Vaccination with Vaxfectin® adjuvanted SIV DNA induces long-lasting humoral immune responses able to reduce SIVmac251 Viremia

Human Vaccines and Immunotherapeutics

Volumn 9, Issue 10, 2013, Pages 2069-2080

  Kulkarni, Viraj ^a   Rosati, Margherita ^a   Valentin, Antonio ^a   Jalah, Rashmi ^a   Alicea, Candido ^a   Yu, Lei ^b   Guan, Yongjun ^b   Shen, Xiaoying ^c   Tomaras, Georgia D ^c   La Branche, Celia ^c   Montefiori, David C ^d   Irene, Carmela ^d   Prattipati, Rajasekhar ^d   Pinter, Abraham ^d   Sullivan, Sean M ^e   Pavlakis, George N ^a   Felber, Barbara K ^a  

^a NATIONAL CANCER INSTITUTE   (United States)

^b UNIVERSITY OF MARYLAND SCHOOL OF MEDICINE   (United States)

^c DUKE UNIVERSITY SCHOOL OF MEDICINE   (United States)

^d UNIVERSITY OF MEDICINE AND DENTISTRY OF NEW JERSEY   (United States)

^e VICAL INC   (United States)

Author keywords

Adjuvant; Antibody; Avidity; HIV; Mucosal immunity; Rectal fluid; Rhesus macaques; Saliva; SIVmac239; neutralizing antibody; SIVsmE660; Systemic immunity; V1 and V2 antibodies

Indexed keywords

DNA VACCINE; GAG PROTEIN; MONOCYTE CHEMOTACTIC PROTEIN 3; PLACEBO; PLASMID VECTOR; VIRUS ENVELOPE PROTEIN;

ANIMAL CELL; ANIMAL EXPERIMENT; ANIMAL MODEL; ANIMAL TISSUE; ANTIBODY PRODUCTION; ANTIBODY RESPONSE; ANTIBODY TITER; ARTICLE; CELLULAR IMMUNITY; CONTROLLED STUDY; CYTOKINE PRODUCTION; CYTOKINE RELEASE; DNA IMMUNIZATION; DRUG DESIGN; DRUG EFFICACY; DRUG FORMULATION; DRUG POTENCY; DRUG TARGETING; DRUG TOLERABILITY; HUMORAL IMMUNITY; IMMUNOGENICITY; IMMUNOSTIMULATION; MOUSE; NONHUMAN; NUCLEOTIDE SEQUENCE; PROVOCATION TEST; RHESUS MONKEY; RISK REDUCTION; SIMIAN ACQUIRED IMMUNODEFICIENCY SYNDROME; SIMIAN IMMUNODEFICIENCY VIRUS; VIREMIA; VIRUS LOAD; VIRUS NEUTRALIZATION;

ADJUVANT; ANTIBODY; AVIDITY; HIV; MUCOSAL IMMUNITY; NEUTRALIZING ANTIBODY; RECTAL FLUID; RHESUS MACAQUES; SALIVA; SIVMAC239; SIVSME660; SYSTEMIC IMMUNITY; V1 AND V2 ANTIBODIES;

ADJUVANTS, IMMUNOLOGIC; ANIMALS; ANTIBODIES, NEUTRALIZING; ANTIBODIES, VIRAL; GENE PRODUCTS, ENV; GENE PRODUCTS, GAG; IMMUNITY, HUMORAL; IMMUNITY, MUCOSAL; MACACA MULATTA; MICE; MICE, INBRED BALB C; PHOSPHATIDYLETHANOLAMINES; SAIDS VACCINES; SIMIAN IMMUNODEFICIENCY VIRUS; VACCINATION; VACCINES, DNA; VIREMIA;

EID: 84885130095     PISSN: 21645515     EISSN: 2164554X     Source Type: Journal    
DOI: 10.4161/hv.25442     Document Type: Article

References (63)

1. Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, Kaewkungwal J, Chiu J, Paris R, et al.; MOPH-TAVEG Investigators. Vaccination with AIVAC and AIDSVAX to prevent HIV-1 infection in Thailand. N Engl J Med 2009;361:2209-20; PMID:19843557; http://dx.doi.org/10.1056/NEJMoa0908492
2. Lai L, Kwa S, Kozlowski PA, Montefiori DC, Ferrari G, Johnson WE, et al. Prevention of infection by a granulocyte-macro phage colony-stimulating factor co-expressing DNA/modified vaccinia Ankara simian immunodeficiency virus vaccine. J Infect Dis 2011;204:164-73; PMID:21628671; http://dx.doi.org/10.10 93/infdis/j ir199
3. Barouch DH, Liu J, Li H, Maxfield LF, Abbink P, Lynch DM, et al. Vaccine protection against acquisition of neutralization-resistant SIV challenges in rhesus monkeys. Nature 2012;482:89-93; PMID:22217938; http://dx.doi.org/10.1038/ nature10766
4. Flatz L, Cheng C, Wang L, Foulds KE, Ko SY, Kong WP, et al. Gene-based vaccination with a mismatched envelope protects against simian immunodeficiency virus infection in nonhuman primates. J Virol 2012;86:7760-70; PMID:22593152; http://dx.doi.org/10.1128/JVI.005 99-12
5. Letvin NL, Rao SS, Montefiori DC, Seaman MS, Sun Y, Lim SY, et al. Immune and Genetic Correlates of Vaccine Protection Against Mucosal Infection by SIV in Monkeys. Sci Transl Med 2011;3:81ra36; PMID:21543722; http://dx.doi.org/10. 1126/scitranslmed.3002351
6. Lai L, Kwa SF, Kozlowski PA, Montefiori DC, Nolen TL, Hudgens MG, et al. SIVmac239 MVA vaccine with and without a DNA prime, similar prevention of infection by a repeated dose SIVsmE660 challenge despite different immune responses. Vaccine 2012;30:1737-45; PMID:22178526; http://dx.doi.org/10.1016/j. vaccine.2011.12.026
7. Patel V, Jalah R, Kulkarni V, Valentin A, Rosati M, Alicea C, et al. DNA and virus particle vaccination protects against acquisition and confers control of viremia upon heterologous simian immunodeficiency virus challenge. Proc Natl Acad Sci U S A 2013;110:2975-80; PMID:23359688; http://dx.doi.org/10.1073/pnas. 1215393110
8. Winstone N, Wilson AJ, Morrow G, Boggiano C, Chiuchiolo MJ, Lopez M, et al. Enhanced control of pathogenic Simian immunodeficiency virus SIVmac239 replication in macaques immunized with an interleukin-12 plasmid and a DNA prime-viral vector boost vaccine regimen. J Virol 2011;85:9578-87; PMID:21734035; http://dx.doi.org/10.1128/JVI.05060-11
9. Kutzler MA, Weiner DB. DNA vaccines: ready for prime time? Nat Rev Genet 2008;9:776-88; PMID: 18781156; http://dx.d0i.0rg/10.1038/nrg2432
10. Mascola JR, Lewis MG, Van Cott TC, Stiegler G, Katinger H, Seaman M, et al. Cellular immunity elicited by human immunodeficiency virus type 1/simian immunodeficiency virus DNA vaccination does not augment the sterile protection afforded by passive infusion of neutralizing antibodies. J Virol 2003;77:10348-56; PMID: 12970419; http://dx.doi.org/10.1128/JVI.77.19.10348- 10356.2003
11. Barouch DH, Santra S, Schmitz JE, Kuroda MJ, Fu TM, Wagner W, et al. Control of viremia and prevention of clinical AIDS in rhesus monkeys by cytokineaugmented DNA vaccination. Science 2000;290:486-92; PMID:11039923; http://dx.doi.org/10.1126/science.290.5491.486
12. Barouch DH, Kunstman J, Glowczwskie J, Kunstman IQ, Egan MA, Peyerl FW, et al. Viral escape from dominant simian immunodeficiency virus epitope-specific cytotoxic T lymphocytes in DNA-vaccinated rhesus monkeys. J Virol 2003;77:7367-75; PMID: 12805435; http://dx.doi.org/10.1128/JVI.77.13.7367-7375. 2003
