WO2010024783A1

WO2010024783A1 - Biarylrhodanine and pyridylrhodanine compounds and their use

Info

Publication number: WO2010024783A1
Application number: PCT/SG2009/000301
Authority: WO
Inventors: Christina L. L. Chai; Paul H. Bernardo; Jin Xu; Victor C. Yu; Kah-Fei Wan; Henry Y. K. Mok; Thirunavukkarasu Sivaraman; Janarthanan Kjrishnamoorthy
Original assignee: Agency For Science, Technology And Research; National University Of Singapore
Priority date: 2008-08-27
Filing date: 2009-08-27
Publication date: 2010-03-04

Abstract

The present invention pertains generally to the field of therapeutic compounds, and more specifically to compounds related to rhodanine, which compounds are inter alia inhibitors and/or binders of antiapoptotic/pro-survival Bcl-2 proteins such as Bcl-XL and/or Mcl-1. More specifically, the present invention is concerned with Rhodanine- based Pan-Bcl-2 inhibitors and Mcl-1 -specific inhibitors as anti-cancer compounds. The present invention also pertains to pharmaceutical compositions comprising such compounds, and the use of such compounds and compositions, both in vitro and in vivo, to inhibit and/or bind Bcl-2 proteins such as Bcl-XL and/or Mcl-1, and in the treatment of diseases and conditions that are mediated by Bcl-2 proteins, that are ameliorated by the inhibition of Bcl-2 protein function (such as Bcl-XL and/or Mcl-1 ) including proliferative conditions such as cancer, optionally in combination with another agent.

Description

BIARYLRHODANINE AND PYRIDYLRHODANINE COMPOUNDS AND THEIR USE

TECHNICAL FIELD OF THE INVENTION

The present invention pertains generally to the field of therapeutic compounds, and more specifically to compounds related to rhodanine, which compounds are inter alia inhibitors and/or binders of antiapoptotic/pro-survival Bcl-2 proteins such as Bcl-X_L and/or Mcl-1.

More specifically, the present invention is concerned with Rhodanine-based Pan-Bcl- 2 inhibitors and Mcl-1 -specific inhibitors as anti-cancer compounds.

The present invention also pertains to pharmaceutical compositions comprising such compounds, and the use of such compounds and compositions, both in vitro and in vivo, to inhibit and/or bind Bcl-2 proteins such as Bcl-X_L and/or Mcl-1 , and in the treatment of diseases and conditions that are mediated by Bcl-2 proteins, that are ameliorated by the inhibition of Bcl-2 protein function (such as Bcl-X_L and/or Mcl-1 ) including proliferative conditions such as cancer, optionally in combination with another agent.

The technical field of this invention deals with the chemical synthesis and biological testing of biologically active compounds, in particular, compounds based on the pyridine alkenyl rhodanine core structure (Structure 1 and 2 - see below) and its reduced forms (Structure 3 and 4 - see below) designed to inhibit the function of pro- survival Bcl-2 proteins.

BACKGROUND

A number of patents and publications are cited herein in order to more fully describe and disclose the invention and the state of the art to which the invention pertains. Each of these references is incorporated herein by reference in its entirety into the present disclosure, to the same extent as if each individual reference was specifically and individually indicated to be incorporated by reference.

Throughout this specification, including the claims which follow, unless the context requires otherwise, the word "comprise," and variations such as "comprises" and "comprising," will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

It must be noted that, as used in the specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a pharmaceutical carrier" includes mixtures of two or more such carriers, and the like.

Ranges are often expressed herein as from "about" one particular value, and/or to "about" another particular value. When such a range is expressed, another embodiment includes from the one particular value and/or to the other particular value. Similarly, when values are expressed as approximations, by the use of the antecedent "about," it will be understood that the particular value forms another embodiment.

Bcl-2 proteins

The Bcl-2 family of proteins have been implicated in the survival of cancer cells. This family of proteins, which includes Bcl-X_Land Mcl-1 , confers protection on cancer cells by sequestering the proteins required for apoptosis induction. The protection offered to cancer cells by the Bcl-2 proteins is by no means trivial. Studies have shown that BCI-X_L can protect a cell from dying even after its DNA has been severely damaged. This may account for the resilience of some cancers in spite of chemotherapy and radiotherapy.

The pro-survival Bcl-2 proteins act by sequestering pro-apoptosis Bcl-2 proteins such as Bak, Bid, Bax and Bad. Inhibiting the function of the pro-survival proteins causes the release of the pro-apoptosis proteins which aggregate to form channels in the mitochondria to allow the release of caspase activators such as cytochrome c. The activation of caspases, the enzymes that promote the degradation and destruction of cellular organelles, ultimately leads to controlled cell death (as illustrated in Figure 1 ).

In 2005, researchers at the Abbot laboratories reported a new compound which specifically targeted the Bcl-2 proteins. This compound, ABT-737, showed potent nanomolar inhibitory activity against the Bcl-X_L protein incubated with a fluorescein- labelled Bid peptide. It also showed nanomolar activity against Bcl-2, and Bcl-w. However, a separate unbiased study showed that ABT-737 has an IC₅₀ value of 64 nM against Bcl-X_L, and 0.12 μM activity against Bcl-2. In the same study, ABT-737 was inactive towards Mcl-1 (>20 μM) when tested against the fluorescein-labelled Bid peptide.

epigallocatechin gallate

The compound GX-15-070 (Obatoclax™) shows micromolar inhibitory activity against various Bcl-2 proteins including Bcl-X_L and Mcl-1. When tested against the fluorescein-tagged Bid peptide, Obatoclax displayed an IC₅O of 4.7 μM against Bcl-X_L and 2.90 μM against Mcl-1. Obatoclax is often touted in the literature as an Mcl-1 specific inhibitor due to its micromolar activity against the proteins and the Bid peptide.

Another compound which has shown activity against the pro-survival Bcl-2 proteins is

BH3I-1. Interestingly, the IC_5O values of BH3I-1 and GX-15 are very similar. In assays involving the fluorescent-labelled Bid peptide, the IC₅₀ value obtained against BcI-X L is 5.9 μM while the activity against Mcl-1 is 2.2 μM. The antiapoptotic Bcl-2 proteins (Bcl-2, Bcl-X_L, Mcl-1 , A1 ) are attractive targets for cancer chemotherapy. It has been shown that cancer cells overexpress one or more of these proteins to prevent the induction of apoptosis or programmed cell death. These proteins confer protection on cancer cells by sequestering the proapoptotic proteins Bax and Bak. Neutralizing these antiapoptotic proteins using natural peptides (e.g. Bim, Bid, NOXA) releases Bax and Bak which then aggregate to form transmembrane channels which result in mitochondrial permeability. The compromised mitochondria can then release apoptogenetic factors such as cytochrome C and apoptosis-inducing factor (AIF) which eventually leads to cell death. Over the years, several molecules such as ABT-737, epigallocatechin gallate and Obatoclax have been reported to inhibit the antiapoptotic Bcl-2 proteins and trigger apoptosis in cancer cells. The compound BH3I-1 is also a well-known inhibitor of the Bcl-2 proteins. Recently, it has been reported that modification of BH3I-1 can result in varied binding profiles to Bcl-X_L with an increase in efficacy.

