Avances

Avances en
en el
el
Diagnóstico
Diagnóstico yy
Tratamiento
Tratamiento del
del HSV
HSV

Prof. Dr. Omar Lupi

omarlupi@globo.com
 Herpes Dx
• Teste Genético “Herpes Dx” permite detectar os
pacientes com risco de desenvolver herpes
simples recorrente.
• Baseado na detecção de uma mutação
específica no gene MBL (mannose-binding
lectin), importante na resposta imunológica
contra infecções virais recorrentes.
• Se o paciente testar positivo a chance de
desenvolver múltiplas crises é de 80%. O custo
do teste nos EUA é de 250 dólares.
• Não é um teste diagnóstico mas sim prognóstico
 Drogas Antiherpéticas
2000 Cidofovir
Docosanol
Valaciclovir
Resiquimod
1990
Famciclovir Aciclovir
Foscarnet
1980
Cytosine-
arabinosida
1970
Trifluridina

1960 Vidarabina

1950 Idoxuridina
 Aciclovir

Membrana Celular

Timidina
Kinase (TK)

Monofosfato
de Aciclovir

Prêmio Nobel
1988 Trifosfato
de Aciclovir
Análogos dos Nucleosídeos
Mecanismo de Ação

A C A G A T C T G A C T G A C
T G T C T A G A C T G A C T A
C
V
 Clássico: Valaciclovir X Aciclovir
• Arch Dermatol. 1998 Feb;134(2):185-91.

– A randomized, placebo-controlled comparison of oral valacyclovir and acyclovir

in immunocompetent patients with recurrent genital herpes infections. The
Valaciclovir International Study Group.

– Tyring SK, Douglas JM Jr, Corey L, Spruance SL, Esmann J.

– OBJECTIVE: To compare valacyclovir with acyclovir in the treatment of recurrent

genital herpes infection. DESIGN: A multicenter, double-blind, placebo-controlled,
randomized study. PATIENTS: One thousand two hundred patients with recurrent
genital herpes simplex infections. INTERVENTIONS: Patients self-initiated oral therapy
with 1000 mg of valacyclovir hydrochloride twice daily, 200 mg of acyclovir 5 times daily,
or placebo for 5 days. RESULTS: Both drugs were significantly more effective than
placebo in speeding resolution of herpetic episodes (median duration, 4.8, 4.8, and 5.9
days, respectively); the hazards ratios for valacyclovir and acyclovir vs placebo were 1.66
(95% confidence interval [CI], 1.38-2.01) and 1.71 (95% CI, 1.41-2.06) (both P < .001).
Similarly, valacyclovir and acyclovir significantly hastened lesion healing (pain duration
was shorter in valacyclovir- and acyclovir-treated patients (median, 2 vs 3 days). Viral
shedding stopped 2.55 times faster in patients treated with valacyclovir and 2.24 times
faster in patients treated with acyclovir than in patients treated with placebo. Aborted
episodes, in which lesions did not progress beyond the macule or papule stage, tended
to occur in more patients treated with valacyclovir (25.9%) or acyclovir (24.8%) than in
patients treated with placebo (19.8%). The nature, severity, and frequency of adverse
events did not differ among the 3 treatment groups. CONCLUSIONS: Twice-daily
valacyclovir was as effective and well tolerated in the treatment of recurrent genital
herpes simplex virus infection as 5-times-daily acyclovir.
acyclovir
Algumas Vantagens do Valaciclovir
Expert Rev Anti Infect Ther. 2006 Jun;4(3): 367-76.

– Valacyclovir for the treatment of genital herpes.

– Brantley JS, Hicks L, Sra K, Tyring SK.