13. Barouch DH, Fu TM, Montefiori DC, Lewis MG, Shiver JW, Letvin NL. Vaccine-elicited immune responses prevent clinical AIDS in SHIV (89.6P) - infected rhesus monkeys. Immunol Lett 2001;79:57-61; PMID:11595290; http://dx.doi.org/10.1016/S0165-2478 (01) 00266-8
14. Chong SY, Egan MA, Kutzler MA, Megati S, Masood A, Roopchard V, et al. Comparative ability of plasmid IL-12 and IL-15 to enhance cellular and humoral immune responses elicited by a SIVgag plasmid DNA vaccine and alter disease progression following SHIV (89-6P) challenge in rhesus macaques. Vaccine 2007;25:4967-82; PMID: 17335943; http://dx.doi.org/10.1016/j.vaccine.2006.11.070
15. Egan MA, Charini WA, Kuroda MJ, Schmitz JE, Racz P, Tenner-Racz K, et al. Simian immunodeficiency virus (SIV) gag DNA-vaccinated rhesus monkeys develop secondary cytotoxic T-lymphocyte responses and control viral replication after pathogenic SIV infection. J Virol 2000;74:7485-95; PMID:10906202; http://dx.doi.org/10.1128/JVI.74.16.7485-7495.2000
16. Belisle SE, Yin J, Shedlock DJ, Dai A, Yan J, Hirao L, et al. Long-term programming of antigen-specific immunity from gene expression signatures in the PBMC of rhesus macaques immunized with an SIV DNA vaccine. PLoS One 2011;6: el9681; PMID:21701683; http://dx.doi.org/10.1371/journal.pone.0019681
17. Boyer JD, Maciag PC, Parkinson R, Wu L, Lewis MG, Weiner DB, et al. Rhesus macaques with high levels of vaccine induced IFN-gamma producing cells better control viral set-point following challenge with SIV239. Vaccine 2006;24:4498-502; PMID:16185790; http://dx.doi.org/10.1016/j.vaccine.2005.08.016
18. Boyer JD, Robinson TM, Kutzler MA, Vansant G, Hokey DA, Kumar S, et al. Protection against simian/human immunodeficiency virus (SHIV) 89.6P in macaques after coimmunization with SHIV antigen and IL-15 plasmid. Proc Natl Acad Sci U S A 2007;104:18648-53; PMID:18000037; http://dx.doi.org/10.1073/pnas.0709198104
19. Haigwood NL, Pierce CC, Robertson MN, Watson AJ, Montefiori DC, Rabin M, et al. Protection from pathogenic SIV challenge using multigenic DNA vaccines. Immunol Lett 1999;66:183-8; PMID: 10203053; http://dx.doi.org/10.1016/S0165-2478 (98) 00156-4
20. Muthumani K, Bagarazzi M, Conway D, Hwang DS, Manson K, Ciccarelli R, et al. A Gag-Pol/Env-Rev SIV239 DNA vaccine improves CD4 counts, and reduce viral loads after pathogenic intrarectal SIV (mac) 251 challenge in rhesus Macaques. Vaccine 2003;21:629-37; PMID:12531331; http://dx.doi.org/10.1016/S0264-410X (02) 00571-6
21. Rosati M, Valentin A, Jalah R, Patel V, Von Gegerfelt A, Bergamaschi C, et al. Increased immune responses in rhesus macaques by DNA vaccination combined with electroporation. Vaccine 2008;26:5223-9; PMID:18468743; http://dx.doi.org/10.1016/j.vaccine.2008.03.090
22. Rosati M, Von Gegerfelt A, Roth P, Alicea C, Valentin A, Robert-Guroff M, et al. DNA vaccines expressing different forms of simian immunodeficiency virus antigens decrease viremia upon SIVmac251 challenge. J Virol 2005;79:8480-92; PMID:15956591; http://dx.doi.org/10.1128/JVI.79.13.8480-8492.2005
23. Yin J, Dai A, Lecureux J, Arango T, Kutzler MA, Yan J, et al. High antibody and cellular responses induced to HIV-1 clade C envelope following DNA vaccines delivered by electroporation. Vaccine 2011;29:6763-70; PMID:21195801; http://dx.doi.org/1 0.1016/j.vaccine.2010.12.055
24. Sardesai NY, Weiner DB. Electroporation delivery of DNA vaccines: prospects for success. Curr Opin Immunol 2011;23:421-9; PMID:21530212; http://dx.doi.org/10.1016/j.coi.2011.03.008