SUMMARY OF THE INVENTION

One aspect of the invention pertains to certain pyridylrhodaπine compounds

(for convenience, collectively referred to herein as "PRD compounds"), as described herein.

A further aspect of the invention pertains to certain diarylrhodanine compounds (for convenience, collectively referred to herein as "DRD compounds"), as described herein.

In another aspect, the present invention also provides a library of compounds based on the pyridine alkenyl rhodanine core structure to inhibit the Bcl-2 family of proteins. As described herein, the biological activity of the compounds has been screened.

In another aspect, the present invention describes the library of compounds based on the pyridine alkenyl rhodanine core structure (formula IV and formula V herein) and its reduced forms (formula Vl and formula VII herein) and their respective biological activities.

Another aspect of the invention pertains to a composition (e.g., a pharmaceutical composition) comprising a PRD or DRD compound, as described herein, and a pharmaceutically acceptable carrier or diluent. Another aspect of the invention pertains to a method of preparing a composition (e.g., a pharmaceutical composition) comprising the step of admixing a PRD or DRD compound, as described herein, and a pharmaceutically acceptable carrier or diluent.

Another aspect of the present invention pertains to a method of inhibiting Bcl-2 protein function (especially one or both of Bcl-X_L and Mcl-1 ), in vitro or in vivo, comprising contacting a Bcl-2 protein (especially one or both of BCI-XL and Mcl-1) with an effective amount of a PRD or DRD compound, as described herein.

Another aspect of the present invention pertains to a method of Bcl-2 protein function (especially one or both of Bcl-X_L and Mcl-1) in a cell, in vitro or in vivo, comprising contacting the cell with an effective amount of a PRD or DRD compound, as described herein.

Another aspect of the present invention pertains to a method of regulating (e.g., inhibiting) cell proliferation (e.g., proliferation of a cell), promoting apoptosis, or a combination of both, in vitro or in vivo, comprising contacting a cell with an effective amount of a PRD or DRD compound, as described herein.

Another aspect of the present invention pertains to a method of treatment comprising administering to a subject in need of treatment a therapeutically-effective amount of a PRD or DRD compound, as described herein, preferably in the form of a pharmaceutical composition.

Another aspect of the present invention pertains to a PRD or DRD compound as described herein for use in a method of treatment of the human or animal body by therapy.

Another aspect of the present invention pertains to use of a PRD or DRD compound, as described herein, in the manufacture of a medicament for use in treatment.

In one embodiment, the treatment is treatment of a disease or condition that is mediated by Bcl-2 protein (especially one or both of Bcl-X_L and Mcl-1 ). In one embodiment, the treatment is treatment of a disease or condition that is ameliorated by the inhibition of Bcl-2 protein function (especially one or both of Bcl-X_L and McM ).

In one embodiment, the treatment is treatment of a proliferative condition.

In one embodiment, the treatment is treatment of cancer.

In one embodiment, the treatment is treatment of: lung cancer, breast cancer, ovarian cancer, CNS cancer or leukemia.

Another aspect of the present invention pertains to a kit comprising (a) a PRD or DRD compound, as described herein, preferably provided as a pharmaceutical composition and in a suitable container and/or with suitable packaging; and (b) instructions for use, for example, written instructions on how to administer the compound.

Another aspect of the present invention pertains to a PRD or DRD compound obtainable by a method of synthesis as described herein, or a method comprising a method of synthesis as described herein.

Another aspect of the present invention pertains to a PRD or DRD compound obtained by a method of synthesis as described herein, or a method comprising a method of synthesis as described herein.

Another aspect of the present invention pertains to novel intermediates, as described herein, which are suitable for use in the methods of synthesis described herein.

Another aspect of the present invention pertains to the use of such novel intermediates, as described herein, in the methods of synthesis described herein.

As will be appreciated by one of skill in the art, features and preferred embodiments of one aspect of the invention will also pertain to other aspects of the invention.

DETAILED DESCRIPTION OF THE INVENTION Compounds

One aspect of the present invention relates to certain pyridylrhodanine compounds

(for convenience, collectively referred to herein as "PRD compounds").

In one embodiment, the compounds are selected from compounds of formula I, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula I wherein:

X¹ is independently selected from CH₂ and C=S;

one of X² and X³ is N and the other is CH;

R¹ is independently selected from H or R^N, wherein R^N is independently branched or unbranched saturated or unsaturated C_1-2OaIKyI and is optionally substituted;

each of R^A and R^B is independently selected from H, C_1-20alkyl, C_1-20alkoxy, C₃-₂₀aryl, C_3-2oaryl-C_1-7alkyl, C_3-20heterocyclyl, halo, amino, OH or R^A and R^B together with the ring atoms to which they are attached form C₃.₇heterocyclyl, and is optionally substituted;

R^c is independently selected from halo and C_3-20aryl, and is optionally substituted;

n is independently 0 to 5;

and is a single or double bond.

*^**** In one embodiment the compounds are selected from compounds of formula II, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula Il wherein:

each of X¹, R¹, X², X³, R^A, R^B and n is independently as defined above with respect to formula I; and

each of R², R³, R⁴, R⁵ and R⁶ is independently selected from H, C_1-20alkyl, C_1- ₂oalkoxy, C_3-2oaryl, C₃.₂oaryl-Ci.₇alkyl, Cs^oheterocyclyl, halo, amino, OH and C_3- yheterocyclyl formed with an adjacent substituent and the ring atoms to which they are attached, and is optionally substituted.

In particularly preferred embodiments, the compound are selected from compounds of formula III, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula

wherein X², X³, R¹, R², R³ and R⁴ are as defined above with respect to formula I or formula II. xl

In embodiments X¹ is independently selected from CH₂ and C=S. In embodiments X¹ is independently CH₂. In embodiments X¹ is independently C=S.

X² and X³

In embodiments one of X² and X³ is N and the other is CH.

In embodiments X² is N and X³ is CH.

In embodiments X² is CH and X³ is N.

Sl

In embodiments R¹ is independently H or R^N. In embodiments R¹ is independently H. In embodiments R¹ is independently R^N.

R^

In embodiments R^N is independently branched or unbranched saturated or unsaturated C_1-20alkyl and is optionally substituted.

