Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108

Contents lists available at ScienceDirect

Bioorganic & Medicinal Chemistry Letters

journal homepage: www.elsevier.com/locate/bmcl

Digest

Improvement in aqueous solubility achieved via small molecular

changes
Michael A. Walker
Dart Neuroscience, 12278 Scripps Summit Dr., San Diego, CA 92131, USA

a r t i c l e i n f o a b s t r a c t

Article history: Overcoming poor solubility is a significant issue in drug discovery. The most common solution is to
Received 4 May 2017 replace carbon atoms with polar heteroatoms such as N and O or by attaching a solubilizing appendage.
Revised 14 September 2017 This approach can lead to other issues such as poor activity and PK or the increased risk for toxicity.
Accepted 17 September 2017
However, there are more subtle structural changes which can be employed that lead to an increase in sol-
Available online 19 October 2017
ubility. These include, excising hydrophobic groups which do not efficiently contribute to binding, mod-
ifying stereo- and regiochemistry, increasing or decreasing the degree of unsaturation or adding small
Keywords:
hydrophobic groups such as fluorine or methyl.
Aqueous solubility
Drug properties
Ó 2017 Elsevier Ltd. All rights reserved.
General Solubility Equation (GSE)
Hydrophobicity
Physicochemical properties

Contents

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5100
Standard approach for improving solubility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5101
Improving solubility through small changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5101
Removal of hydrophobic functionality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5102
Modification of molecular geometry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5102
Reduction or increase in the degree of unsaturation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5104
Carbon-carbon based degree of unsaturation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5105
Carbon-heteroatom based degree of unsaturation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5105
Addition of small hydrophobic groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5106
Addition of fluorine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5106
Addition of methyl . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5107
Summary and conclusion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5108
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5108

Introduction tein. Hydrophobicity can play a positive role as a non-specific driv-

Discovering a new drug is very challenging since the molecular
properties which drive optimal biology and pharmacology are
often in opposition to those which lead to favorable physicochem-
ical properties such as solubility. For example, drug binding can be
viewed as a distribution equilibrium between the surrounding
aqueous solution and the hydrophobic cavity of the targeted pro-
E-mail address: mwalker@dartneuroscience.com
ing force for the partitioning of the drug into the binding site by
raising its free energy in water.1–3 Additional binding affinity
comes from specific interactions of the compound with the target.
However, from a physicochemical property point of view,
hydrophobicity is undesirable since it leads to decreased solubility
as well as other issues. Thus, finding the appropriate balance can
be difficult.
Thermodynamic solubility of a solid is defined as its concentra-
tion in solution when it is at equilibrium with a stable crystalline