– Genital herpes is the most prevalent STD infection. While sometimes mild in severity, it can
be a distressing and painful chronic condition. While there is no cure for these conditions,
treatment to alleviate symptoms, suppress recurrences and reduce transmission has been
drastically improved over the past 20 years with the use of guanine nucleoside antivirals,
such as valacyclovir, the highly bioavailable prodrug of acyclovir, and famciclovir, a highly
bioavailable prodrug of penciclovir. Clinical trials involving approximately 10,000 patients
(including patients from nongenital herpes studies, such as herpes zoster) have assessed
the safety and efficacy of valacyclovir in the treatment of initial genital herpes outbreaks,
episodic treatment of recurrent episodes and daily suppressive therapy. It was shown that
valacyclovir has similar efficacy to acyclovir in the episodic and suppressive treatment of
genital herpes. Valacyclovir is the only antiviral drug approved for a once-daily dose
of suppressive therapy for genital herpes,
herpes as well as the only antiviral drug US FDA
approved for a 3-day regimen of episodic treatment of recurrent genital herpes. herpes In
addition, valacyclovir is also indicated in the reduction of the sexual transmission of herpes
simplex virus infection and for the treatment of herpes labialis. In herpes zoster,
valacyclovir is more effective than acyclovir or placebo (and as equally effective as
famciclovir) in shortening the length and severity of herpes zoster-associated pain
and postherpetic neuralgia.
neuralgia Valacyclovir has an acceptable safety profile in patients with
herpes simplex and herpes zoster.
 Famciclovir para Herpes Genital
Recorrente
Clin Infect Dis. 2006 Jan 1;42(1):8-13.

– Single-day, patient-initiated famciclovir therapy for recurrent genital herpes: a

randomized, double-blind, placebo-controlled trial.

– Aoki FY, Tyring S, Diaz-Mitoma F, Gross G, Gao J, Hamed K.

– BACKGROUND: Orally administered antiviral therapy for genital herpes improves the time
to lesion healing and resolves symptoms during an outbreak. Although traditional therapy
for a recurrent episode for healthy adults has consisted of twice-daily dosing for 5 days,
recent studies have indicated that shorter courses of antiviral therapy are effective.
METHODS: This multicenter, randomized, double-blind, placebo-controlled study
compared single-day, patient-initiated oral famciclovir (1000 mg given twice daily) with
placebo for the treatment of recurrent genital herpes. Patients were instructed to initiate
therapy within 6 h after onset of prodromal symptoms or genital herpes lesions. RESULTS:
Famciclovir reduced (P < .001) the time to healing of non-aborted lesions, compared with
placebo. The proportion of patients with aborted lesions was larger in the famciclovir group
than in the placebo group (23.3% vs. 12.7%; P = .003). Adverse events in the famciclovir
group were infrequent overall; most were of mild-to-moderate severity and were similar to
adverse events in the placebo group. CONCLUSIONS: A single-day regimen of patient-
initiated famciclovir treatment was well tolerated and safe,
safe and the healing of recurrent
genital herpes lesions occurred approximately 2 days faster than with placebo. Moreover,
single-day famciclovir treatment stopped the development or progression of lesions beyond
the papule stage.
 DOSES CLÁSSICAS DE
ANTIVIRAIS SISTÊMICOS
HERPES SIMPLES HERPES ZOSTER
 Aciclovir (200mg 5xd)  Aciclovir (400-800mg 5
3-5d(R) e 7-10d(P) x dia) 7-10 dia
 Valaciclovir (500mg  Valaciclovir (500mg-
2xd) 3-5d(R) e 7- 1gr 3 x dia) 7-10 dia
10d(P)  Famciclovir (250mg 3
 Famciclovir (125mg x dia) 7-10 dia
2xd) 3-5d(R) e 7-
10d(P)
 Avaliação por Meta-Análise da
Terapia Supressiva Anti-HSV
J Am Acad Dermatol. 2007 Aug;57(2):238-46.

– A meta-analysis to assess the efficacy of oral antiviral treatment to

prevent genital herpes outbreaks.

– Lebrun-Vignes B, Bouzamondo A, Dupuy A, Guillaume JC, Chosidow O.