In embodiments R^N is independently branched or unbranched saturated or unsaturated C_1-15alkyl, preferably C_1-10alkyl, more preferably C_1-7alkyl and most preferably C_1-5alkyl, and is optionally substituted.

In embodiments R^N is branched. In embodiments R^N is saturated.

Preferably R^N is independently Ci_-2oalkyl and is substituted by R^AN, wherein R^AN is an anionic group or a group that is converted to an anionic group when metabolised (e.g. by enzymatic action) (e.g., in vivo). In embodiments, a group that is converted to an anionic group when metabilised is a prodrug, as defined herein.

Suitably anionic groups include: -COO^", -SO₃ ^", -SO₂ ^", -PO₃H^" and -B(OH)O^".

The anionic functionality can be provided by an acid, for example: -COOH, -SO₃H, - SO₂H, -P(O)(OH)₂ and -B(OH)₂. Acordingly, in embodiments R^AN is independently an acid, preferably independently selected from: -COOH, -SO₃H, -SO₂H, -P(O)(OH)₂ and -B(OH)₂.

Similarly, the anionic functionality can be provided by a salt, for example a Na or K salt, for example: -COONa, -SO₃Na, -SO₂Na, -P(O)(OH)ONa, -B(OH)ONa, -COOK, -SO₃K, -SO₂K, -P(O)(OH)OK and -B(OH)OK. Acordingly, in embodiments R^AN is independently an acid salt, preferably independently selected from: -COONa, - SO₃Na, -SO₂Na, -P(O)(OH)ONa, -B(OH)ONa, -COOK, -SO₃K, -SO₂K, -P(O)(OH)OK and -B(OH)OK.

Suitable groups that are converted to an anionic group when metabolised include esters, especially alkyl esters. Acordingly, preferably R^AN is independently an ester, preferably alkyl ester.

Preferably R^AN is independently selected from: -CO₂R, -SO₃R, -SO₂R, - P(O)(OH)(OR) and -B(OH)(OR), wherein R is an ester substituent as defined herein, preferably akyl, preferably R^E, wherein each R^E is independently selected from H, Ci- ₇alkyl and C₃:₁₂aryl.

Preferably R^N is independently Ci_-20alkyl and is substituted by R^AN, wherein R^AN is independently an acid group or an ester group. Suitably the acid group is selected from carboxylic acid, phosphonic acid, sulfonic acid and boronic acid. Suitably the ester group is selected from an ester of carboxylic acid, an ester of phosphonic acid, an ester of sulfonic acid and an ester of boronic acid.

Thus, preferably R^N is independently C_1-2Oalkyl and is substituted by one or more groups independently selected from -C(0)0R^E, -P(O)(OR^E)₂, -S0₃R^E and -B(OR^E)₂; wherein each R^E is independently as defined herein. Suitably R^AN (preferably being an acid group or ester group, more preferably selected from -C(O)OR^E, -P(O)(OR^E)₂, -SO₃R^E and -B(OR^E)₂) is on the α-carbon with respect to the nitrogen of the oxothioxothiazolidinyl ring.

Thus, preferably R^N is:

such that the NR^N group has the form:

wherein R^AN is as defined herein, preferably being an acid group or an ester group, more preferably selected from -C(O)OR^E, -P(O)(OR^E)₂, -SO₃R^E and -B(OR^E)₂; wherein each R^E is independently as defined herein; and R^Nα is selected from H and branched or unbranched, saturated or unsaturated C_1-10alkyl and is optionally substituted.

Thus, in embodiments, the NR^N group is an amino acid, amino phosphonic acid, amino sulfonic acid or amino boronic acid, or an ester of such acids.

It is particularly preferred that R^N is independently branched or unbranched, saturated or unsaturated C_1-2oalkyl and is substituted by -C(O)OR^E, i.e. R^N is - C(O)OR^E-C_1-20alkyl, preferably C(O)OR^E-C₁.₇alkyl and more preferably C(O)OR^E-C_1- salkyl.

In the preferred configuration wherein the -C(O)OR^E group is attached to the α- carbon, preferably R^N is:

C(O)OR¹ E wherein R^E is as defined herein, preferably H or C_1-3alkyl; and R^Nα is independently selected from H and branched or unbranched, saturated or unsaturated C_1-5alkyl and is optionally substituted.

In embodiments R^Nα is unsubstituted.

In embodiments R^Nα is substituted, preferably with C_3-2oaryl ,as discussed in more detail below.

Preferred embodiments of R^N are:

C(O)OR^ε C(O)ORE C(O)OR^E

and

wherein R^E is as defined herein, preferably H or C_1-3alkyl.

Particularly preferred embodiments of R^N are:

C(O)OH C(O)OH C(O)OH C(O)OH

C(O)OH C(O)OMe C(O)OMe C(O)OMe

C(O)OMe _and C(O)OMe As discussed in embodiments R^N is substituted with C_3-2oaryl, preferably C_5-i₂aryl, preferably C₅.₇aryl and more preferably Cβaryl, which aryl substituent is itself optionally substituted. A particularly preferred aryl substituent is phenyl, which is optionally substituted.

Thus, preferably R^N is independently selected from branched or unbranched saturated or unsaturated C_1-2oalkyl and C^oaryl-C^oalkyl and is optionally substituted.

Preferably R^N is independently selected from branched or unbranched saturated or unsaturated C_1-20alkyl substituted by R^AN and C₃._2oaryl-Ci-₂oalkyl substituted by R^AN and is optionally further substituted. Particularly preferred is C_1-10alkyl or C_5-Ba^l-C_1- ioalkyl substituted by R^AN and is optionally further substituted.

In embodiments R^N is substituted by C₃.₂₀aryl and R^AN. That is, in embodimets R^N is independently C_3-2oaryl-Ci_₂₀alkyl substituted by R^AN and is optionally further substituted.

Thus, in embodiments, R^N is aryl substituted alkyl, preferably C₅.₇aryl-Ci_-7alkyl, and is preferably substituted with R^AN as defined herein.

It is particularly preferred that R^N is :

_RNα

RAN

wherein R^AN is an acid group or an ester group, preferably selected from -C(O)OR^E, - P(O)(OR^E)₂, -SO₃R^E and -B(OR^E)₂; wherein each R^E is independently selected from H, C_1-7alkyl and C₃.i₂aryl; and R^Nα is selected from branched or unbranched, saturated or unsaturated d.ioalkyl optionally substituted with C_5-7aryl, which aryl is itself optionally substituted.