25. Hutnick NA, Myles DJ, Bian CB, Muthumani K, Weiner DB. Selected approaches for increasing HIV DNA vaccine immunogenicity in vivo. Curr Opin Virol 2011;1:233-40; PMID:22440782; http://dx.doi.org/10.1016/j.coviro.2011.08. 003
26. Patel V, Valentin A, Kulkarni V, Rosati M, Bergamaschi C, Jalah R, et al. Long-lasting humoral and cellular immune responses and mucosal dissemination after intramuscular DNA immunization. Vaccine 2010;28:4827-36; PMID:20451642; http://dx.doi.org/10.1016/j. vaccine.2010.04.064
27. Luckay A, Sidhu MK, Kjeken R, Megati S, Chong SY, Roopchand V, et al. Effect of plasmid DNA vaccine design and in vivo electroporation on the resulting vaccine-specific immune responses in rhesus macaques. J Virol 2007;81:5257-69; PMID: 17329330; http://dx.doi.org/10.1128/JVI. 00 05 5-07
28. Jalah R, Patel V, Kulkarni V, Rosati M, Alicea C, Ganneru B, et al. IL-12 DNA as molecular vaccine adjuvant increases the cytotoxic T cell responses and breadth of humoral immune responses in SIV DNA vaccinated macaques. Hum Vaccin Immunother 2012;8:1620-9; PMID:22894956; http://dx.doi.org/10.4161/hv. 21407
29. Rosati M, Bergamaschi C, Valentin A, Kulkarni V, Jalah R, Alicea C, et al. DNA vaccination in rhesus macaques induces potent immune responses and decreases acute and chronic viremia after SIVmac251 challenge. Proc Natl Acad Sci U S A 2009;106:15 831-6; PMID: 19717425; http://dx.doi.org/10.1073/pnas. 0902628106
30. Kulkarni V, Jalah R, Ganneru B, Bergamaschi C, Alicea C, Von Gegerfelt A, et al. Comparison of immune responses generated by optimized DNA vaccination against SIV antigens in mice and macaques. Vaccine 2011;29:6742-54; PMID:21195080; http://dx.doi.org/10.1016/j.vaccine.2010.12.056
31. Cox KS, Clair JH, Prokop MT, Sykes KJ, Dubey SA, Shiver JW, et al. DNA gag/adenovirus type 5 (Ad5) gag and Ad5 gag/Ad5 gag vaccines induce distinct T-cell response profiles. J Virol 2008;82:8161-71; PMID: 18524823; http://dx.doi.org/10.1128/JVI.00620-08
32. Kalams SA, Parker S, Jin X, Elizaga M, Metch B, Wang M, et al.; NIAID HIV Vaccine Trials Network. Safety and immunogenicity of an HIV-1 gag DNA vaccine with or without IL-12 and/or IL-15 plasmid cytokine adjuvant in healthy, HIV-1 uninfected adults. PLoS One 2012;7:e29231; PMID:22242162; http://dx.doi.org/10. 1371/journal.pone.0029231
33. Sullivan SM, Doukas J, Hartikka J, Smith L, Rolland A. Vaxfectin: a versatile adjuvant for plasmid DNA-and protein-based vaccines. Expert Opin Drug Deliv 2010;7:1 433-46; PMID:21118032; http://dx.doi.org/10.1517/17425247.2010. 538047
34. Hartikka J, Bozoukova V, Ferrari M, Sukhu L, Enas J, Sawdey M, et al. Vaxfectin enhances the humoral immune response to plasmid DNA-encoded antigens. Vaccine 2001;19:1911-23; PMID: 11228361; http://dx.doi.org/10.1016/S0264-410X (00) 00445-X
35. Locher CP, Witt SA, Ashlock BM, Levy JA. Evaluation of genetic immunization adjuvants to improve the effectiveness of a human immunodeficiency virus type 2 (HIV-2) envelope DNA vaccine. DNA Cell Biol 2004;23:107-10; PMID:15000750; http://dx.doi.org/10.1089/10445490432275 9911
36. Reyes L, Hartikka J, Bozoukova V, Sukhu L, Nishioka W, Singh G, et al. Vaxfectin enhances antigen specific antibody titers and maintains Th1 type immune responses to plasmid DNA immunization. Vaccine 2001;19:3778-86; PMID:11395213; http://dx.doi.org/10.1016/S0264-410X (01) 00090-1
37. Porter KR, Ewing D, Chen L, Wu SJ, Hayes CG, Ferrari M, et al. Immunogenicity and protective efficacy of a vaxfectin-adjuvanted tetravalent dengue DNA vaccine. Vaccine 2012;30:336-41; PMID:22085548; http://dx.doi.org/1 0.1016/j.vaccine.2011.10.085