– BACKGROUND: Efficacy of oral antiviral therapies, ie, acyclovir, valacyclovir

(VACV), and famciclovir, for suppression of recurrent genital herpes was studied
at different doses and regimens. METHODS: The selected trials were: parallel
randomized clinical trials testing prophylactic oral antiviral treatment of genital
herpes versus placebo in immunocompetent and nonpregnant patients.
RESULTS: Fourteen randomized clinical trials were selected, including a
total of 6158 patients.
patients The global relative risk of developing at least one
recurrence during the study was reduced by 47% (95% confidence interval
45%-49%) in antiviral drug groups compared with the placebo. The best
evaluated regimens, with comparable efficacies, were given twice daily, ie,
acyclovir (400 mg twice day), VACV (250 mg twice daily), and famciclovir (250
mg 2x day), or once day. CONCLUSION: The results of this meta-analysis
confirmed the high clinical efficacy of oral acyclovir, valacyclovir, or
famciclovir for prophylaxis against recurrent genital herpes.herpes
 Atividade Anti-HSV da Lisina
J Inherit Metab Dis. 1998 Apr;21(2):103-11.

– Varicella and varicella immunity in patients with lysinuric protein intolerance.

– Lukkarinen M, Salonen K, Ruuskanen O, Lauteala T, Nuutinen M, Simell O.

– Two patients with lysinuric protein intolerance (LPI) had near-fatal

generalized varicella infection with severe interstitial pneumonitis, hepatitis,
decreased platelet count, bleeding and hypoalbuminaemia. Active haemolysis
resulted in anaemia and massive haemoglobinuria. Serum lactate dehydrogenase
activity and ferritin concentration, which in patients with LPI in normal
circumstances exceed the upper reference values 3-folds to 10-fold, increased to >
10,000 U/L and > 10,000 micrograms/L, respectively. The patients were treated
with fresh frozen plasma, red-cell transfusions and intravenous acyclovir for 14
days, and recovered clinically in a month. Retrospectively, 3 of the 32 other
known Finnish patients with LPI had had varicella infection that had been
more severe than that in the other children in the family or in subjects in the
neighbourhood and had led to hospital admission.
admission We suggest that patients
with LPI who have no varicella zoster antibodies should be treated with acyclovir if
exposed to varicella and should be (re)vaccinated against chickenpox.
 L-Lisina Ajuda a Prevenir
Recorrências do HSV
Altern Med Rev. 2006 Jun;11(2):93-101.

– Natural remedies for Herpes simplex.

– Gaby AR.

– Herpes simplex is a common viral infection of the skin or mucous

membranes. The lesions caused by this infection are often painful,
burning, or pruritic, and tend to recur in most patients. Short-term
treatment with acyclovir can accelerate the healing of an acute outbreak,
and continuous acyclovir therapy is often prescribed for people with
frequent recurrences. While this drug can reduce the recurrence rate by
60-90 percent, it can also cause a wide array of side effects, including
renal failure, hepatitis, and anaphylaxis. Safe and effective alternatives
are therefore needed. There is evidence that certain dietary
modifications and natural substances may be useful for treating
active Herpes simplex lesions or preventing recurrences.
recurrences Treatments
discussed include lysine,
lysine vitamin C, zinc, vitamin E, adenosine
monophosphate, and lemon balm (Melissa officinalis).
 Manejo das Recorrencias do
HSV
• ACV 600 mg/dia • L-Lisine 125 mg/dia
• ACV 400 mg/dia • L-Lisine 250 mg/dia
• ACV 200 mg/dia • L-Lisine 500 mg/dia
• ACV 200 mg seg-sex • L-Lisine 1g/dia
• ACV 200 mg 3 x sem • L-Lisine 1g/dia

Cada etapa do tratamento deve ser continuada por pelo menos 30 dias.

Restrição Alimentar: Arginina – chocolate, sementes e abacaxi. Aumentam a

Disponibilidade de lisina: carne vermelha e laticínios.
 Publicação Original Sobre o
Uso de IFN + ACV
Antiviral Res. 1998 May;38(2):95-106.

