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 ADVANCES in
PULMONARY
DRUG DELIVERY
 ADVANCES in
PULMONARY
DRUG DELIVERY

edited by
PHILIP CHI LIP KWOK • HAK-KIM CHAN
CRC Press
Taylor & Francis Group
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Boca Raton, FL 33487-2742
© 2017 by Taylor & Francis Group, LLC
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Version Date: 20161012

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Dedication

To my students

Hak-Kim Chan
To my family

Philip Chi Lip Kwok

Contents
Advances in Pulmonary Drug Delivery �������������������������������������������������������������������ix
Editors �����������������������������������������������������������������������������������������������������������������������xi
Contributors ����������������������������������������������������������������������������������������������������������� xiii

Chapter 1 Inhaled Proteins and Peptides ��������������������������������������������������������������1

Ron K. Wolff

Chapter 2 Inhaled Insulin: More Compelling than Ever ������������������������������������ 23

John S. Patton and Samantha R. Miller

Chapter 3 Inhaled Therapeutic siRNA for the Treatment of Respiratory

Diseases ���������������������������������������������������������������������������������������������� 31
Dongmei Cun, Lan Wu, and Mingshi Yang

Chapter 4 New Molecules to Treat Asthma and COPD ������������������������������������� 49

Amir Hakim and Omar S. Usmani

Chapter 5 Inhaled Anticancer Agents ����������������������������������������������������������������� 67

Rajiv Dhand

Chapter 6 Inhaled Countermeasures for Respiratory Tract Viruses������������������� 93

Ralph A. Tripp and Jarod M. Hanson

Chapter 7 Pulmonary Delivery of Antibiotics for Respiratory Infections�������� 131

Qi (Tony) Zhou, Li Qu, and Hak-Kim Chan

Chapter 8 Inhaled Liposomes ��������������������������������������������������������������������������� 151

David Cipolla

Chapter 9 Inhaled Traditional Chinese Medicine for Respiratory Diseases ���� 175
Yun Zhao, Jing Zhou, Yong-Hong Liao, and Ying Zheng

vii
viii Contents

Chapter 10 Bronchoprovocation Tests for the Evaluation of Drug Efficacy

in Asthma ����������������������������������������������������������������������������������������� 199
John Paul Oliveria, Brittany M. Salter, and Gail M. Gauvreau
Index ���������������������������������������������������������������������������������������������������������������������� 227
Advances in Pulmonary
Drug Delivery
The respiratory tract has been used to deliver biologically active chemicals into
the human body for centuries� Inhaled volatile oils and herbal smokes have been
employed by ancient civilizations for therapeutic and recreational purposes [1]� This
demonstrates the efficiency of the lungs to absorb drugs into the systemic circulation,
due to its large surface area and profuse blood supply� Inhalation is also the most
effective route of administration for local diseases such as asthma, chronic obstruc-
tive pulmonary disease (COPD), and lung infections� The inhaled drug particles can
target the site of action rapidly with a relatively low dose, thus reducing systemic
adverse effects� However, the lungs are complex in their anatomy and physiology,
which poses challenges to drug delivery� Therefore, inhaled formulations are gener-
ally more sophisticated than those for oral and parenteral administration� This book
highlights the latest developments in this field�
Most classical inhaled drugs are small molecules (e�g�, beta-2 agonists, anticho-
linergics, corticosteroids) for pulmonary diseases, but increasingly more biopharma-
ceuticals (e�g�, proteins, peptides, nucleic acids) have been investigated or marketed
for inhalation for both local and systemic delivery� This is a reflection of the global
trend in the increasing use of biopharmaceutical formulations via other routes of
administration� Wolff [2] provides an overview on various inhaled proteins and pep-
tides such as insulin, alpha-1 antitrypsin, cyclosporine, human growth hormone,
measles vaccine, and anti-IgE antibody for a variety of diseases� Of these agents,
insulin and alpha-1 antitrypsin are further discussed by Patton and Miller [3]� The
challenges of siRNA delivery and the use of nonviral vectors to enhance transfec-
tion efficiency are covered by Cun and Yang [4]� Advances in the understanding
of the pathophysiology of asthma and COPD have led to the development of novel
anti-inflammatory molecules (e�g�, phosphoinositide-3-kinase-delta, dual phosphodi-
esterase-3 and -4, p38 mitogen-activated protein kinase, humanized antibodies, and
chemokine receptor antagonists) [5]� Likewise, novel biopharmaceutical anticancer
agents involving immunological and genetic agents are actively being investigated
for lung cancer treatment [6]� Inhalation delivery is ideal for lung cancers to achieve
local drug targeting and reduce potential systemic toxicity� The same rationale also
applies to inhaled vaccines and antibiotics for pulmonary infections [7,8]� Liposomes
have been used to effectively modify the release, and enhance the pharmacokinetics
and pharmacodynamics, of inhaled antibiotics [9]� A novel trend for treating pulmo-
nary diseases in China is the inhalation of traditional Chinese herbal formulae, which
are usually administered orally or parenterally [10]� Although some effectiveness of
inhaled traditional Chinese medicine has been demonstrated, more rigorously con-
trolled clinical trials are required� Besides inhaling drugs to treat particular diseases,
chemical trigger agents (e�g�, mannitol, adenosine monophosphate, allergens) can also

ix
x Advances in Pulmonary Drug Delivery

be delivered into the airways for asthma diagnosis and antiasthma drug evaluation in
bronchoprovocation tests [11]�
Pulmonary drug delivery is a specialized field because of its many unique issues
and challenges� Rapid progress is being made and novel solutions are being offered
to existing treatment problems� The major direction in which this field is headed is
the use of biologically active macromolecules� These new drugs are generally more
fragile and potentially more immunogenic than traditional small molecular drugs�
Therefore, active research on their stability, safety, and efficacy is anticipated in the
near future� Despite the setback caused by the withdrawal of Exubera® in 2007, the
approval in 2014 of yet another dry powder inhaled insulin (Afrezza® by MannKind
Corporation) [12] is likely to kindle more interests in the field�

Philip Chi Lip Kwok

Hak-Kim Chan

REFERENCES
1� Sanders M� Inhalation therapy: An historical review� Primary Care Respiratory
Journal� 16(2), 71–81 (2007)�
2� Wolff R� Chapter 1� Inhaled proteins and peptides�
3� Patton JS, Miller SR� Chapter 2� Inhaled insulin: More compelling than ever�
4� Cun D, Yang M� Chapter 4� Inhaled therapeutic siRNA for the treatment of respiratory
diseases�
5� Hakim A, Usmani OS� Chapter 5� New molecules to treat asthma and COPD�
6� Dhand R� Chapter 6� Inhaled anti-cancer agents�
7� Tripp RA, Tompkins M� Chapter 7� Inhaled influenza vaccines�
8� Zhou Q, Chan H-K� Chapter 8� Pulmonary delivery of antibiotics for respiratory
infections�
9� Cipolla D� Chapter 9� Inhaled liposomes�
10� Zhou J, Liao Y-H� Chapter 10� Inhaled traditional Chinese medicine for respiratory
diseases�
11� Oliveria JP, Watson BM, Gauvreau GM� Chapter 11� Bronchoprovocation tests for eval-
uation of drug efficacy in asthma�
12� MannKind Corporation� About MannKind� http://www�mannkindcorp�com/� Accessed
March 18, 2016�
Editors
Dr. Philip Chi Lip Kwok is an assistant professor at the Department of Pharmacology
and Pharmacy, University of Hong Kong, Pokfulam, Hong Kong� He earned his
BPharm (Hons) and PhD in 2002 and 2007, respectively, from the Faculty of
Pharmacy, University of Sydney, Sydney, New South Wales, Australia� He was a
research associate in the Advanced Drug Delivery Group in this faculty from 2007
to 2011� He has 45 peer-reviewed publications and 2 joint patents in pharmaceuti-
cal aerosols and respiratory drug delivery� He is an editorial board member of four
scientific journals, including the Journal of Aerosol Medicine and Pulmonary Drug
Delivery, which is the official journal of the International Society for Aerosols in
Medicine� Dr Kwok co-hosted and chaired the inaugural Inhalation Asia conference
in Hong Kong in June 2013� This was the first international conference of its kind
in the Asian region� Following its success, he was an organizing committee member
for the second one held in Shenyang, China, in September 2015�

Hak-Kim Chan, professor in pharmaceutics, is leading the Advanced Drug Delivery

Group and Respiratory Disease Theme at the Faculty of Pharmacy, University of
Sydney� He graduated from the NDMC in Taiwan (BPharm) and the University
of Sydney (PhD, DSc) and did his postdoc training at the University of Minnesota�
He worked as a scientist at Genentech Inc� His research focuses on inhalation drug
delivery, ranging from powder production by novel processes, particle engineering
and aerosol formulation to scintigraphic imaging of lung deposition and clinical out-
comes� He has over 300 scientific publications on pharmaceutical formulation and
drug delivery (with over 9500 citations) and holds seven patents in these areas� He is
an executive editor of Advanced Drug Delivery Reviews and on the editorial advi-
sory boards of various pharmaceutical journals, including Pharmaceutical Research
and International Journal of Pharmaceutics� He is a fellow of the American
Association of Pharmaceutical Scientists and of the Royal Australian Chemical
Institute (RACI) and was chair of the NSW Pharmaceutical Science Group of the
RACI and vice president of the Asian Federation for Pharmaceutical Sciences�

xi
Contributors
Hak-Kim Chan Yong-Hong Liao
Faculty of Pharmacy Institute of Medicinal Plant
The University of Sydney Development
Sydney, New South Wales, Australia Chinese Academy of Medical Sciences
and Peking Union Medical College
David Cipolla Haidian District, Beijing, People’s
Aradigm Corporation Republic of China
Hayward, California
Samantha R. Miller
Dongmei Cun Dance Biopharm Inc�
Department of Pharmaceutical Science Brisbane, California
Shenyang Pharmaceutical University
Shenyang, Liaoning, People’s Republic John Paul Oliveria
of China Department of Medicine
McMaster University
Rajiv Dhand Hamilton, Ontario, Canada
Department of Medicine
The University of Tennessee Graduate John S. Patton
School of Medicine Dance Biopharm Inc�
Knoxville, Tennessee Brisbane, California

Gail M. Gauvreau Li Qu
Department of Medicine Monash Institute of Pharmaceutical
McMaster University Sciences
Hamilton, Ontario, Canada Monash University
Parkville, Victoria, Australia
Amir Hakim
National Heart and Lung Institute Brittany M. Salter
Imperial College London and Royal Department of Medicine
Brompton Hospital McMaster University
London, United Kingdom Hamilton, Ontario, Canada

Jarod M. Hanson Ralph A. Tripp

Department of Infectious Diseases Department of Infectious Diseases
University of Georgia University of Georgia
Athens, Georgia Athens, Georgia

Lan Wu Omar S. Usmani

Department of Pharmaceutical Science National Heart and Lung Institute
Shenyang Pharmaceutical University Imperial College London and Royal
Shenyang, Liaoning, People’s Republic Brompton Hospital
of China London, United Kingdom

xiii
xiv Contributors

Ron K. Wolff Ying Zheng

Safety Consulting Inc�
Carbondale, Colorado
Mingshi Yang
Department of Pharmaceutical Science
Shenyang Pharmaceutical University
Shenyang, Liaoning, People’s Republic
of China
and
Department of Pharmacy
University of Copenhagen
Copenhagen, Denmark
State Key Laboratory of Quality
Research in Chinese Medicine
University of Macau
Taipa, Macao SAR, People’s Republic
of China
Jing Zhou
Institute of Medicinal Plant
Development
Chinese Academy of Medical Sciences
and Peking Union Medical College
Haidian District, Beijing, People’s
Republic of China

Yun Zhao Qi (Tony) Zhou

Institute of Medicinal Plant Development Department of Industrial and Physical
Chinese Academy of Medical Sciences Pharmacy
and Peking Union Medical College College of Pharmacy
Haidian District, Beijing, People’s Purdue University
Republic of China West Lafayette, Indiana
 1 Inhaled Proteins
and Peptides
Ron K. Wolff

CONTENTS
Introduction ����������������������������������������������������������������������������������������������������������������1
Mechanisms of Absorption and Clearance ����������������������������������������������������������������2
Immunogenicity ����������������������������������������������������������������������������������������������������2
Nonclinical Development Considerations �������������������������������������������������������������4
Examples of Development Candidates ����������������������������������������������������������������������5
Inhaled rhDNase (Pulmozyme) �����������������������������������������������������������������������������5
Inhaled Insulin�������������������������������������������������������������������������������������������������������6
Inhaled Human Growth Hormone �������������������������������������������������������������������������9
PTH1-34 ����������������������������������������������������������������������������������������������������������������9
Cyclosporine ������������������������������������������������������������������������������������������������������� 11
Alpha-1 Antitrypsin��������������������������������������������������������������������������������������������� 12
Inhaled Vaccines �������������������������������������������������������������������������������������������������� 13
Anti-IgE��������������������������������������������������������������������������������������������������������������� 14
Inhaled Gene Therapy ����������������������������������������������������������������������������������������� 14
Formulation Approaches ������������������������������������������������������������������������������������� 16
Future Directions ����������������������������������������������������������������������������������������������������� 18
Summary ������������������������������������������������������������������������������������������������������������������ 19
References ���������������������������������������������������������������������������������������������������������������� 19

INTRODUCTION
Inhaled proteins and peptides have undergone a great deal of study in the last two
decades� However, at present, there is only one approved inhaled protein, Pulmozyme
(DNase), for the treatment of cystic fibrosis� Exubera (inhaled insulin) was approved
but was withdrawn from the market primarily due to a lack of commercial success,
not due to a lack of efficacy� Pulmozyme is an example of using inhalation for local
lung treatment, while Exubera is an example of using lung delivery for systemic
administration� A number of nonclinical and clinical studies for either local treatment
or systemic delivery have shown varying degrees of success for a range of proteins
and peptides� Summary information is provided for inhalation programs with DNase,
insulin, growth hormone, cyclosporine, alpha-1-antitrypsin (AAT), anti-IgE, and

1
2 Advances in Pulmonary Drug Delivery

vaccines� Examination of the features of these various programs provides insights for
future activities� This chapter will provide high-level information related to inhaled
proteins and peptides� More details are available in recent review papers [1–6]�

MECHANISMS OF ABSORPTION AND CLEARANCE

The major mechanisms for clearance of proteins from the lung have been reviewed
by Patton and Byron [3] and Wolff [4]� Proteins that deposit on ciliated epithelium
are not absorbed to a significant extent and are primarily cleared by mucociliary
transport up the airways and then eliminated via the gastrointestinal tract where they
are degraded and eliminated� Proteins that deposit in the alveolar region are cleared
from the lung primarily via four routes: (1) phagocytosis by alveolar macrophages,
(2) paracellular diffusion through tight junctions, (3) vesicular endocytosis or pino-
cytosis, and (4) receptor-mediated transcytosis�
Phagocytosis by alveolar macrophages does not appear to be as important a clear-
ance mechanism as absorption� This may occur because phagocytosis is most effi-
cient for uptake of relatively insoluble particles� Therefore, this clearance pathway is
likely to be of most importance if there is degradation of proteins to insoluble forms�
It appears that soluble proteins effectively dissolve in lung fluids and distribute them-
selves in the surfactant and mucus layers of the lung�
For soluble proteins, clearance of protein from the lung closely parallels absorp-
tion into blood� The inverse dependence of absorption versus protein molecular
weight, as shown in Figure 1�1, has been used to suggest that diffusion across alveolar
epithelial membranes through tight junctions is a major absorption mechanism [3]�
The available data also support the view that absorption of high-molecular-weight
proteins the size of albumin (68 kDa) or greater is not likely to be extensive or rapid
because they are generally too large to be absorbed via tight junctions� Antibodies
that are frequently in the >150 kDa also show little absorption from the lung into
blood�
These data (Figure 1�1) indicate for proteins less than or equal to the MW of
human growth hormone (hGH; 22 kDa) that absorption is adequate enough that sys-
temic delivery can be considered� However, the aerosol delivery system needs to be
considered to maximize the overall efficiency� For larger proteins, certainly those
greater than 68 kDa, absorption into the blood is low, half-life in the lung is relatively
long, and so potential for utility in local lung treatment is enhanced�
More research is needed in this area, however, and it is clear that at present, the
absorption and disposition of each protein being considered for therapeutic use must
be studied individually, because there is not sufficient knowledge for accurate pre-
dictions with currently available data [4]�

ImmunogenIcIty
In the case of a biopharmaceutical, antidrug antibodies (ADAs) may be raised
against the biomolecule� If the ADA response is sufficiently robust, this may result in
immune complex formation and subsequent toxicity� Typically immune complexes
Inhaled Proteins and Peptides 3

120

Bioavailability (% of dose deposited in lung)

100

Interferon-alpha
Leuprolide, DDAVP

G-CSF

Growth hormone
80

Insulin
Calcitonin

60

Erythropoietin
PTH(1–84)

40
Interferon-gamma
PTH(1–34)

20

0
0 10,000 20,000 30,000 40,000
MW (Da)