https://doi.org/10.1016/j.bmcl.2017.09.041
0960-894X/Ó 2017 Elsevier Ltd. All rights reserved.
 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108
form. Therefore, thermodynamic solubility is not only affected by
hydrophobicity but also by the crystal lattice stability of the com-
pound in the solid state. Kinetic solubility is defined as the concen-
tration at which a compound separates from solution. It is
generally assumed that kinetic solubility is driven mainly by
hydrophobicity. In the experience of the author, solubility is some-
times defined in the literature according to the experimental
method used to measure it and might not be an accurate reflection
of the intrinsic thermodynamic or kinetic solubility. There is a large
variety of methods employed which differ in the form of the ana-
lyte (crystalline solid, amorphous-solid, concentrated stock solu-
tion, or dried semi-solid from stock-solution) the aqueous phase,
temperature, and equilibration time. When reporting solubility, it
is highly recommended that it be accompanied by a description
of the physical state of the solute, the make-up of the aqueous
phase (pH and buffer concentration), temperature and equilibra-
tion time.
The General Solubility Equation (GSE, Eq. (1))4,5 equates the sol-
ubility of a solid organic compound to its log P (hydrophobicity)
and melting point (solid state stability). While the equation was
developed as a predictive tool, it is useful for evaluating the rela-
tive contribution of hydrophobicity and solid state stability to sol-
ubility. Poorly soluble compounds, where the log P is larger than
the melting point term, are said to display solvation limited solu-
bility. Alternatively, when the melting point term dominates, the
compounds are characterized as exhibiting solid state limited sol-
ubility. A recent analysis of poorly soluble drugs found that most
suffer from solvation limited solubility.6 This is to be expected
given the positive correlation between affinity and hydrophobicity
mentioned above. Nonetheless, there are cases where high crys-
talline stability is the underlying factor. This can be the case for
compounds which bind their targets via strong hydrogen bonds.
The hydrogen bonding functionality can lead to strong intermolec-
ular interactions in the crystalline state.
Log SðMÞ ¼ 0:5  log P  0:01ðMPð CÞ  25Þ
The GSE also offers insight into the expected change in solubil-
ity associated with changes in log P or melting point. Assuming a
constant melting point, a decrease in log P of 1 log unit will result
in a 10-fold increase in solubility. Likewise, holding log P constant
and reducing the melting point by 100 °C will result in the same
change in solubility. However, it is usually not possible to indepen-
dently change either log P or melting point without affecting the
other. Moreover, they often change in opposite directions. This is
usually because functionality which improves one property will
often have the opposite effect on the other. For example, groups
such as H-bond donors and acceptors, which lead to lower log P,
raise the possibility of intermolecular H-bonding in the solid state.
Other modifications which lead to an increase in compound polar-
ity can have a similar effect due to more favorable intermolecular
interactions in the solid state. In contrast, structural modifications
which weaken crystalline stability can sometimes increase
hydrophobicity.
Table 1
Molecular properties affected by the standard approach for improving solubility.
a b b
Property Method a Method b
Log P decrease decrease
mol wt NA increase
#HBD/#HBA increase increase
Basicity/acidity increase increase
PSA increase increase
a Find more soluble stereo/region-isomer
Abbreviations: mol wt; molecular weight, #HBD; hydrogen bond donor count,
#HBA; hydrogen bond acceptor count, PSA; polar surface area
b Add F or Me
Defined in the text.
5101
Standard approach for improving solubility
Since hydrophobicity is the most often encountered factor driv-
ing poor solubility, increasing polarity is usually the path most
medicinal chemists follow for overcoming it. Thus, as a simplified
generalization, the standard approach, as defined here, involves
one of two types of structural modifications. In the first method
(method a) hydrophobic carbon atoms are replaced by polar atoms,
most commonly nitrogen or oxygen. This can be accomplished by
replacing phenyl rings with heterocycles such as pyridine, or ali-
phatic CH2’s with oxygen or nitrogen resulting in an ether or
amine. Numerous other substitutions are possible. In most cases
these modifications alter the core-template of the molecule and
as such can have an impact on drug binding. In order to avoid this
issue, an alternative approach (method b) is to attach a polar or
ionizable solubilizing appendage to the molecule. In a majority of
cases this is a basic nitrogen containing group but it can also be
a carboxylic acid or neutral-polar moiety. The appendage is
attached to a site on the molecule which does not bind to the target
and which, theoretically should not interfere with affinity. This
method can be applied as a late stage fix following the optimiza-
tion of activity.
While these tactics are effective, the changes to the physical
properties are significant enough that one risks unfavorably mod-
ifying in vivo behavior. A small sample of the physicochemical
properties altered by employing the standard approach are shown
in Table 1. The favorable decrease in log P is accompanied by an
unfavorable increase in molecular weight, hydrogen bond donor
or acceptor count, acidity or basicity, and polar surface area. There
are upper bounds to these properties, above which, drug absorp-
tion and blood brain barrier permeability are significantly ham-
pered.7,8 In addition, carboxylic acids can be glucuronidated to
form reactive intermediates while basic amines are recognition
elements for P-glycoprotein efflux,9 inhibition of hERG and
phospholipidosis.10
ð1Þ
Improving solubility through small changes
Subtle structural modifications which reduce log P or melting
point can also lead to improved solubility and are available as an
alternative to the methods described in the previous section. As
shown in Table 2, these modifications include removing hydropho-
bic groups, modifying geometry by altering the position of func-
tional groups, changing stereochemistry, reducing or increasing
the degree of unsaturation, or adding substituents such as methyl
or fluorine. These changes increase solubility by reducing molecu-
lar size, H-bond acceptor/donor count or by altering the shape of
the molecule. Basicity/acidity, polar surface area and other param-
eters are not negatively impacted.
As illustrated in the following examples, alterations to these
properties can have a significant effect on solubility. Since a num-
ber of publications covering this topic have appeared in the litera-
ture in the last few years11–13 the current review is focused on
more recent examples and should be considered a snapshot of
the most recent literature. Also, the primary goal here is to demon-
strate solubility SAR, therefore the compounds were ‘cherry picked’
Table 2
Small molecular changes which can improve solubility.
Structural change Effect on molecular weight
Remove unnecessary hydrophobic groups reduce
maintain
Change degree of unsaturation reduce/increase by 2
increase by 18 or 15
5102 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108

from larger data-sets and may not be representative of the optimal Table 4
progression of the chemotype discussed in the reference. In cases Log D, pKa and kinetic solubility (pH 7.4) of the 2,3-anti and 2,3-syn isomers 3a-e and
4a-e.
where previously reviewed topics are presented, the goal is to aug-
ment, through additional examples the original findings. The OH OH
examples are ordered according to the net effect the modification O O
has on the molecular weight of the parent (Table 2) as a measure N +/- N +/-
of the relative size of the change. Where possible the conditions N 3
N 3
used in the solubility assay are reported. Activity data is included 2 2
O O
in order to monitor the effect the structural change has on target N N
engagement. Finally, for the sake of brevity, the principal ideas of H H N
O N O
this review are illustrated using a minimal number of examples
and compounds.