Suitably, as noted above, R^AN is -C(O)OR^E. In the preferred configuration wherein the -C(O)OR^E group is attached to the α-carbon, preferably R^N is: V

C(O)OR^E wherein R^E is as defined herein; and R^Nα is independently branched or unbranched, saturated or unsaturated C_1-5alkyl and is optionally substituted with C₅.₇aryl, which aryl is itself optionally substituted.

A preferred embodiment is:

wherein R^E is as defined herein, preferably H or C_1-3alkyl.

Particularly preferred embodiments are:

_□AN

In embodiments R^AN is an anionic group or a group that is converted to an anionic group when metabolised (e.g. by enzymatic action) (e.g., in vivo). In embodiments, a group that is converted to an anionic group when metabilised is a prodrug, as defined herein.

In embodiments R^AN is independently selected from an acid, an acid salt or an ester. In embodiments R^AN is independently selected from an acid and an ester. In embodiments R^AN is independently an acid. In embodiments R^AN is independently an ester. In embodiments R^AN is independently an acid salt. In embodiments R^AN is independently selected from: -COOH, -SO₃H, -SO₂H, - P(O)(OH)₂ and -B(OH)₂.

In embodiments R^AN is independently selected from: -CO₂R, -SO₃R, -SO₂R, -

P(O)(OH)(OR) and -B(OH)(OR), wherein R is an ester substituent as defined herein, preferably akyl, preferably R^E, wherein each R^E is independently selected from H, C_1- ₇alkyl and C₃.₁₂aryl.

In embodiments R^AN is independently selected from: -COONa, -SO₃Na, -SO₂Na, - P(O)(OH)ONa, -B(OH)ONa, -COOK, -SO₃K, -SO₂K, -P(O)(OH)OK and -B(OH)OK.

Rf

In embodiments R^E is independently selected from H, C_1-7alkyl and C_3-12aryl.

In embodiments R^E is independently selected from H, C_1-7alkyl and C_5-6aryl.

In embodiments R^E is independently selected from H and C_1-7alkyl.

In embodiments R^E is independently selected from H and C_1-3alkyl. In embodiments R^E is independently selected from H and dalkyl.

Preferably R^E is H.

R^A and R^B

In embodiments each of R^A and R^B is independently selected from H, C_1-2oalkyl, C_1- 2oalkoxy, C_3-20aryl, C_3-20aryl-C_1-7alkyl, C_3-2oheterocyclyl, halo, amino, OH or R^A and R^B together with the ring atoms to which they are attached form C_3-7heterocyclyl, and is optionally substituted.

In embodiments each of R^A and R^B is independently selected from H, C_1-7alkyl, C_1- ₇alkoxy, C_5-7aryl, C_5-7aryl-C_1-7alkyl, C_5-7heterocyclyl, halo, amino, OH or R^A and R^B together with the ring atoms to which they are attached form C_5-7heterocyclyl, and is optionally substituted. In embodiments each of R^A and R^B is independently selected from aryl, alkyl, arylalkane, halogen, amino, hydroxyl, hydrogen and a heterocyclic group, and is optionally substituted.

Preferably R^A is independently H. Preferably R^B is independently H. Preferably R^A is H and R^B is H.

R^

In embodiments R^c is independently selected from halo and C_3-2oaryl, and is optionally substituted;

In embodiments R^c is independently halo. In embodiments R^c is independently C_3-2oaryl and is optionally substituted.

In embodiments R^c is selected from F, Cl and Br, preferably Cl or Br and most preferably Br.

In embodiments R^c is C_5-12aryl, preferably C_5-7aryl, more preferably C_5-6aryl, more preferably, and most preferably phenyl and is optionally substituted.

Suitably R^c is substituted, preferably with one or more groups selected from the groups as defined herein with respect to each of R², R³, R⁴, R⁵ and R⁶, more preferably in respect of each of R², R³ and R⁴.

R², R³, R⁴, R⁵ and R⁶ In embodiments each of R², R³, R⁴, R⁵ and R⁶ is independently selected from H, C_1- ₂₀alkyl, Ci_-2oalkoxy, C_3-20aryl, C₃.₂₀aryl-C_1-7alkyl, C^oheterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached. In embodiments wherein any one of R², R³, R⁴, R⁵ and R⁶ is Ci_-2oalkyl, preferably each alkyl is independently C^salkyl, more preferably C_1-10alkyl, more preferably Gi- ₇alkyl, and most preferably C_halky!.

In embodiments wherein any one of R², R³, R⁴, R⁵ and R⁶ is C_1-2oalkoxy, preferably each alkoxy is independently Ci-i₅alkoxy, more preferably Ci_-10alkoxy, more preferably Ci_-7alkoxy, and most preferably C_1-5alkoxy.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is C_3-2oaryl, preferably each aryl is independently C_5-20aryl, more preferably C_5-i₂aryl, more preferably C_5-7aryl, more preferably C_5-6aryl and most preferably C₆aryl. A particularly preferred C₆aryl is phenyl, suitably unsubstituted phenyl.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is amino, preferably each amino is independently NR^N1R^N2, wherein R^N1 and R^N2 are independently amino substituents, suitably selected from H, C_1-7alkyl, C_3-20heterocyclyl, and C_5-20aryl, preferably H or C_1-7alkyl, or R^N1 and R^N2, taken together with the nitrogen atom to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms.

In embodiments wherein any one of R², R³, R⁴, R⁵ and R⁶ is halo, preferably each halo is independently selected from Cl, Br and F, more preferably Cl and Br and most preferably Cl.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is C₃.₂₀heterocyclyl, preferably each heterocyclyl is independently C_5-20heterocyclyl, more preferably C_5-12 heterocyclyl, more preferably C_5-7heterocycle, more preferably C_5-6heterocyclyl and most preferably C₆heterocyclyl.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is C_5-2oaryl-C_1-7alkyl, preferably each aryl-alkyl is independently C_5-i₅aryl-Ci_-7alkyl, more preferably

C_5-10aryl-C_1-7alkyl, more preferably C₅.₇aryl-C_1-7alkyl, more preferably C₅-₆aryl-Ci_-7alkyl and most preferably C₆aryl-Ci_-7alkyl.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is C_3-7heterocycle formed with an adjacent substituents and the ring atoms to which they are attached, preferably the C₃.₇heterocycle comprises one or two heteroatoms, preferably two heteroatoms.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is C_3-7heterocycle formed with an adjacent substituents and the ring atoms to which they are attached, preferably the C_3-7heterocycle comprises one or two oxygen ring atom, preferably two oxygen ring atoms.

In embodiments where any one of R², R³, R⁴, R⁵ and R⁶ is Cs-_T-heterocyclyl formed with an adjacent substituents and the ring atoms to which they are attached, preferably the heterocyclyl is C₃-₅heterocyclyl, more preferably C₅heterocyclyl.

