Embed Size (px)
Citation preview
1
Dr. Susana Garcia de ArribaScientific & Medical Expert - Regulatory Affairs
Track 1: Toxicology Approaches
Human Safety of free Hydroquinone derived from Herbal medicinal products
containing Uvae ursi folium
2
1. Herbal Medicinal Product:
• HMPC – Monograph
2. Arctostaphylos uva-ursi folium
3. Pharmacokinetics – Elimination
4. Human Exposure to Free HQ
5. Exposure limits for use in risk identification
Permitted Daily Exposure of free HQ
6. Assessment Genotoxic Potential of Free HQ
• AMES
• Micronucleous in mice
7. Safety Data – Cystinol akut
8. Conclusions
Content
3
National markets for Herbal medicines
annual sales per capita consumption
Germany -- $2.2 billion --- $36.55 million
Denmark -- $ 30.2 million -- $ 26.6 million
The Global Herbal Medicine Market is estimated* to be US$ 60 billion, which is poised to grow to $5 trillon by the year 2050
EUJapan
Asia + Australia
USA
Rest EU
Others
*World Health Organization (WHO) / Secretariat of the convention on Biological Diversity (2008)
28 US$
11 US$
10 US$
7 US$
(US$ billion)
Increase in herbal medicine sales 10 % per Year
Herbal Medicine Market
Global Herbal Medicine Market
4
1. Herbal Medicinal Products (HMPs)
*HMPs are defined as any medicinal product, exclusively containing as active ingredients one or more herbal substances or one or more herbal
preparations, or one or more such herbal substances in combination with one or more such herbal preparations.
HERBAL SUBSTANCE (HS) - dry /fresh: whole or parts of plants, algae, fungi and lichens. HERBAL PREPARATION (HP)- obtained by subjecting HS to treatments such as extraction, distillation,expression, fractionation, purification, concentrationor fermentation.
• Require a Marketing Authorization or Registration before placement on the market In the EU community
• Drug Law on pharmaceutical products for human use - - -> *Directive 2001/83/EC applies to HMP
Protect public health Harmonized authorization across the EU Facilitate the authorization/ registration
procedure of HMP
5
http://ec.europa.eu/health/human-use/herbal-medicines/index_en.htmhttp://www.ema.europa.eu/ema/index.jsp?curl=pages/regulation/general/general_content_000208.jsp&mid=WC0b01ac05800240cf
1.1 Market access for HMP in the EU
under European medicines legislation, HMPs must fall within one of the following 3 categories to reach the market:
1. Traditional use registration (simplified registration procedure) Directive 2004/24/EC amending Directive 2001/83/EC.
- HMP with sufficient evidence of medical use >30 years, >15 years within EU. Sufficient safety data and plausible efficacy. THMP with Traditional medicinal use based upon long-standing use.
2. Marketing authorisation (Bibliographic application) HMP with well-established medicinal use on basis of recognized efficacy, at least one controlled clinical study of good quality is required to substantiate efficacy, and an acceptable level of safety.
3. Marketing authorisation – (full/ mixed application) a product with safety and efficacy data from the company’s own development (‘stand alone’) or a combination of own studies and bibliographic data (‘mixed application’). As a result the product is granted a marketing authorisation.
1. Herbal Medicinal Products
6http://ec.europa.eu/health/human-use/herbal-medicines/index_en.htm
EMA
Med
ical
Pro
duct
s hu
man
use
Com
mitt
ee o
n H
erba
l Med
icin
al P
rodu
cts
HM
PC
HM
PC M
onog
raph
Regulation (EC) No 726/2004 and Directive 2004/24/EC
Responsible for Agency's opinions on herbal medicines Experts in the field of herbal medicines. Advisers28 EU Member States (+ Iceland and Norway)Scientific evaluation of non-clinical and clinical data; safety and efficacy data; quality of the herbal substance /preparations intended for medicinal use and license Evaluation of long-standing medicinal use in the EU
HS/HP complies qualitative - quantitative declaration, therapeutic indication and daily recommended dose
efficacy and safety of the HS or HP are ensured
• Marketing Autorisation• Registration
Sept. 2004
Euro
pean
Med
ical
Age
ncy
1.2 EMA, HMPC and HMPs
1. Herbal Medicinal Products
Listi
ng!!
