вход по аккаунту


Digitalis Research in BerlinЦBuchЧRetrospective and Perspective Views.

код для вставкиСкачать
Digitalis Research in Berlin -Buch-Retrospective and Perspective Views**
Kurt R. H. Repke,* Rudolf Megges, Jiirgen Weiland, and Rudolf Schon
“The following remarks consist partially
of matter of fact, and partially of opinion. The former will be permanent; the
latter must vary with the detection of
error, o r the improvement of knowledge. I hazard them with diffidence, and
hope they will be examined with candour.” These declarations, which stem
from the famous book “An Account of
the Foxglove and some of its Medical
Uses” by physician William Withering
in 1785 in which he introduced preparations from digitalis leaves in the therapy
of dropsy (cardiac failure), are cited
here by the senior author because of his
awareness of the difficulties in present-
ing a balanced report on his life-long research project on the further development of digitalis. His decision to devote
himself to digitalis research originated
at the bedside, when as a physician he
experienced the grim final stages of cardiac failure in which no real help for the
patients is possible. Unfortunately. his
research project did not fit into the research program decreed by the Ministry
of Science of the German Democratic
Republic, so that he was ordered to stop
the digitalis project in favor of biomembrdne studies. Fortunately, he got round
the ban simply by labeling the digitalislike acting steroids as probes for the cell
membrane-located Na+/K+-transporting ATPase which he had just recognized as the digitalis target (receptor) enzyme. These and other ventures by the
authors are collated here for the first
time. The aim of this review is to foster
straightforward research for solving a
major challenge: the development of
steroidal drugs for the prevention and
cure of cardiac failure.
Keywords: digitalis . drug design .
medicinal chemistry . pharmaceuticals .
1. Introduction: The Challenge of Digitalis Research
1.1. The Underlying Medical Demand
The challenge of digitalis research derives from the need
for new drugs for the prevention of cardiac failure and the
successful treatment of the failing heart. Specifically, the
need follows from: the prevalence and mortality of heart
failure in developed countries; the fact that the cardiac drugs
available are clearly inadequate to restore health or even to
minimize discomfort and disability; and the condition that
previous research efforts to provide drugs satisfying the above
requirements have widely failed. Selected facets of the latter
studies were previously reviewed by the authors of this review.“. 2l
The incidence and prevalence of congestive heart failure have
increased in recent years,’31and as the average age of the population increases, it is expected to continue to increase. The prognosis
after first diagnosis is
grim; less than half of the
patients survive more
than five years.[41 The
survival rate depends on
several variables, essentially defined in Figure 1.
The therapeutic value
of preparations from
Fig. 1. Clinical manifestations of myocardial disease. The Scheme shows the posDigitalis plants was dissible coincidences of left ventricular dyscovered by Withering
function (LVD). exercise intolerance, and
ventricular arrhythmias as the most commore than 200 years
mon manifestations of the failing heart
ago.@] Digitoxin (1) and
syndrome. Clinical heart failure (CHF)
digoxin (2) (Fig. 2), the
corresponds to the coexistence of LVD and
exercise intolerance. Sudden death (SD)
predominant active mateoccurs in patients with arrhythmias and
rials, still belong today to
predominantly in those with LVD and
C H F (from ref. [ 5 ] ) .
the drugs most common-
Prof. Dr. K. R. H . Repke, Dr. R. Megges. Dr. J. Weiland. Dr. R. Schon
Max-Delbruck-Centrum fur Molekulare Medizin
Robert-Rossle-Strasse 10. D-13125 Berlin-Buch (Germany)
Telefax: Int. code +(30) 949-4161
Definitions and abbreviations: Digitalis: generic term to cover all steroids
which act cardlotonic by inhibiting Na+/K’-ATPase by occupancy of its digitalis recognition matrix and binding cleft; Nat;Ki-ATPase, the N a + / K i transporting adenosine triphosphate phosphohydrolase (EC 3.6.1 3 7 ) . residing
in the plasma membrane of all mammalian cells. which effects the conversion
of scalar phosphoryl energy of ATP into vectorial electrochemical energy: for
the molecular mechanism see K. R. H. Repke. R. Schon, B i d . Res. 1992, (57,
31-78; Biochirn. Biophrs. A l m 1992. 1154. 1 -16.
Digitalis and Digitalis-Like Drugs
1.2. Paradise Postponed
Fig. 2. Structural lortnulae ofdigitoxin ( I ) and digoxin (2) (R = tridigitoxosyloxy).
Compound 1 is shown in 1422-conformation and 2 in 14/21-conformation in which
the C22 proton o r the C11 protons, respectively, are localized opposite t o the
C14P-OH group High potency is preserved in conformationally semi-rigid cdrdenolide derivative.; In which the hutenol~deC21 protons, due to bulky substituents at
the C22 atom (ci. Fig 9). are forced into a potential energy well near C14C(-OH.
Hencc. the 14,21-conformer appears to be the receptor-bound conformer [ 7 ] .Note
the cls-,junctron o f t h e rings C and D of the steroid framework is not achieved in the
hiosynthettc pathways of humans and animals (except for a few toads) (cf. Sections 2 5 and 2.6)
ly prescribed for the treatment of cardiac failure.[3s81In 1930,
the famous heart specialist Wenckebach[’’ concluded: “Digitalis treatment is one of the most important and serious duties of
the general physician; it demands a great deal of skill, power of
observation, keen interest, and experience. A long life is too
short to learn enough about this wonderful drug.” More recently, however. considerable controversy has arisen about the role
and value of digitalis in treating patients with cardiac failure.[3]
The open question at present, is that one cannot draw any
conclusions about the effects of digitalis on survival.” Physicians are looking forward to additional first-line agents that are
relatively safe and that can be used as monotherapy to improve
the quality of life, retard progression of the disease, and prevent
One may expect in the coming years that
every effort must and will be made to develop new compounds
that will not only beneficially affect the hemodynamic and
functional impairment of patients with cardiac failure, but will
also hopefully contribute to reaching a n achievable goal, namely, the prevention of the clinical manifestation of cardiac insufficiency.“. ’1
During the last decade many new drugs have been developed
and tested on patients with heart failure at a rate of one new
agent every month. An overview of the more interesting agents
and their putative major mechanisms has been presented in a
detailed, but balanced way by G. Grupp.[”l Phosphodiesterase
inhibitors such as amrinone and milrinone seemed to hold great
promise, but finally yielded disappointing results. In contrast to
the digitalis compounds such cardiotonic drugs lose their effectiveness when they are needed most, that is in severe heart fail~ r e . [ Moreover,
reports on increased mortality[’4- “1 have
dampened the initial enthusiasm for the use of phosphodiesterase inhibitors: “paradise postponed”.[”] The return on the
huge investment of money, time, manpower, and resources to
develop an orally administrable inotropic drug to supplement o r
replace digitalis compounds has generally been disappointing.
This has had a devastating effect on the further development of
this class of non-digitalis inotropic drugs.”’]
Only digitalis, diuretics, and inhibitors of the angiotensinconverting enzyme each fulfill some of the criteria of a first-line
agent in the treatment of chronic heart failure. However, none
of these drugs satisfies all of the desired characteristics, and
none can optimally manage the heart failure state when used
alone.[”] Unfortunately, converting enzyme inhibitors and diuretics have the tendency to impair renal function, while digitalis compounds tend to improve renal function. However, digitalis intoxication has continued to be one of the most prevalent
adverse drug reactions encountered in clinical practice due to
the narrow margin between the therapeutic and toxic doses and
the marked variations in individual sensitivity.[201Nevertheless,
there has been little decline in the use of digitalis over the last
five years, indicating that the mentioned newer agents for the
treatment of cardiac failure have not replaced the widespread
use of digitalis.[31
1.3. Apparent Causes for the Previous Lack of Success
The search for cardiotonic steroids with an increased therapeutic range by modification of the structures of known digitalis
Kurt R. H . Repke, born in 1919, obtained his medical doctor’s degree in 1945. Between 1945
and 1950 he trained in various medical disciplines at hospitals in Hamburg and Potsdam as well
as, under Professor Gerhardt Katsch, at the Clinic for Internal Medicine of the University in
Greifwald. To acquire a profound understanding of the pharmacological and biochemical
foundations o f drug therapy, he worked and habilitated under Paul We1.y at the Institute of
Pharmacology of the University in Grelfiwald (1950- 1955), andsubsequently until f 963 under
Professor Karl Lohmann at the Institute of Biochemistry of the Academy of Sciences in
Berlin-Buch. He followed Lohmann as the director of this institute (1964-1971) and then
headed the Biometnbrane Section of the newly formed Central Institute of Molecular Biology
of the Academy (197f - 1984). Since he has been an emeritusprofessor he haspurstred Ivith full
vigor his research aims which include in addition to digitalis research the elucidation of the
complex mechanism of the interconversion of the scalar phosphoryl energy of A T P into the
vectorial energy of cation gradients through hiomembranes. Professor Repke has been a member of rhe Deutsche Akademie der Naturforscher Leopoldina and a honorary member of the
Cardiac Muscle Society of USA. He devotes his leisure time to the study of Gorthe’s genius.
C h m lrir Ed. Engl. 1995, 34, 282 -294
K. R. H. Repke et al.
compounds appeared unsuccessful for several decades. This has
electrostatic requirements for binding to the catalytic center of
been interpreted as probably reflecting an already insuperably
a target enzyme than it has been to elaborate these lead struchigh degree of structural optimization in the therapeutically
tures into potent inhibitors with a small number of
used natural compounds of herbal origin.[”] However, the lack
The superiority of that “direct” strategy, as described for the
of progress might have followed from conceptual limitations or
discovery of drugs, over existing methods remains to be investiexperimental weaknesses in the research endeavors.