Removal of hydrophobic functionality 3a-e (2,3-anti) 4a-e (2,3-syn) OPh

OPh

Improving solubility via molecular size reduction is very attrac-

tive since it can potentially help avoid other issues later in the Property 3a–e 4a–e
development process.14 Although hydrophobicity drives potency, a
IC50 nM 1–5 16–450
compounds sometimes possess lipophilic groups or domains which Log D 4.3–4.5 3.5–3.9
make only a moderate contribution to the overall affinity of the pKa.(N15) 6.08–6.18 7.07–7.16
b
molecule. The presence of such groups can be identified by evalu- sol7.4 mg/mL 0.5–1.2 51–58
ating the binding affinity of the molecule (pKi or pIC50) relative to a
Inhibition of T. cruzi growth in mouse fibroblasts.
its hydrophobicity (log P). The Lipophilic Ligand Efficiency (LLE) b
DMSO stock solution with pH 7.4 PBS buffer/1% DMSO, 18 h at room
index (Eq. (2)) is one method for doing this.15 Compounds which temperature.
bind efficiently will have a pKi (or pIC50) which is significantly
higher than log P. A net difference of 5 is generally considered
the minimal benchmark for efficient binding. Thus, a higher LLE, kinase and low solubility (sol. < 0.5 mg/mL). Both of these issues
indicates that affinity is driven more by specific interactions with were solved by truncating the left-hand portion of the molecule
the target (pKi), and less by hydrophobic binding to the target leading to compound 2 which is less hydrophobic (clog P = 2.03
(log P). It is possible that a highly potent molecule could exhibit versus 0.74 for 1 and 2 respectively) and more soluble (sol. =
a low LLE if the log P is high due to being excessively large. One 10.6 mg/mL). In addition, the corresponding increase in activity
remedy for this is to look for portions of the molecule which can (IC50 = 0.012 mM) combined with the lower clog P resulted in much
be paired down without affecting binding affinity. more efficient binding (LLE = 4.32 versus 7.18 for 1 and 2
LLE ¼ pK i ðor pIC 50 Þ  log P ð2Þ respectively).

A recent example from the literature illustrates this approach

nicely. Compound 1 (Table 3) was identified in a cell-based screen Modification of molecular geometry
as a Hedgehog pathway inhibitor.16 Although it was potent (IC50 =
0.45 mM) it suffered from both poor selectivity against p38 MAP Stereochemistry and regiochemistry can affect solubility by
altering the shape and geometry of a molecule in the solid state
and in solution, leading to changes in melting point and/or log P.
Table 3 For example, according to Carnelly’s rule, isomers with higher sym-
Solubility, clog P, IC50 and LLE of 1 and 2. metry exhibit higher melting points.17 Therefore, solubility can
sometimes be improved by breaking the symmetry of the
O
molecule. This approach has been recently reviewed.18 Molecular
R N O geometry can also strengthen or weaken inter- and intra-molecu-
H
N O lar H-bonds in the solid state which influence crystalline stability.
H Stereochemistry also affects solution state behavior (i.e. Log P)
N
as illustrated by a recent example (Table 4).19 The antiparasitic
compounds 3 and 4 contain 3-stereocenters resulting in 8 possible
O N ∗ diastereomers (3a–e, 4a–e). Examination of the corresponding log
N ∗ D, pKa, and solubility at pH 7.4 revealed an interesting trend
N NH
H related to the relative stereochemistry at positions 2 and 3. The
2 four 2,3-anti diastereomers 3a–e where found to be more
F hydrophobic (log D = 4.3–4.5) and less soluble at pH 7.4 (sol7.4 =
0.5–1.2 mg/mL) than the corresponding 2,3-syn isomers, 4a-e (log
1
D = 3.5–3.9, sol7.4 = 51–58 mg/mL). In addition, the basicity of the
a
IC50/bEC50 mM 0.04/2.3 0.01/0.01 benzylic amine was also dependent on the relative 2,3-geometry.
p38a IC50 mM 0.03 2.7
Conformational analysis indicated that the benzylic amine is favor-
mw 448 308
c
sol. mg/mL <0.5 10.6 ably positioned to form an intramolecular H-bond with the nicoti-
IC50 mM 0.45 0.012 noyl amide NH. This type of intramolecular interaction is known to
clog P 2.03 0.74 increase hydrophobicity.20 Intramolecular H-bonding can increase
LLE 4.32 7.18
hydrophobicity in two ways. First, it stabilizes the unprotonated
a
Inhibition of Luciferase reporter gene in NIH-3T3 cells. form of the C2-benzylic-methylamine thereby reducing the degree
b
Inhibition of Hedgehog dependent differentiation of C3H10T1/2 cells. of ionization in solution and second it blocks the amine from H-
c
Method not reported. bonding to water. In the case of the syn isomers, steric strain
 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108 5103

Table 5 Table 7
Solubility, log P and clog Kb of compounds 5–7. Solubility of compounds 12–15.

R
N Het
N N CO2 H
CN
N
O O
N CN

a
compd pEC50 (S1P1) sol. mg/mL
N N N O O N
Property
Het ∗
O N N ∗
∗ ∗ ∗
O N
12 9.7 46
5 6 7
∗
a
sol.7.4 mg/mL 63 2.5 5.3 ∗ N
Log P 2.6 3.3 3.1
N N
13 8.5 187
clog Kb 2.9/2.6 1.8/0.8 1.6/1.6
∗
a
pH 7.4, 24 h at 25 °C ∗ O