!*
*list of herbal substances, preparations and combinations thereof for use in traditional herbal medicinal products
7
http://ec.europa.eu/health/human-use/herbal-medicines/index_en.htmhttp://www.ema.europa.eu/ema/index.jsp?curl=pages/regulation/general/general_content_000208.jsp&mid=WC0b01ac05800240cf
All HMPs in the EU market have to fulfill the demands of HMPC and competent national authorities
EU legislation on pharmaceutical products for human use
Pharmaceutical quality – Eur. Pharmacopoeia/NfG
Standardized production process: GMP (GDP, GACP)
Drug Safety: Preclinical: Genotoxic/Mutagenic potential (GLP) Pharmacovigilance (GVP)
Proved Well Established/ Traditional long-standing medicinal use in the EU
1.3 HMPs in the EU market
1. Herbal Medicinal Product
8 http://www.ema.europa.eu/ema/index.jsp?curl=pages/regulation/general/general_content_000208.jsp&mid=WC0b01ac05800240cf
1.4 EMA - HMPC 1. Herbal Medicinal Product
9http://www.ema.europa.eu/docs/en_GB/document_library/Other/2009/12/WC500017724.pdf
1.5 Committee on Herbal Medicinal Products
1. Herbal Medicinal Product
10 http://www.ema.europa.eu/ema/index.jsp?curl=pages/medicines/landing/herbal_medicines_search_landing_page.jsp&mid=
1.6 HMPC Monograph
1. Herbal Medicinal Product
Latin name of the genus Arctostaphylos
Latin name of herbal substance Uvae ursi folium
Botanical name of plant Arctostaphylos uva-ursi (L.) Spreng.English common name of herbal substance Bearberry Leaf
Status F: Assessment finalisedDate added to the inventory 23/11/2005Date added to priority list 23/11/2005Outcome of European assessment Community herbal monograph
11
1.6 HMPC Monograph
1. Herbal Medicinal Product
Final Assessment
report
Final community
herbal monograph
12
1.6 HMPC Monograph
1. Herbal Medicinal Product
13
1.7 Assessment Report - HMPC Monograph
1. Herbal Medicinal Product
14
• HMP -
• Herbal Preparation ethanolic (60% v/v) dry extract of
Arctostaphylos uva-ursi folium
• Composition: 1 Tablet
238.7-297.5 mg ethanolic (60% v/v) dry extract Arctostaphylos uva-ursi folium (DER 3.5 – 5.5:1)
corresponding to 70 mg arbutin anhydrous
• Posology adults and children over 12 years
Take 2 coated tablets 3 times daily
• Recommended Daily Dose (RDD):
≈ 1.6 g Extract = 420 mg arbutin
• Traditionally used to treat symptoms of lower urinary tract infections
2. Arctostaphylos uva-ursi folium (Cystinol® akut)
Cystinol® akut
15
COMPLIANT with the qualitative and quantitative composition, posology as well as with the therapeutic area of
application
2.1
16
Information about HMP on the market in the Member States - Regulatory status overview
Country Date – Regulatory status
Formulation 1 Dose Daily Dose, RDD
Powdered herbal substance in solid dosage forms
Germany >1976 MA coated tablets 500 mg 6 tablets 4 times - 12 g
Spain >1991 R capsule 270 mg 1 capsule 3 times - 810 mg
France >1982 R hard capsules 350 mg 2 capsules 2 times - 1.4 g
Germany, Poland, Spain, Estonia, Slovenia
R/MA Herbal Tea 1.5-3 g/150ml 3 - 6 times - 4.5 - 18 g
Herbal preparations - Dry extracts
Germany > 1976 MA (39 y) coated tablets 238.7 – 297.5 mg 2 tablets 3 times - 1.6 g
Poland > 2000 R film-coated tablets
215 mg of 4 tablets 3 times - 2.6 g
France > 1992 R hard capsules 200 mg 1 capsule 2 times - 400 mg
MA - Marketing authorisationR - Registration
http://www.ema.europa.eu/docs/en_GB/document_library/Herbal_-_HMPC_assessment_report/2011/07/WC500108750.pdf
2.1 HMPC monograph Arctostaphylos uva-ursi folium
17
1. high pharmaceutical quality2. widely documented drug safety3. Medicinal use proven by HMPC /National-International Monographs
Quality of the extract
„Good LaboratoryPractice“ GLP No systemic toxic
effects
Medicinal use Therapeutical use
Safety Data - preclinical -
Safety Data - Human -
Medicinal use Therapeutical experience
Ethanolic Uvae ursi folium extract