38. Hartikka J, Bozoukova V, Morrow J, Rusalov D, Shlapobersky M, Wei Q, et al. Preclinical evaluation of the immunogenicity and safety of plasmid DNA-based prophylactic vaccines for human cytomegalovirus. Hum Vaccin Immunother 2012;8:1595-606; PMID:22922766; http://dx.doi.org/10.4161/hv. 21225
39. Veselenak RL, Shlapobersky M, Pyles RB, Wei Q, Sullivan SM, Bourne NA. A Vaxfectin (®)-adjuvanted HSV-2 plasmid DNA vaccine is effective for prophylactic and therapeutic use in the guinea pig model of genital herpes. Vaccine 2012;30:7046-51; PMID:23041125; http://dx.doi.org/10.1016/j.vaccine. 2012.09.057
40. Lin WH, Vilalta A, Adams RJ, Rolland A, Sullivan SM, Griffin DE. Vaxfectin adjuvant improves antibody responses of juvenile rhesus macaques to a DNA vaccine encoding the measles virus hemagglutinin and fusion proteins. J Virol 2013;87:6560-8; PMID:23552419; http://dx.doi.org/10.1128/JVI.00635-13
41. Smith LR, Wloch MK, Ye M, Reyes LR, Boutsaboualoy S, Dunne CE, et al. Phase 1 clinical trials of the safety and immunogenicity of adjuvanted plasmid DNA vaccines encoding influenza A virus H5 hemagglutinin. Vaccine 2010;28:2565-72; PMID:20117262; http://dx.doi.org/10.1016/j.vaccine.2010.01.029
42. Karasawas N, Billings E, Rao M, Williams C, Zolla-Pazner S, Bailer RT, et al.; MOPH TAVEG Collaboration. The Thai Phase III HIV Type 1 Vaccine trial (RV144) regimen induces antibodies that target conserved regions within the V2 loop of gpl20. AIDS Res Hum Retroviruses 2012;28:1444-57; PMID:23035746; http://dx.doi.org/10.1089/aid.2012.0103
43. Rolland M, Edlefsen PT, Larsen BB, Tovanabutra S, Sanders-Buell E, Hertz T, et al. Increased HIV-1 vaccine efficacy against viruses with genetic signatures in Env V2. Nature 2012;490:417-20; PMID:22960785; http://dx.doi.org/10.103 8/nature 11519
44. Haynes BF, Gilbert PB, McElrath MJ, Zolla-Pazner S, Tomaras GD, Alam SM, et al. Immune-correlates analysis of an HIV-1 vaccine efficacy trial. N Engl J Med 2012;366:1275-86; PMID:22475592; http://dx.doi.org/10.1056/NEJMoal 113425
45. Boyer JD, Robinson TM, Kutzler MA, Parkinson R, Calarota SA, Sidhu MK, et al. SIV DNA vaccine coadministered with IL-12 expression plasmid enhances CD8 SIV cellular immune responses in cynomolgus macaques. J Med Primatol 2005;34:262-70; PMID:16128921; http://dx.doi. Org/10.1111/j.1600-0684.2005. 00124.x
46. Schadeck EB, Sidhu M, Egan MA, Chong SY, Piacente P, Masood A, et al. A dose sparing effect by plasmid encoded IL-12 adjuvant on a SIVgag-plasmid DNA vaccine in rhesus macaques. Vaccine 2006;24:4677-87; PMID: 16288822; http://dx.doi.org/10.1016Zj.vaccine.2005.10.035
47. Schadeck EB, Sidhu M, Egan MA, Chong SY, Piacente P, Masood A, et al. A dose sparing effect by plasmid encoded IL-12 adjuvant on a SIVgag-plasmid DNA vaccine in rhesus macaques. Vaccine 2006;24:4677-87; PMID:16288822; http://dx.doi.org/10.1016/j.vaccine.2005.10.035
48. Robinson TM, Sidhu MK, Pavlakis GN, Felber BK, Silvera P, Lewis MG, et al. Macaques co-immunized with SIVgag/pol-HIVenv and IL-12 plasmid have increased cellular responses. J Med Primatol 2007;36:276-84; PMID:17669216; http://dx.doi.org/10.111 l/j.1600-0684.2007.00245.x
49. Halwani R, Boyer JD, Yassine-Diab B, Haddad EK, Robinson TM, Kumar S, et al. Therapeutic vaccination with simian immunodeficiency virus (SIV)-DNA + IL-12 or IL-15 induces distinct CD8 memory subsets in SIV-infected macaques. J Immunol 2008;180:7969-79; PMID:18523260