S N
14 8.6 160
∗
between the C3-methyl and C2-benzylic–methylamine disfavors ∗ N
the intramolecular H-bonded conformation reducing its propensity
N N
15 8.9 256
to exist in solution. As a result the amine is available to interact ∗
with water leading to higher solubility compared to the anti ∗ S
isomers. a
Method not provided.
In a separate publication it is revealed that the more soluble anti
diastereomers are less active than the syn analogues.21 However,
this example illustrates two points. First, intramolecular H-bond- probably due to the higher dipole moment associated with the
ing often leads to an increase in hydrophobicity. Second, the H- 1,3,4- compared to the 1,2,4-substitution pattern.23, In addition,
bond can be disrupted without having to remove the donor or the authors suggest that the lower log P is also the result of the
acceptor groups. It is feasible that if one had a molecule where stronger H-bonding potential of the nitrogen atoms of the 1,3,4-
the donor and/or acceptor group directly engaged the target that oxadiazole. The calculated H-bond basicities (clog Kb)24 of the
both activity and solubility could be improved by hindering nitrogen atoms of the parent oxadiazoles are shown in the table.
intramolecular H-bonding through conformational control. It is assumed that higher H-bond acceptor basicity leads to a more
The regioisomers of multi-heteroatom containing heterocycles favorable H-bonding interaction with water.
often exhibit large differences in physicochemical properties such A broader analysis of oxadiazole compounds using matched
as log P, melting point and solubility despite the fact that they molecular pair analysis (MMPA) was conducted by researchers at
are structurally very similar. In a recent paper22 the properties of AstraZeneca.25 They found that in accordance with the results in
a series of oxadiazole regioisomers were examined (Table 5). For Table 5, 1,3,4-oxadiazoles tend to be less hydrophobic and more
this set of compounds the 1,3,4-oxidiazole 5 was found to be more soluble than 1,2,4-oxadiazoles. The difference in log D between
soluble than its 1,2,4-isomeric counterparts 6 and 7. The solubility the two regioisomers is on the order of 1 log unit. However, despite
difference is likely the result of the lower log P of 5 (log P = 2.6) rel- the lower log D, some exceptions to the general trend of higher sol-
ative to 6 and 7 (log P = 3.3 and 3.1, respectively). This suggests ubility for the 1,3,4-regioisomers were noted. A separate example
that 5 is better solvated by water compared to 6 and 7. This is from the literature was found which demonstrates this behavior
in a series of soluble epoxide hydrolase inhibitors and suggests
one possible reason for the outliers.26 As seen in Table 6, while
Table 6
Solubility, clog P and melting point of compounds 8–11. compounds 8 and 9 are predicted to have lower clog P than 10
and 11, they are slightly less soluble at pH 7.4. The reason for this
Het is that the lower clog P for the 1,3,4-isomers is offset by corre-
spondingly higher melting points. It can hypothesized that the lar-
O
ger dipole moment and stronger H-bond basicity of 8 and 9 lead to
N stronger intermolecular interactions in the solid state. In this way
H
the improved solution behavior imparted by the 1,3,4-oxadiazole
R is accompanied by an unfavorable increase in crystalline stability
leading to lower solubility. This illustrates that changes in log P
a
and melting point are often negatively correlated with one
compd R clog P mp °C sol.7.4 mg/mL
Het ∗ another. The activity of the compounds against soluble epoxide
∗ hydrolase was measured using a fluorescence based assay. Deriva-
tives 8–11 inhibited the enzyme by 46–64% at 1 nM. Thus, the
N N
8 H 3.54 228 5.89
structure of the oxadiazole core appeared to have had little impact
9 ∗ F 3.37 235 5.00
O
on activity.
∗ Interestingly, there is evidence from the literature that the
O N
10 H 4.41 152 6.12
regioisomeric effect seen for the oxadiazoles might apply to other
11 ∗ F 4.62 173 5.23
N 5-membered ring heterocycles. Shown in Table 7, during the
∗ course of optimizing a series of S1P1 agonists the oxadiazole
a
pH 7.4, 25 ± 1.0 °C. regioisomers, 12 and 13 and thiadiazoles 14 and 15 were synthe-
5104 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108

Table 8 Table 10
Solubility difference between pyridazine (16) and pyrimidine (17) containing Solubility, clog P and melting point of hydroxyl-quinoxaline (21),- quinazoline (22)
inhibitors. and -cinnoline (23) heterocycles.

Cl OH OH OH
N N N
HN SO2 N
N N het N
O N
S 22 23
21
F3 C SO2 HN
sol. mg/mL 6.6 1.3 0.6
mp °C 102–103 149–150 185–186
N

