evaluation, Australian Army Malaria Institute, Enoggera, Brisbane, Queensland, Australiaa; Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, Australiab; Jacobus Pharmaceutical Company, Princeton, New Jersey, USAc
P
reviously, we reported JPC-2997 [4-(tert-Butyl)-2-((tertbutylamino)methyl)-6-(6-(trifluoromethyl)pyridin-3-yl)phenol] to possess high in vitro antimalarial activity against Plasmodium falciparum lines, low cytotoxicity in mammalian cell lines, a long blood elimination half-life in Aotus monkeys (10.8 days), and potent in vivo efficacy against animal malaria models (1). Herein, as planned by Birrell et al. (1), we report on the in vivo efficacy of JPC-2997 against the chloroquine- and quinineresistant P. falciparum FVO strain in Aotus monkeys (2). As part of a malaria vaccine study of chemically attenuated parasites (CAP) in Aotus monkeys (3), various regimens of JPC2997 given alone (1- and 3-day treatments) and JPC-2997 coadministered with artesunate (1- and 3-day treatments) were used to treat monkeys that developed an FVO infection. Briefly, nonsplenectomized Aotus monkeys (n ⫽ 9; body weight range, 0.98 kg to 1.55 kg) were inoculated intravenously with 1 ⫻ 105 to 4 ⫻ 107 parasites of the P. falciparum FVO strain. Three to 13 days after the onset of patency, monkeys were treated by orogastric intubation with either JPC-2997 alone or JPC-2997 plus artesunate. After treatment of the monkeys, thick and thin blood smears were examined daily by counting parasitized erythrocytes against 200 leukocytes or 10,000 erythrocytes. When no parasites were detected in 100 microscopic high-power (⫻1,000) fields, follow-up blood thick smears were examined twice a week for 3 weeks after parasite inoculation and then approximately once a week up to at least day 60 after parasite inoculation. The study was approved by the AMI Animal Ethics Committee (protocol AEC 10-13).
Regimens evaluated and parasite responses after treatment with JPC-2997 alone and in combination with artesunate are summarized in Table 1. The 1-day JPC-2997 (20 mg/kg of body weight) treatment cleared parasites as rapidly as the two 3-day treatment courses of JPC-2997 (10 or 20 mg/kg/day), with ⬎99% reduction in parasitemia by day 3 after starting treatment. None of the 4 naive monkeys had a recrudescence after administration of the three JPC-2997 regimens. Previously, we reported that artesunate administered at 10 mg/kg daily for 3 days rapidly cleared parasites in Aotus monkeys infected with the P. falciparum FVO strain but that recrudescence occurred between days 9 and 20 (4). In the present study, a 1-day treatment of JPC-2997 (20 mg/kg) plus artesunate (10 mg/kg) administered to 4 monkeys (2 naive and 2 infected with CAP) with rising high parasitemia (range, 264.4 to 512.8 parasites ⫻ 103/l) cleared parasites by day 4 postdose, with no recrudescence observed over a 60-day follow-up period. Although the 1-day treat-
Accepted manuscript posted online 14 December 2015 Citation Mccallum F, Harris I, van Breda K, De SL, Stanisic DI, Good MF, Jacobus DP, Edstein MD. 2016. Evaluation of the 2-aminomethylphenol JPC-2997 in Aotus monkeys infected with Plasmodium falciparum. Antimicrob Agents Chemother 60:1948 –1949. doi:10.1128/AAC.02799-15. Address correspondence to Michael D. Edstein, [email protected]. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
TABLE 1 Parasitological responses of Aotus monkeys infected with the FVO strain of Plasmodium falciparum following 1-day and 3-day oral treatments with JPC-2997 or 1-day and 3-day treatments with JPC-2997 plus artesunatea Parasitemia (no. of parasites ⫻ 103/l) on day:
Regimen (1-day and 3-day treatments)
No. of monkeys
Immune status
0
1
2
3
4
7
JPC-2997
20 mg/kg on days 0, 1, and 2 10 mg/kg on days 0, 1, and 2 20 mg/kg on day 0
1 1* 1** 1
Naive Naive Naive Naive
110.9 68.0 59.5 6.3
83.1 3.1 2.1 0.5
1.7 1.6 1.4 NPS
0.7 0.5 0.2 NPS
ND ND ND NPS
NPS NPS NPS NPS
AS ⫹ JPC-2997
10 mg/kg AS on days 0 and 1, and 20 mg/kg JPC-2997 on day2 10 mg/kg AS ⫹ 20 mg/kg JPC-2997 on day 0
1
Naive
188.8
76.4
1.2
NPS
NPS
NPS
1 1 1 1 1* 1**
Naive Naive CAP vaccine CAP vaccine Semi-immne Semi-immne
512.8 418.4 264.4 309.8 16.0 3.6
219.9 29.3 10.6 17.7 1.0 0.1
26.0 4.5 0.4 0.3 0.1 NPS
2.7 1.2 NPS 0.2 NPS NPS
ND NPS NPS NPS NPS NPS
NPS NPS NPS NPS NPS NPS
Drug
a AS, artesunate; CAP, monkeys were exposed to chemically attenuated parasites 101 days before being challenged with the FVO strain; semi-immune, monkeys had been infected 101 days previously with the FVO strain and treated with JPC-2997 (10 mg/kg daily for 3 days or a single dose of 20 mg/kg); ND, not determined; NPS, no parasites seen. Day 0 is the day of treatment. Other days are the days after treatment was started. Nine Aotus monkeys were tested. Single and double asterisks indicate the same monkey. Parasite recrudescence was nil for all drugs. The follow-up period was at least 60 days after starting treatment.