GMP, GDP, GACP
Eur. Pharmacopoeia
2.2 Cystinol® akut - HMP Documentation -
Quality, DER, HPHMPC Monograph
No genotoxic-mutagenic Risk
Pharmacovigilance Data
HMPC monographWHO, ESCOP, Comm. E
Monographs
18
National and International herbal monographs on Uvae ursi folium
Commission E, 1994; WHO, 20021; ESCOP2, 2012 and HMPC, 20123
HMPC Monograph on Arctostaphylos uva-ursi (L.) Spreng, folium
There is sufficient evidence of medicinal use throughout a period of at least 30 years, including at least 15 years in European Community
Uvae ursi folium extract - Cystinol® akut
long-standing medicinal use and experience testified by bibliographic and expert evidence
1World Health Organization. Folium Uvae ursi. Monografie 2 (2002): 342-3512European Scientific Cooperative on Phytotherapy (ESCOP) Uvae ursi folium, bearberry leaf. (2012), online series. www.escop.com3HMPC Community herbal monograph on Arctostaphylos uva-ursi (L.) Spreng., folium (28.02.2012) http://www.ema.europa.eu/docs/en_GB/document_library/Herbal_-_Community_herbal_monograph/2011/04/WC500105350.pdf
2.3 The therapeutic efficacy and safety
19
Commission E1994
WHO (World Health Organisation)
2002
ESCOP
(European Scientific Cooperative on
Phytotherapy) 2012
HMPC
(Herbal Medicinal Products Committee
of EMA) 2012Indication Inflammatory
diseases of the lower urinary tract
Moderate inflammatory conditions of the urinary tract and bladder, such as cystitis, urethritis and dysuria
Uncomplicated infections of the lower urinary tract, when antibiotic treatment is not considered essential
Treatment of symptoms of mild recurrent lower urinary tract infections (traditional use)
Dosage 3 g drug or 400 - 840 mg hydroquinone derivates, calculated as arbutin, up to 4x daily
3 g drug or 400 - 850 mg hydroquinone derivates, calculated as arbutin, up to 4x daily
400-800 mg of arbutin/day,equivalent preparations
Single dose corresponding to 100-210 mg hydroquinone derivates calculated as anhydrous arbutin, 2-4x daily
Duration of use
Without medical advice: up to 5x per year, not longer than 1 week in each case
No long-term use, patients with persistent symptoms should consult a physician
Until complete disappearance of symptoms (up to a maximum of 1 week)
Not longer than 1 week
Contra-indictions
Pregnancy, lactation, children < 12 years
Pregnancy, lactation, children, patients with kidney disorders
Pregnancy, lactation, children < 12 years
Hypersensitivity, kidney disorders
Interactions
Urinary acidifying substances*
Urinary acidifying substances
None reported None reported
Side effects
Nausea and vomiting possible in persons with a sensitive stomach
Nausea and vomiting may occur due to stomach irritation from the high tannin content
Nausea and vomiting may occur due to stomach irritation from the high tannin content
Nausea, vomiting, stomach-ache
2.4 International /national Monographs
Þ Positive Benefit-Risk Profileofficially approved
20
Extract = active principle
Hydroquinon (HQ) derivatives: arbutin to 23.5 - 29.3% free HQ < 0.3%
Phenolic acidsgallic acid, Ellagic acid
Polyphenols (tannins) gallotannins, ellagitannins
Flavonoids hyperoside
Triterpenesursolic acid, uvaeol
Arbutin HQ
Gallic acid Ellagic acid
2.5 Relevant Constituents : Qualitative -Quantitative
21Siegers et al. 1997
3. Pharmacokinetics
• 12 healthy volunteers (6 males - 6 females)• Dose 3 x 2 Tablets Cystinol® akut
1.6 g Extract/day
420 mg Arbutin (169 mg HQ)
• Urine samples collected after 36h
• Free and Total HQ quantified in individual urine samples – HPLC
STUDY 1 - Pharmacokinetics
Elimination
Free HQ hydroquinoneTotal HQ = Free HQ + conjugated HQ
22
3.1 Single Data Elimination
Individual urine samples n=12
n=11 Free HQ
(mg)To
tal H
Q (m
g)HMP intake
Free
HQ
(mg)
Time (hours)
Subject 3 had an extreme high level of free HQ before HMP intake.