O P31,32 progressively change as the nitrogens draw closer together

in the ring. Therefore, the higher solubility of 16 is consistent with
a
the higher dipole moment, pKHBX, pKa, and lower log P of the pyri-
Compd sol.7.4 mg/mL
∗ ∗ dazine compared to the corresponding pyrimidine of 17.
Het
As with the 5-membered ring heterocycles, the solubility of 6-
16 N N 35 membered ring heterocycles is determined by the net balance of
∗ ∗ solution (log P) and solid state (melting point) properties. The com-
pounds shown in Table 10 illustrate this point.34–37 Solubility is
17 N 3
driven more by crystalline stability than hydrophobicity for this
∗ ∗
N series. Compound 21 has a much lower melting point (mp = 102–
103 °C) than 22 and 23 (mp = 149–150 and 185–186 °C, respec-
a
Dried from DMSO sample, pH 7.4, 24 h, 25 °C.33 tively) which correlates well with their observed solubilities.
The observed trend in melting point for 21, 22 and 23 is oppo-
site to what one would expect from the monocyclic heterocycles in
sized. Similar to the oxadiazoles the 1,3,4-thiadiazole regioisomer Table 9. This can be rationalized based on the putative nitrogen H-
15 (sol. = 256 mg/mL) was more soluble than the corresponding bonding basicities of the isomers and the presence of an H-bond
1,2,4-thiadiazole analogue 14 (sol. = 160 mg/mL).27 While the donor group (OH). According to Etter’s rule38 which states that
1,2,4-oxadiazole is nearly 10-fold more potent than the 1,3,4-iso- ‘‘all good proton donors and acceptors are used in hydrogen bond-
mer, the thiadiazole regioisomers are similar in activity. A more ing”, one can assume that compounds 21–23 will be stabilized by
thorough investigation of the literature is needed in order to see the formation of such bonds in the solid state. If true, the inter-
how general this phenomenon is for other 5-membered ring molecular H-bonding in 23 would be expected to be stronger than
heterocycles. 21 and 22 due to the higher H-bond basicity. This would lead to
Six membered nitrogen-containing heterocycles also display higher crystalline stability and melting point. The correlation of
interesting solubility SAR related to their regiochemistry. For solubility with H-bond basicity would also account for the differ-
example in the course of optimizing inhibitors of Bcl-2/Bcl-xL both ence in solubility between 21 and 22.
the pyridazine and pyrimidine analogues 16 and 17 were synthe-
sized (Table 8).28 Although 16 was found to be approximately
Reduction or increase in the degree of unsaturation
10-fold less active than 17 (data not shown), it exhibited a 10-fold
improvement in solubility at pH 7.4 (sol. = 35 and 3 mg/mL for 16
The textbook definition for the total number of p-bonds plus
and 17 respectively). This is relatively large compared to their
rings in a molecule is ‘‘degree of unsaturation”.39 In this section
structural similarity and the fact that pyrimidine and pyridazine
both sp2-hybridized atoms and ring-count are considered. A dou-
make up only a small portion of the structures.
ble bond or ring are each equivalent to one degree of unsaturation
Examination of the physical properties of the parent heterocy-
and reflect the loss of 2 hydrogen atoms. The addition or subtrac-
cles offers some interesting insights. The three possible arrange-
tion of 2 hydrogen atoms is perhaps one of the smallest changes to
ments of 2-nitrogen atom containing, 6-membered ring
the constitutional make-up of a molecule one can make. This
heterocycles, are shown in Table 9. In this series the dipole
increases or decreases the total number of rings and/or p-bonds
moment,29 H-bond basicity (pKHBX),30 pKa, melting point and log
contained in the molecule. This seemingly small change in struc-
ture can result in large improvements in solubility.
A number of retrospective analyses have appeared in the last
Table 9
Dipole moment, pKHBX, pKa, melting point and log P of compounds pyrazine (18), few years evaluating the effect of unsaturation (sp2-hybridization
pyrimidine (19) and pyridazine (20). and aromaticity) on the solubility of drug like molecules. The gen-
eral consensus is that solubility is reduced as the relative-number
Compd N N N
N of unsaturated atoms increases.40 For example, Lovering et al.
N found that solubility was positively correlated with the fraction
N
of sp3-hybridized carbon atoms (Fsp3).41 In addition, Ritchie and
19 20
18 Mcdonald reported an increase in hydrophobicity and decrease in
a
solubility as the number or aromatic rings increased.42 Although
DM 0.0D 2.4D 3.9D
pKHBX 0.92 1.07 1.65
these studies and others show a clear correlation between solubil-
b
pKa 0.7 1.3 2.3 ity and the level of unsaturation one should be cautious in not
mp °C 52 20 -8 over-interpreting the results to suggest that this reflects the funda-
log P -0.26 -0.44 -0.72 mental properties of sp3 and sp2 carbon atoms. The effect of unsat-
a
Dipole moment (DM) measured in dioxane. uration on solubility is context dependent, especially in the case of
b
pKa of the conjugate acid hydrocarbon-aromatic versus heteroaromatic systems. The follow-
 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108
ing sections describe situations under which unsaturation
decreases solubility and others where it increases it.
Carbon-carbon based degree of unsaturation
The correlation of physical properties of hydrocarbons or hydro-
carbon-moieties accompanying alterations in the degree of unsat-
uration can be appreciated by examining the series of C8-
hydrocarbons shown in Table 11. Removal of 2 hydrogen atoms
from n-octane (25) increases the degree of unsaturation by 1 and
yields an alkene or a ring (26 and 27). Both of these modifications
lead to a reduction in lipophilicity (log P) which is sufficient to
yield an improvement in solubility. The lower lipophilicity of 1-
octene (log P = 4.57) compared to octane (log P = 4.93) is likely
the result of increased polarity. On the other hand one would not
expect cyclooctane to be more polar than n-octane but it displays
a lower log P of 4.07 nonetheless. The effect of cyclization is to
reduce the size of the molecule, and thus the solvation cavity rel-
ative to the acyclic form, which translates to a lower entropic pen-
alty. Therefore, cyclooctane is more soluble than n-octane because
it is smaller in size.
The degree of unsaturation plays a role in melting point as well.
Although the C8-alkanes in Table 11 are liquids at room tempera-
ture the effects of sp2/sp3-hybridization and cyclization on melting
point can be extrapolated to compounds which are solids. In the
case of isolated carbon-based sp2 centers saturation usually leads
to an increase in melting point. In the case of aromatic systems sat-
uration will lead to a reduction in melting point. In addition to sat-
uration, melting point is usually positively correlated with the
cyclization. The melting point of n-octane (mp = 57 °C) versus
cyclooctane (mp = 13 °C) illustrates this point. Moreover, melting
point continues to increase as the number of rings increases such
that bicyclo[3.2.1]octane (28) has a melting point of 135 °C.43
Therefore, in certain cases solubility might be improved by dean-
nulation in order to lower the melting point even though this
might increase log P.
28
Although the C8-compounds in Table 11 are simple mole-
cules they reflect the fundamental properties of hydrocarbon
systems and the same effects can be seen in more drug-like
molecules. For example, cholesterol (29) and cholestanol (30)
are naturally occurring compounds differing from one another
by a single site of saturation. Similar to the octane example
in Table 11 the saturated derivative cholestanol is significantly
less soluble in water (sol. = 0.009 mg/mL at 35 °C)44 than choles-
terol (sol. = 0.026 mg/mL at 30 °C).45 Likewise, in Table 12 the
relative solubilities of the steroid pair testosterone 31 (monohy-
drate, sol. = 23 mg/mL)46,47 and stanolone 32 (sol. = 5 mg/mL)48
follow the same trend.
Table 11
Degree of unsaturation, solubility, clog P and melting point (mp °C) of n-octane (25),
1-octene (26) and cyclooctane (27).
25 26
deg. unsat. 0 1
sol. mg/mL 1 3
log P 4.93 4.57
mp °C 57 102
5105
H H
H H H H
HO HO
29 30
sol. = 0.026 μg/mL (30 oC) sol. = 0.009 μg/mL (35 oC)
In summary, for non-aromatic hydrocarbon based molecules or
moieties, increasing the degree of unsaturation through sp2-carbon
atoms or cyclization can lead to a reduction in lipophilicity. On the
other hand lowering the degree of unsaturation via deannulation
can lead to improved solubility via a lowering the melting point.
With some caveats these principles can also be extended to aro-
matic hydrocarbons when used as a substitute for Fsp3. Carbo-
cyclic aromatic compounds tend to be more soluble than their
saturated counterparts due to a more favorable enthalpy of solva-
tion.49 For example, benzene has a clog P = 2.14 while cyclohexa-
triene, cyclohexene and cyclohexane have clog P = 2.45, 2.87 and
3.35 respectively. On the other hand unlike isolated sp2 functional-
ity, aromatic compounds tend to have higher melting points due to
better crystal packing. The increase in log P associated with satu-
rating a carbocyclic-aryl might be offset by a reduction in melting
point.
This is interesting given the conclusions of the empirical studies
mentioned above which suggest that increasing Fsp3 yields
improved solubility. However, since the effect of unsaturation on
solubility is context dependent, this would suggest that the results
of these studies are biased by the make-up of the compounds in
the data sets. This seems plausible since, heterocycles make up a
large portion of the compounds encountered in medicinal chem-
istry. As seen in the next section, the observations in these studies
most likely reflect the properties of the heteroatoms contained
within these molecules. However, since phenyl is one of the most
commonly encountered rings found in drug-like molecules50 the
modifications described in the current section should prove useful.
Carbon-heteroatom based degree of unsaturation
As mentioned above, there is an important distinction between
all-carbon unsaturated functionalities versus those containing het-
eroatoms. The change in polarity of the heteroatom accompanying
the change in the degree of unsaturation (ie. sp2 to sp3) needs to be
considered. For example, as shown in Table 13, the partially satu-
rated heterocycle tetrahydroisoquinoline 33 is less hydrophobic
than the fully aromatic isoquinoline, 34 (log P = 1.71 and 2.09
respectively). This is the opposite trend observed for the all-carbon
templates where saturation results in increased hydrophobicity.
Table 12
Solubility, clog P and melting point of testosterone (31) and stanolone (32).
OH OH
H H
H H H H
O O
H
27 31
32
1
7 sol. mg/mL 23 5
4.07 log P 3.32 3.55
13 mp °C 155 181
5106 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108