– Combined effects of interferon-alpha and acyclovir on herpes simplex virus type 1

DNA polymerase and alkaline DNase.

– Taylor JL, Tom P, O'Brien WJ.

– Treatment of cells with combinations of human interferon-alpha (IFN-alpha) and the

nucleoside analog, acyclovir (ACV), leads to the synergistic inhibition of herpes simplex
virus type 1 (HSV-1) replication. We have examined the effect of these agents on the
replication of HSV-1 DNA and the synthesis of early viral enzymes to understand the
mechanism(s) responsible for this synergistic activity. Combination treatment with 100
IU/ml IFN-alpha and 5 microM ACV led to HSV-1 DNA levels more than 8-fold lower than
in cells treated with ACV alone, while IFN-alpha treatment alone had no detectable effect
on viral DNA synthesis. Steady state levels of DNA polymerase were reduced
approximately 50% by IFN-alpha and 25% by ACV, but combination treatment did not
decrease enzyme levels to an extent greater than the sum of these effects. In contrast, the
activity of another early viral enzyme, alkaline DNase, was reduced less than 20% by IFN-
alpha alone or combination treatment and was unaffected by ACV treatment. No decrease
in the level of mRNA encoding either enzyme was detected in IFN-alpha-treated cells
although ACV treatment reduced polymerase mRNA levels. These studies suggest that
the synergistic anti-HSV activities of IFN-alpha with ACV could lead to the reduction
in production of viral early enzymes,
enzymes especially DNA polymerase.
 Ação Sinérgica Anti-HSV
do IFN2 + ACV
• Chem Biol Drug Des. 2007 Jun;69(6):429-34.

– Cell metabolism of acyclovir phosphonate derivatives and

antiherpesvirus activity of their combinations with alpha2-
interferon.
– Skoblov YS, Karpenko IL, Jasko MV, Kukhanova MK,
Andronova VL, Galegov GA, Sidorov GV, Myasoedov NF.

– The combinational use of acyclovir (ACV) phosphonate

esters and alpha(2)-interferon was shown to produce a
synergistic effect on inhibition of HSV-1 replication in Vero
cell cultures. Unlike other acyclovir phosphonate derivatives
studied earlier, ACV H-phosphonate is not an ACV prodrug. On
penetrating into the cells, it may be directly converted into ACV
monophosphate escaping dephosphonylation-phosphorylation
steps.
Novas Drogas
 Inibidores da Primase-
Helicase do HSV
 BAY 57-1293 is a member of the thiazolylsulfonamides, a recently
discovered class of nonnucleosidic compounds with potent
antiherpetic activity in vitro and in vivo, based on a novel
mechanism of action. It inhibited the replication of HSV.
HSV BAY 57-
1293 targets the viral primase-helicase complex and inhibits its
ATPase activity.
 BAY 57-1293 is also active against acyclovir-resistant mutant
strains which carry mutations in the tk or DNA pol genes. The
compound showed favorable pharmacokinetics in all species
investigated (mouse, rat, and dog), with an oral bioavailability of
>60% and an elimination half-life of >6 h.
 Reading Suggestion: Kleymann G. Helicase primase: targeting the
Achilles heel of HSV. Antivir Chem Chemother 2004;15 (3): 135-40.
 Inibidores da Helicase Primase
• Antiviral Res. 2007 Jul;75(1):30-5.

– The helicase primase inhibitor, BAY 57-1293 shows potent therapeutic antiviral
activity superior to famciclovir in BALB/c mice infected with herpes simplex virus
type 1.

– Biswas S, Jennens L, Field HJ.