The synthetic modifications were based on equating the natuDespite impressive advances in X-ray crystallography of
ral compounds with lead structures. Specifically, the derivatizamacromolecules, availability of high-quality crystals remains
tions were focussed 011 the side chain at C17 in 1 and 2, errothe major limiting factor in the exploitation of this technique.
neously taken to be the pharmacophoric lead s t r ~ c t u r e . [ ~ ’ - ~ ~ ]Otherwise, modeling based on the knowledge of homologous
The activity of the derivatives was primarily evaluated in animal
proteins is now generally held to provide an insufficiently stable
model systems. They allow the measurement of inotropic and
base for rational drug design.[331In addition, for the majority of
toxic effects, but not the discrimination between the diverse
protein sequences with little significant homology to known
molecular mechanisms underlying the observed effects (cf. Secstructures, the problem of predicting secondary and tertiary
tion 2.4). Such observations preclude the derivation of self-conaccurately enough for drug design applications is
sistent structure-activity relationships, as emerges from pertifelt to be still insurmountable.[331
nent review^.^'^-'^^ Thus, the initially untargeted strategy for
As the “direct” strategy in drug design has been mostly not
the chemical modification necessarily resembled a “molecular
applicable, “indirect” routes based on the analysis of structureroulette”. However, for a rational approach to the development
activity relationships have more generally been pursued. These
of drugs, an understanding of the effect-triggering interaction
routes had to be followed also in digitalis research (cf. ref. [1, 2,
between active agent and receptor on the molecular level is
35-71]), As long as neither the site nor the mechanism of digineeded, as emphasized by Ariens in 1976.[271He felt that basic
talis action were known, we had to discover by “untargeted”
research into physiological and biochemical processes would
strategy the target enzyme (receptor) to clear the path to the
become the cornerstone for new drug development in the near
targeted strategy (see Sections 2.1 and 2.2). The ensuing multifuture.
stage development (cf. ref. [72]) of digitalis research in BerlinBuch led to the targeted strategy, comprising the following
stages: the Na+/K+-ATPasewas identified as the digitalis target
1.4. The Early Illusion and the Present Realities
(receptor) enzyme and its role in controlling cardiac contractility was derived (Section 2.2) ; the “microscopic” digitalis mechanism of Na+/K+-ATPase inhibition was clarified (Section 2.3);
As reviewed by Parascandola1281very early some attempts
the capabilities and limitations of biological and molecular
were made to relate the pharmacological action of organic compounds to their structure while structural organic chemistry was
screening test systems were evaluated (Section 2.4); the lead
still in its infancy. Nevertheless, Brown and Fraser began their
structures for digitalis-like action were discovered in the course
of an extended “chemical hunt”“. 2 3 3 5 - 7 1 1 (Sections 2.5 and 3);
promising work in 1869 with the following declaration of faith:
“There can be no reasonable doubt that a relation exists bethe mystery surrounding the chemical identity of the putatween the physiological action of a substance and its chemical
tive endogenous digitalis was attributed to the lack of specificity
composition.” In 1877, Brunton even speculated that physicians
of the various analytical procedures applied (Section 2.6).
would soon be able to predict the pharmacological action of any
Based on the achieved insights, some facets of the most
compound from its constitution. This early hope turned out to
recent developments of the research will be delineated in Secbe an illusion that still applies. Consequently, in 1992 K ~ n t z [ ‘ ~ ] tion 3.
stated that we cannot design drugs with a specific mode of
action and acceptable biological properties from first principles.
What we can reliably expect is to be able to design competitive
2. The Stages of Digitalis Research
enzyme inhibitors, and to begin the long process of drug development from such a sensible starting point.[291
Over thirty years, digitalis research has been stimulated by
The cornerstone of the newest strategy for design of novel
mutual supplement and criticism of the research groups in Syddrugs has been proclaimed to be the accurate knowledge of the
ney and Berlin-Buch. Although both studying the digitalis
three-dimensional structure of the potential target proteins,
problem, their research efforts differed in many ways due to
which are complexed with a great variety of ligands [inhibitors)
different experiences and preferences as well as different
in their recognition and triggering p l a ~ e . [ ’ ~ As
- ~ ~stated
by the
methodological and conceptional approaches. The work in
staff of a specialist drug design company in 1991[321-knowlSydney has focussed on the isolation and structure determinaedge of the three-dimensional structure of pharmacologically
tion of natural substances and, above all, the synthesis of digitalis analogues by substitution of the lactone group at C17 by
significant receptor-ligand complexes at the level of resolution
different open side chains or/and substitution of the sugar comachieved by X-ray crystallography has the potential for proponent at C 3 by different ester groups as well as the determinafoundly influencing and speeding the discovery and development of lead compounds. Since the desolvation of ligand and
tion of the inotropic activity of the compounds on surviving
protein in their going from free to bound form are not considpreparations from the cardiac muscle of the guinea-pig. The
ered in this procedure, it has been easier to discover various
voluminous results of the Australian groups were comprehennovel lead compounds by satisfying the evident nonpolar and
sively published by Wright in 3960;[731Thorp and Cobbin in
Angew. Chem. Inr. Ed. Engl. 1995, 34, 282-294
Digitalis and Digitalis-Like Drugs
1967;[741Thomas, Boutagy, and
Gelbart in 1974;r7s1 Thomas,
Gray. and Andrews in 1990[241
and Thomas in 1991.[251Thus,
their brilliant contributions will
not be reviewed here for the sake
of brevity. too.
2.1. The Role of
The metabolic transformation
of digitalis glycosides (reviewed
i n refs. [76, 771) can sometimes
render the interpretations of biological potency data in terms of
structure- activity relationships
more difficult. This becomes evident from the following examples. When orally applied,
lanatoside C undergoes deglucosylation and deacetylation by microorganisms in the intestinal
tract before the glycoside becomes absorbed in the form of
digoxin (2) .17x~- 801 Lanatoside C
is thus a prodrug. Pentaacetylgitoxin requires deacetylation to
16-acetylgitoxin by microbial and
animal hydrolases before it becomes effective; it presents thus
an example of drug latentiation.i8i - 8 3 1
As illustrated in Figure 3, digitoxin (1) and digoxin (2) undergo,
by cleavage of the tridigitoxoside
chains, a slow, stepwise degradation to the aglycones which are
rapidly inactivated by epimerization and conjugation of the
released hydroxy group at
C3.[84-401Taking into account
the locus and speed of biotransformation on the one hand and
the biological activities of the
metabolites on the other (cf.
Fig. 3), the following conclusions
can be drawn: As a rule. the ultimately active agents are not
formed by biotransformation.
The biotransformations
react i o n ~ . [ ~ ~Due
. ~ ' ]to its slow removal, the sugar side chain prolongs the half time of the
active molecules in the animal
body.'84, 8 5 . 9 3 1 However, the
large variations in acute potency
Angrtt Ciwni. in(. Ed. Drgl 1995. 34, 282-294
2 (46.6)
9 (39.7)
10 (37.5)
11 (41.1)
12 (32.5)
Fig. 3. Biotransformation of lanatoside A (3) in the animal body. The deacetylation and deglucosylation (3 --t 1) are effected
by microbial enzymes in the intestinal tract, allowing absorption of 1 [78-801. Then, liver enzymes bring about C12fl-hydron9), conjugation with sulfuric acid (8 + 10)
ylation (1 + 2). stepwise separation of the three digitoxose residues (1 + 8; 2
or epimerization of C3B-OH (8 4 12) [84-901. The latter two reactions are the decisive detoxification steps as the drop in
the Gibbs energy of interaction with the receptor enzyme (given below the formulae in parentheses, expressed in - kJmol-')
indicates. Scrutiny of the AGO' values suggests that the sugar side chain for the most part remains outside the digitalis
recognition matrix and binding cleft of the receptor and that the sugar residue next to the steroid nucleus lies at the mouth
of the receptor cleft. If so, the lactone function is positioned at the bottom of the cleft such that the cleft length amounts to
about 1.88 nm (91, 921.
shown by digitalis glycosides of different constitution and configuration cannot be explained by differences in the biotransformation.[76. 7 7 . 9 3 -961
The 12j-hydroxylation of digitoxin (1) to form digoxin (2)[971
is no prerequisite of cardiotonic action;[971as indicated in Figure 3, it rather involves a small loss of potency. The double bond
in the butenolide side chain can be hydrogenated by microorganisms in the intestinal tract.[76.7 7 , 791 Since this appears to be
irreversible in the animal tissues, the lactone residue cannot
serve as hydrogen carrier in the intermediary metabolism[76]as
suggested earlier. The double bond orients the H-binding acceptor oxygen atoms of the butenolide towards the H-donating
amino acid residues of the recognition matrix of Na+/K'-ATPase involved in long-range attraction and high potency binding['] (cf. Fig. 9). Moreover, the double bond is important for
stabilizing the lactone side chain against hydrolysis at physiological p H which is accompanied by loss of potency.[981
2.2. The Target Enzyme and Its Role in Controlling
The results of our studies on the metabolic fate of digitalis
compounds led us to the inviting conclusion that the question as
to the biochemical mechanism of digitalis action is not related to
the biotransformation of the herbal compounds in the mammalian body, but that this question appears to coincide with the
question as to the nature and function of the digitalis receptor.[761This conception appeared in 1961 rather daring as it
emerges from a look back at a symposium on enzymes and drug
action held at that time.[99]When asked to comment on receptor
models, R. J. P. Williams replied that he understood the word
"receptor" as a synonym for lack of knowledge, and Chain
added to not believe that the enzyme approach is the right way
to discover new drugs nor to understand in a general way the
mode of action of drugs.