Total HQ = Free HQ + HQ-Conjugate
Elimination of Free HQ and Total HQ Time-dependent and gender independent
6/12 eliminate Free HQ3/6 high levels free HQ at t=0High Free HQ elimination no related To increase of Metabolism/elimination
0.99
109.4
23
N= 11
• 0.6% Arbutin Dose as Free HQ
= 0.99 mg Free HQ
• 70% Arbutin Dose as conjugate HQ to glucuronic and sulfuronic acid
= 109.4 mg total HQ
3.2 Quantitative Elimination of Free and Total HQ
• Mean values ± SD• Parallel and progressive elimination
STUDY 1 - RESULTS
Time (h)
24
Pharmacokinetic profile HQ; HydroquinoneHQ-Glu: HQ-glucuronide; HQ-Sulf: HQ-sulphate
3.3 Metabolism - Elimination
Major elimination:HQ-glucuronide >> HQ-sulphate (≈ 70% of total conj.)
0.6% Free HQ
70% Total HQ
25
3.4 Comparison with Bibliographic Data
Subjects, urine collection (h)
ArbutinDaily Dose
% Arbutin Dose
Free HQ Total HQ
Siegers et al. 1997 n=11, 36 420 mg 0.6% 70%
Paper et al. 1993 n=5, 24 100 mg 0 85%
Schindler et al. 2002 n=16, 36 210 mg 0.1% 66.7%
Quintus et al. 2005 n=3, 24 150 mg 0.6% 75%
* Detection limit (1µg/ml)Arbutin MW 272.25HQ MW 110.11
26
20fold
0.6% HQ
>70%
3.5 Bacterial deconjugation
• 4 healthy volunteers (2 males - 2 females)• Dose 3 x 2 Tablets Cystinol® akut
420 mg Arbutin
• Urine samples collected after 24h
• Free and Total HQ quantified
STUDY 2 – Bacterial deconjugation
24h 37°C
100 µl/ml Percloric acid
HPLC quantification
HQ; HydroquinoneHQ-Glu: HQ-glucuronide; HQ-Sulf: HQ-sulphate
Siegers et al. 2003; Garcia et al. 2010; Garcia et al. 2013
27
3.6 Summary Elimination /deconjugation
Garcia et al. Int J Toxicol. 2013
The elimination of arbutin metabolites after oral intake of 420 mg arbutin Only 0.6 % of the administered arbutin-dose was excreted as free HQ in the urine
In 6 out of 12 volunteers free HQ was not detected
70% of the administered arbutin-dose was eliminated as HQ-conjugates (HQ-glucuronide; HQ-sulphate)
Free HQ deconjugation/accumulation inside of the bacteria Uropathogenic bacteria such as E. coli have the ability to deconjugate the HQ-
conjugates eliminated in the human urine to a high extent
20fold higher amount of free HQ was detected in bacteria sediments as compared to the supernatant
HQ-conjugates found in urine are ingested by the uropathogenic bacteria and transformed into free HQ inside of the bacteria
Intracellularly formed free HQ is responsible for the bactericide effect
28
0.99 mg Free HQ
420 mg Arbutin
3.7 Summary Elimination process
Recovered in Urine
29
• Elimination of Free HQ in 36 h: 0.99 mg
• Level of Free HQ in urine after 36 h: 0.6 – 0.5 µg/ml
• Corresponding to Human exposure level:11 µg/Kg b.w. day *
* 60 kg body weight
4. Human exposure to Free HQ
*Garcia et al. Int J Toxicol. 2013
30
4.1 Food sources of Free HQ
Product Arbutin(µg/kg)
Free Hydroquinone (HQ)
Per day LEVEL OF EXPOSUREµg free HQ/kg b.w./day*
Coffee and Tea 20 - 90 µg /glass1 2-3 glass/day
0.66 - 4.5
Red wine 562 µg / glass1,2 1 glass/ day 9.3
non-filter cigarettes
110 - 300 µg /cigar.3 18 cigar./day**
33 - 90
Wheat products 0.015 - 0.13
0.2 µg/g1 250 g/day 0.833
Broccoli 0.14 µg/g1 250 g/day 0.6
Pears 0.037 - 0.18
0.02 - 0.05 µg/g1 100 g/day 0.033 - 0.083
High-HQ diet(n=4)
4 mg/kg1 125 -2751
Cystinol RDD 420 mg/day
0.99 mg in urine5 11
*Body weight of 60 kg; 1Deisinger et al. 1996; 2IPCS, 1996; 3NTP, 2009; 4Carlson and Brewer (1953); 5Siegers et al. 1997. **Mean value in EU.