– BAY 57-1293 represents a new class of potent inhibitors of herpes simplex virus (HSV) that
target the virus helicase primase complex. The present study was conducted using the
zosteriform infection model in BALB/c mice. The helicase primase inhibitor, BAY 57-1293
was shown to be highly efficacious in this model. The beneficial effects of therapy were
obtained rapidly (within 2 days) although the onset of treatment was delayed for 1 day after
virus inoculation. The compound given orally, or intraperitoneally once per day at a dose of
15 mg/kg for 4 successive days was equally effective or superior to a much higher dose of
famciclovir (1mg/ml, i.e. approximately 140-200mg/kg/day) given in the drinking water for 7
consecutive days, which, in our hands, is the most effective method for administering
famciclovir to mice. In contrast to the vehicle-treated infected mice, all mice that received
antiviral therapy looked normal and active with no mortality, no detectable loss of weight and
no marked change in ear thickness. BAY 57-1293 and famciclovir reduced the virus titers in
the skin to below the level of detection by days 3 and 7 post infection, respectively. In both
BAY 57-1293 and famciclovir-treated mice, infectious virus titers in the ear pinna and
brainstem remained below the level of detection. Consistent with these findings, BAY 57-
1293 also showed a potent antiviral effect in an experiment involving a small number
of severely immunocompromised athymic-nude BALB/c mice.
 Inibidores da Helicase Primase
Antivir Chem Chemother. 2007;18(1):35-48.
– Superior efficacy of helicase-primase inhibitor BAY 57-1293 for herpes
infection and latency in the guinea pig model of human genital herpes
disease.

– Baumeister J, Fischer R, Eckenberg P, Henninger K,

Ruebsamen-Waigmann H, Kleymann G.

– The efficacy of BAY 57-1293, a novel non-nucleosidic inhibitor of herpes simplex

virus 1 and 2 (HSV-1 and HSV-2), bovine herpesvirus and pseudorabies virus, was
studied in the guinea pig model of genital herpes in comparison with the licensed
drug valaciclovir (Valtrex). Early therapy with BAY 57-1293 almost completely
suppressed the symptoms of acute HSV-2 infection, and reduced virus shedding
and viral load in the sacral dorsal root ganglia by up to three orders of magnitude,
resulting in decreased latency and a greatly diminished frequency of subsequent
recurrent episodes. In contrast, valaciclovir showed only moderate effects in this
set of experiments. Compared with valaciclovir, BAY 57-1293 halved the time
necessary for complete healing. Moreover, the onset of action was fast, so that
only very few animals developed new lesions after treatment commenced. Finally,
in a study addressing the treatment of recurrent disease in animals whose primary
infection had remained untreated BAY 57-1293 was efficient in suppressing the
episodes. In summary, superior potency and efficacy of BAY 57-1293 over
standard treatment with valaciclovir was demonstrated in relevant animal
models of human genital herpes disease in terms of abrogating an HSV infection,
reducing latency and the frequency of subsequent recurrences.
VACINAS ANTI-HERPÉTICAS:
MITO OU REALIDADE?

Prof. Dr. Omar Lupi

 CMV SV40

plasmídeo
Escherichia coli
A Engenharia Genética Deve Muito
aos Vírus Bacteriófagos
 Plasmídeo
pcDNA3.1/HisA,B,C

PCMV T7His6EK BGH f1 SV40 N SV40 ColE1 Amp

pA ori ori pA
PCMV= Permite a expressão em qualquer célula de mamíferos
T7= Fornece a orientação correta para a transcrição do plasmídeo
His6= Permitirá a purificação da proteína recombinante com Nq++
EK=Permite remover a etiqueta de polihistidina c/ enteroquinase
BGHpA=Transcrição eficiente do RNAm
F1ori = Permitirá o resgate do DNA de hélice simples
SV40ori-N-SV40pA=transcrição do gene p/ resistência neomicina
ColE1 = É replicon que permitirá replicação eficaz na E.coli
Amp = Gene que codifica resistência à β-lactamase na E.coli
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Sobrevida à infecção Herpética
Vaginal