The increase in hydrophilicity for tetrahydroisoquinoline can be
correlated to its higher H-bond acceptor basicity (pKBHX) = 2.04)
compared to isoquinoline (pKBHX) = 1.94).27 In addition the more
basic sp3-nitrogen of tetrahydroisoquinoline (pKa = 9.3 at 37 °C)51
will be more highly protonated than the sp2-hybridized nitrogen
of isoquinoline (pKa = 5.4 at 25 °C) at neutral pH further adding
to the increased solubility.52 For heteroatom containing templates
the effect of unsaturation on melting point is variable. In some
cases, like 33, the fully aromatic analogue is higher melting com-
pared to its saturated counterpart, while for others, for example
quinolone (mp = 15 °C) and tetrahydroquinoline (mp = 14 °C),
the opposite is true.
A recent example demonstrates the large effect that partial sat-
uration of a heterocyclic ring can have on aqueous solubility. In a
report describing the optimization of a series of androgen receptor
down regulators, hERG-inhibition and poor solubility arose as crit-
ical issues.53 Lead compound 35 was found to be highly hydropho-
bic (log D7.4 = 3.2) and poorly soluble (sol. = 3 mg/mL, crystalline
solid, 0.1 M PBS, 24 h). Partial saturation of the triazolopyridazine
ring yields 36 resulting in a significant reduction in hydrophobicity
(log D7.4 = 2.5) and improved solubility (sol. = 299 mg/mL, crys-
talline solid, 0.1 M PBS, 24 h). Although compound 36 (pIC50 =
5.75) was less active than 35 (pIC50 = 6.4) it showed much lower
hERG-inhibition as well as higher plasma free fraction and lower
intrinsic clearance in human hepatocytes. The overall profile was
such that compound 36 was advanced as a candidate (AZD3514)
for clinical trials.
activity in the kinesin spindle assay was also improved by
deannulation.
Addition of small hydrophobic groups
The final area of structural modification considered is the addi-
tion of small hydrophobic substituents such as methyl or fluorine.
The effect of replacing hydrogen by either of these groups has been
evaluated using matched molecular pair analysis (MMPA). The
results from three separate literature reports are summarized in
Table 15.55–57 The mean change in solubility (Dlog S) associated
with replacing hydrogen with F or CH3 is given while the percent-
age of matched pairs where an increase was observed is provided
in parenthesis. As will be discussed below, the context of the sub-
stitution is important. The MMPA studies in Table 15 were carried
out using different collections of compounds, and the observations
reflect the structural make-up of these sets. Nonetheless, the stud-
ies yield interesting observations that provide hints to some under-
lying trends.
Addition of fluorine
As shown in the Table 15, while the mean effect of replacing H
by F results in a reduction in solubility (Dlog S = 0.22, 0.45 and
0.10 respectively) there appears to be a significant percentage of
instances where the reverse is observed. Leach and Zhang reported
an increase in solubility for 22% and 34% of the compounds respec-
tively, while Gleeson observed it for 9%. Taken together this sug-