These foods are consumed several
times a day and for a lifetime
without causing AE or safety
concerns
Garcia et al. Int J Toxicol. 2013
31
5. Exposure limits for risk identification
Estimates the dose below which there is a negligible risk to human health
*- Guideline on setting health based exposure limits for use in risk identification in the manufacture of different medicinal products in shared Facilities EMA/CHMP/ CVMP/ SWP/169430/2012
*- Appendix 3 of ICH Q3C (R4) “Impurities: Guideline for Residual Solvents”
Represents a substance-specific dose that is unlikely to cause an adverse effect if an individual is exposed at or below this dose every day for a lifetime
Permitted Daily Exposure Definitions*
32
5.1 Calculation of PDE
Toxicological parameters of free HQ
CAS N.: 123-31-9 (IPCS, 1994; IPCS, 1996; OECD/ SIDS, 2002; Williams et al. 2007; NTP, 2009).
PDE for free HQ =
(25 mg/kg b.w./day x 50 kg b.w.) (5 x 10 x 5)
PDE free HQ = 5 mg/day or 100 µg/kg b.w./ day
33
PDE for Free HQ = 5 mg/day or 100 µg/kg b.w./ dayExposure level of Free HQ derived from HMP = 11 µg/k.g. b.w./day
• PDE level is 9-times higher than the maximum exposure level to free HQ in a “daily usual” scenario after ingestion of a therapeutic RDD (420 mg of arbutin) of a herbal medicinal product containing Uvae ursi folium
• PDE level corresponds to 8.3-times RDD of Cystinol HMP (6 tables/day). A patient should take 50 Tablets/day in order to be exposed to the PDE value of free HQ
Accordingly,
the level of free HQ produced by the administration of a RDD of Uvae ursi folium has a negligible probability of any toxicological risk to human health*
5.2 Results - PDA
*Garcia et al. Int J Toxicol. 2013
34
5.3 Results - PDA
Supporting this low risk human exposure level, the HMPC monograph on Uvae ursi folium concluded that concentrations reached in the human body by monographed Uvae ursi folium-products are below the most conservative Thresholds of Toxicological Concern (TTC) (HMPC, 2012).
In agreement with the above statement, the benefit-risk ratio of the medicinal use of Uvae ursi folium extract has been considered favourable for the claimed indication and dosage by all national and international current Monographs on Uvae ursi folium.
• Commission E (1994)
• WHO monograph (2004).
• HMPC monograph (2012),
• ESCOP (2012)
35
6. Assessment of genotoxicity
• Guidelines for genotoxicity testing of pharmaceuticals have been established by HMPC, OECD and ICH
• Testing of medicinal products involves a battery of genotoxicity tests, in which pro- and eukaryotic systems in in vitro and in vivo experimental setups with and without metabolic activation are employed
• EMEA/HMPC/107079/2007 into effect 01.12.2008 Guideline on the assessment of genotoxicity of herbal substances / preparations
• Note for guidance on genotoxicity: guidance on specific aspects of regulatory genotoxicity tests for pharmaceuticals (CPMP/ICH/141/95) = ICH guideline S2 (R1)
• OECD (1997). OECD Guidelines for the Testing of Chemicals No. 471: Bacterial Reverse Mutation Test [the Ames test].
• OECD (1997). OECD Guidelines for the Testing of Chemicals No. 474: Mammalian Erythrocyte Micronucleus Test [in vivo micronucleus test].
36
6.1 Scheme Mutagenic-Genotoxic Studies
RDD 6 tablets Cystinol/ day
1.6 g Uvae ursi folium extract =
7 mg/kg/day arbutin
N= 12 Urine pool
total HQ: 36.00 µg/ml
free HQ: 0.512 µg/ml
In vitro
AMES test
Risk????