FIGURE 1.1 Bioavailability from the lung as a function of molecular weight calculated
on the basis of the percentage of protein that is deposited in the lung and absorbed into the
circulation�

are bound by red blood cells and then degraded in the spleen� However, immune
complexes may result in inflammation and tissue damage following kidney depo-
sition� ADA can have a direct effect on potency by preventing the antibody from
binding to its ligand as well as a reduction in half-life via the clearance mechanisms
involving neutralizing antibodies�
Assays for immunogenicity are able to measure the formation of ADA� However,
these assays are typically unable to determine the impact of ADA on the potency of
the drug; that is, they cannot differentiate between ADA and neutralizing antibody�
For demonstration of neutralizing antibody, a cell-based potency assay may be nec-
essary to clearly demonstrate a loss of drug potency in the presence of neutralizing
antibody�
Nonclinical immunogenicity is not necessarily predictive of clinical immunoge-
nicity but there are frequent parallels in relative immunogenicity between nonhuman
primates (NHPs) and humans, particularly if there is close homology between these
species� It is important to assess the pharmacokinetic/pharmacodynamic (PK/PD) expo-
sure in terms of observed toxicities [7]� In practical terms, if the PK/PD profile of the
biopharmaceutical is different between the first and last dose and/or there is evidence
of immune-mediated toxicity, then ADA assessments can aid in the interpretation of
the data� A dramatic decrease of Cmax and AUC values after repeated doses is a major
red flag that there may be antibody effects reducing the levels of the circulating
protein� In this instance ADA measurements are definitely recommended� However,
4 Advances in Pulmonary Drug Delivery

if the PK/PD profile is unchanged, then ADA measurements are not necessarily
warranted� However, because it is not possible to predict events in advance, it is good
practice to collect plasma samples that may be used for later analysis if needed�
For vaccine products, immunogenicity is the intended pharmacology and immune
responses are an important part of the nonclinical toxicity studies� Because vaccine
dosing is general single doses or a few repeat doses at infrequent intervals, it is often
difficult to measure TK� Therefore, assessment of the vaccine-induced immune anti-
body response can also serve as a measure of the exposure of the animals to the test
material�

nonclInIcal Development consIDeratIons

Nonclinical studies to support human dosing should be conducted according to Good
Laboratory Practices (GLP) and must reflect the intended clinical use (e�g�, dose
level and frequency of administration) of the biopharmaceutical� However, studies
for individual proteins or peptides are developed on a case-by-case approach to gen-
erate the appropriate data for clinical development� Consultations with regulatory
agencies are recommended�
The ICH S6 guidelines [8] define a relevant species as “one in which the test
material is pharmacologically active due to the expression of the receptor or an epit-
ope (in the case of monoclonal antibodies)�” Unlike small molecules, a single species
may be sufficient if a second species is not suitable to predict risk in humans� For
example, for many biopharmaceuticals, the human therapeutic target is expressed in
the NHP and not in other species or target binding and pharmacology is only pres-
ent within the NHP� However, where two pharmacologically relevant species exist
(one rodent and one nonrodent), both should be used for initial safety assessment
studies� The ICHS6 R1 addendum allows chronic studies to proceed only in rodents
rather than nonrodents where adequate long-term exposure and pharmacology can
be maintained�
The ICH S6 guidelines recommend that dose levels should be selected to provide
information on a dose response that includes a toxic dose and a no observed adverse
effect level (NOAEL)� However, for some types of biologics where toxicity is a result
of exaggerated pharmacology, the pharmacologically active dose, or PAD, may be a
better indicator of observations of potential toxicity than NOAEL [9]� The high dose
level in toxicity studies should ideally include a safety margin of at least 10× the
maximum anticipated clinical dose; however, sometimes exaggerated pharmacologi-
cal responses preclude dosing this high�
In repeat dose nonclinical safety studies, dosing frequency should be similar to
that of proposed clinical trial� However, more frequent dosing may be required if
the test item is more rapidly cleared in animals than humans [9]� Repeat dose stud-
ies should include toxicokinetics and a recovery phase (to detect regression of any
pathological change and/or detect potential delayed toxic effects)� Repeat dose tox-
icity studies of 6-month duration in rodents and nonrodents have been shown to be
generally sufficient to predict potential human risks [10]�
Pharmacodynamic endpoints: Since the toxicity of most biopharmaceuticals
is related to their mechanism of action, then high doses may be associated with
Inhaled Proteins and Peptides 5

exaggerated pharmacology� Therefore, pharmacodynamic endpoints should be mea-

sured on studies to better understand adverse effects, such as glucose monitoring
following insulin administration (discussed later)�

EXAMPLES OF DEVELOPMENT CANDIDATES

InhaleD rhDnase (pulmozyme)
There is currently only one marketed inhaled protein, recombinant human(rh)
DNase (Pulmozyme), developed for the treatment of cystic fibrosis� Inhalation toxi-
cology studies were conducted in both rats and monkeys for durations of 28 days and
6 months [11]� The exposure concentrations and daily exposure durations spanned
inhaled doses of 1�3–69 times the expected daily clinical inhaled dose� In the
28-day rat study, mild to moderate alveolitis was observed in the high-dose group�
This lesion was not evident after 4 weeks of recovery� Bronchiolitis was observed
in one of eight high-dose monkeys after 4 weeks of exposure, but, again, there
were no lesions observed after 4 weeks of recovery� In the 6-month rat study, bron-
chiolitis was observed at the end of the treatment period but at a somewhat lower
incidence than in the 4-week study� These data suggest that the mild lesion was not
progressive in rats� In the 6-month monkey study, respiratory rates measured during
aerosol exposure to monitor for anaphylactic or irritant responses were unchanged
compared to preexposure values� Positive serum antibody titers to rhDNase were
observed beginning at week 4 and persisting through the treatment period� Serum
concentrations of rhDNase at 24 h postdose indicated that there was no accumu-
lation of rhDNase throughout the 6-month treatment period� Histopathologically,
there was increased perivascular lymphocytic cuffing, peribronchial lymphoid
hyperplasia, terminal airway-related bronchiolitis/alveolitis with eosinophilic
infiltrates, and increased siderophages� There appeared to be a close relationship
between the severity of the pulmonary lesions and the antibody titer to rhDNase
measured in serum� The lesions were consistent with an allergic or hypersensitiv-
ity (type I) response to a foreign protein� This is not unexpected, because there are
considerable differences between animal and human DNases� There is only an 80%
homology between the rat and human DNases; monkey DNase, although it has not
been completely sequenced, also appears to be highly dissimilar to the human form
(S� Shak, personal communication, 1997)�
Some of the most persuasive data related to challenges of assessing immune
reactions to inhaled therapeutic proteins in humans versus animals come from
the development of rhDNase (Pulmozyme) for the treatment of cystic fibrosis�
Pulmozyme involves the inhalation of a relatively large amount of rhDNase, which
is an enzyme that cleaves DNA� The clinical summary portion of the Summary
Basis of Approval for Pulmozyme (FDA, 1993) provides information related to
immune responses� It was concluded that antibodies to rhDNase were of little con-
sequence to the safety profile of rhDNase on patients because levels were generally
low and there was no correlation between antibody levels and clinical responses� As
noted previously, there were immune responses found in the rat and monkey inhala-
tion toxicology studies in which the animals were exposed to a highly heterologous
6 Advances in Pulmonary Drug Delivery

protein compared to their native DNases� There was no evidence of anaphylaxis in

the animal studies; however, lung lesions that were observed were consistent with
only mild alveolitis and were judged to be related to the immunological response
to a foreign protein� Thus, the findings in the animal studies were deemed to be
the result of immunological reaction to a nonhomologous foreign protein and not
relevant to results in people� The clinical data indicate little concern for the homolo-
gous protein in humans [4]�

InhaleD InsulIn
Inhaled insulin is covered in detail in Chapter 7, and so it will be dealt with briefly
here, concentrating mainly on safety issues� Inhaled insulin is attractive for the treat-
ment of type 2 diabetes because it avoids the use of needles and may offer better
outcomes through improved compliance� A large number of clinical trials have been
carried with inhaled insulin by a number of companies, as thoroughly reviewed by
Siekmeier and Scheuch [12] and Mastrandrea [13]� In controlled clinical trials, more
than 13,000 patients have been treated with inhaled insulin on average for 1 year
with some patients on Exubera® for up to 8 years� Overall, clinical trials have dem-
onstrated that inhaled insulin is comparable to subcutaneous insulin for improv-
ing glycemic control� Also pharmacokinetics and pharmacodynamics were roughly
similar, with the exception that inhaled regular insulin has faster absorption than
subcutaneous regular insulin and it is comparable to fast-acting insulins�
Immunogenicity was studied extensively� In all major clinical programs, delivery of
insulin by inhalation induced higher antibody levels in some patients than comparators�
However, these antibodies were not shown to decrease the effectiveness, safety, or toler-
ability of inhaled insulin over time and did not affect clinical outcomes [14]�
A number of inhaled insulin formulations have been developed but only Exubera
received marketing authority� Most efforts were discontinued after Pfizer with-
drew Exubera from the market but MannKind’s inhaled insulin Afrezza continued
in development� The major insulin development programs are cited in the follow-
ing list, but a more complete list can be found in the publication of Siekmeier and
Scheuch [12]:

• The only inhaled insulin that is approved is Afrezza (MannKind), a dry

powder formulation, precipitated from fumaryl diketopiperazine solution�
• Exubera (insulin human [rDNA origin]) Inhalation Powder, a spray-dried
dry powder formulation—withdrawn from the market by Pfizer�
• AERx (Novo Nordisk), an aqueous solution of insulin—discontinued after
phase 3 studies�
• AIR (Eli Lilly), spray-dried dry powder formulation of insulin, dipalmitoyl
phosphatidylcholine (DPPC), and sodium citrate—discontinued after phase
3 studies�
• Aerodose (Aerogen), liquid aerosol—development discontinued�

Preclinical study design considerations have to include both pharmacokinetics

(measurement of serum insulin levels) and pharmacodynamics (onset and duration
Inhaled Proteins and Peptides 7

of the hypoglycemic effect)� Hypoglycemia is a significant concern and has been

mitigated by feeding animals immediately prior to insulin aerosol exposure, careful
monitoring of the glucose response, and supplementation with food and/or glucose
in episodes of severe hypoglycemia�
Of the major inhaled insulin toxicology programs, four have significant toxicol-
ogy data available in the public domain [15–20]� In all four cases no adverse effects
on lung were noted�
Pfizer/Nektar (two 1-month rat, one 6-month rat, one 1-month cynomolgus mon-
key, and one 6-month cynomolgus monkey studies): These studies, in addition to
assessing standard toxicological parameters, also evaluated the potential toxic-
ity of the complete formulation and the excipients alone to the respiratory tract�
Investigations included respiratory and pulmonary function, lung cell proliferation
indices, insulin antibody titers, and histopathology� There were no toxicological find-
ings relevant to human systemic or pulmonary risk with the excipients or with the
complete formulation at up to 40 times for rats and four times for monkeys compared
to the clinical starting dose of 0�15 mg/kg/day� Maximum tolerated doses (MTDs)
were limited by hypoglycemia� No evidence of exposure-related inflammatory or
immune-mediated hypersensitivity reactions were observed in the respiratory tract�
A weak antibody response to the insulin powder was observed in rat serum but anti-
insulin antibodies were not detected in monkey serum or bronchoalveolar lavage
fluid� Rat insulin differs from human insulin by three amino acids, pig and dog
differ by one amino acid, and cynomolgus monkey insulin is identical to human
insulin� A comparable weak antibody response in rats was also observed following
subcutaneous injection� Assessment of respiratory and pulmonary function param-
eters revealed no exposure-related effects� No exposure-related histopathological
responses were observed in the respiratory tract or in lung-associated lymph nodes
in either species� Staining the bronchioles and alveoli with markers for cell prolifera-
tion showed no biologically significant differences in proliferation indices attribut-
able to the complete formulation or excipients alone in either species [15,18]�
AIR/Lilly (6 months, dogs): The purpose of this study was to characterize the
toxicity, pharmacokinetics, and pharmacodynamics of daily human-inhaled insu-
lin powder (HIIP) in beagle dogs via head-only exposures� The formulation was a
dry powder composed of insulin, DPPC, and sodium citrate� Dogs were exposed
15 min/day to an air control, placebo, maximal placebo (3× the placebo dose), or
one of three doses of HIIP (mean inhaled doses of 0�80, 0�24, or 0�70 mg/kg/day for
the HIIP-low, HIIP-mid, and HIIP-high dose, respectively)� Dose-related exposure
(Cmax, AUC) to inhaled insulin was observed with rapid absorption and there were no
apparent gender differences or accumulation after repeated inhalation exposures for
26 weeks� The expected pharmacological effect of insulin was observed with dose-
related decreases in serum glucose levels following HIIP administration� There were
no toxic effects observed including no HIIP or placebo treatment–related effects
on mean body weights, absolute body weight changes, clinical observations, food
consumption, respiratory function parameters, ophthalmic examinations, electrocar-
diograms, heart rates, clinical pathology, or urinalysis� Similarly, there were no HIIP
or placebo treatment–related effects on pulmonary assessments that included respi-
ratory function parameters, bronchial alveolar lavage assessments, organ weights, or
8 Advances in Pulmonary Drug Delivery

macroscopic and microscopic evaluations, including lung cell proliferation indices�

HIIP was considered to have either low or no immunogenic potential in dogs� The
NOAEL and MTD were the average inhaled dose of 0�70 mg insulin/kg/day [16],
approximately fivefold greater than a typical clinical dose�
Aerogen (1 month, dogs): The inhalation safety of Humulin R U500 liquid insu-
lin formulation was evaluated in a 28-day repeat dosing study in dogs� In addition to
500 IU/mL of human DNA-derived insulin, Humulin R contains glycerin 16 mg/mL,
metacresol 2�5 mg/mL, and zinc oxide to supplement the endogenous zinc to obtain a
total zinc content of 0�017 mg/100 units and water for injection� The pH was 7�0–7�8�
Two pulmonary doses of insulin, a low dose, and the MTD were evaluated against water
and placebo controls� After 28 days of daily exposure, the animals were killed and
examined at necropsy� Respiratory tissues were examined microscopically� The mean
pulmonary doses achieved were 2�3 and 8�3 U/kg/day for the low and MTD dose levels,
respectively� The aerosols were highly respirable with 78% of the insulin aerosols being
<3�8 μm� The treatments were well tolerated� No adverse in-life, necropsy, or histologi-
cal findings were detected that were related to insulin or inhalation treatment [17]�
MannKind/Sanofi (26-week rat, 39-week dogs; carcinogenicity studies, 104-week
inhalation, rats; 26-week transgenic mice): In inhalation toxicology studies of
AFREZZA, rats exposed to doses up to 1�91 mg/kg/day for 26 weeks and 1�23 mg/kg/day
for 104 weeks, and dogs exposed to doses up to 1�92 mg/kg/day were well tolerated in
all animals� There were no adverse findings including no microscopic findings in the
lungs or evidence of carcinogenicity or proliferation� The main findings were, in rat
26-week study, goblet cell hyperplasia and eosinophilic globule accumulation of mini-
mal severity in the olfactory/respiratory epithelium and, in dogs, neutrophil infiltration
of minimal severity in lung� The TechnoSphere carrier controls at doses of 11�7 mg/kg
also had similar lack of findings in the rat 26-week study, but also a slight increase in
proliferating cell nuclear antigen in the bronchioles was observed [19]�
In a 26-week carcinogenicity study, transgenic mice (Tg-ras-H2) exposed to
doses up to 5 mg/kg/day of AFREZZA and to 75 mg/kg/day of TechnoSphere
carrier had no increased incidence of tumors� AFREZZA also was not genotoxic
in Ames bacterial mutagenicity assay and in the chromosome aberration assay,
using human peripheral lymphocytes with or without metabolic activation� The
TechnoSphere carrier alone was not genotoxic in the in vivo mouse micronucleus
assay [20]�
In a reproduction toxicity study, female rats given sc doses of 10, 30, and 100 mg/
kg/day of TechnoSphere carrier starting 2 weeks before mating until gestation day 7
had no adverse effects on male fertility at doses up to 100 mg/kg/day (a systemic
exposure 14–21 times that following the maximum daily AFREZZA dose of 99 mg
based on AUC)� Female rats at 100 mg/kg/day did have increased pre- and postim-
plantation loss but not at 30 mg/kg/day (14–21 times higher systemic exposure than
the maximum daily AFREZZA dose of 99 mg based on AUC) [20]�
In all of these extensive preclinical studies, at substantial multiples of clinical
doses, no adverse effects on lung were found related to either insulin or the excipi-
ents� Potential still exists for inhaled insulin since it has been shown to be both safe
and effective� However, its main advantage over sc delivery is convenience, which
may result in better compliance� Two programs are currently conducting clinical
Inhaled Proteins and Peptides 9

trials (Mannkind and Dance) and so there should be indications in the near future if
inhaled insulin will be part of the diabetes treatment arsenal�

InhaleD human growth hormone

Inhaled hGH was under development by Alkermes/Lilly [5]� One-month and
6-month inhalation toxicology studies were carried out in cynomolgus monkeys
with no adverse effects in the lung observed� These studies supported clinical
trials in normal volunteers� The trial was a crossover design in 12 young adult
subjects, each subject receiving both subcutaneous and inhaled hGH on separate
days� Inhaled hGH was well tolerated with no coughing or adverse taste issues�
Pulmonary function and vital signs were measured with no apparent changes of
clinical significance� The PK profiles for sc injection (4 mg hGH) and inhalation
delivery (92 mg hGH in 4 dry powder capsules) were quite similar as seen in
Figure 1�2 Overall delivery efficiency was approximately 5% compared to sc deliv-
ery, somewhat less than the approximately 10% values observed for a similar dry
powder formulation and delivery system for inhaled insulin� The decreased effi-
ciency with respect to inhaled insulin was expected because of the greater molecu-
lar weight of hGH�
Following the demonstration of safety in normal young adults, a similar crossover
study was carried out in pediatric patients� Again, there were no adverse clinical
outcomes for inhaled delivery, PK profile was similar to that from sc delivery, but
overall delivery efficiency was less than the business development goal of 5% rela-
tive to sc delivery� With this result, the program was terminated�
If manufacturing costs of hGH can be reduced, inhaled delivery of hGH could be
considered to improve compliance in children and increase the willingness of par-
ents to start therapy in their children since reluctance to start injections is a negative
factor in initiating growth hormone therapy�

pth1-34
Development of inhaled PTH1-34 to treat osteoporosis was also considered� Feasibility
data from intratracheal instillation studies in rats had shown that PTH1-34 was well
absorbed from the lung into blood with similar bioavailability to that of insulin
(Figure 1�1)� Then a single-dose PK study in monkeys was carried out� The PK of
PTH1-34 delivery by inhalation and subcutaneous (sc) administration was studied in
a crossover manner in three anesthetized rhesus monkeys (Figure 1�3)� Aerosol was
delivered from an ultrasonic nebulizer through an endotracheal tube to intubated mon-
keys� The PK profiles from the sc and inhalation routes were similar and indicated a
bioavailability of approximately 40% for inhaled delivery compared to sc� Tmax val-
ues were approximately 0�5 h and half-lives were approximately 1 h for each delivery
mode� This type of pulsatile delivery is important to provide the desired anabolic effect
on bone since constant delivery actually results in hypercalcemia and bone resorption�
Subsequently, a 2-week inhalation toxicity study was conducted in rhesus mon-
keys� No adverse effects were noted in any of the measured endpoints, including no
test-article-related effects on lung histopathology� PK profiles were similar on the
10 Advances in Pulmonary Drug Delivery