Nasal

Pele

Cerebral

0 10 20 30 45
Período de Acompanhamento
(dias)
 Desenho do Estudo
• O protocolo envolveu 284 pacientes HIV e HSV-2 negativos, checados
por Western-blot (WB).
• Os pacientes foram contactados por rádio e jornais após aprovação
do Comitê de Ética da University of Texas.
• Receberam imunização de forma randomizada e duplo-cega: Vacina
(gB2-gC2-gD2+ MF59) ou Placebo (tampão citrato + MF59).
• Imunização IM no início do acompanhamento e reforços c/ 1 e 3 meses
(30µg de cada glicoptna). Tempo total de acompanhamento de 9
meses, reavaliados a cada 3 sem.
• Soroconversão analisada através da mensuração de Ac neutralizantes
para o HSV-2 (Reação de microneutralização dependente de
complemento) e de Ac ligantes para gB-C-D do HSV-2 (ELISA).
• Cultura viral foi realizada nos casos clínicos suspeitos.
Vacinação Antiherpética
Resposta Imune Celular e
Humoral
 Títulos de Acs Neutralizadores
Mulheres
vacinadas
Mulheres
80 Placebo
Homens
70 vacinados
60 Homens
50 placebo
40
Homens placebo
30
Homens vacinados
20
Mulheres Placebo
10
Mulheres vacinadas
0
1Mes 2Mes 3Mes 4Mes
Situação Atual – Vacinas anti-HSV
A vacina é muito imunogê-nica
e bem tolerada. A tec-nologia
envolvida é segura e bem
dominada.
A excelente resposta no sexo
feminino parece dever-se a
fisiopatogenia própria do HSV-
2 nas mulheres.
Uma vacinação antiherpéti-ca
eficaz deverá ser feita, com
pelo menos, 3 doses.
Há proteção cruzada para
herpes labial.
A vacina parece eficaz no
controle das crises mas não
na proteção contra novos
casos de herpes genital.
A via IM pode não ser a
mais adequada. Imunização
via mucosa (intra-nasal)
pode ser mais efetiva.
Imunidade humoral parece
não ser suficiente, isolada-
mente, para conferir
imunidade ao HSV-2.
 Update em Vacina Anti-HSV
• Curr Opin Investig Drugs. 2006 Feb;7(2):136-41.

– Vaccines for herpes simplex virus infections.

– Koelle DM.

– Infections with herpes simplex virus (HSV) type 1 (HSV-1) and type 2
(HSV-2) can have serious medical consequences. Although antiviral
medications can suppress symptomatic disease, asymptomatic
shedding and transmission, they neither cure nor alter the natural
history of HSV infections. Manipulation of the immune response is one
potential method to decrease disease burden. Current research on
prophylactic and therapeutic vaccination approaches is discussed in this
review, with a focus on compounds that have entered clinical trials or
that display novel compositions or proposed mechanisms of action. One
such vaccine is an alum and monophosphoryl lipid A-adjuvanted subunit
glycoprotein D2 vaccine that has demonstrated activity in the
prevention of HSV-2 infection and disease in HSV-uninfected
women in a phase III clinical trial. Further confirmatory clinical trials of
this vaccine are currently underway. Other vaccine formats also in
development include attenuated live or replication-incompetent HSV-2
strains and technologies that target virus-specific CD8 T-cell responses.
 Update em Vacina Anti-HSV
• Hum Vaccin. 2007 Jun 3;3(6).

– Effectiveness and Efficiency of Imperfect Therapeutic HSV-2 Vaccines.

– Schwartz EJ, Bodine EN, Blower S.