Nevertheless, our group wanted to test whether the Na'/K+ATPase, only just shown to be inhibited by the digitalis glycoside ouabain,[l""l exhibits the characteristics to potentially
serve as the digitalis target (receptor) enzyme. To this end, we
extensively compared the many conditions under which digitalis
compounds act on contractility, N a + / K + transport, and Na'/
K'-ATPase activity. At each level, correlations were demonstrated for all 20 characteristics of digitalis action examined, for
example locus of action, species differences of digitalis sensitivity, structure-activity relationships, ionic antagonism and synergism, cation requirements for action, time course for development of effects, dependence on specific functional states, and
range of effective digitalis concentrations.[76.l o ' - 'O'] Since the
early sixties, our findings and conclusions have been confirmed
and extended in several ways by various research groups." O S - 31
At long last, after decades of controversy, a consensus has
emerged that the inotropic effects of digitalis compounds result
from binding to and inhibition of Na+/K+-ATPase as stated by
T. W Smith in 1988.[201Okita et al. reported in 1990 that, in
accordance with its receptor function, digoxin (2), administered
therapeutically in the clinical setting, inhibits the N a + / K '
pump in the human heart.["41
The N a + / K +-ATPase is the biochemical machinery in the
Na '/K pump which maintains the normally high intracellular
K . R. H. Repke et al.
concentration of K ' ions and low intracellular concentration of
Na' ions in cardiac muscle and other excitable cells. As deduced
by Repke in 1964," ' 'I the actual Na' concentration regulates
indirectly the Ca2+concentration in cell interior which has been
known to be the final regulator of cardiac contractility. More
specifically, the integrating assessment of the then available experimental data on the relationship between the contractility of
the cardiac muscle cell and the flows of K', Na', and C a + ions
across the cell membrane led him to the hypothesis that a transient increase of the intracellular Na' concentration, resulting
from the digitalis-produced reduction of the N a + / K + pumping
capacity of the cell by inhibition of a portion of the N a + / K f ATPase total, plays the primary role in the control of cardiac
force.["'- ' I 6 ] The indicated interrelationship has often been
questioned, but in 1985 it was finally fully corroborated in using
Nat -selective microelectrodes. Lee[' ' established the essential
role of the increase in the intracellular Na' activity for the
action of digitalis at low and high concentrations. Specifically,
he showed that digitalis produces a parallel increase in Na'
activity and contractile force, and that these parameters remain
closely correlated during the positive inotropic effect. The increase of the chemical activity of Na' ions leads, via the N a + /
Ca'+ exchanger, to an increase of the chemical activity of the
Ca2+ ions through enhanced Ca2+ influx into and reduced
Ca" efflux from the cardiac muscle cell as reviewed elsewhere."]
2.3. The "Microscopic" Mechanism of Na+/K+-ATPase
In 1953, Wilbrandt et a1.[1181accounted for the strong dependence of the cardiac action of digitalis compounds on contraction frequency with the hypothesis that these steroidal agents
interfere with cation transmembrane flows occurring during excitation and recovery. This suggestion was based on the knowledge that other steroids, especially corticosteroids, play an important role, possibly as cation carriers (Fig. 4), in cation
Fig. 4. Assumed structural analogy between a strong inhibitor of N a + / K ' transport (digitoxigenin; left) and a presupposed chelate formed between 1l-deoxycorticosterone and an alkali metalcation (M:right). O n this basis, the action of digitalis
compounds was suggested by Wilbrandt [119] to be caused by competitive antagonism towards cation-laden corticosteroids which were thought to be complexed with
Na+/K+-transportingATPase and to serve as cation carriers across the membrane
of the cell (from ref. [119]).
transport. In support of the above hypothesis, Wilbrandt" ''1
reported that corticosteroids antagonize the inhibitory action of
cardiotonic steroids on cellular Na '/K
transport. Consequently, Wilbrandt" 2 1 1 considered the possibility that the digitalis effect may be on a corticosteroid-carrier part of N a + / K + ATPase rather than on the enzyme protein. However. our
group[1221showed that corticosteroids do not attenuate a digi+
Angeu. Cheni. lnt. Ed. Engl. 1995, 34. 282-294
Digitalis and Digitalis-Like Drugs
talis-induced Na + / K +-ATPase inhibition, and the observations
appeared to exclude a direct, functionally antagonistic competition between corticosteroids and cardiotonic steroids for binding to a common site on that enzyme. The antagonistic action of
corticosteroids rind cardiotonic steroids found by Wilbrandt
and we is^['^^' for intact cells appears to be based on the induction of biosynthetic increases of the number of Na+/K+-ATPase moleculesr’L 4 1 compensating for the portion of digitalis-inhibited enzyme molecules.
The inhibitory effect of digitalis compounds on Na+/K+-ATPase activity arises from their interaction with the recognition
part and effect-triggering binding site on the enzyme protein
which we have further characterized by the following findings.
Large excesses of ATP over digitalis with respect to the concentration d o not antagonize digitalis binding to the enzyme.[1251
There is a competitive K effect on digitalis binding under certain conditions. but, as shown by us.[1251its molecular basis is
not competition for a common binding site on the enzyme, but
competition for the ATP- o r digitalis-binding enzyme conformation. Using definitions given by Monod et a1.,[1261digitalis
compounds are thus noncompetitive, heterotropically acting allosteric inhibitors which produce the transition from an active
to an inactive conformational state of enzyme protein. More
specifically, a common denominator of the inhibitory action of
digitalis compounds has been shown by our group[t271to be a
large gain in entropy in the enzyme protein associated with a
loss of high ATP affinity to the catalytic center.
The search for the proper locality of the digitalis recognition
and binding protein matrix has intensively been approached by
several groups through elucidation of the connection between
distinctions in digitalis affinity (ouabain taken as a prototype) and those in the primary structure of Na+/K+-ATPase
(Fig. 5 ) . However, with increasing knowledge all hitherto published proposals (see review in ref. [2]) have turned out to be
untenable. As we shortly shall show elsewhere. the digitalis
binding cleft is formed by two membrane-penetrating peptide
sequences (H1 in Fig. 5 ) which through their association bring
about the interplay between the two catalytic a subunits in the
diprotomeric holoenzyme (E,!I)~
such that, by intercalation of a
digitalis molecule, the catalytic cycle becomes interrupted.
2.4. The Comparison of Biological and Molecular Screening
Test Systems
The most suitable method for the quantitative determination
of the positive-inotropic (cardiotonic) effect of a cardioactive
compound is the assessment of its influence on the isometric
contraction curve measured in isolated cardiac muscle preparations from some animal species. However, to obtain reliable
data for the comparison of the inotropic potency of the various
cardioactive compounds, it is essential to perform the experiments under identical conditions.[1311Because of the variety of
test conditions used, it is not easy or is even impossible to integrate in terms of structure-activity relationships the numerical
values reported in the literature.[t32.1 3 3 1 A comprehensive survey of the structure-activity relationships found in guinea-pig
cardiac preparations under identical conditions has recently
been presented by R. Thomas et al.[241
Important for the realization of the message of this review is
the understanding that there are two basic possibilities for misinterpretation of observations with animal model systems. First,
all inhibitors of the N a + / K + transport system irrespective of
their microscopic inhibitory mechanism (see below) produce
positive inotropic effects if they do not additionally affect other
biochemical systems which preclude the manifestation of the
inotropic action.[”’] Second, some inhibitors of N a + / K f transport, even when effective through occupancy of the digitalis
binding cleft, can fail to produce a positive inotropic action
when they additionally affect biochemical systems which preclude the manifestation of their inotropic faculty. Prominent
examples for both cases were diagnosed[’’ by successive use of
the “macroscopic” and “microscopic” test systems characterized below.
The macroscopic screening with N a + / K +-ATPase proved to
be an only initially useful test. It restricts itself to the assessment
Fig. 5 . Schematic representation of
the primary structure and spatial organization of the catalytic subunit of
Na ‘/K+-ATPase. The amino acid
chain penetrates the membrane of the
cell eight times. The membrane crossing sequences HI H4 carry the extracellularly disposed HI - H2 junction
(Q111 N122) and H3-H4 junction
(E3077E312) which were supposed to
interplay in building the digitalis
recognition and binding matrix [2]
The H4 stretch directly mediates the
communication of the matrix with the
phosphorylatable aspartyl residue
D369 in the catalytic center of the enzyme. The scheme shown here is a
combination of the figures in refs. [I28
K. R. H. Repke et al.
of the apparent Gibbs energy change (AGO') in the formation of
the equilibrium complex of an inhibitor with the enzyme under
standardized conditions; the experimental details were published elsewhere by us.[9z.134, i 3 5 1 The availability of the AGO'
values allowed us to apply the extrathermodynamic approach to
the quantitative analysis of structure-activity relationships.[", 134, ' 3s1 This procedure clearly requires that a uniform
macroscopic and microscopic mechanism underlies the observed inhibitory effectiveness. However, fulfilment of this prerequisite cannot be derived from studies on the inhibition of
Na+/K+-ATPase (or N a + / K + transport): therefore the investigation is named a macroscopic screening test. Specifically, many
inhibitory-active ligands of any structure (cf. ref. [136]) can interrupt the catalytic cycle of Na+/K+-ATPase by binding to
quite different binding sites than those occupied by digitalis
compounds (Fig. 6). Hence, agents which inhibit the enzyme
system through a digitalis-unlike microscopic mechanism will
necessarily lead to a null hypothesis concerning structure-activity relationships.
Digitalis compound
P 'K
Fig. 6. Reaction and function scheme of Na+/K+-ATPase. Noteworthy are the
large differences in the association rate constants for the formation of the inhibitory
complex between various digitalis compounds and the phosphorylated enzyme intermediate on the one hand, and for the formation of the productive complex
between ATP and the same intermediate on the other. during maximum activity of
the enzyme. The letters E denote the two catalytic subunits, and the superscripts
indicate the locations of the N a + or K' ions on the intracellular (i) or extracellular
(ej side of the enzyme or the cell. respectively. Compared with the diffusional rate
constant of slowly diffusing reactants (lo9 M - ' s - l j , the digitalis association rate
consrants were found to be4 7orders ofmagnitude smaller[127], but similar to the
isomerization constant known to characterize the protein conformation change in
enzyme-substrateinteractionsrangingfrom1U2-104 s-'.Thedurationofthe halfcycle at 37 C IS 6-7 ms (from ref. [1371).