80

70

60
hGH conc (ng/ml)

50

40

30

20

10

0
0 2 4 6 8 10 12 14 16 18
(a) Time (h)

80

70

60
hGH conc (ng/ml)

50

40

30

20

10

0
0 2 4 6 8 10 12 14 16 18
(b) Time (h)

FIGURE 1.2 Pharmacokinetic profiles in healthy volunteers following Inhalation (a) and sc
(b) administration of human growth hormone�

first and last day of exposure, indicating no accumulation of the compound and also
no indication of neutralizing antibody production� Further, there was clear indication
of efficacy� The time scale was too short for increases in bone density but there were
clear significant increases in 1,25 di-hydroxyvitamin D3 levels indicating a positive
bone formation response� Despite these positive findings, further development was
not carried out because there was no clear therapeutic benefit of PTH1-34 inhalation
Inhaled Proteins and Peptides 11

1500

1000
Serum PTH (pg/mL)

Subcutaneous dose (10 µg/kg)

500

Lung dose
(10 µg/kg)

0
0 30 60 90 120 150 180 210 240
Time after dosing completed (min)

FIGURE 1.3 Single-dose inhalation studies of PTH1-34 in monkeys show good absorption
and similar pharmacokinetics to sc delivery�

delivery over sc administration other than more convenience� In a motivated popu-

lation suffering from osteoporosis, it was deemed that there was limited additional
market potential for an inhaled product (Figure 1�4)�

cyclosporIne
Cyclosporine is a cyclic peptide with potent immunosuppressive properties� Inhaled
cyclosporine has been investigated over a large number of years to aid in the postop-
erative treatment of lung transplant patients to reduce the incidence of rejection [21]�
The inhaled formulation had no unexpected systemic toxicity or clinically limiting
findings in the respiratory tract in 28-day rat and dog studies [22] and 9-month dog
studies with exposures 3 days a week [23]� In phase 2 clinical trials, the inhaled
formulation improved overall survival [24]� Recent data have shown that inhalation
of cyclosporine in solution with propylene glycol given in addition to conventional
immunosuppression appeared to improve important pulmonary function param-
eters in lung transplant recipients compared to patients receiving aerosol placebo
or conventional immunosuppression alone� It was also shown to improve overall
survival in phase 2 clinical trials [24]� However, recently completed phase 3 tri-
als showed no efficacy beyond that of standard of care when used as supplemental
targeted therapy [25]� The authors noted that “administering a cyclosporine aerosol
12 Advances in Pulmonary Drug Delivery

Control
PTH
200
Serum conc’n (pg/mL)

100

0
0 7 14
Time (days)

FIGURE 1.4 1,25 Dihydroxyvitamin D3 levels show positive biological response to inhaled
PTH1-34�

to this highly vulnerable patient population is not without challenges and this may
have influenced the study outcome” [17]� Other investigators are looking into pos-
sible uses to treat asthma [1]�

alpha-1 antItrypsIn
Alpha-1 antitrypsin is an enzyme that can be used to treat protease–antiprotease imbal-
ance in the lung� For patients with alpha-1 antitrypsin deficiency, this is a serious issue
since because of the low levels of antiproteases in the lung, proteases predominant
and lead to lung inflammation, tissue destruction, and a predisposition to emphysema�
Currently, there are three FDA-approved alpha-1 antitrypsins for IV administration:
Prolastin (Grifols), Zemaira (Aventis-Behring), and Aralast (Alpha Therapeutic Corp�)�
All of these products are derived by purification from human serum�
Inhaled alpha-1 antitrypsin has been studied as a potential therapy with gener-
ally encouraging results [26]� Siekmeier [27] summarized that “the data demon-
strate the feasibility of alpha-1 antitrypsin inhalation for restoration of the impaired
protease antiprotease balance, attenuation of the inflammation and neutralization
of the excess activity of neutrophil elastase�” Inhaled alpha-1 antitrypsin may well
be useful to COPD and CF as well as pure alpha-1 antitrypsin deficiency since
protease–antiprotease imbalances have been suggested to contribute to both these
respiratory diseases� Siekmeier [27] also noted that potentially the inhalation route
may provide cheaper therapy than that for IV administration since only about 2%
is delivered to the lung following IV delivery as compared to possibly 20%–30%
that could be delivered to the lung following inhalation� The 20%–30% aerosol
deposition value can only be achieved with optimized devices and formulations�
Kamada [28] is developing inhaled alpha-1 antitrypsin as potential therapy for AAT
Inhaled Proteins and Peptides 13

deficiency and announced results from their European phase 2/3 clinical study in
September 2014� These failed to meet either the primary “time to the first mod-
erate or severe exacerbation event” or secondary exacerbation endpoints with the
intent to treat (ITT) population� However, there were clinically relevant changes
in various lung function measurements in the ITT population, as well as in the
Most Frequent Exacerbators population, of which some were statistically signifi-
cant� Inhaled AAT was also safe and well tolerated in the patients� Based on the
strength of the lung function changes, especially in the Most Frequent Exacerbators
population, Kamada still plans to file for approval in Europe and the United States�
The complete data set from the phase 2/3 clinical study was presented in May 2015
[28]� The benefit of inhaled AAT treatment is also being evaluated in other patient
populations including cystic fibrosis [28]�
One additional opportunity that could enhance inhaled alpha-1 antitrypsin usage
would be the availability of a recombinant human protein to reduce immunogenicity
potential, increase product reproducibility, and potentially reduce costs if an appro-
priate manufacturing process can be developed�

InhaleD vaccInes
Inhaled vaccines, particularly measles vaccines, have received considerable atten-
tion� There has been the thought that delivery of the vaccine to the respiratory tract,
the natural entry route for the pathogen, might provide an attractive therapy by
engaging mucosal immunological mechanisms� Inhaled delivery is attractive for use
in developing countries because of lack of suitably trained staff to administer injec-
tions and problems in disposal of used needles and syringes� Early work by Albert
Sabin [29] demonstrated the feasibility followed by a mass vaccination campaign in
4 million children between 1988 and 1990 [30]�
In 2002 the Measles Aerosol Vaccine Project was initiated by the WHO, CDC,
and American Red Cross in order to develop a practical inhaled measles vaccine
using liquid aerosol delivery [31]� Phase 1 studies showed that the aerosolized vac-
cine was safe and well tolerated and produced an appropriate immune response�
Phase 2/3 studies were completed and showed results equivalent to sc delivery for
children ages 10–35 months, but for ages 9–10 months, immune response was not
as good as for sc delivery [32]� Challenges with getting good lung deposition in
nose-breathing infants likely contributed to these findings� The conclusion by WHO
was that for children greater than 10 months aerosol delivery should be effective�
In addition, there have been efforts to develop a dry powder aerosolized vaccine
using carbon dioxide–assisted bubble drying [33]� This effort was spearheaded by
Aktiv-Dry with key support from a Grand Challenges grant from the Gates founda-
tion� Preclinical studies in monkeys with the dry powder vaccine showed no adverse
effects and the production of an immune response was similar to that from sc deliv-
ery [34]� Phase 1 clinical trials have been completed in healthy men with preexist-
ing immunity to measles [35]� No adverse events were reported and the inhaled dry
powder vaccine produced serologic responses generally similar to subcutaneous vac-
cination; however, these results are difficult to interpret because of the high baseline
antibody levels�
14 Advances in Pulmonary Drug Delivery

One of the key hopes of the inhaled measles vaccines efforts is to provide access
to therapies in developing countries where infrastructure for needle injection deliv-
ery systems is problematical� Another key hope is that ultimately more cost-effective
therapies might also be possible [36]� This may be challenging, but with continued
technological innovation, it appears to be feasible�

anti-Ige
In the late 1990s there was a very interesting drug development plan by Genentech to
investigate the possibility of using inhaled anti-IgE to treat asthma� The rationale was
that there was evidence for both local and systemic effects of IgE in the etiology of
asthma� The hypothesis was that local delivery of anti-IgE to the lung would inhibit
IgE-mediated inflammation in the lung and provide improved asthma therapy� Both
nonclinical [37] and clinical studies [38] were undertaken� The definitive data from
the clinical trials showed that inhaled anti-IgE were well tolerated� It was concluded
“that aerosol administration of an anti-IgE monoclonal antibody does not inhibit the
airway responses to inhaled allergen in allergic asthmatic subjects�” The nonclinical
studies had demonstrated that aerosol delivery did result in good deposition of the
anti-IgE in lungs� It was also found that only <0�1% of the IgE delivered to lungs was
absorbed into blood� These data confirm that local deposition was indeed achieved
and the low absorption into blood, consistent with the high molecular weight of anti-
IgE, suggested a potential long-term residence in lung� In this case, systemic delivery
of anti-IgE by subcutaneous injection produces superior therapeutic results for treat-
ing asthma than local delivery by inhaled administration�
The results of this program suggest that for new initiatives with inhaled antibod-
ies, there needs to be careful consideration of target receptors, receptor affinities,
and relative influence of systemic and local effects� Although not discussing inhaled
therapies, Catley et al� [39] have reviewed potential use of monoclonal antibodies to
treat asthma� There has been some conjecture that future targets with relatively high-
affinity receptors in the lung and high lung specificity might be attractive opportu-
nities� Another factor supporting inhaled use for future therapies would be if there
were no systemic delivery alternative for the proposed therapy or if the only systemic
therapy is intravenous delivery�

InhaleD gene therapy

Inhaled gene therapy approaches to treat cystic fibrosis have been largely unsuc-
cessful using approaches such as attenuated adenovirus vectors to deliver the CFTR
gene [40]� There is broad consensus that treatment of CF lung disease will require
prolonged expression of CFTR for many months� However, only three trials using
adenovirus and adeno-associated virus vectors [41–43] have assessed duration in
the lung� These vectors were shown to be unsuitable because of adaptive immune
responses to these viral vectors [44]�
However, newer approaches using lipid-based carriers of CFTR DNA are under-
way� An inhalation study was carried out in mice to determine the efficacy and safety
of administering of cationic lipid formulation GL67A complexed with pGM169,
Another random document with
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particulièrement à vous être agréable et m’a recommandé de vous
faire un long article d’avant-première.
L’Auteur. — Excusez-moi, je suis en pleine répétition.
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quelques heures avant la générale.
L’Auteur, avec un grand effort de bonhomie. — Allons, allons,
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Omer et sa partenaire reprennent la scène, d’abord avec une

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Édouard Audoir. — Édouard Audoir, rédacteur à l’Espoir.
Maître, pourrais-je vous interroger ?
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 Édouard Audoir. — C’est que la copie du courrier doit être
donnée avant sept heures à l’imprimerie…
(L’Auteur se résigne et Édouard Audoir tire son carnet.)
Le Directeur, à l’Auteur. — Est-ce que vous leur avez donné le
mot de sortie pour Claire ?
L’Auteur, redescendant à l’avant-scène en essayant de mettre
le plus de dignité possible dans son oubli des injures. — Je l’ai
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Madorec, le régisseur, apportant le manuscrit. — Voilà.
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(Claire dit son mot de sortie. L’auteur fait entendre un petit

grognement d’approbation.)

Le Directeur, d’une voix profonde. — Il vaut mieux qu’elle sorte

sans rien dire.
L’Auteur. — C’est vous qui avez demandé un mot de sortie ?
Le Directeur. — Je n’ai pas demandé celui-là. Mes enfants,
répétons, répétons. La scène des deux hommes… Est-ce qu’on a
fait des coupures ?
L’Auteur. — J’ai pensé, je crois qu’il serait dangereux de faire
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bon, disons tout le texte, tout le texte.
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pas risquer d’être obscur.
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donc travailler.
 L’Auteur, se rebiffant. — Que je vous laisse travailler !
Le Directeur. — Eh bien ! oui, nous passons demain. Vas-y
Omer, vas-y Sénéchal. Toute votre scène, toute votre scène, sans
en perdre un mot…
L’Auteur. — C’est intolérable, je m’en vais…
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L’auteur irrité passe dans la salle sans savoir exactement où il

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Maître, si vous avez un moment…
L’Auteur, de guerre lasse. — Asseyez-vous là, sur cette
banquette… Qu’est-ce que vous voulez savoir ?

Il commence à dicter, les yeux clos, d’une voix de revenant. Il

faut parler de sa pièce mais il ne veut pas en déflorer le sujet. Alors il
insiste sur ses intentions et sur ses principes de théâtre qu’il
découvre d’ailleurs au fur et à mesure qu’il dicte. Il dit aussi que la
pièce a été reçue d’enthousiasme et que tout s’est bien passé aux
répétitions, dans la cordialité et la belle humeur.
 Le régisseur et les doubles

Le régisseur est d’ordinaire un acteur, remarquable surtout par

ses qualités d’ordre et de sérieux et qui manque peut-être de la
séduction frivole que les brillantes vedettes exercent sur le public.
Parmi ses fonctions, la plus honorifique est de mettre en scène
aux premières répétitions, quand le metteur en scène attitré n’est
pas encore arrivé à son poste. On appelle cela débrouiller. C’est un
travail très utile sans doute, mais dont rien ne subsistera aussitôt
que le seigneur de ces lieux aura pris l’affaire à son compte. Tel
personnage qui, selon les premières indications, entrait lentement au
premier plan à gauche, fait maintenant irruption par la porte du fond.
Le régisseur assiste avec une inconscience apparente à cette faillite
de ses conceptions. Son emploi a changé maintenant : il consistera
à remplacer, le manuscrit en mains, tous les artistes qui manquent à
la répétition, à soupirer les plaintes de l’ingénue encore en
déplacement sur la Côte d’Azur ou à hurler les grossièretés du
hobereau, pour qui on n’a pas encore trouvé l’interprète idéal ou
simplement convenable.
Il a encore d’autres fonctions délicates. Le matin, à 8 heures, il se
met en campagne pour s’occuper de tous les meubles que le
magasin du théâtre n’est pas en état de fournir. C’est lui également
qui découvre les phénomènes : l’homme qui doit, en coulisse, imiter
le rossignol, le montreur d’ours et son ours pour la fête villageoise du
« un ». Il s’occupe en général du recrutement des artistes de second
plan.
Ce pouvoir souverain que, dans les premières répétitions, il avait
à l’avant-scène, il le retrouvera quelques jours après le succès,
quand il dirigera, cette fois en maître absolu, les répétitions des
doubles.
Souvent, pas toujours, ces répétitions se font un peu à la flan,
comme le maniement d’armes dans la cour du quartier, quand c’est
un gradé modeste qui surveille l’exercice et que l’officier de semaine
n’est pas en vue.
Les doubles, c’est un certain nombre de personnages
secondaires qu’une angine subite des protagonistes appellera un
soir à un rôle capital. Honneur éphémère et surtout obscur, car les
notes de presse, pendant tout le temps que durera le remplacement,
resteront singulièrement muettes sur cette substitution. C’est
seulement quand le titulaire aura repris son rôle que l’on rendra un
tardif hommage à « M. Troupanel qui, pendant la maladie de son
camarade, etc. ». L’important est que les spectateurs soient
désormais assurés de trouver la vedette à son poste et ne soient
plus exposés à y rencontrer M. Troupanel.
M. Troupanel sait tout cela et témoigne de peu d’ardeur à
apprendre le rôle de son glorieux camarade. Parfois, pour doubler la
prima donna, on est obligé d’engager une artiste en dehors du
théâtre. Celle-là, si elle est jolie et si elle a une bonne réputation de
talent, ne s’y met pas non plus avec beaucoup de fougue. Elle sait
très bien que si le bruit se répand qu’elle fera de l’effet dans le grand
rôle, la vedette la plus fragile de santé est capable de retrouver une
robustesse à toute épreuve et ne manquera pas une représentation.
 L’accessoiriste

L’accessoiriste, comme son nom l’indique, est chargé du service

des accessoires. C’est lui qui s’occupe des verres, des assiettes,
des filets de sole en banane, du blanc de poulet en biscuit, du
champagne en eau gazeuse, du cognac en eau teintée, des lettres,
des télégrammes, des poignards, des revolvers.
Il ne se manifeste pas dès les premières répétitions. Les artistes
boivent encore dans des verres absents et savourent avec délices
des mets impondérables. Le vicomte, au moment où il doit exhiber le
document révélateur, a plongé la main dans une des poches
intérieures de sa jaquette, en a sorti rien du tout, qu’il a tendu
fièrement à la comtesse. Celle-ci a lu la lettre fatale dans la paume
de sa main ouverte, comme une femme pour qui la chiromancie a
des secrets imprévus.
Peu à peu des accessoires provisoires ont fait leur apparition. La
table du souper, que doit recouvrir un jour une nappe somptueuse,
est pour le moment un pauvre meuble en bois blanc. On sable un
champagne invisible dans des gobelets cabossés.
L’accessoiriste a dans son tiroir des billets de la Sainte-Farce,
auxquels il tient comme à des vrais billets. Quand, à la
représentation, Serge tendra au vicomte une liasse des fausses
banknotes, celui-ci les mettra à l’abri dans son pardessus boutonné.
Mais il ne les possédera pas longtemps. Car, à peine sorti de scène,
il verra se dresser devant lui l’accessoiriste, qui lui dira simplement :
« Mes billets ». Et le vicomte, sans piper, lui remettra le produit de
son chantage.
Le vicomte, depuis les répétitions, a beaucoup changé. Il s’est
vieilli, ou plutôt il a cru se vieillir. Il a étendu sur ses cheveux une
sorte de confiserie argentée. Ainsi, il paraît à peine l’âge qu’il a
vraiment dans la vie.
Pour garantir les personnages de la pièce contre un froid
imaginaire, on a allumé une lampe à verre rouge dans la cheminée,
derrière un solide feu de bois en tôle. C’est là que la comtesse
jettera le paquet de lettres, que l’accessoiriste recueillera
pieusement de l’autre côté du décor, pour la représentation suivante.
Il y a des accessoires qui ne peuvent resservir, tels que les
enveloppes à cinq cachets rouges, que l’on décachète violemment.
Ces enveloppes, c’est l’orgueil de l’accessoiriste, qui les prépare
chaque soir avec amour. Et pourtant, il est bien rare qu’on le félicite
de son zèle. Mais sa satisfaction intime lui suffit. Il aime son métier
et serait très heureux s’il n’avait pas un souci cruel : les armes à feu,
dont il a la garde et dont il doit assurer le bon fonctionnement.
Quand le revolver, qui devait abattre le traître, rate sinistrement,
à la joie indécente du public, il y a quelqu’un, en coulisse, qui blêmit
et qui chancelle, atteint au plus profond de son honneur.
 Brec