O O
N N
N Me N Me

N O N O

N N
N N N N
N N
CF3 35 CF3 36
logD7.4 = 3.2 logD7.4 = 2.5
sol7.4 = 3 μg/mL sol 7.4 = 299 μg/mL

The kinesin spindle inhibitors 37 and 38 (Table 14) demonstrate gests that there might be a special chemical property of fluorine
the effect of ring opening on solubility.54 The carbazole based which imparts improved solubility when deployed in certain struc-
derivative 37 contains 4-rings and is very insoluble. Its solubility tural contexts.
in water (pH 7.4 PBS) was less than 1 mg/mL leading the research- Much earlier than this Hansch made some pioneering observa-
ers to use a solution of 50% EtOH in pH 7.4 PBS in their measure- tions concerning fluorine’s effect on hydrophobicity (log P or log D)
ments. Under these conditions the solubility of 37 was 0.5 mg/
mL. It also exhibits a very high melting point (mp > 300 °C) sug-
Table 14
gesting that the poor solubility is the result of strong crystal pack-
Solubility, HPLC retention time and melting point of kinesin spindle inhibitors 37 and
ing. Deannulation yields compound 38 which has a lower melting 38.
point (mp = 190 °C) and is 3-fold more soluble. Interestingly, ring-
opening appears to increase hydrophobicity as indicated by the O O
NH NH
longer retention time via reverse phase HPLC (logP not reported).
This might be expected given the C8-examples in Table 11. Finally,

N CF3 HN
H
Table 13 CF3
Solubility, clog P and melting point of tetrahydroisoquinoline (33) and isoquinoline
37
(34). 38
a
IC50 mM 0.18 0.045
b
sol. mg/mL 0.5 1.8
NH N c
HPLC-RT min. 21.7 24.2
mp °C >300 190
33 34
a
Inhibition of microtubule-activated KSP ATPase assay.
sol. mg/mL NA 5.66 b
50% EtOH/7.4 PBS, 48 h, 25 °C.
pKBHX 2.04 1.94 c
HPLC analysis was carried out on a Cosmosil 5C18-ARII column (4.6  250 mm),
log P 1.71 2.09
and the material eluted by a linear MeCN gradient (70:30–30:70 over 40 min) in
mp °C 30 25
0.1% TFA; flow rate of 1 mL/min.
 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108 5107

Table 15 F R F O F R
Matched molecular pair analysis (MMPA) for replacement of H by F, and CH3. O R
OH
R Dlog S (% comps with increased sol.) F F O F
a
Leach et al. Zhang et al. a
Gleeson et al. O
OH
F 0.22 (34)[Ar] 0.45 (22) 0.10 (9)[Ar]
CH3 0.21 (33)[Ar] 0.50 (26) 0.11 (11)
a Fig. 1. Examples of fluorinated functionality leading to reduced log D.
[Ar]; substitution of an aromatic H by R.