The microscopic screening is based on our experience[66, 301
that the competent occupancy of the digitalis binding cleft by an
inhibitor of Na+/K+-ATPase, no matter of structure, and hence
its quality as a mechanistically digitalis-like compound is proved
by its capacity to promote the synthesis of the energy-rich carboxyphosphate-phosphoenzyme from orthophosphate in presence of Mg2+ ions. Microscopic screening has therefore been
proven as a final test for selecting o r rejecting compounds for
uncovering the pharmacophoric lead
structure in bimolecular
recognition studies.[66,I 3 O 1
As well-known, man is in many respects a unique species.
Therefore it is not really surprising that for the receptor-effec288
tor parameters, there are considerable distinctions between the
isoforms of Na+/K+-ATPase from man and other specie~."'~.,'21 1 3 4 , ' 381 Fortunately, the Na+/K+-ATPase preparations from human tissues, suitable for the use in the above
molecular test systems, can easily be made.[92s1 3 4 ] The times are
over, in which a surviving heart cut out of a decapitated man
had to be used to study the action of digitoxin on human cardiac
muscle.[i 391
2.5. The Chemical Hunt for the Discovery of Lead
Structures and the Functional Classification of the Other
Structural Components in Digitalis-Like Acting Inhibitors
The lead structure in complex biologically active compounds
is defined as that structural component which shows the minimal, specific recognition requirements and is still able to evoke
the biological effect of interest although its potency may be
weak.['40*I 4 l 1 The discovery of the lead structure has been regarded as an essential step in reducing the gamble in drug design.[1421In fact, the development of a novel drug can demand
the synthesis of 10000-20000 new compounds and cost more
than 200 million German Marks."431 Also in case of cardiac
glycosides such as digitoxin (1) or digoxin (2) (Fig. 1) the detection of their lead structure proved to be difficult. Digitalis compounds are noncompetitive, heterotropically acting allosteric
inhibitors of Na+/K+-ATPase as deduced in Section 2.3. This
means that they bear no resemblance to the substrate ATP and
thus d o not provide a sensible starting point in the search for the
lead structure. This appears to explain why it took until 1985
before our group detected the proper lead component in cardiac
921 Our systematic screening of about seven hundred essentially congeneric digitalis derivatives has been made
possible through the donation of numerous rare compounds by
many eminent chemists such as K. Meyer, G. R. Pettit, T. Reichstein, D. Satoh, F. Sondheimer, C. Tamm, R. Tschesche, K.
Wiesner, and W. W. Zorbach to name but a few. The given rich
fund has been supplemented in our laboratory by targeted partial synthesis of steroid derivatives not available from herbal
source^.[^^-^^^ The structure of the compounds and the pertinent test data have fully been presented in references [I, 21.
Thus, we have travelled a long distance down the road that
has led us to the functional apportionment of the three structural components of ordinary digitalis compounds. As concisely
substantiated in Figure 7, the steroid skeleton, 58,148-androstane-3/l,14-diol (16), is the inherently active lead structure
in the natural cardiac glycosides. The removal of the hydroxyl
group at C3p only lessens the potency slightly and the replacement of Cl4p-OH by C148-NH2 can even considerably increase
the inhibitory effect.L2]Hence, the basic lead structure is 58,148androstane (cf. Figs. 7 and 20). The C 3 p - 0 and C17p side
chains, when separated from the steroid skeleton, are without
intrinsic inhibitory action of their own ; however, when linked
with the steroid lead, the C3p-0 and C17p side chains contribute moderately or powerfully, respectively, to the integral
interaction energy. This is summarized below.
The tridigitoxoside chain of digitoxin (1) and digoxin (2) can
more than isoenergetically be replaced by several monosides as
exemplified in Figure 8. The butenolide substituent of 1 and 2
Angenr. Chem. In(. Ed. E t ~ g l .1995, 34, 282-2Y4
Digitalis and Digitalis-Like Drugs
, - i t
OH (6.7)
(5.1) OH
15 (30.7)
Fig. 8. Differences in the increments of Gibbs energy of the interaction between
Na+;K+-ATPase from human cardiac muscle and digitoxigenin (14) produced by
introduction of various substituents at C3/I-OH 121. The conformational llexibility
of the sugar residues. arising from the two degrees of rotation;il freedom about the
glycoside bonds. can help to account for the finding that both p-u- and 2-L-sugars
with C1 or 1C conformation, respectively. increase the interaction energy. I n addition, the amino acid sequences involved in forming the sugar binding subsite may
have some conformational adaptability. The numbers in parentheses indicate the
6AC" values in -kJinol-'.
17 (38.1)
19 (--)
16 (22.6)
20 (38.7)
21 (36.3)
Fig. 7 The impact of stripping the side chains of 3/(-O-(x-~-rhamnosyl)14. 13 + 15, 13 + 16) reveals their distinct contributions
digitoxigenin (13) (13
to the integral Gibbs energy of interaction with Na'/K+-ATPase from human
cardiac muscle 191.921. While the sugar and lactone residues are devoid of inhibitory effcct on their own. the steroid skeleton 16 exhibits an intrinsic potency not
different from unity [130] and acts thus as the lead structure in the digitalis glycosidec 191. 92, 1301. Conversion of the A:B-cis- into the A/B-lruns-ring junction
(14 + 17) reduces the potency of the aglycon and also diminishes the strengthening
13 with 17 + 18). However,
of Ihe inhibitory efficacy by glycosidation (cf. 14
conversion of the C/D-cr,s- into the C/D-fruns-ring junction (14 + 19) annuls the
potency of the precursor compound. A comparison of the Gibbs interaction energies of the aglycons 14, 17.20. and 21 shows that 14 with a strongly bent, essentially
inflexible steroid skeleton fits best into the digitalis binding cleft. The numbers in
parenthew indicate the AGO' values in -kJmol-'; 19 is inactive.
At~,qi'w. ( ' h r n i .
Ini. Ed. EngI. 1995. 34. 282 -294
also can isoenergetically or even more than isoenergetically be
replaced by several heterocycles shown in Figure 9. Attempts
have been made to utilize our complete data for the derivation
of the chemotopography of the digitalis recognition matrix and
binding cleft in the Na+/K+-ATPase protein as a step in the
rational design of new inotropic steroids.[']
As shown in Figure 7, the powerful inhibitor digitoxigenin
14 loses its potency already by conversion of the C/D-cis ring
junction into the C/D-transjunction (14 -+ 19). The latter geometry is common in the endogenous hormonal steroids. Due to
this early experience, C/D-trans steroids were regarded by us as
irrelevant in considerations of the relationship between structure and inhibitory potency, although we had found some compounds without lactone substituent at C178 as active o r even
more active than their C/D-cis relatives[921(cf. the AGO' values
for 16 and 23 in Figures 7 and 10).
As pointed out by Marshall and Cramer r e ~ e n t l y , 1 ' ~at-~ ]
tempts to correlate biological activity with the structure of a
compound often have had limited success unless the study has
confined itself to congeneric series. However. the inclusion of
structurally diverse molecules, interacting at the same receptor
site, promises more basic solutions to the problem of structureactivity relationships, which may lead to the development of
novel therapeutic agent^.^'^^' Hence, we resumed and systematically extended our studies with C/D-frcms steroids of hormonal
Fundamental for this endeavor was our detection that in the
macroscopic and microscopic screening tests (cf. Section 2.4)
some progestins such as megestrol acetate (25) and chlormadino1 acetate (26) proved to be effective inhibitors of N a + / K + ATPase (Fig. 10) operating through a digitalis-like microscopic
mechanism, that is, competent occupancy of the digitalis binding cleft.[661Linkage of C38-OH with a monosaccharide either
K. R. H. Repke et al.
23 (23.3)
I -
24 (23.3)
H 3 C H T
H 3 C H q y
Fig. 9. The kinetic role and energetic contribution of substituents at C178 of
5b,14b-androstane-3,!i,14-diol in the interaction of the inhibitors with N a + / K + ATPase from human cardiac muscle. The grossly dipolar distribution of the
molecular electrostatic potential field in the outer space of digitoxigenin (14) shown
in the upper part of the figure (from ref. [911), appears to control the long- and
medium-range attraction of the molecule in forming the diffusion complex and to
promote the pull of the aglycon, with the lactone ring ahead, into the receptor
binding cleft [91. 921. The formation of the inhibitory complex appears to involve
hydrogen bonds between the electron-rich oxygen, sulfur, or nitrogen atoms in the
different side chains on the one hand and H-donating amino acid side chains in the
pertinent receptor binding subsite on the other. A perusal of the very different
contributions of the C17fi-substituents to the interaction energy reveals that the
sizes of the measured integrals result from an intricate superposition of both attractive and repelling electronic and steric components of their partial structures [I, 21.