1948
aac.asm.org
Antimicrobial Agents and Chemotherapy
March 2016 Volume 60 Number 3
Letter to the Editor
ment with JPC-2997 plus artesunate cured the FVO infections, parasite clearance was more rapid when a 3-day course of artesunate at 10 mg/kg daily was used, with no parasites seen on day 3 after starting treatment of an initial parasite load of 145 to 515 parasites ⫻ 103/l (4). The lack of recrudescence reported in this study following treatment of FVO infections with either JPC-2997 alone or JPC2997 in combination with artesunate provides further evidence of the in vivo potency and long prophylactic activity of JPC-2997. These findings suggest that JPC-2997 is a promising long-acting candidate antimalarial drug. ACKNOWLEDGMENTS We thank Joanne Beckett and John Hunter for veterinary support. We also acknowledge the excellent technical support of Stephen McLeodRobertson and Nerissa Walpole for animal husbandry and handling during drug administration and blood collections, Kerryn Rowcliffe for preparation of parasite inoculums, and Thomas Travers and Donna MacKenzie for microscopy analysis. The opinions expressed are those of the authors and do not necessarily reflect those of the Australian Defense Organization or any extant policy. The study was supported and funded by the National Health and Medical Research Council (Australia Fellowship and Program Grant), the Australian Defense Organization, and the Jacobus Pharmaceutical Company. With the exception of D.P.J., who is the Director of Jacobus Pharmaceutical Company, we have no conflicts of interest to declare.
March 2016 Volume 60 Number 3
FUNDING INFORMATION Australian Defence Organisation provided funding to Fiona McCallum, Ivor Harris, and Michael D. Edstein. Jacobus Pharmaceutical Company provided funding to Karin van Breda and David P. Jacobus. Department of Health | National Health and Medical Research Council (NHMRC) provided funding to Sai Lata De, Danielle I. Stanisic, and Michael F. Good under grant number AAP1037304.
REFERENCES 1. Birrell GW, Chavchich M, Ager AL, Shieh HM, Heffernan GD, Zhao W, Krasucki PE, Saionz KW, Terpinski J, Schiehser GA, Jacobus LR, Shanks GD, Jacobus DP, Edstein MD. 2015. JPC-2997, a new aminomethylphenol with high in vitro and in vivo antimalarial activities against blood stages of Plasmodium. Antimicrob Agents Chemother 59:170 –177. http://dx.doi .org/10.1128/AAC.03762-14. 2. Schmidt LH. 1978. Plasmodium falciparum and Plasmodium vivax infections in the owl monkey (Aotus trivirgatus). II. Responses to chloroquine, quinine, and pyrimethamine. Am J Trop Med Hyg 27:703–717. 3. De SL, Stanisic DI, van Breda K, Bellete B, Harris I, McCallum F, Edstein MD, Good MF. 2015. Persistence and immunogenicity of chemically attenuated blood stage Plasmodium falciparum in Aotus monkeys, poster no. LB5144 for poster session 79. 64th Annual Meeting of the American Society of Tropical Medicine and Hygiene (ASTMH) (25 to 29 October 2015). ASTMH, Philadelphia, PA. 4. Haynes RK, Fugmann B, Stetter J, Rieckmann K, Heilmann HD, Chan HW, Cheung MK, Lam WL, Wong HN, Croft SL, Vivas L, Rattray L, Stewart L, Peters W, Robinson BL, Edstein MD, Kotecka B, Kyle DE, Beckermann B, Gerisch M, Radtke M, Schmuck G, Steinke W, Wollborn U, Schmeer K, Romer A. 2006. Artemisone—a highly active antimalarial drug of the artemisinin class. Angew Chem Int Ed Engl 45:2082–2088. http: //dx.doi.org/10.1002/anie.200503071.
Antimicrobial Agents and Chemotherapy
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1949