– Background: Efforts are currently underway to develop therapeutic vaccines for Herpes
Simplex Virus type 2 (HSV-2). Methods: We use a mathematical model to predict the
potential public health impact of imperfect, therapeutic HSV-2 vaccines. We evaluate
vaccine effectiveness and efficiency for the general population in the United States where
HSV-2 prevalence is currently 22%. We assume that therapeutic vaccines will produce two
therapeutic benefits in vaccinated infected-individuals: (1) the rate of viral reactivation will
decrease (hence infected-individuals will experience fewer viral shedding episodes), and (2)
the average length of the viral shedding episodes will be shortened. In addition, we assume
that therapeutic vaccines will benefit uninfected individuals by reducing viral shedding in
(and hence transmission from) vaccinated infected-individuals. Results: Our predictions
show that therapeutic vaccines could substantially reduce HSV-2 epidemics by
reducing new infections by 77% and preventing 0.84 new infections for each
vaccinated individual. These vaccines could prevent 212,600 (median; IQR, 156,064-
288,558) new infections after only one year. We show that increased effectiveness and
efficiency are more strongly correlated with a vaccine-induced reduction in transmission
probability than with either of the two therapeutic benefits that accrue directly to the infected
individuals (specifically, the reduction in episode length and number of episodes).
Conclusions: We suggest that current vaccine development efforts target mechanisms that
reduce viral shedding (thereby reducing transmission) thus providing both a beneficial
therapeutic and a beneficial epidemic-level impact. Our results also demonstrate that
therapeutic vaccines would be substantially more useful than prophylactic vaccines for
epidemic control.
HERPESVIRUS, CÉLULAS-TRONCO E
TERAPIA GENÉTICA
 Células-Tronco

Cultura células-tronco

skin liver neurons heart eyes etc

feto
 HSV como Vetor de Engenharia
Genética em Dçs Cardíacas
Cell Transplant. 2006;15(1):67-74.

– Engineered cell therapy for sustained local myocardial delivery of nonsecreted proteins.

– Bian J, Kiedrowski M, Mal N, Forudi F, Penn MS.

– Novel strategies for the treatment of congestive heart failure have taken the form of gene and cell
therapy to induce angiogenesis,
angiogenesis optimize calcium handling by cardiac myocytes, or regenerate
damaged myocardial tissue. Arguably both gene- and cell-based therapies would be benefited by having
the ability to locally deliver specific transcription factors and other usually nonsecreted proteins to cells in
the surrounding myocardial tissue. The herpes simplex virus type 1 (HSV-1) has been shown to
mediate protein intercellular trafficking to mammalian cells and finally localize into the nucleus,
which makes it a useful cargo-carrying functional protein in cell-based gene therapy. While VP22
has been studied as a means to modulate tumor growth, little is known about the distribution and
transport kinetics of VP22 in the heart and its potential application in combination with autologous cell
transplantation for the delivery of proteins to myocardial tissue. The aim of this study was to evaluate the
efficacy of VP22 fusion protein intercellular trafficking combined with autologous cell transplantation in
the heart. In an in vitro study untransfected rat heart cells were cocultured with stably transfected rat
cardiac fibroblasts (RCF) with fusion constructs of VP22. The control experiment was untransfected rat
heart cells co-plated with RCF stably transfected with enhanced green fluorescence protein (eGFP). The
Lewis rat model was selected for in vivo study. In the in vitro studies there was a 14-fold increase in the
number of GFP-positive cells 48 h after initiating coculture with VP22-eGFP RCF compared to eGFP
RCF. The VP22-eGFP area consisted of eGFP-positive endothelium, smooth muscle cells, and cardiac
myocytes with delivery to an area of approximately 1 mm2 of myocardial tissue. Our data suggest a
viable strategy for the delivery of proteins that are not naturally secreted or internalized, and provide the
first insight into the feasibility and effectiveness of cell-penetrating proteins combined with cell
transplantation in the heart.
Benthocodon pedunculata

Aequorea vitoria

Green Fluorescent
protein (GFP)
 Infecçáo Experimental com
Trypanosoma cruzi = Dç Chagas
Dç Chagas
Trypanosoma cruzi cronica CT fluorescentes
modificdas pelo
HSV

BALB/c mice
C-T Modificadas pelo HSV Detectadas no
Tecido Cardíaco (15 dias)

Anti GFP + Anti Myosine

 Reduçáo da Miocardite em Ratos
Tratados com C-T modificadas pelo HSV

INFLAMACAO FIBROSE
500
50
Inflammatory cells/mm2

400

Fibrotic area/mm2
40
300 Saline Saline
30
SC SC
200 20

100 10

0 0
0 2 4 6 8 0 2 4 6 8
Months after Treatment Months After Treatment
 HSV Auxilia Tratamento de Gliomas
Malignos Cerebrais sem Cirurgia
• Mol Ther. 2007 Jul;15(7):1373-81.