The numbers in parentheses indicate the &A@' values in -kJmol-'.
decreases (26 -P 27) or increases (28 + 29) the inhibitory potency. This roughly parallels the response of some C/D-cis
steroids (cf. 17 -+ 18 with 14 + 13 in Fig. 7). The mentioned
differential impact of glycosidation reflects the unwieldy or fitting spatial disposition of the sugar side chain in the sugar subsite of the digitalis binding cleft that is guided by the geometry
of the A/B ring junction.[651
Most remarkably, glycosidation of chlormadinol acetate
26 -P 27 transformed its negative-inotropic
into a
surprisingly continual, positive-inotropic action.r641The effects
of 27 on cardiac contractility and other circulatory parameters
in cats are more favorable than those of digitoxin (1). The pos290
30 (18.9)
Fig. 10. The Gibbs energy of interaction between Na+/K+-ATPasefrom human
cardiac muscle and digitalis-like acting C/D-transsteroids of hormonal type: impact
of changes in configuration and substitution [2. 64-66, 1451. Applying the definition cited in Section 2.5 [141, 1421, the lead structure in these steroids is Sfi-androstane when occupying the digitalis binding cleft. The numbers in parentheses
give the AGO' values in -kJmol-'.
sibility that this functional distinction is related to the C/D-trans
or C/D-cis ring junction in the two inotropic agents still needs
to be verified. The way to the answer of this question will be
addressed in Section 3. Here, the primary, most important conclusion from our studies with 27 is that the C/D-cis ring junction, known to occur in steroids of some plants and toads only,
is---contrary to the present dogma-not an indispensable requirement for cardiotonic action; likewise animal hormone
steroids with a C/D-trans ring junction can have the capability
Angew. Chem. Inl. Ed. Engl. 1995.34. 282-294
Digitalis and Digitalis-Like Drugs
of eliciting digitalis-like, positive inotropic actions.[641The prerequisite for the realization of this potential has been recognized
to be the elimination of the hormonal potency by steroid glycosidation which prevents interaction with the intracellularly
located hormone receptors.ih41A further potential case for such
connection will be considered in Section 3.
in mammals. Our doubt about the identity of the endogenous
inhibitor of the Na+/K+-ATPase with ouabain has most
recently been reinforced.['541In this context. we are tempted to
speculate that some CID-rrans hormone steroids. such as the
progestins listed in Section 2.5, could serve as models for the
chemical nature of endogenous digitalis.[661
2.6. The Mystery of Endogenous Digitalis
3. The Most Recent Development Stage:
Data-Directed Optimization of the Lead Steroid
Our interest in the exploration of C/D-trans steroids of horThe major limiting factor of the maximum digitalis dose
mone type with regard to possible digitalis-like actions has also
which can be administered in the treatment of cardiac failure, is
been stimulated by their potential relation to endogenous digithe development of ventricular arrhythmias. An essential eletalis. The methodical search for its nature was pioneered by
ment contributing to the origin of arrhythmogenic action of
S ~ e n t - G y o r g y i " ~who
' ~ reported in 1953 that normal blood
cardiac glycosides is cerebral neuroexcitation.[' '"The imporserum contains substances which exert a digitalis-like action.
tance of neural digitalis actions follows from the various funcAmong 22 steroids probed, deoxycorticosterone and progestions of Na+/K+-ATPasein neural tissues.['"] Thus. inhibition
terone were found to share with serum and cardiac glycosides
of neural Na +/K -ATPase by digitalis compounds plays the
the action on the contractility of the heart. Their potency, howcausal role in triggering both the therapeutically useful and
ever, was too low to account for the serum activity. In the above
detrimental, initially neural actions of digitalis."571 Hence, a
context, Szent-Gyorgyi reasoned that "the digitalis glycosides
promising way to increase the ratio of doses for the therapeutic
are no drugs at all : they are substitutes for a missing screw in our
and toxic digitalis effects appears to be the design of digitalis
machinery. which had a cardinal role in one of the most basic
derivatives with exceptionally different affinities for the isophysiological regulations".
forms of Na+/K+-ATPasein cardiac muscle and brain cells.['. 21
More recently. the search for endogenous digitalis was fuelled
An initiating comparison of Na+/K'-ATPase preparations
by the discovery of endorphins, the endogenous ligands of the
opiate receptor. Thus, it became tempting to conclude that a
from human cardiac muscle and brain cortex with regard to
parallel situation might apply to an endogenous analogue of
their affinity to 76 C/D-cis steroids revealed on average a 1.5fold lower affinity to the cerebral enzyme preparation.['341The
digitalis. This conception appears to be supported by the fact
C/D-trans compound 27 showed a 3.2-fold lower affinity to the
that a specific digitalis-binding site exists on the Na+/K'-ATPsame enzyme
This possibly explains why 27 did
ase of all animal species.
not evoke arrhythmias in cats as observed with 2.[641However,
For theoretical and practical reasons, the search for endogeboth enzyme preparations apparently consisted of a mixture of
nous digitalis has become a challenge to many people in medical
three Na '/K+-ATPase isoforms[' 581 of unknown individual
and basic research as well as in the pharmaceutical industry. As
reviewed by Goto et al.[1481and Hamlyn and M a n ~ n t a , [ ' ~ ~ ]affinities. The clearcut decision about the relative affinities of
numerous affirmative reports have provided circumstantial and
C/D-cis and C/D-trans steroids to particular isoenzymes from
substantial evidence for the occurrence of endogenous incardiac muscle and neural cells requires studies with the isolated
hibitors of Na+/K+-ATPase,but the clarification of their chemisoforms not available as yet. In the light of the knowledge that
ical identity has remained elusive. Many candidates for an enthe steroid skeleton serves as the pharmacophoric lead structure
dogenous digitalis-like factor have been proposed on the basis
not only for Na+/K+-ATPases from human tissues but also
of the application of nine criteria." 481 However, all underlying
from animal tissues of widely differing digitalis affiniassays say little or nothing about the precise binding site of the
1341it appears almost certain to suppose that a proporcandidates and the microscopic mechanism of observed effects.
tion of Na '/K+-ATPase isoforms in human contractile and
The most suitable criterion for classifying a substance or an
neural cells differs in the chemotopography of the steroid bindisolate as digitalis-like acting is by proving its occupancy of the
ing subsite in the digitalis binding cleft.
Our endeavor to clarify the possible suitability of C/D-trans
digitalis binding matrix'". 1301 as evidenced by the microscopic
test technique (cf. Section 2.4). However, it has hitherto been
hormone steroids as parent structures for the design of novel
utilized in one study only.['
Therein, the well-characterized
cardiac drugs has most recently centred on the aldosterone anhypothalamic inhibitor of Na+/K+-ATPase has shown, unlike
tagonists spironolactone (30) and canrenone (31) (see Fig. 10).
digitalis. a negative response in the microscopic assay. ApparOur interest in these spirosteroids was raised by reports from
ently, some physiological inhibitors of Na+/K+-ATPaseexist in
clinicians and general practitioners that these CID-trans steroids
the animal body which have not necessarily a structural relaexert a direct positive-inotropic action in the treatment of pationship to digitalis compounds.['511
tients with severe congestive heart failure." s 9 - 1641 However,
The reports by Hamlyn et al.11521and Goto et a1.i1531
the spirosteroids have been fully effective only in combination
therapy with a diuretic, an inhibitor of the angiotensin-activatsome N a + / K +-ATPase inhibitors isolated from various sources
appeared to be identical with ouabain or digoxin were, in fact,
ing enzyme or/and digoxin. Hence, the mechanism of the remost compelling. The serious problem with the acceptance of
markably beneficial cardiac action of the aldosterone antagonists has remained not well u n d e r ~ t o o d . ~ ' ~ ~For
their identity is that a biosynthetic pathway for such C/D-cis
steroids. as exists in some rare plants and toads, is unknown
A n p w . Chrm. I n l . Ed. Enpl. 1995. 34. 282-294
29 1
K. R. H. Repke et al.
been analyzed by us; it turned out to be one of the rare cases of
“clinical feedback”.
Our analysis of the interaction of 30 and 31 with human
Na /K +-ATPase preparations in the macroscopic and microscopic test system (cf. Section 2.4) uncovered that both spirosteroids are digitalis-like acting inhibitors of the receptor enzyme. Their low inhibitory potency (cf. their AGO’ values shown
in Fig. 10 with those of digitoxin (1) o r digoxin (2) given in
Fig. 3) accounts for the need of the mentioned combination
therapy. Derived from our experience on the impact of glycosidation on the potency of various progesterone derivatives[64,h 5 1
we succeeded in increasing the efficacy of 31 twenty-fold by
The potency of 33 is even somewhat
glycosidation 31 -+ 33.11451
higher than that of 27 which we had shown to exert a favorable
positive-inotropic action in cats.[641Of course, additional studies with 33 and compounds of this type will have to be performed to establish their pharmacological profile and to select
those derivatives not with top potency, but with the largest
therapeutic dosage range. The experience with the pharmacological profile of 27L641
gives reasons for the expectation that the
adverse hormonal reactions of the aldosterone antagon i s t ~ [ ’ ~ ~are reduced or eliminated by their glycosidation,
when their interaction with the hormone receptors can be prevented. The probe on the generality of the postulated interrelationships and the further structural and functional optimization
of aldosterone antagonists may be studied by exploiting the
innovative potential invested in the more comprehensive insight
in those problems which is derivable from our studies with C/Dcis steroidsr’.’’ (cf. Figs. 7-9).
4. Summary and Prospect
The article starts by outlining the reasons why cardiotonic
steroids constitute a long-standing and still continuing research
challenge, and by disclosing the motivation of the senior author
to engage himself in this demanding field 30 years ago. This is
followed by detailing the stages and levels o f d i g i t a h research in
which the authors have extensively been involved, and by outlining the most recent developmental stage opened by their discoveries. The article reveals that even the elementary stages of drug
research as treated here are a multifaceted and intricate domain
which at last should include pharmacological and clinical feedback “loops” to test the limits of chemical and biochemical
patterns of thought suggested by the initiating work of the authors. Any major development as envisaged here is necessarily
a long haul and involves extensive investment of resources in the
pursuit of the designated concept, the therapeutic merit of which
can be judged only at the end of the road (cf. ref. [168]).
The most recent work has been supported by grant5 from the
Deutsche Forschungsgemeinschaft f Re 87811 - 4 ) und the Fonds
der Chemischen Indus t r ie .
Received September 15, 1993 [A 25 IE]
German version Angen C h m 1995. 107, 308
[I]K . R . H. Repke. W. Schonfeld, J. Weiland, R. Megges, A. Hache in Diqqn of’
En:.yme lnhibriors US Drugs (Eds.: M. Sandler. H . J. Smith), Oxford Universi-
ty Press, Oxford. 1989, pp. 435-502.
121 K. R. H . Repke. J. Weiland, R. Megges, R. Schon. Prog Med. C1w.m. 1993.
30. 135-202.
S . Yusuf, R. Garg, P. Held. R. Gorlin, Am. J. Curdiol. 1992, 69, 64G- 70G.