50. Cristillo AD, Weiss D, Hudacik L, Restrepo S, Galmin L, Suschak J, et al. Persistent antibody and T cell responses induced by HIV-1 DNA vaccine delivered by electroporation. Biochem Biophys Res Commun 2008;366:29-35; PMID:18036339; http://dx.doi.org/10.1016/j.bbrc.2007.11.052
51. Nasioulas G, Zolotukhin AS, Tabernero C, Solomin L, Cunningham CP, Pavlakis GN, et al. Elements distinct from human immunodeficiency virus type 1 splice sites are responsible for the Rev dependence of env mRNA. J Virol 1994;68:2986-93; PMID:8151769
52. Schneider R, Campbell M, Nasioulas G, Felber BK, Pavlakis GN. Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/pro tease and particle formation. J Virol 1997;71:4892-903; PMID:9188551
53. Schwartz S, Campbell M, Nasioulas G, Harrison J, Felber BK, Pavlakis GN. Mutational inactivation of an inhibitory sequence in human immunodeficiency virus type 1 results in Rev-independent gag expression. J Virol 1992;66:7176-82; PMID:1433510
54. Schwartz S, Felber BK, Pavlakis GN. Distinct RNA sequences in the gag region of human immunodeficiency virus type 1 decrease RNA stability and inhibit expression in the absence of Rev protein. J Virol 1992;66:150-9; PMID: 1727477
55. Jalah R, Rosati M, Kulkarni V, Patel V, Bergamaschi C, Valentin A, et al. Efficient systemic expression of bioactive IL-15 in mice upon delivery of optimized DNA expression plasmids. DNA Cell Biol 2007;26:827-40; PMID:17979522; http://dx.doi.org/10.1089/dna.2007.0645
56. Kulkarni V, Rosati M, Bear J, Pilkington GR, Jalah R, Singh AK, et al. Comparison of Intradermal and Intramuscular Delivery of SIV Env DNA by in vivo Electroporation in Macaques, in preparation 2013.
57. Del Prete GQ, Scarlotta M, Newman L, Reid C, Parodi LM, Roser JD, et al. Comparative characterization of transfection-and infection-derived simian immunodeficiency virus challenge stocks for in vivo nonhuman primate studies. J Virol 2013;87:4584-95; PMID:23408608; http://dx.doi.org/10.1128/JVI.03507-12
58. Guan Y, Sajadi MM, Kamin-Lewis R, Fouts TR, Dimitrov A, Zhang Z, et al. Discordant memory B cell and circulating anti-Env antibody responses in HIV-1 infection. Proc Natl Acad Sci U S A 2009;106:3952-7; PMID: 19225108; http://dx.doi.org/10.1073/pnas. 0813392106
59. Moore JP, Wallace LA, Follett EA, McKeating JA. An enzyme-linked immunosorbent assay for antibodies to the envelope glycoproteins of divergent strains of HIV-1. AIDS 1989;3:155-63; PMID:2540772; http://dx.doi.org/10.1097/ 00002030-198903000-00006
60. Montefiori DC. Evaluating neutralizing antibodies against HIV, SIV, and SHIV in luciferase reporter gene assays. Curr Protoc Immunol 2005; Chapter 12:Unit 12 1.
61. Tomaras GD, Yates NL, Liu P, Qin L, Fouda GG, Chavez LL, et al. Initial B-cell responses to transmitted human immunodeficiency virus type 1: virion-binding immunoglobulin M (IgM) and IgG antibodies followed by plasma anti-gp4l antibodies with ineffective control of initial viremia. J Virol 2008;82:12449-63; PMID: 18842730; http://dx.doi.org/10.1128/JVI.01708-08
62. Bolton DL, Song K, Wilson RL, Kozlowski PA, Tomaras GD, Keele BF, et al. Comparison of systemic and mucosal vaccination: impact on intravenous and rectal SIV challenge. Mucosal Immunol 2012;5:41-52; PMID:22031182; http://dx.doi.org/10.1038/mi.2011.45
63. Romano JW, Shurtliff RN, Dobratz E, Gibson A, Hickman K, Markham PD, et al. Quantitative evaluation of simian immunodeficiency virus infection using NASBA technology. J Virol Methods 2000;86:61-70; PMID: 10713377; http://dx.doi.org/10.1016/S0 166-0934 (99) 00184-6