An example of such a heterocycle formed by R³, R⁴ and the ring atoms to which they are attached is:

In embodiments each of R², R³, R⁴, R⁵ and R⁶ is independently selected from H, C_1- ₂oalkyl, C_1-20alkoxy, C_3-2oaryl, halo, amino, OH and C₃.₇heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

Preferably each of R², R³, R⁴, R⁵ and R⁶ is independently selected from H, Ci.₂oalkyl, C_1-20alkoxy, halo and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

Preferably each of R², R³, R⁴, R⁵ and R⁶ is independently selected from H, C_1- ₂₀alkoxy, halo and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

In embodiments each of R², R³, R⁴, R⁵ and R⁶ is independently selected from aryl, alkyl, arylalkane, halogen, amino, hydroxyl, hydrogen and a heterocyclic group, and is optionally substituted. In embodiments at least one of R², R³, R⁴, R⁵ and R⁶ is not H. In embodiments at least two of R², R³, R⁴, R⁵ and R⁶ are not H.

In embodiments at least one of R², R³ and R⁴ is not H. In embodiments at least two of R², R³ and R⁴ are not H.

In embodiments (i) R² is not H and R³ is not H, or (ii) R³ is not H and R⁴ is not H.

Thus, suitably the phenyl is mono- or di-substituted. In the case of di-substitution, 2,3-substitution or 3,4 substitution is particularly preferred.

In embodiments each of R⁵ and R⁶ is independently H.

In particularly preferred embodiments R⁵ is H and R⁶ is H and at least one, preferably at least two, of R², R³ and R⁴ is/are not H.

In particularly preferred embodiments each of R², R³ and R⁴ is selected from H, C_1- 2oalkyl, C_1-2OaIkOXy, halo and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

In particularly preferred embodiments two of R², R³ and R⁴ are independently selected from Ci_-20alkyl, C_1-2oalkoxy and halo or together (if adjacent) form C_3- ₇heterocycle with the ring atoms to which they are attached, and the other one is H.

In embodiments, R², R³ and R⁴ are selected as follows: (a) R² is Ci-₂₀alkoxy, R³ is C_1- ₂₀alkoxy and R⁴ is H; or (b) each of R³ and R⁴ is independently C_1-20alkyl, C_1-2oalkoxy, halo or together form a -0-CH₂-O- group, and R² is H. Suitably in such embodiments each of R⁵ and R⁶ is H.

Particularly preferred embodiments of R², R³, R⁴, R⁵ and R⁶ are:

and

In embodiments R² is independently selected from H, Ci_-2oalkyl, Ci_-2oalkoxy, C_3-2oaryl,

C3-₂oaryl-C_1-7alkyl, C_3-20heterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

Jn embodiments R² is independently selected from H, C₁.₂oalkyl, C_1-20alkoxy and halo.

In embodiments R² is independently selected from H, C_1-7alkyl, Ci_-7alkoxy and halo.

In embodiments R² is independently selected from H and C_1-7alkoxy.

In embodiments R² is independently selected from H and C_1-5alkoxy.

In embodiments R³ is independently selected from H, Ci_-20alkyl, C_1-20alkoxy, C_3-2Oaryl,

C_3-20aryl-Ci_-7alkyl, C_3-20heterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached.

In embodiments R³ is independently selected from H, C_1-20alkyl, C_1-20alkoxy, halo and

-0-CH₂-O- formed together with R⁴.

In embodiments R³ is independently selected from C_1-7alkyl, C_1-7alkoxy, halo and -O-

CH₂-O- formed together with R⁴.

In embodiments R³ is independently selected from d_-5alkoxy, halo and -0-CH₂-O- formed together with R⁴.

In embodiments R³ is independently selected from C_1-5alkoxy, Cl and -0-CH₂-O- formed together with R⁴. In embodiments R³ is independently selected from OCH₃, Cl and -0-CH₂-O- formed together with R⁴.

R⁴.

In embodiments R⁴ is independently selected from H, C_1-2oalkyl, C_1-2oalkoxy, C₃,₂oaryl,

C_3-2oaryl-Ci.₇alkyl, C₃.₂₀heterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached. In embodiments R⁴ is independently selected from H, C_1-20alkyl, C_1-2oalkoxy, halo and

-0-CH₂-O- formed together with R³.

In embodiments R⁴ is independently selected from Ci_-7alkyl, C_1-7alkoxy and -0-CH₂-

O- formed together with R³.

In embodiments R⁴ is independently selected from Ci_-5alkyl, C_1-5alkoxy and -0-CH₂- O- formed together with R³.

In embodiments R⁴ is independently selected from -C(CH₃)₃, -OCH₃, -OCH(CH₃)₂ and -0-CH₂-O- formed together with R³.

Rf

In embodiments R² is independently selected from H, Ci_-2oalkyl, C_1-2oalkoxy, C_3-20aryl, C_3-20aryl-C_1-7alkyl, C_3-20heterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached. In embodiments R⁵ is H.

Rf

In embodiments R² is independently selected from H, Ci_-20alkyl, C_1-20alkoxy, C_3-20aryl, C_3-20aryl-Ci-₇alkyl, C_3-20heterocyclyl, halo, amino, OH and C_3-7heterocycle formed with an adjacent substituent and the ring atoms to which they are attached. In embodiments R⁶ is H. In embodiments n is independently 0 to 5.

In embodiments n is 0 to 3, preferably 0 to 2, more preferably 0 or 1 and most preferably 0.

In embodiments is independently a single bond or a double bond In embodiments is a double bond.

As discussed above, the present invention pertains to compounds based on the pyridine alkenyl rhodanine core structure (see formula IV and formula V below) and its reduced forms (see formula Vl and formula VII below) designed to inhibit the function of pro-survival Bcl-2 proteins.

Thus, preferably the compound is selected from one of Formula IV, V, Vl and VII:

Formula IV

Formula V

suitably wherein R², R³, R⁴, R⁵, R⁶, R⁷ and R⁸ = various substituents including but not limited to aryl, alkyl, arylalkanes, halogens, amino, hydroxyl, hydrogen, heterocyclic groups;

NR = compounds including but not limited to amino acids, aminosulfonic acids, aminophosphonic acids, amino boronic acids;

X²=N, X³ = C; or X² = C, X³ = N; and (where present) n = 1 , 2, 3...

In a further aspect, the present invention relates to certain pyridylrhodanine compounds, wherein the compounds are selected from compounds of formula IV and pharmaceutically acceptable salts, hydrates, and solvates thereof.

In a further aspect, the present invention relates to certain pyridylrhodanine compounds, wherein the compounds are selected from compounds of formula V and pharmaceutically acceptable salts, hydrates, and solvates thereof.