W. M. Smith, Am. J. Curdiol. 1985, 55, 3A-XA.
J. N. Cohn. Circirhrrion 1988, 78, 1099-1107.
W. Withering. An Account o/”he Fosglove, andsomr of its Mrdicul ( I S E S , itith
Pructirul Remurks on Drupsy, und other Disruses, Robinson, London, 1785.
[7] R . Hintsche. R. Megges. D. Pfeiffer. H. J. Portius, W. Schdnfeld, K. R. H.
Repke, Eur. J. Med. Cliem. 1985. 20, 9 15.
[8] E. Erdmdnn in Curdiuc Glycosides f785-1985 (Eds.: E. Erdmann, K . Greeff,
J C . Skou). Steinkopff, Darmstadt. 1986. p. V.
[9] K. F. Wenckebach, Br. Med. J. (Suppl. Epitome) 1930. J , 181.
[lo] K. Swedberg. Circulution 1993, 87 (Suppl. IV), 1V-126-IV-129.
[l 11 J. J. Kellermann, A m . Heurt J. 1990, 120, 1529-1531.
1121 G. Grupp. Mol. Cdl. Bimham. 1987. 76, 97-112.
[l3] E. Erdmann. M . Biihm in lnorropic Stiinulurion and Myocardial Energetics
(Eds.: H. Just, C. Hohbdrsch, H. Scholz), Steinkopff, Darmstadt, 1989,
pp. 125- 133.
(141 R. DiBianco, R. Shabetai, W. Kostuk. J. Mordn, R.-C. Schlant, R. Wright, N.
Engf. J. Med. 1989, 320, 677-683
[15] B. F. Uretsky. M . Jessup. M. A. Konstdm. G . W. Dec. C. V. Leier, J. Benotti,
S . Murali, H. C. Herrmann, J. A. Sandberg, Circulation 1990, 82, 774--780.
[16] A. D. Goldberg, J. Nicklas. S. Goldstein, ,407. J. Curdid. 1991, 68, 631 -636.
1171 J. N. Cohn. N. Engl. J. Mrd. 1989, 320, 729-731
(181 C . V. Leier. Am. J. Curdiol. 1992, 69, 120G-129G.
(191 M. Packer. Circulution 1989, 79, 198--204.
(201 T. W. Smith, N. E q l . J: Med. 1988. 318, 358-365.
[21] V. Austel. E. Kutter in Ar-neimirtelenhcifklun~:Grundlugen - Strutrgien Perspvklrwn (Ed.: E. Kutter), Thieme, Stuttgart. 1978. pp. 113-139.
(221 H. J. Portius. K R. H. Repke, Ar:neim. Furxh. 1964, f4, 1073-1077.
[23J D. S. Fullerton. K. Ahmed, A. H. L. From, R. H. McParland, D . C . Rohrer,
J F. Griffin in Mdeculur Graphics und Drug Design (Eds.: A. S . V. Burgen,
G . C. K. Roberts. h4. S. Tute), Elsevier, Amsterdam. 1986, pp. 257-284.
[24] R . Thomas. P. Gray. J. Andrews, Adv. Drug Res. 1990, 19, 313-562.
[25] R. E. Thomas in Molwulur S t r u c ~ u rund
Biobgicul Acrivif? rfSreroids (Eds.:
M. Bohl. W. L. Dudx), CRC. Bocd Raton, FL, 1992. pp. 399-464.
1261 T. W. Guntert. H. H. A. Linde, Curdiuc Giycosidrs, Purr I : Esperrmentul
Phunnucolog~(Ed.: K. Greeff) (Hundh. E.1,. Phurn?uco/. 1981.56if, 13-24).
[27] E. J. Ariens. MFIL Chew. Proc. In/. SJmp. 5th 1976 1977. 409-412.
[28] J. Parascandola. Phurm. Hirt. 1971, 13, 3-10.
(291 I D. Kuntz, Science 1992. 257, 1078-1082.
[30] W. G. J. Hol. Angcir. Chem. 1986,98,765 -777; Angeir, Chem. f n f . Ed, Engi.
1986. 25. 761-778.
[31] N. C. Cohen. J. M Blaney. C. Humblet, P. Gund, D. C. Barry, J. Med. Chem.
1990, 33. 883 894.
1321 K . Appelt. R. J. Bacquet, C. A. Bartlett, C . L. J. Booth, S. T. Freer, M. A. M .
Fuhry. M . R. Gehring. S. M. Herrmann, E. F. Howland. C. A. Janson. T. R.
Jones. C:C. Kan, V. Knthardekar, K. K. Lewis, G. P. Marzoni, D. A.
Matthews, C . Mohr. E. W. Moomaw, C. A. Morse, S. J. Odtley, R. C. Ogden,
M. R. Reddy. S. H. Reich. W. S. Schoettlin. W. W. Smith, M. D . Varney, J. E.
Villafrdnca. R. W. Ward, S. Webber, S. E. Webber, K. M. Welsh. J. White, J
Med. Ch1.m. 1991,34, 1925-1934.
[33] J. Hodgson. Bio/Turhno/ogy 1991. Y, 19-21.
[34] R. Y. Yada. R. L. Jackman, S. Nakai. fn1. J. Pept. Protein Rrs. 1988. 31,
98- 108.
[35] R. Megges, R. Franke, B. Streckenbach. K. Repke, H.-J. Schmidt (VEB
Arzneimittelwerk Dresden), DE-OS 1939173. 1970 [Chew!. A h r r . 1970, 73.
[36] R. Megges, K. Repke. B Streckenbach. R. Franke, G. Kammann (VEB
Arzneimittelwerk Dresden), DE-OS 2019967, 1970 (Chem. Ahsrr. 1971, 74,
319 15j] .
I371 F. Dittrich. R. Megges. H. J. Portius, K. Repke (Akademie der Wissenschaften der DDR), DD-P 94616, 1972 [Chem. Ahstr. 1973, 79, 42775tJ.
(381 R. Megges, H. Timm, F. Dittrich. H. J. Portius, K. Repke (Akademie der
Wissenschaften der DDR). DD-P 109869. 1974 (Chem. Ahslr. 1975, 83.
[39J R. Megges. H . Timm, P. Thiemann. F. Dittrich, P. Franke, H. J. Portius. K .
Repke (Akademie der Wissenschaften der DDR). DD-P 109622.1974 [Chern.
Ah.\/,.. 1975. 8.7. 59166al.
[40] C . Lindig, P. Franke. K . R. H . Repke, J. Prukr. Chem. 1975, 317, 17-28.
[41] P. Franke. C. Lindig. K. R. H. Repke. J. Prukr. Chem. 1975. 317. 86-98.
[42] R. Megges. H. J. Portius. K. R. H. Repke, Phurmuzre 1979.34, 328-329.
1431 R. Megges. H. Kreissl, H. J. Portius. K. Repke(Akademiede1 Wissenschaften
der DDR). DD-P 144060, 1980 [Chrm. Ahsrr. 1981, 95. 6260821.
[44] C. Lindig, K. R H. Repke, J. Prukr. Chrm. t980,322,991-1002.
1451 F. Theil, C . Lindig. K. R. H. Repke. J. Prukt. Chrm. 1980. 322, 1003-1011.
[46] F. Theil. C. Lindig, K . R. H. Repke. J: Prukt. Chern. 1980, 322, 1012-1020,
1471 R. Megges. H. Timm, P. Franke (Akademie der Wissenschaften der DDR),
DD-P 148222. 1981 [Chrm. Ahsir. 1982. 96, 35645~1.
148) A. Messerschmidt. R. Megges, H. Schrauber. Cryst. Siruct. Comrnun. 1981,
10. 1041 - 1051.
[491 E Hfihne. A. Messerschmidt. R. Megges. Cryst. Siruct. tommun. 1981. / ( I ,
407 -41 4
Angot,. Cltcm. In[. Ed. EnfI. 1995, 34, 282-294
Digitalis and Digitalis-Like Drugs
[50] A . M e s s c r d m i d t , E. Hiihne. R . Megges, Cryst. Strucr. Cominun. 1981. f0,
399 406.
1511 R. Megges. I . Eschholz. R. Hintsche, R. Schwensow (Akademie der Wissenschafteii der D D R ) , DD-P 154899.1982 [Chrm. Absrr. I983,98,54299w].
1521 B. Streckenbiich. P. Franke. R. Hintsche, H. J. Portius. K . R. H . Repke, J.
f r < / l t / . Cliwli. 1983. 325. 599- 606.
(S3l C. Lindig. K R . H. Repke, J. Prukr. Chem. 1983, 325. 574-586.
(541 C'. Lindig. J. Prukr. Chem. 1983, 325. 587-598.
[55] J. Wicha. M . Masnyk. W Schiinfeld. K. R. H. Repke. Helerocvcles 1983, 20,
231 234.
[56] R. Megges, H. Timm. P Frdnke, R. Hintsche (Akademie der Wissenschaften
der DDR). D D - P 216242. 1984 [Chem. Absrr. 1985, /03, 142275111,
[57] 1.Weilnnd. R. Schwensow, R. Megges, W. Schonfeld, M. M . Kabat, A. Kurek,
J. Wicha (Akademie der Wissenschaften der D D R ) . D D - P 237170, 1986
[ ~ ' h i w.4hrrr. 1988. /OX, 75780mI.
[SX] C . Lindig. J. Prukr. Chmi. 1986, 328. 682-694.
[59] C'. Lindig. K. R. H. Repke. J. Prukr. Chem. 1986. 328. 695-704.
[60] R. Megges. H. Timm. W. Rollka. P. Franke, J. Weiland, K. R. H. Repke
(Akademieder Wissenschaften der DDR), DD-P 233570, 1986 [Chem. Absrr.
1987, 106. 50557ql.
[61] K. Schwabe, J. Weiland. D . Hiibler. K . R. H. Repke (Akademie der Wissenschaften der DDR). DD-P 249273, 1987 [Chem. Ahsfr. 1988, 109.
2 1 1 39OjJ.