Dans ce théâtre où l’on répétait ma pièce, Brec était depuis

trente-cinq ans un employé minuscule, par la taille et par la fonction.
Employé à quoi ? Aux accessoires, peut-être ; mais il n’était pas
accessoiriste en pied. C’était lui qui faisait les commissions de tout le
monde, quand le groom de la direction était absent ou trop
nonchalant. Brec allait chercher une voiture pour l’auteur, un petit
pain pour la comédienne affamée, ou, sur l’injonction du régisseur,
apportait un crayon presque toujours sans mine. Cinq ou six fois
dans sa vie, à des changements de directeur, on l’avait essayé
comme souffleur aux répétitions, mais son registre vocal se
composait de deux voix, une imperceptible et l’autre formidable,
capable de couvrir l’artillerie d’une armée en pleine préparation
d’attaque. Quand on l’avait entendue, cette voix étrange, on
regardait Brec avec une telle stupeur qu’il n’osait plus la sortir.
Le groom de la direction, la femme de ménage du concierge, le
chasseur de la poste étaient tous, et de très haut, les supérieurs de
Brec sur qui, de tous les degrés de la hiérarchie, des injures
tombaient sans relâche, mais elles ne lui arrachaient même pas un
sourire. Il attendait la fin du chapelet, puis faisait un signe de tête,
comme pour dire à l’insulteur : « Vous pouvez disposer ».
Brec était le plus indifférent des martyrs. C’est à peine si parfois,
de la plus petite de ses deux voix, il disait au chasseur : « Ferme ! »
ou : « Ça va ! » au groom de la direction. Des artistes et des auteurs
il recevait parfois un petit pourboire qu’il accueillait sans grâce et
sans hostilité, d’un simple hochement de tête approbateur.
Les légendes les plus romanesques et les plus contradictoires
circulaient sur sa vie privée. Personne ne les avait contrôlées, car il
habitait dix minutes plus loin que le point terminus de son métro. On
prétendait qu’il était de noble famille et d’autres racontaient qu’il
avait une femme énorme et onze enfants perpétuellement en bas
âge.
Les auteurs nouveaux de la maison ne manquaient pas, le soir
de la générale, de demander à Brec : « C’est un succès ? » De
mémoire d’homme, il avait toujours répondu : « Pour sûr », ayant
remarqué d’instinct qu’il n’y avait pas pour lui de réponse plus
profitable.
Brec aimait-il le spectacle ? On ne l’a jamais su. Entré dans la
maison à l’âge de onze ans, Brec, qui avait entendu aux répétitions
les bribes de cent vingt pièces inédites, vu périr ou prospérer douze
directeurs, monter et décliner autant de gloires d’artistes, Brec n’était
jamais allé au théâtre.
 Le prix de Diane en matinée

Le marquis de Hottebrède, ses cheveux noirs cachés sous d’épais

cheveux blancs, le visage soigneusement ridé, est sur le point
d’entrer en scène, pour sa grande scène du troisième acte. A ce
moment le traître Fourval, qui vient de poser ses conditions à la
marquise, sort de scène, laissant Claire de Hottebrède effondrée
sur un beau canapé ancien, prêté au théâtre par un grand
tapissier.

Le Marquis, au traître. — Ça doit être couru à cette heure-ci. Il

est quatre heures dix… Puisque tu n’es pas de la fin du trois, tu
devrais faire un saut jusque chez le bistro, où les résultats sont
affichés…
Le Traître, timidement. — Oh ! mon vieux, maquillé comme
ça…
Le Marquis. — Ça n’a aucune importance. Il n’y a personne
dans les rues… Tout Paris est à Chantilly ou à la campagne. Non,
mon vieux, vas-y. J’ai six louis d’engagés dans la course, tu
donneras le résultat à Fauvel. Il me l’apportera avant mon agonie.
Le Traître. — Oh ! tu fais de moi ce que tu veux ! (Il sort,
pendant que le marquis, après s’être fortement voûté, entre avec
trente années de plus pour la grande scène qui a fait le succès de la
pièce (déjà 234 représentations), et s’avance jusqu’au canapé.)
 La Marquise, levant la tête, et suffoquée de surprise pour la
deux cent trente-cinquième fois. — Renaud !
Le Marquis. — Je vois que vous ne m’attendiez pas. (Elle se
lève. Il s’approche du canapé et s’y assied. Elle est en face de lui. Ils
se regardent en silence.)
La Marquise, bas. — T’as l’résultat ?
Le Marquis, entre ses dents. — On va l’apporter. (Haut.) Je ne
suis pas seul, madame… Notre fils me suit à quelques pas. L’heure
des explications a sonné.
La Marquise. — Gérald, Gérald va m’être rendu !… Ah ! quoi
que vous fassiez de moi, je serai trop heureuse !… Il me semble
entendre son pas… Ah ! quelle émotion !
Le Marquis, bas. — Je comprends. (Entre Gérald (Fauvel). Il se
jette dans les bras de sa mère, qui l’embrasse avec frénésie.)
Gérald, bas, dans le cou de sa mère. — Pellsie première.
La Marquise, bas. — Et Frisky ?
Gérald, de même. — Nulle part.
La Marquise, de même. — Crotte !
Le Marquis a suivi anxieusement cette scène. Il est un peu loin
d’eux. Il se lève et, changeant pour ce jour-là la mise en scène,
s’avance jusqu’au couple. (Noblement.) — Quels que soient vos
torts et vos fautes, je n’ai pas voulu mourir sans vous avoir réunis. (Il
perd la respiration. Dans un souffle)… Eh bien ?
La Marquise, bas. — Pellsie.
Le Marquis, de même. — Pellsie ! (Haut, mais faiblement.) Il y a
des châtiments trop inhumains pour qu’une créature humaine les
prononce… (Bas.) Et je l’avais l’autre jour à Saint-Cloud. (Haut.) Je
suis soulagé parce que mes forces m’ont porté jusqu’ici. Mais…
mais… (Il s’abat lourdement et adroitement sur le sol. Gérald et la
marquise se précipitent sur son corps.)
Gérald. — Mon père !
La Marquise. — Pardonne-moi, Renaud !
 Gérald. — Son cœur a cessé de battre.
Le Marquis, bas. — Où est Esmée ?
Gérald. — Troisième.
Le Marquis. — Chouette. Je l’avais en couverture… Ça va faire
du six contre un au moins… (Il meurt heureux.)
 Loisirs

J’ai reçu la lettre suivante que je tiens à transcrire fidèlement :

« Monsieur,

« Vous avez publié, il y a quelques semaines, un petit article,

intitulé : Le Prix de Diane en matinée, où vous montriez des artistes
dramatiques qui, pendant qu’ils interprétaient un drame des plus
austères, demeuraient préoccupés par les paris qu’ils avaient faits
aux courses. L’inconvénient de ces fantaisies est de laisser croire
qu’il n’y a place dans l’esprit de certains comédiens que pour des
soucis aussi frivoles. Je tiens à protester au nom de ceux qui,
comme moi, ne parient jamais aux courses, et consacrent le meilleur
de leur soirée à jouer au poker.
« Ma protestation, si vous le voulez bien, restera anonyme…
Aussi bien, hélas ! garderait-elle ce caractère, vis-à-vis de vos
lecteurs, même si je signais de mon nom… Je suis peu connu, mon
rôle se bornant la plupart du temps à faire part à des personnages
de la pièce, titrés ou riches, de l’arrivée, dans une antichambre
supposée, d’autres personnages importants, et de quitter la scène
sur la réplique : « Faites entrer. »
« Je suis parfois en habit noir, d’autres fois en livrée à boutons de
cuivre. Depuis quelques semaines, sans doute parce que mes
maîtres sont de nouveaux riches (je ne connais pas exactement la
pièce) ou bien parce que mon auteur a des idées un peu
conventionnelles sur la tenue des domestiques du grand monde, je
porte une culotte et une chaude perruque blanche, qui ne reste
heureusement sur ma tête que pendant les courts instants de ma
présence en scène.
« Le reste du temps — et il en reste — je suis installé dans la
loge somptueuse d’une jeune fille ruinée et persécutée, en
compagnie d’un vieux duc qui, lui non plus, n’est ni du « deux » ni du
« trois ». Nous jouons au poker.
« Là, les différences de classe, établies arbitrairement par
l’auteur, disparaissent tout à fait, et chacun reprend le rang qu’il
mérite… L’humble serviteur que je suis domine de cent coudées le
vieux duc (qui n’a d’ailleurs que 22 ans). J’ai l’habit d’un valet, mais
j’ai l’âme d’un bluffeur de premier ordre…
« Ci-joint mon nom et celui de mon théâtre, que vous voudrez
bien ne pas publier. Mais venez nous voir un soir, vers 9 h. 1/2. Vous
ferez le quatrième.
« Mes sentiments les meilleurs. X… »