In a further aspect, the present invention relates to certain pyridylrhodanine compounds, wherein the compounds are selected from compounds of formula Vl and pharmaceutically acceptable salts, hydrates, and solvates thereof.

In a further aspect, the present invention relates to certain pyridylrhodanine compounds, wherein the compounds are selected from compounds of formula VII and pharmaceutically acceptable salts, hydrates, and solvates thereof.

_*****

In a further aspect, the present invention provides compounds according to any one of structures 1 to 4 and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Ri . R₂. R3. R4. R5. Re. R7=various substituents including but not limited to aryl, alkyl, arylalkanes, halogens, amino, hydroxy, hydrogen, heterocyclic groups

NR=compounds including but not limited to amino acids aminosulfonic acids, aminophosphonic acids, amino boronic acids

R₁, R₂, R₃, R₄, R₅, R₆, R₇=various substituents including but not limited to aryl, alkyl, arylalkanes, halogens, amino, hydroxy, hydrogen, heterocyclic groups

NR=compounds including but not limited to amino acids

aminosulfonic acids, aminophosphonic acids, amino boronic acids Structure 2

Structure 3

R-i - R2. R3, R₄. R5. Re. R₇=various substituents including but not limited to aryl, alkyl, arylalkanes, halogens, amino, hydroxy, hydrogen, heterocyclic groups

Structure 4

In a further aspect the present invention relates to certain diarylrhodanine compounds (for convenience, collectively referred to herein as "DRD compounds").

In one embodiment, the compounds are selected from compounds of formula VIII, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula VIII wherein:

each of X , R ₁ R ₁ R and n is independently as defined herein; and

R^D is independently C_3-2oaryl, and is optionally substituted.

In a particularly preferred embodiment, the compounds are selected from compounds of formula IX, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula IX

wherein each of R , Rr, R , R^ά, R , R^&, R and R are as defined herein.

In embodiments R^D is C_3-20aryl and is optionally substituted. In embodiments R^D is C_5- i₂aryl and is optionally substituted, preferably C_5-7aryl and is optionally substituted, more preferably C_5-6aryl and is optionally substituted, more preferably C₆aryl and is optionally substituted, and most preferably phenyl and is optionally substituted. Suitably R^D is substituted, preferably with one or more groups selected from the groups as defined herein with respect to each of R², R³, R⁴, R⁵ and R⁶, more preferably in respect of each of R², R³ and R⁴.

In an especially preferred embodiment, the compounds are selected from compounds of formula X, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula X

wherein each of R², R³, R⁴ and R^N are as defined herein.

In a yet more preferred embodiment the compounds are selected from compounds of formula Xl, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

wherein each of R , R , R , R and R is independently as defined herein. Substituents The phrase "optionally substituted" as used herein, pertains to a parent group which may be unsubstituted or which may be substituted.

Unless otherwise specified, the term "substituted" as used herein, pertains to a parent group which bears one or more substitutents. The term "substituent" is used herein in the conventional sense and refers to a chemical moiety which is covalently attached to, or if appropriate, fused to, a parent group. A wide variety of substituents are well known, and methods for their formation and introduction into a variety of parent groups are also well known.

Examples of substituents and preferred forms for such substituents are described in more detail below.

Alkyl: The term "alkyl," as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of a hydrocarbon compound having from 1 to 20 carbon atoms (unless otherwise specified), which may be aliphatic or alicyclic, and which may be saturated or unsaturated (e.g., partially unsaturated, fully unsaturated). Thus, the term "alkyl" includes the sub-classes alkenyl, alkynyl, cycloalkyl, cycloalkyenyl, cylcoalkynyl, etc., discussed below.

In the context of alkyl groups, the prefixes (e.g., C_1-4, C_1-7, C_1-20, C_2-7, C_3-7, etc.) denote the number of carbon atoms, or range of number of carbon atoms. For example, the term "Ci_₄alkyl," as used herein, pertains to an alkyl group having from 1 to 4 carbon atoms. Examples of groups of alkyl groups include C_1-4alkyl ("lower alkyl"), C_1-7alkyl, and C^alkyl. Note that the first prefix may vary according to other limitations; for example, for unsaturated alkyl groups, the first prefix must be at least 2; for cyclic and branched alkyl groups, the first prefix must be at least 3; etc.

Examples of (unsubstituted) saturated alkyl groups include, but are not limited to, methyl (C₁), ethyl (C₂), propyl (C₃), butyl (C₄), pentyl (C₅), hexyl (C₆), heptyl (C₇), octyl (C₈), nonyl (C₉), decyl (C₁₀), undecyl (C₁₁), dodecyl (C₁₂), tridecyl (C₁₃), tetradecyl (C₁₄), pentadecyl (C₁₅), and eicodecyl (C₂₀). Examples of (unsubstituted) saturated linear alkyl groups include, but are not limited to, methyl (C₁), ethyl (C₂), n-propyl (C₃), n-butyl (C₄), n-pentyl (amyl) (C₅), n-hexyl (C₆), and n-heptyl (C₇).

Examples of (unsubstituted) saturated branched alkyl groups include iso-propyl (C₃), iso-butyl (C₄), sec-butyl (C₄), tert-butyl (C₄), iso-pentyl (C₅), and neo-pentyl (C₅).

Alkenyl: The term "alkenyl," as used herein, pertains to an alkyl group having one or more carbon-carbon double bonds. Examples of groups of alkenyl groups include C_2-4alkenyl, C_2-7alkenyl, C_2-20alkenyl.

Examples of (unsubstituted) unsaturated alkenyl groups include, but are not limited to, ethenyl (vinyl, -CH=CH₂), 1-propenyl (-CH=CH-CH₃), 2-propenyl (allyl, -CH-CH=CH₂), isopropenyl (1-methylvinyl, -C(CH₃)=CH₂), butenyl (C₄), pentenyl (C₅), and hexenyl (Ce)-

Alkynyl: The term "alkynyl," as used herein, pertains to an alkyl group having one or more carbon-carbon triple bonds. Examples of groups of alkynyl groups include C_2-4alkynyl, C_2-7alkynyl, C_2-20alkynyl.

Examples of (unsubstituted) unsaturated alkynyl groups include, but are not limited to, ethynyl (ethinyl, -C≡CH) and 2-propynyl (propargyl, -CH₂-CMDH).

Cycloalkyl: The term "cycloalkyl," as used herein, pertains to an alkyl group which is also a cyclyl group; that is, a monovalent moiety obtained by removing a hydrogen atom from an alicyclic ring atom of a carbocyclic ring of a carbocyclic compound, which carbocyclic ring may be saturated or unsaturated (e.g., partially unsaturated, fully unsaturated), which moiety has from 3 to 20 carbon atoms (unless otherwise specified), including from 3 to 20 ring atoms. Thus, the term "cycloalkyl" includes the sub-classes cycloalkyenyl and cycloalkynyl. Preferably, each ring has from 3 to 7 ring atoms. Examples of groups of cycloalkyl groups include C_3-20cycloalkyl, C₃.₁₅cycloalkyl, C_3-1ocycloalkyl, C_3-7cycloalkyl.