1621 C Lindig, K. R. H. Repke. J. Prukr. Chem. 1987. 329, 841-858.
1631 J. Weiland, D. Pfeiffer. M. Wunderwald, R. Schwensow, R. Megges
(Akademie der Wissenschaften der D D R ) , DD-P 245879, 1987 [Chem. Ahsrr.
1988. 109. hX91g].
[64] J. Wriland. K. Schwahe. D. Hiihler, W. Schonfeld, K. R . H. Repke. J. Enz,mr
Inhih. 1987. 2, 31 -36.
[65] J. Weiland. W Schonfeld. K.-H. Menke, K . R. H. Repke. Phurmucol. R ~ A .
1991. 33. 27 32.
[66] K. R. H. Repke. J. Weiland, K.-H. Menke, J. En:wneInhih. 1991, 5. 25-32.
1671 J. Weiland. P. Franke. D. Tresselt, M . Nitz, W. Schonfeld (Akademie der
Wissenschaften. Berlin). DD-P 290891, 1991 [Chem. Ab.srr. 1991, //5,
1681 J. Weiland. 0 . Tresselt, M. Nitz. R. Megges. W. Schonfeld (Akademie der
Wisscnuchallen. Berlin). DD-P 290892, 1991 [Chem. Ahsrr. 1991. /IS.
[69] K. R . H. Rcpke. W. Schonfeld, Trends Phurmucol. S1.i. 1984, 5. 393-397.
(701 W. Schbnfeld. J. Weildnd. K . R. H. Repke in Curdiuc G1ycoside.s 1785-fY85
(Eds.: E. Erdmann, K . Greeff. J. C. Skou). Steinkopff, Darmstadt. 1986.
pp. 127 - 134.
171) K . R H. Repke, J. Weiland, Phurniacol. Res. Commun. 1988. 20. 425450.
1721 T. A. Krenitsky. G . B. Elion in Srrutegy in Drug Reseurch (Ed.: J. A. K.
Buisman). Elsevier, Amsterdam, 1982, pp. 65 -87.
[73] S. E. Wright. The Metabolism of Curdiuc Glycosides. Thomas, Springfield, IL.
[74] R. H Thorp, L. B. Cobbin. Currliuc S/imulunt Suhsrunces, Academic Press,
Ncu York. 1967.
1751 R. Thomas, J. Boutagp. A. Gelbart. J. Pharm. Scr. 1974. 63. 1649-1683.
1761 K R ti. Repke. N m Aspeers of Curdiuc Glvcosrtles (Eds.: W. Wilbrandt. P.
Lindgren) ( P r o . . In/. Phurmacol. Mcer. / A t / 9 6 / 1963, 3, 47-73).
1771 K. R. H Rcpke, O A Z Orsterr. Aporh. Ztg. 1970, 24, 515-522.
[78] F. Lnuterbach. K. R . H . Repke, D. Nitz, Nauri??n-Schmrrdehrrg.sArch. Eup.
furhol Phurnui~ol 1960, 240. 45- 71
1791 1. Herrmann. K. R. H. Repke. Sjmpasium uhrr biochemische Aspekte der
S r c r o r ~ I / ~ ~ r . ~(Ed.:
c / i i r ~K.
i ~Schubert) (Ahh. Dr.rch. Akud. Wiss. Berlin KI. Med.
196812)). Akndemie-Verlag. Berlin, 1969, pp. 115 5 1 19.
[XO] I. Herrmnnn. K. R . H . Repke, Acru Microbiol. Acud. Sci. Hung. 1975, 22.
481 485
[ X i ] K. R. H . Repke. R. Megges. Dtsch. Grsundherfswrs. 1963. 18, 1325-1333.
[82] K. R. H. Repkc. R Megges. Theropimwhe 1973.23. 2314-2318.
[83] K:O. Hauatein. C. Pachaly, D. Murawski, Int. J. Clin. Phurmucol. Biaphurm.
1978. 16. 285 789.
[X4] K. R. H. Repke. ~~uunpii-Schmrad~,h~~r~.s
Arch. Eup. Puthal. Phurmukol. 1958.
233, 271 283.
[85] K. R. H . Repkc. L. Roth, S. Klesczewski. Nuuii~n-Sclimirdi~her~.s
Arch. E.vp.
Prrthol. PharmaXiil. 1959. 237, 155- 170.
(861 E Lauterbach. K. R. H. Repke, D. Nitz, Nuun~n-Schmiedcher~.\
Arch. Exp.
Pirrhol. Phurmukol. 1960. 239. 196-218.
[87] I Herrmann. K. R. H. Repke. Nuun!,n-SchmretlrbrrRs Arch. E.up. Purhol.
Phurnrakol. 1964, 24X. 351 -369.
(881 I . Herrmann. K. R. H. Repke, Nuiin,sn-SehmiedeberR(~r~.\
Arch. Exp. Purhol.
Phormukol. 1964. 34X. 370- 386.
[89] K. R. H. Repke. I.. T. Samuels, Biochemirrry 1964, 3. 685-689.
[90] K. R. H. Repkc. L. T. Samuels, Biochemi.srrj 1964. 3. 689-695.
[Yl] K. R. ti. Repke, P m d s Phurniucol. Sci. 1985, 6, 275-278.
[92] W. Schiinfcld, J. Wciland. C. Lindig. M. Masnyk, M. M. Kabat, A. Kurek, J.
Wicha. K . R. H Repke. ~ ~ u u n ~ n - S i ~ h r n r e d e hArch.
~ r g ' sPhurniucol. 1985.329,
Angiw. C'lwnr. l n f . Ed. Gig/. 1995. 34. 282-294
1931 K . R. H . Repke, Nuun)n-Schmii,debergs Arch. €.up. Purhol. Pharmukol. 1959.
236, 242-245.
(941 I. Herrmann. H . J. Portius, K. R. H. Repke, Nuunl-r7-Schnii~dehi,rg.\ Arch.
Exp. Pulhol. Phurmukol. 1964, 247. 1 18.
1951 I. Herrmann. K. R. H . Repke. Nuirnyn-Sthmredeher,U., Arch. E.up. Purhol.
Phurmukol. 1964, 247, 19-34.
[96] I. Herrmann. K. R. H. Repke, Nuun,vn-Schmiedeher~~.s
Arch. E.xp. Puihoi.
Phurmukol. 1964, 247. 35-48.
[97] K . R. H. Repke, S. Klesczewski, L. Roth,
Arch. ESP.
P d i o l . Phurmukol. 1959, 237, 34-48.
[98] C. Lindig. K. R. H . Repke. Actu B i d . Med. Ger. 1971. 26. 501 ~ 5 1 2 .
[99] Cihu Foundation Svmpasium: Enzymes und Drug Acrion. Punel Discussion.
Churchill. London, 1962, p. 396, 429.
[loo] R . L. Post. C . R. Merritt. C. R. Kinsolving. C. D. Albright, J Biol. Chptn
1960, 235, 179661802,
[loll K. R. H. Repke, H. J. Portius. Experienfiu 1963, (9. 452-458.
[I021 K. R. H. Repke. H. J. Portius, Nuunyn-SchmiedeherR.s Arch. E x p . Purhol.
Phurmukol. 1963. 245, 59 -61.
[lo31 K. R. H. Repke. H . J. Portius. Folru Huematol. ( L e r p g i 1965. 83. 28-38.
[lo41 K. R. H. Repke, M. Est, H. J. Portius, Biochem. Phurmucol. 1965, 14, 17851802.
I1051 A. Schwartz, J. C . Allen. W. B. van Winkle. R. Munson. J. Phurmiicol. Exp.
Ther. 1974, 191. 119-127.
[I061 A. Schwartz, G. E. Lindenmayer. J. C. Allen, Phurmiaiitl. K1.e. 1975, 27. 3134.
[lo71 A. Schwartz. K. Whitmer. G. Grupp. I. Grupp, R. J. Adams. S.-W. Lee, Ann.
N . K Acud. Sci. 1982, 402. 253.~271.
[lo81 H. Flasch. N. Heinz. N u u n ~ i i - S c I 7 m i ~ ~ d eArch.
b ~ r ~Phrirmacol.
1978, 304,
37 -44.
(1091 L. H . Michael. A. Schwartz, E. T. Wdllick. Mol. Phurmrrrol. 1979. 16, 135146.
[I101 T. Akera. Curdiac Glrcosrdec. Purr I - Erperrmmrul Pliurmu~~olugy
(Ed.: K .
Greeff) (Hundb. €.up. Phurmui~ol.1981. 5611. 287-336).
[ I 111 E. Erdmann, Cardiac Glwosides, Purr I . E.xperimc.nru1 PhurmucoIoXy (Ed :
K . Greeff) (Hundb. E.rp. Phurmucol. 1981. 5611, 337- 380).
(1121 L. Brown, E. Erdmann, R. Thomas. Biochern. Piiurmui~nl.1983, 32, 27672774
[I131 T. W. Smith, E. M. Antman, P. L. Friedman. C. M. Blatc. 1. D Marsh. Prug.
Curdioeusc. Dis. 1984, 26, 495.- 523.
[I 141 H. H. Rasmussen. G . T. Okita, R. S. Hartz. R. E. ten Eick. J. Phurmucol. Exp.
Thrr. 1990, 252, 60-64.
11151 K. R. H . Repke. Klin. Wo<henJchr. 1964, 42, 157 -165.
[116] K. R. H. Repke, Drugs and Enz,vme.s (Eds.: B. B. Brodie. J. R. Gillette. R.
Capek) (Proc. I n r . Phurmucol. M e e r . 2nd 1963 1965, 65-87)
[I171 C. 0. Lee, Am. J. PI7,~siol.1985. 249, C367-C378.
[I 181 W. Wilbrandt, K . Brawand, P. N. Wilt, Nuun~.n-Schmirdi,h~r,~s
Arch. E . Y ~ .
Purhol. Phumiukol. 1953, 219. 397-407.