shedding light onto structural contexts where its dual nature is
manifested. It was found that fluorine behaves differently when
attached to an aryl ring versus an alkyl group. A summary of these
results are shown in Table 16.58–61 Using X = H as a reference
(entry 1), fluorine yields a lower log P when attached to an alkyl
chain (log P = 2.95 and 3.68 for X = F and H respectively) versus
phenyl (log P = 2.27 and 2.13 for X = F and H respectively). The
order of magnitude in the reduction in log P is 0.73 log units
which could potentially translate into a 10-fold increase in solu-
bility. Interestingly, this behavior is shared, to a certain degree, by
chlorine and bromine (entries 3 and 4) where log P is either
reduced (Cl) or left unchanged (Br) when attached to an alkyl-site.
Therefore, the unusual behavior noted for fluorine might be better
classified as a halogen atom property.
While this would explain Zhang’s results (Table 15) it does not
account for Leach and Gleeson studies where aryl substitution was
examined. Interestingly, an earlier report by Böhm62 examined the
effect of fluorine substitution on log D of compounds from the
Roche compound collection. While the mean effect was an increase
in log D there were certain structural configurations associated
with a decrease. As shown in Fig. 1, in addition to alkyl-based
groups, a number of aryl based sub-structures were found. It is
not certain whether these structural motifs were present in the
Leach and Gleeson data sets but it reinforces the idea that the
behavior of fluorine is context dependent.
The solubility enhancing effects of fluorine have been demon-
strated in a number of drug discovery programs. As an example,
fluorination of the BRaf inhibitor 39 (sol. = 20 mg/mL, from solid,
pH 6.5 PBS, room temperature, 24 h) provides a modest improve-
ment in solubility for 40 (sol. = 60 mg/mL, from solid, pH 6.5 PBS,
room temperature, 24 h).63 Both compounds exhibit favorable
inhibition of B-Raf (IC50 = 3.8 and 0.6 nM for 39 and 40, respec-
tively)
F F
O O
O S O S F
N O N O
H H
NH Cl NH Cl
N N
N N 39 N N 40
H H
clog P = 2.19 clog P = 1.61
sol. = 20 μg/mL sol. = 60 μg/mL
A much more profound example comes from a series of com-
pounds designed as potential agents for the treatment of amy-
otrophic lateral sclerosis (ALS).64 The lead compound 41 was
found to be potent, exhibiting good oral bioavailability and brain
penetration. Fluorination was examined as one of two approaches
to further optimize the solubility and stability in microsomes and
plasma. Introduction of F onto both of the phenyl rings (42) yielded
no change in solubility (from DMSO stock solution, PBS buffer, 45
min.–16 h). In contrast bis-fluorination of the ethyl-spacer group
provided a 10-fold increase in solubility for 43 and 44. In this
case, despite the improved solubility, the clog P predicted an
increase in hydrophobicity. This could represent the inability of
the calculation protocol to accurately predict the log P of this
chemotype. Fluorination of the alkyl portion of the molecule also
resulted in increased stability in plasma and human microsomes.
O O O O
N N F N N F
O O
41 42
clog P = 3.42 clog P = 3.70
sol. = 10 μg/mL sol. = 12 μg/mL
O O O O
F F F F F F F F
N N F N N F
O O
43 44
clog P = 4.61 clog P = 4.89
sol. > 200 μg/mL sol. = 111 μg/mL
Addition of methyl
The MMPA analysis in Table 15 also indicates that methylation
can result in an increase in solubility. One mechanism by which
methylation can increase solubility is by blocking the formation
of intra- and intermolecular hydrogen bonds. Intramolecular H-
bonding hampers interactions with water while intermolecular
H-bonds lead to higher crystalline stability. In the final example,
the Mycobacterium tuberculosis inhibitor 45 (Table 17) features a
primary amide which is positioned in close proximity to two H-
bond acceptors, the adjacent ether-oxygen and the distal pyridine
nitrogen.65 As demonstrated in Table 4, it can be hypothesized that
intramolecular H-bonding leads to higher lipophilicity (log P not

Table 16
Hansch’s analysis of the effect of halogenation at alkyl or aryl positions.

entry X
X X

Log P Dlog P Log P Dlog P

1 H 3.68 – 2.13 –
2 F 2.95 -0.73 2.27 0.14
3 Cl 3.55 -0.13 2.84 0.71
4 Br 3.72 0.04 2.99 0.86
5108 M.A. Walker / Bioorganic & Medicinal Chemistry Letters 27 (2017) 5100–5108
Table 17
Solubility of 45–47.
O 2007;50:5858.
O R
N O
N R
O 9. Hitchcock SA. J Med Chem. 2012;55:4877.
N 2013;23:4587.
a b
compd R R MIC mM
45 H H 0.6 2 14. Hann MM. Med Chem Commun. 2011;2:349.
46 H Me 1.2 95
47 Me Me >75 327
a 18. Ishikawa M, Hashimoto Y. J Med Chem. 2011;54:1539.
M. tuberculosis H27Rv.
b
Dried-from solvent isolate, PBS buffer, 24 h, 25 °C
reported). Invoking Etter’s rule as above (examples 21–23), one
would also expect that the presence of both an H-bond donor
and acceptor in a molecule will result in the formation of inter-
molecular H-bonding in the solid state. The introduction of a single
methyl group (46) results in a significant increase in solubility (sol.
= 95 mg/mL,). Assuming that the remaining hydrogen of the sec-
ondary amide in 46 is capable of acting as an intramolecular H-
bond donor, solubility should increase by adding an additional
methyl. Presumably, tertiary amide 47 blocks both intramolecular
an intermolecular H-bonding which would explain its improved
solubility (sol. = 327 mg/mL) over 45 and 46. The presence of an
amide-NH appears to be important for activity since 47 is com-
pletely inactive. However, compound 46 achieves a more favorable
balance between activity and solubility compared to 45.
Summary and conclusion
In summary, the methods described in this review provide an
alternative to the standard approach which medicinal chemists
resort to in order to improve solubility. In theory a drug only needs
to be soluble enough to allow dissolution of the targeted dose.
Since the dose is likely a multiple of the in vitro activity of the com-
pound high potency will lower the bar for the required solubility.
However, the problem is that activity usually comes at the price
of increased hydrophobicity which leads to lower solubility. The
solution to this dilemma is to take advantage of H-bonding interac-
tions and to employ lipophilic functionality parsimoniously so that
LLE is maximized. This will not necessarily result in high solubility.
For example, a compound with a Ki = 1 nM can have a log P of 4 and
still be considered to bind efficiently (i.e. LLE = 5). Alternatively,
optimizing LLE by building in H-bonding functionality to increase
activity and lower log P often leads to increases in melting point.
Rather, LLE ensures that the gap between the solubility of the lead
compound and that needed to deliver the targeted dose is narrow.
Under such circumstances the best approach for optimizing solu-
bility further would be to employ minor structural changes that
do not disturb potency and PK. This review identifies several minor
structural changes which lead to improved solubility and can be
applied to diverse templates.
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