Examples of cycloalkyl groups include, but are not limited to, those derived from: saturated monocyclic hydrocarbon compounds: cyclopropane (C₃), cyclobutane (C₄), cyclopentane (C₅), cyclohexane (C₆), cycloheptane (C₇), methylcyclopropane (C₄), dimethylcyclopropane (C₅), methylcyclobutane (C₅), dimethylcyclobutane (C₆), methylcyclopentane (C₆), dimethylcyclopentane (C₇), methylcyclohexane (C₇), dimethylcyclohexane (C₈), menthane (Ci₀); unsaturated monocyclic hydrocarbon compounds: cyclopropene (C₃), cyclobutene (C₄), cyclopentene (C₅), cyclohexene (C₆), methylcyclopropene (C₄), dimethylcyclopropene (C₅), methylcyclobutene (C₅), dimethylcyclobutene (C₆), methylcyclopentene (C₆), dimethylcyclopentene (C₇), methylcyclohexene (C₇), dimethylcyclohexene (C₈); saturated polycyclic hydrocarbon compounds: thujane (C₁₀); carane (C₁₀), pinane (C₁₀), bornane (C₁₀), norcarane (C₇), norpinane (C₇), norbomane (C₇), adamantane (C₁₀), decalin (decahydronaphthalene) (C₁₀); unsaturated polycyclic hydrocarbon compounds: camphene (C₁₀), limonene (C₁₀), pinene (C₁₀); polycyclic hydrocarbon compounds having an aromatic ring: indene (C₉), indane (e.g., 2,3-dihydro-1H-indene) (C₉), tetraline (1 ,2,3,4-tetrahydronaphthalene) (C₁₀), acenaphthene (C₁₂), fluorene (C₁₃), phenalene (C₁₃), acephenanthrene (C₁₅), aceanthrene (C₁₆), cholanthrene (C₂₀).

Alkylidene: The term "alkylidene," as used herein, pertains to a divalent monodentate moiety obtained by removing two hydrogen atoms from an aliphatic or alicyclic carbon atom of a hydrocarbon compound having from 1 to 20 carbon atoms (unless otherwise specified). Examples of groups of alkylidene groups include C_1-2oalkylidene, d^alkylidene, C_1-4alkylidene.

Examples of alkylidene groups include, but are not limited to, methylidene (=CH₂), ethylidene (=CH-CH₃), vinylidene (=C=CH₂), isopropylidene (=C(CH₃)₂), cyclopentylidene, and benzylidene (=CH-Ph).

Alkylidyne: The term "alkylidyne," as used herein, pertains to a trivalent monodentate moiety obtained by removing three hydrogen atoms from an aliphatic or alicyclic carbon atom of a hydrocarbon compound having from 1 to 20 carbon atoms (unless otherwise specified). Examples of groups of alkylidyne groups include Ci_-2oalkylidyne, Ci_-7alkylidyne, C^alkylidyne. Examples of alkylidyne groups include, but are not limited to, methylidyne ( ≡CH), ethylidyne ( ≡C-CH₃), and benzylidyne ( ≡C-Ph).

Carbocyclyl: The term "carbocyclyl," as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a carbocyclic compound, which moiety has from 3 to 20 ring atoms (unless otherwise specified). Preferably, each ring has from 3 to 7 ring atoms.

In this context, the prefixes (e.g., C_3-20, C_3-7, C_5-6, etc.) denote the number of ring atoms, or range of number of ring atoms. For example, the term "C₅.₆carbocyclyl," as used herein, pertains to a carbocyclyl group having 5 or 6 ring atoms. Examples of groups of carbocyclyl groups include C_3-2ocarbocyclyl, C₃.₁₀carbocyclyl, C_5-10carbocyclyl, Cs^carbocyclyl, and C_5-7carbocyclyl.

Examples of carbocyclic groups include, but are not limited to, those described above as cycloalkyl groups; and those described below as carboaryl groups.

Heterocyclyl: The term "heterocyclyl," as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a heterocyclic compound, which moiety has from 3 to 20 ring atoms (unless otherwise specified), of which from 1 to 10 are ring heteroatoms. Preferably, each ring has from 3 to 7 ring atoms, of which from 1 to 4 are ring heteroatoms.

In this context, the prefixes (e.g., C_3-20, C_3-7, C_5-6, etc.) denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms. For example, the term "C_5-6heterocyclyl," as used herein, pertains to a heterocyclyl group having 5 or 6 ring atoms. Examples of groups of heterocyclyl groups include C_3-20heterocyclyl, C_5-2oheterocyclyl, C_3-15heterocyclyl, C_5-15heterocyclyl,

Cs-ioheterocyclyl, C_5-10heterocyclyl, C₃.₇heterocyclyl, C_5-7heterocyclyl, and Cs-εheterocyclyl.

Examples of (non-aromatic) monocyclic heterocyclyl groups include, but are not limited to, those derived from: N₁: aziridine (C₃), azetidine (C₄), pyrrolidine (tetrahydropyrrole) (C₅), pyrroline (e.g., 3-pyrroline, 2,5-dihydropyrrole) (C₅), 2H-pyrrole or 3H-pyrrole (isopyrrole, isoazole) (C₅), piperidine (C₆), dihydropyridine (C₆), tetrahydropyridine (C₆), azepine (C₇);

O₁: oxirane (C₃), oxetane (C₄), oxolane (tetrahydrofuran) (C₅), oxole (dihydrofuran) (C₅), oxane (tetrahydropyran) (C₆), dihydropyran (C₆), pyran (C₆), oxepin (C₇);

S₁: thiirane (C₃), thietane (C₄), thiolane (tetrahydrothiophene) (C₅), thiane (tetrahydrothiopyran) (C₆), thiepane (C₇);

O₂: dioxolane (C₅), dioxane (C₆), and dioxepane (C₇);

O₃: trioxane (C₆);

N₂: imidazolidine (C₅), pyrazolidine (diazolidine) (C₅), imidazoline (C₅), pyrazoline (dihydropyrazole) (C₅), piperazine (C₆);

N₁O₁: tetrahydrooxazole (C₅), dihydrooxazole (C₅), tetrahydroisoxazole (C₅), dihydroisoxazole (C₅), morpholine (C₆), tetrahydrooxazine (C₆), dihydrooxazine (C₆), oxazine (C₆);

N₁S₁: thiazoline (C₅), thiazolidine (C₅), thiomorpholine (C₆);

N₂O₁: oxadiazine (C₆);

O₁S₁: oxathiole (C₅) and oxathiane (thioxane) (C₆); and,

N₁O₁S₁: oxathiazine (C₆).