[I 191 W. Wilbrandt, Hels. PhpioI. Pliurmucul. Actu 1958. 16. 31 -43.
[120] W. Wilbrandt, Drsch. Mcd. Wochmschr. 1957. 82, 1153- 1158.
(1211 W. Wilbrandt. Nrw Aspecfs of Curdiur G/jco.siclrs (Eds.: W. Wilhrdndt, P.
Lindgren) (Proc. fnt. Phurmucol Meer. 1st 1961 1963, 3. 3 9).
[I221 H. J. Portius, K . R. H. Repke, Symposium iiher hiochemischr A.spekre dcr
Srrroi~lflforschung(Ed.: K. Schubert) (Ah17. Drsch. Akud. Wrss. Berlin KI. Mcd.
1968(2)). Akademie-Verlag. Berlin, 1969, p. 180 -183.
[123] W. Wilbrandt, E. M. Weiss, Armeim. Farsch. 1960. 10. 409 -412.
[124] B. C Rossier. L. G. Palmer in The Kidnc?. Phj'srology untl Purhophpolog~.
2nd ed. (Eds.: D . W. Seldin. G. Giebisch). Raven. New York. 1992, pp 1373 1409
[I251 W. Schonfeld. R. Schon. K.-H. Menke. K. R. H. Repke. A ( . / u Brol. M c d . Grr.
1972, 28, 935-956.
11261 J. Monod. J. Wyman, J.-P. Changeux. J. Mu/. B i d . 1965. 13. 88 -118.
[I271 J. Beer. R. Kunze. I. Herrmann. H. J. Portius, N. M. Mirsalichova, N. K.
Abubakirov. K. R. H. Repke, Biuchim. Brophw. A i r a 1988. Y37, 335-346.
(1281 Y. A. Ovchinnikov. N. M . Arzdmazova. E. A. Arys~irkhova. N. M.
Gevondyan. N. A. Aldanova. N. N. Modyanov. FEBS Lrrr 1987,217,269274.
[129] J. M. Capasso. S. Hoving, D. M. Tal. R. Goldshleger, S. .I.0. Karlish. .I: Brol.
Chem. 1992, 267. 1150--1158.
[130] W. Schonfeld, K.-H. Menke. R. Schonfeld, K . R. H. Repke. I Enzyme Inhih.
1987, 2. 37-45.
[131] M. Reiter. <urdiuc Glwiridrs, Purr I : Ewperimiv7tul Phurnnrcology (Ed.: K.
Greeff) (Hundh. E.xp. Phurmucol. 1981, %:I, 153- 159).
[I321 L. Brown. R. Thomas. Arznerm. Forsch. 1983, 33. 814-817.
11331 L. Brown, R . Thomas. Armeim. For.rch. 1984. 34. 572-574
[ I 341 W. Schonfeld, R. Schdnfeld, K.-H. Menke. J. Weiland, K . R . H. Repke,
Biochem. Phurmacol. 1986, 35. 3221 -3231
(1351 W. Schonfeld. K. R. H. Repke. Qlrunt. Sri-ucr. A i r . Relur. 1988. 7. 160-165.
[I361 H . H. Shlevin. Drug Dee. RrJ. 1984, 4. 275- 284.
[137] K. R. H. Repke. F. Dittrich. Trends Pharmucol. Sci. 1980, 1. 398-402.
[I381 K. R . H. Repke, W. Schonfeld, R. Schonfeld. K.-H. Menke. Phurmuzir 1990,
45. 237-239.
K. R. H. Repke et al.
[I531 A. Goto. T. Ishiguro. K. Yamada, M. Ishii. M. Yoshioka. C. Eguchi. M.
11391 T. Deneke. H. Adam, Z . Exp. Pathul. Ther. 1906. 2. 491.
Shiniora. T. Sugimoto, Biuch~m.Biophys. R1.r. Conimun. 1990. 173, 10931140) G. R. Marshall, C . D. Barry. H. E. Bosshard. R. A. Dammkoehler, D. A.
Dunn. Compurer-Assisrcd Drug Design (Eds.: E. C . Olson. R. E. Christof[154] A. A. Tymidk, J. A. Norman, M. Bolgdr. G. C . DiDonato. H. Lee, W. L.
fersen) ( A C S Symp. Ser. 1979. f12.205-226).
Parker. L.-C. Lo, N. Berovd. K. Nakanishi. E. Haher. G . T. Haupert. Jr.,
(1411 A. J. Stuper, W. E. Briigger, P. C. Jurs. Cornpurer AssisledStridi~~s~~~Cliemnicul
Proc. Nutl. Acud. Sci. U S A 1993, 90. 8189-8193.
Slructur[, and Biologicul Function. Wiley, New York. 1979. p. 1.
T / w . 1982. 255. 103-116.
11551 D. G. Pace, R. A. Gillis, Arch. I n t . Phurmrrcod~~n.
11421 K. R. H. Wooldridge, M e d . Cheni. Proc. I n l . Sytnp. 5th 1976 1977. 427--432.
1156) W. L. Stahl, NaurochPm. In[. 1986, 8. 449-476.
[I431 M. Bohl in M o l ~ u l u rSfrucfuwund Bicrlogicul Acriviry u/Sreroidr (Eds. : M.
11571 R. A. Gillis. J. A. Quest in Curdiuc GIycosides 17x5- IY85 (Eds.: E. Erdmann,
Bohl. W. L. Duax), CRC, Boca Raton, FL, 1992, pp. 91 -155.
K. Greeff, J. C. Skou). Steinkopff, Darmstadt, 1986, pp. 341-356.
11441 G. R. Marshall. R. D. Cramer 111. Trends Phurmucol. Sci. 1988, Y. 285-289.
[I581 R. M. Young, J. B. Lingrel. Biuduw. Biuplzj's. Res. Commun. 1987, 145. 52[I451 K. R. H. Repke. J. Weiland, R. Megges, R. Schon, M . Nitz (Max-Delhruck58.
Centrum fur Molekuldre Medizin). DE-OS 432 1937, 1995.
[I591 R. Schroder. B. Ramdohr. U. Huttemann. K. P. Schiiren, Dtsch. M e d .
[146] F. S. LaBella. I. Bihler. J. Templeton, R.-S. Kim. M. Hnatovich. D. Rohrer.
WorlJmschr. 1972, 97, 1535- 1538.
F d . Proi. 1985. 44, 2806-283 1.
11601 U. Ehrlich, H. Kleprig, Med. Klin. / M u n i c h ) 1976, 71. 1546-1554.
11471 A. Szent-Gyorgyi. C/~en7ictrlP/iy.siology of Cunlruction in Body rind H i w t
(Ed.: E . Mutschler).
[161] U. Lindner in Tilerupie mir A/dusferon-Antugoni,s/~~n
Muscle, Academic Press. New York. 1953. pp, 79-88.
Urban & Schwarzenherg, Miinchen, 1988. pp. 71 -82.
[148] A. Goto, K. Ydmdda. N . Ydgi, M. Yoshioka. T. Sugimoto, Phurmucol. Rev.
1162) A. A. van Vliet. A. J. M . Donker, J. J. P. Nauta, F. W. A. Verheugt, Am. J.
1992. 44, 377 399.
Curdiol. 1993, 71. 21A-28A.
[I491 J. M. Hamlyn. P. Manunta. .IH.i'per/en.r. 1992. 10 (Suppl. 7). S9Y-Slll.
11631 U. Dahlstrom, E. Karlsson. Am. J. Curdid. 1993, 7 f 29A
~ -33A.
1150) C. T. Carilli, M. Berne. L. C. Cantley, G. T. Haupert, Jr.. J. B i d . Chem. 1985.
ICurdioL 1993. 7/. 34A-39A.
11641 F. Zannad, Am. .
260, 1027-1031.
[165] D. J. Greenblatt, J. Koch-Weser. J A M A J; A m . M r d Assoc. 1973.225.40-43.
[I511 G. T. Haupert, Jr. in The N u + , K +Pump., Purr B: Cdlulur Aspcrs (Eds.: J. C.
11661 H. R . Ochs. D. J. Greenblatt. G. Bodem, T. W. Smith. Am. H m r f J. 1978, 96,
Skou, J. G. Nmhy, A. B. Maunsbach, M. Esmann), Liss, New York. 1988,
389- 400.
pp. 297-320.
[167] Y. Nishino, H. Schroder. M . F. El Etreby. A r x e i m . Fursch. 1988. 38, 18001152) J. M. Hamlyn, M. P. Blaustein. S. Bow. D . W. DuCharme. D . W. Harris. F.
Mandel, W. R. Mathews, J. H . Ludens, Proc. Nutl. Acud. Sci. U S A 1991, 88,
(1681 M. Weatherall. Nuture iLondon) 1982. 296. 387 -390.
Ingham, J. et al.
Chemical Engineering Dynamics
Modelling with PC Simulation
1994. XX, 701 pages
with experimentation with the pro- vision of 85 accompanying
430 figures.
Hardcover. DM 276.4
computer-based simulation
examples (Part 2) supplied
OS 2153.4 sFr. 256.-.
on diskette.
ISBN 3-527-28577-6
In this book, the reader is
guided through the complex study of dynamic
chemical engineering systems by the unique combination of a simplified presentation of the fundamental theory (Part 1) and direct hands-on computer
The treatment employed in
this book is well tried and
tested, based on over 20
years experience in teaching an international postexperience course. Whether
The ISIM digital simulation for the teacher, the student,
language is very simple to the chemist or engineer, this
use and its powerful interac- book serves as the key to a
tive nature enables the read- greater understanding of
ers to create their own chemical engineering dysimulations, based on their namics through the fun and
own specific problems. This enjoyment of active learnpowerful dynamic ISIM soft- ing.
ware is ready to run on any
DOS personal computer.
Angaw. Chem. I n [ . Ed. Engl. 1995, 34, 282 -294
Без категории
Размер файла
1 585 Кб
research, digitalis, berlinцbuchчretrospective, view, perspectives
Пожаловаться на содержимое документа