Oui, monsieur X…, j’irai certainement vous voir un soir. Et je vous

rappellerai le souvenir glorieux de Joliet, de la Comédie-Française,
qui, lorsqu’il n’était pas d’un acte, n’hésitait pas à quitter rapidement
son costume, à reprendre pour une demi-heure ses habits de ville
pour aller continuer une partie d’échecs au café de la Régence. Les
vrais joueurs ont ainsi de ces traits magnifiques, dont les âmes
mesquines de ceux qui ne jouent pas ne peuvent comprendre la
beauté.
 Le feu sacré

Ce grand garçon est toujours bien habillé. Donnons-lui vingt-cinq,

trente ou trente-cinq ans, sans préciser, car il n’est pas seul de son
espèce.
Il a certainement la vocation du théâtre, et c’est d’autant plus
méritoire qu’il n’a aucune espèce de dons… Il est en bois et met une
voix sourde et monotone au service d’une intelligence médiocre.
Mais sa taille avantageuse, l’élégance de ses costumes
permettent de l’utiliser et de lui confier le rôle de monsieur en
smoking qui, au début d’un acte, écoute, le cigare à la bouche, un
autre fumeur visiblement plus important. Ou bien encore, il est le
jeune homme, en pantalon et chemise blancs, qui passe au fond du
théâtre, une raquette à la main. Parfois il a la gloire d’être censé
faire la cour, à la cantonade, à l’héroïne, et sera même honoré, dans
la pièce, des soupçons d’un jeune premier à 10.000 francs par mois.
Mais, d’une façon générale, ce qui lui arrive se passe en
coulisse. Il case donc assez vite son rôle dans sa mémoire et peut
employer ses loisirs à écrire des lettres sans nombre aux auteurs
qui, bien entendu, sont toujours des « maîtres ».
Au nouvel an, la première carte que ces écrivains reçoivent est
celle de ce fidèle.
Il s’éloigne passablement de l’ancienne conception des acteurs
errants, compagnons sans souci, méprisés par les marchands
drapiers. C’est un garçon sobre, réservé d’allures, mesuré dans ses
propos. Il n’aime ni l’aventure, ni la fantaisie, ni même le théâtre. Il
lui plaît seulement d’exercer la profession d’artiste dramatique. Voilà
en quoi consiste son feu sacré.
Flamme point négligeable, certes, puisqu’elle le stimule sans
cesse et le pousse à écrire ses lettres inlassables aux auteurs, à leur
faire, à des intervalles courts et réguliers, de persévérantes visites.
« Maître, ce n’est pas mon genre de me mettre en avant et de
venir parler de moi. Il faut que je me force… »
C’est qu’il est sincère. Il faut qu’il se force. Mais il se force bien.
 Les Attributs du pouvoir

On s’illusionne souvent sur la puissance des gens, sous prétexte

qu’ils semblent occuper un emploi d’une haute importance, et qu’ils
sont vêtus d’un costume impressionnant.
Il y a une quinzaine d’années, un dimanche, je passais, vers
deux heures, devant un grand théâtre du Boulevard. On y jouait le
Bossu, une pièce que je ne me lasserai jamais de revoir… J’y
retrouve, à chaque représentation, ma haine fidèle du tortueux
Gonzague et mon éternelle sympathie pour Cocardasse et
Passepoil… Je suis les péripéties du drame avec angoisse, comme
si ce qui est écrit et que je sais par cœur n’était pas définitif, et
comme si le traître allait échapper, ce jour-là, à la fatalité du texte.
J’entrai par l’escalier des artistes, salué par un geste amical du
concierge qui me connaît très bien (pas sous mon nom d’ailleurs, car
il m’a donné un jour le nom d’un de mes confrères ; mais ceci n’est
qu’un détail, et l’important est d’être connu).
La lumière du jour pénètre rarement dans les coulisses des
théâtres, même en matinée. Je me dirigeai vers la Régie, où un
artiste d’un certain âge était assis devant un petit bureau. Il jouait
dans la pièce et se trouvait tout habillé et tout maquillé pour entrer
en scène.
— Bonjour, comment ça va ? Y a-t-il moyen d’assister à la
matinée ?
 — Oh ! monsieur… je suis désolé… J’ai les ordres les plus
sévères. En l’absence du directeur et du secrétaire, on me défend
de donner des entrées à qui que ce soit… Je vous prie de ne pas
m’en vouloir. Mais je n’ai aucun pouvoir pour cela, et c’est une
consigne formelle à laquelle je suis forcé d’obéir…
Celui qui parlait ainsi n’était autre que le Régent de France, vêtu
d’un habit somptueux, magnifique, couvert de pierreries, orné par
surcroît d’un large cordon bleu clair qui ne se donnait pas à tout le
monde.
 Le texte de Mme de Juxanges

La personne qui, dans la pièce précédente, avait répété —

jusqu’au troisième jour avant la générale — le rôle de la concierge
du deux avait reçu cette fois la mission d’incarner, dans la pièce
nouvelle, une femme du grand monde, Mme de Juxanges.
La liste des personnages féminins comprenait d’abord l’héroïne,
puis la mère de l’héroïne, puis la rivale d’amour de l’héroïne, puis
toutes les bonnes, puis Mme de Livrac, puis Mme de Juxanges.
Il n’y avait pas, entre ces derniers noms, la petite conjonction
« et » qui eût donné à l’interprète de Mme de Juxanges une
apparence de « vedette américaine ».
Mme de Livrac, elle, était chargée de deux phrases : « Je le crois
galant homme », et, plus loin : « Vous verrez qu’il fera sa
soumission. » Mme de Juxanges n’avait rien à dire. Le jour de la
lecture, on lui remit une feuille de papier, qui portait dans le coin, en
grosses lettres de ronde : Mme de Juxanges. On indiquait
simplement sur quelles répliques de ses camarades elle devait
entrer, puis sortir.
Le directeur avait dit à l’auteur : « Il faudra lui donner une petite
phrase. » Puis il avait dit à sa pensionnaire : « On vous piquera
quelques mots. »
Chaque jour, à midi trois quarts pour une heure, Mme de
Juxanges arrivait la première à la répétition dans une longue six-
cylindres, don de M. Roibourg, grand brasseur d’affaires, et
commanditaire de la maison. L’automobile stationnait devant l’entrée
des artistes jusqu’à cinq heures. Pendant ce temps, Mme de
Juxanges, assise sur une chaise rustique dans l’ombre du plateau,
entretenait une conversation un peu stagnante avec la cuisinière du
trois et le facteur du deux.
D’ordinaire, M. Roibourg arrivait, ses affaires brassées, vers cinq
heures, et prenait livraison de Mme de Juxanges.
Le directeur, à deux ou trois reprises au cours des répétitions,
avait dit à l’auteur : « Lui avez-vous mis sa phrase ? » L’auteur
répondait toujours : « Demain. » A vrai dire, il éprouvait une difficulté
singulière à prêter la moindre réflexion ou remarque à un
personnage aussi peu défini.
Pourtant il dit de lui-même, quelques jours avant la générale, en
voyant répéter Mme de Juxanges : « Il faut que je lui donne sa
phrase. » Mais le directeur, par esprit de contradiction, avait dit à mi-
voix : « Il y a des choses plus pressées. Établissez-moi d’abord votre
baisser de rideau du deux, qui n’y est pas du tout. »
Ce qui ne l’avait pas empêché de demander sévèrement, le
lendemain :
— Et la phrase de cette petite ?
Enfin, visité par l’inspiration, l’auteur trouva : Mme de Juxanges
dirait à Mme de Livrac :
— Je ne suis pas très tranquille sur le destin de ce petit ménage.
Le jour de la générale, ce propos de mondaine avertie fut proféré
très au-dessous du ton par une personne étranglée d’épouvante. On
entendit cependant ceci :
— Je ne suis pras tès tancrille sur le dessin de ce petit mén…
La finale age, ce serait pour une autre fois.
Il y eut simplement aux fauteuils un petit : Oh ! oh !… isolé, qui, si
discret qu’il fût, fut perçu de la salle entière.
A l’entr’acte, dans un coin obscur du plateau, le directeur passait
quelque chose de sérieux à M. Roibourg.