– Bystander Killing of Malignant Glioma by Bone Marrow-derived Tumor-Infiltrating

Progenitor Cells Expressing a Suicide Gene.
– Miletic H, Fischer Y, Litwak S, Giroglou T, Waerzeggers Y, Winkeler A, Li H,
Himmelreich U, Lange C, Stenzel W, Deckert M, Neumann H, Jacobs AH, von Laer
D.
– Adult stem cells are promising cellular vehicles for therapy of malignant gliomas as they
have the ability to migrate into these tumors and even track infiltrating tumor cells.
However, their clinical use is limited by a low passaging capacity that impedes large-
scale production. In the present study, a bone marrow-derived, highly proliferative
subpopulation of mesenchymal stem cells (MSCs)-here termed bone marrow-derived
tumor-infiltrating cells (BM-TICs)-was genetically modified for the treatment of malignant
glioma. Upon injection into the tumor or the vicinity of the tumor, BM-TICs infiltrated solid
parts as well as the border of rat 9L glioma. After intra-tumoral injection, BM-TICs
expressing the thymidine kinase of herpes simplex virus (HSV-tk) and enhanced green
fluorescent protein (BM-TIC-tk-GFP) were detected by non-invasive positron emission
tomography (PET) using the tracer 9-[4-[(18)F]fluoro-3-hydroxymethyl)butyl]guanine
([(18)F]FHBG). A therapeutic effect was demonstrated in vitro and in vivo by BM-TICs
expressing HSV-tk through bystander-mediated glioma cell killing. Therapeutic efficacy
was monitored by PET as well as by magnetic resonance imaging (MRI) and strongly
correlated with histological analysis. In conclusion, BM-TICs expressing a suicide gene
were highly effective in the treatment of malignant glioma in a rat model and
therefore hold great potential for the therapy of malignant brain tumors in humans.
TERAPIA GENÉTICA COM HSV

HSV

ACV/GCV
 HSV Auxilia C-T a Prevenir AVC
• J Cell Physiol. 2007 Nov;213(2):341-7.

– Adult mesenchymal stem cells for tissue engineering versus regenerative medicine.

– Caplan AI.

– Adult mesenchymal stem cells (MSCs) can be isolated from bone marrow or marrow aspirates
and because they are culture-dish adherent, they can be expanded in culture while maintaining
their multipotency. The MSCs have been used in preclinical models for tissue engineering of
bone, cartilage, muscle, marrow stroma, tendon, fat, and other connective tissues. These
tissue-engineered materials show considerable promise for use in rebuilding damaged or
diseased mesenchymal tissues. Unanticipated is the realization that the MSCs secrete a large
spectrum of bioactive molecules. These molecules are immunosuppressive, especially for T-
cells and, thus, allogeneic MSCs can be considered for therapeutic use. In this context, the
secreted bioactive molecules provide a regenerative microenvironment for a variety of injured
adult tissues to limit the area of damage and to mount a self-regulated regenerative response.
This regenerative microenvironment is referred to as trophic activity and, therefore, MSCs
appear to be valuable mediators for tissue repair and regeneration. The natural titers of MSCs
that are drawn to sites of tissue injury can be augmented by allogeneic MSCs delivered via the
bloodstream. Indeed, human clinical trials are now under way to use allogeneic MSCs for
treatment of myocardial infarcts, graft-versus-host disease, Crohn's Disease, cartilage and
meniscus repair, stroke, and spinal cord injury. This review summarizes the biological basis for
the in vivo functioning of MSCs through development and aging.
DESENHO DO ESTUDO 1
DESENHO DO ESTUDO 2
Comparando a inteligencia dos animais tratados com C-T
com aqueles tratados com placebo
• Peso corporal
• Consumo de alimento e água