In vivo
Mice Micronucleus testSTUDY 1 - Pharmacokinetics
STUDY 3 – AMES test
STUDY 4 – Micronucleus test
37
6.2 AMES Test
Þ Salmonella typhimurium LT2 : Four different histidine auxotrophic strains.
Identify two basic classes of point mutations: base pair substitutions (TA 1535, TA 100) frameshift mutations (TA 1537, TA 98). All strains contain GC base pairs at the site of the histidine mutation
Fifth strain for detecting point mutations at adenine-thymine (AT) Do not comply with genotoxicity guidelines
Þ Investigation: Urine samples according to OECD-AMES guideline by two independent experiments:
(I) the plate incorporation test (II) the pre-incubation test
Þ Evaluation: Mutagenic Potential it induces a > 2-fold increase in the number of revertant colonies over negative control (spontaneous reversion rate)
38
6.2 AMES Test: Preliminary Toxicity Test
• Concentrations from 36.00/0.511 µg/ml to 0.240/0.0026 µg/ml of total HQ and free HQ• Bacterial strains TA 98 and TA100 • Plate incorporation method with or without addition of S9 fraction. • Two independent assays (n=3).
• 4-nitro-o-phenylene-diamine (4-NOPD, 10 μg/plate)
• sodium azide (NaN3, 10 μg/plate)
• 2-aminoanthracene (2-AA, 2.5 μg/plate)
• control urine sample + 5 µg/plate free HQ
39
6.2 AMES Test: Plate Incorporation Test
- Plate incorporation test with/without metabolic activation. - Two independent assays (n=3).
• sodium azide (NaN3, 10 μg/plate)
• 2-aminoanthracene (2-AA, 2.5 μg/plate)
• control urine sample + 5 µg/plate free HQ
40
• Pre-incubation test with /without metabolic activation. • Two independent assays (n=3). Mean±SD
6.2 AMES Test: Pre-incubation Test• sodium azide (NaN3, 10 μg/plate)
• 2-aminoanthracene (2-AA, 2.5 μg/plate)
• control urine sample + 5 µg/plate free HQ
41
6.3 Micronucleus Test: in bone marrow cells
• 4 NMRI mice (2 females and 2 males)
• Under identical conditions as in the in vivo micronucleus assay concerning test item, animal strain, vehicle, route, frequency and volume of administration.
• Administration undiluted pooled urine (D1) i.p., 10 ml/kg b.w.
• Examination of acute toxic symptoms (e.g. death, reduced spontaneous activity, eyelid closure, apathy etc.) at intervals of 1, 6, 24 and 48 hours
Preliminary study on acute toxicity
RESULTS: No signs of toxic reactions after 1, 6, 24 and 48 h were observed The undiluted urine D1 was considered as a suitable test item
42
6.3 Micronucleus Test: in bone marrow cells
UrineUndiluted
D1
Urine ½ D1
Urine 1/10 D1
10 NMRI mice /test group
ControlUrine
ControlUrine + free HQ
Control0.9% NaCl
+ Control
CPA*
*cyclophosphamide (CPA) at 30 mg/kg b.w
24 – 48 h
(10 ml/kg b.w.) i.p.
mice bone marrow micronuclei stained with May-Grünwald-Giemsa
Cellular target: Polychromatic erythrocytes (PCE) in the bone marrow mouse.
Preparation of the marrow and cells
Each group: 5 male/5 female
43
Þ 2000 polychromatic erythrocytes (PCEs, reticulocytes; immature erythrocytes) were scored per animal for frequency of micronucleated cells
Þ in 1000 erythrocytes the ratio of PCEs/normochromatic erythrocytes (NCE) in the bone marrow is scored for each dose group as an indicator of chemical-induced toxicity
• A test item is considered mutagenic, if it induces either a dose-related increase in the number of micronucleated PCE or a statistically significant positive response for at least one of the test points
• The cytotoxic effect of the test item due to the treatment was estimated as the ratio between PCE and NCE and reported as the number of NCE per 1000 PCE
• Statistical significance p< 0.05 was evaluated by means of non-parametric Mann-Whitney test
6.3 Micronucleus Test
Evaluation:
44
Test Item Dose(ml/kg b.w.)
Sampling time (h)
PCE with micronuclei (%)
Range ratio PCE/NCE
Control urine 10 24 0.13 0-4 1000/726Control urine + HQ (50µg/ml)
10 24 0.12 0-2 1000/745
0.9% NaCl-solution 10 24 0.13 0-4 1000/722Urine 1/10 D1 diluted 10 24 0.14 0-4 1000/736Urine ½ D1 diluted 10 24 0.10 0-3 1000/747Urine undiluted (D1) 10 24 0.09 0-3 1000/790Cyclophosphamide 30 mg/kg
b.w.24 0.97* 4-25 1000/736
Urine 1/10 D1 diluted 10 48 0.11 0-4 1000/718Urine ½ D1 diluted 10 48 0.09 0-2 1000/738Urine undiluted (D1) 10 48 0.12 0-4 1000/756
• Frequencies of micronucleated PCE in 2000 PCEs per animal • Ratio PCEs/ NCE in 1000
6.3 Micronucleus Test: DATA -RESULTS
*increase micronucleus frequency statistically significant (p<0.0001, non-parametric Mann-Whitney test
45
• NO dose-related increase in the number of micronucleated PCE cells were observed
• NO clear increase in the number of micronucleated cells in a single dose group at a single sampling time were observed
• NO Cytotoxicity = No change in the ratio polychromatic erythrocytes (PCEs) /normochromatic erythrocytes (NCE) was observed
Results :
6.3 Micronucleus Test:
46
A multicenter survey with 75 registered doctor’s offices located in Germany was conducted in 1992-1993 reporting their clinical experience with Cystinol mono (Cystinol akut since February 1996) for the treatment of diseases of the lower urinary tract
A total of 186,122 patients, (117,282 women)
56 doctors reported experience with a total of 17,430 children and adolescents.
Physicians with >30 years of experience (average of 10 years)
34.6%
Thera
peuti
c In
dic
ati
ons
Cyst
inol akut
7. Safety 7.1 Cystinol acut – Medicinal use
47
Uncomplicated cystitis
Irritable bladder
Urinary tract infection
Prevention of recurrence bacterial cystitis
88 90 92 94 96 98 100 102
Judgment of Therapy Effect by the Doctors (%) Good -
very Good
98.6%
Uncomplicated cystitis
Irritable bladder
Urinary tract infection
Prevention of recurrence bacterial cystitis
0 25 50 75 100
Urine alkalinization (%)
Cha
nge
of U
rine
pH
is n
ot req
uier
ed
Uncomplicated cystitis
Irritable bladder
Urinary tract infection
Prevention of recurrence bacterial cystitis
Tolerability (%)
very PoorPoorvery Good -Good
Mean duration
Treatment (days)
Time to reach Effect
(days)
Uncomplicated cystitis
9 3
Irritable bladder 10 4Urinary tract infection
10 4
Prevention of recurrence bacterial cystitis
28
7. Safety 7.2 Cystinol akut– Medical use
48
Wide-spread OTC medicinal use Pharmacovigilance Data
Post-marketing patient exposure Cystinol Akut from all over the world
Cumulative period of 10 years ≈ 7,000,000 patients
In this period of time A total of 161 cases (275 AEs)
SPC/PIL Side effects, adverse reactions
≥ 1/10,000 to < 1/1,000 gastrointestinal complaints (nausea, stomachache
and vomiting) < 1/10,000 allergic reactions
7. Safety 7.2 Pharmacovigilance
49
Relevant beneficial Properties
Antibacterial/anti-adherent /anti-inflammatory properties
Broad spectrum of antibacterial activity
NO development of antibiotic resistant bacteria
Local antibacterial effect Urine + Bladder
NO effect on intestinal, vaginal flora/immuno defenses
Safe and well-toleratedBenefit – Risk Balance clearly positive
50
8. Conclusions
Free und conjugate HQ were detected and quantified in human urine samples.
Human exposure to Free HQ was below its Permitted Daily Exposure
No risk for human health exposed at or below this dose every day for a lifetime
In vitro and in vivo mutagenic and genotoxic studies were negative
Accordingly, urine samples obtained from healthy volunteers receiving a regular recommended dose as proposed for therapeutic use in the HMPC monograph bears no mutagenic risk.
Pharmacovigilance Data from more than 30 years do not found AE different as reported in SPC/PIL
In agreement with HMPC monograph
Uvae ursi folium preparations are safe
51
Thank you for your attention
Don’t worry…We can eat him.
He took Uva ursi only!!
52