Examples of substituted (non-aromatic) monocyclic heterocyclyl groups include those derived from saccharides, in cyclic form, for example, furanoses (C₅), such as arabinofuranose, lyxofuranose, ribofuranose, and xylofuranse, and pyranoses (C₆), such as allopyranose, altropyranose, glucopyranose, mannopyranose, gulopyranose, idopyranose, galactopyranose, and talopyranose. Examples of heterocyclyl groups which are also heteroaryl groups are described below with aryl groups.

Aryl: The term "aryl," as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound, which moiety has from 3 to 20 ring atoms (unless otherwise specified). Preferably, each ring has from 5 to 7 ring atoms.

In this context, the prefixes (e.g., C_3-20, C_5-7, C_5-6, etc.) denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms. For example, the term "C₅.₆aryl," as used herein, pertains to an aryl group having 5 or 6 ring atoms. Examples of groups of aryl groups include C_3-20aryl, C_5-2oaryl, C_5-15aryl, C_5-i₂aryl, C_5-10aryl, C₅.₇aryl, C_5-6aryl, C₅aryl, and C₆aryl.

The ring atoms may be all carbon atoms, as in "carboaryl groups." Examples of carboaryl groups include C₃.₂ocarboaryl, C_5-20carboaryl, C_5-15carboaryl, C_5-12carboaryl, C_5-iocarboaryl, C_5-7carboaryl, C_5-6Ca rboary I₁ C₅carboaryl, and C₆carboaryl.

Examples of carboaryl groups include, but are not limited to, those derived from benzene (i.e., phenyl) (C₆), naphthalene (Ci₀), azulene (C₁₀), anthracene (C₁₄), phenanthrene (C₁₄), naphthacene (C₁₈), and pyrene (C₁₆).

Examples of aryl groups which comprise fused rings, at least one of which is an aromatic ring, include, but are not limited to, groups derived from indane (e.g., 2,3- dihydro-1H-indene) (C₉), indene (C_g), isoindene (C₉), tetraline

(1 ,2,3,4-tetrahydronaphthalene (C₁₀), acenaphthene (C₁₂), fluorene (C₁₃), phenalene (C₁₃), acephenanthrene (Ci₅), and aceanthrene (C₁₆).

Alternatively, the ring atoms may include one or more heteroatoms, as in "heteroaryl groups." Examples of heteroaryl groups include C₃.₂₀heteroaryl, C_5-20heteroaryl, C_5-15heteroaryl, C₅.₁₂heteroaryl, C_5-i₀heteroaryl, C_5-7heteroaryl, C_5-6heteroaryl, C₅heteroaryl, and C₆heteroaryl.

Examples of monocyclic heteroaryl groups include, but are not limited to, those derived from:

N₁: pyrrole (azole) (C₅), pyridine (azine) (C₆); O₁: furan (oxole) (C₅); S₁: thiophene (thiole) (C₅); N₁O₁: oxazole (C₅), isoxazole (C₅), isoxazine (C₆); N₂O₁: oxadiazole (furazan) (C₅); N₃O₁: oxatriazole (C₅);

N₁S₁: thiazole (C₅), isothiazole (C₅);

N₂: imidazole (1,3-diazole) (C₅), pyrazole (1 ,2-diazole) (C₅), pyridazine (1 ,2-diazine) (C₆), pyrimidine (1 ,3-diazine) (C₆) (e.g., cytosine, thymine, uracil), pyrazine (1 ,4-diazine) (C₆); N₃: triazole (C₅), triazine (C₆); and, N₄: tetrazole (C₅).

Examples of heterocyclic groups (some of which are also heteroaryl groups) which comprise fused rings, include, but are not limited to: Cgheterocyclic groups (with 2 fused rings) derived from benzofuran (O₁), isobenzofuran (Oi), indole (N₁), isoindole (N₁), indolizine (N₁), indoline (N₁), isoindoline (N₁), purine (N₄) (e.g., adenine, guanine), benzimidazole (N₂), indazole (N₂), benzoxazole (N₁O₁), benzisoxazole (N₁O₁), benzodioxole (O₂), benzofurazan (N₂O₁), benzotriazole (N₃), benzothiofuran (S₁), benzothiazole (N₁S₁), benzothiadiazole (N₂S);

C₁₀heterocyclic groups (with 2 fused rings) derived from chromene (O₁), isochromene (O₁), chroman (O₁), isochroman (O₁), benzodioxan (O₂), quinoline (N₁), isoquinoline (N₁), quinolizine (N₁), benzoxazine (N₁O₁), benzodiazine (N₂), pyridopyridine (N₂), quinoxaline (N₂), quinazoline (N₂), cinnoline (N₂), phthalazine (N₂), naphthyridine (N₂), pteridine (N₄);

C_Tiheterocylic groups (with 2 fused rings) derived from benzodiazepine (N₂); C₁₃heterocyclic groups (with 3 fused rings) derived from carbazole (N₁), dibenzofuran (O₁), dibenzothiophene (S₁), carboline (N₂), perimidine (N₂), pyridoindole (N₂); and, C₁₄heterocyclic groups (with 3 fused rings) derived from acridine (N₁), xanthene (O₁), thioxanthene (S₁), oxanthrene (O₂), phenoxathiin (O₁Si), phenazine (N₂), phenoxazine (NiO₁), phenothiazine (N₁S₁), thianthrene (S₂), phenanthridine (N₁), phenanthroline (N₂), phenazine (N₂).

Heterocyclic groups (including heteroaryl groups) which have a nitrogen ring atom in the form of an -NH- group may be N-substituted, that is, as -NR-. For example, pyrrole may be N-methyl substituted, to give N-methylpyrrole. Examples of N- substitutents include, but are not limited to Ci-7alkyl, C₃.₂oheterocyclyl, C_5-2oaryl, and acyl groups.

Heterocyclic groups (including heteroaryl groups) which have a nitrogen ring atom in the form of an -N= group may be substituted in the form of an N-oxide, that is, as -N(→O)= (also denoted -N⁺(→O^")=). For example, quinoline may be substituted to give quinoline N-oxide; pyridine to give pyridine N-oxide; benzofurazan to give benzofurazan N-oxide (also known as benzofuroxan).

Cyclic groups may additionally bear one or more oxo (=0) groups on ring carbon atoms.

Monocyclic examples of such groups include, but are not limited to, those derived from:

C₅: cyclopentanone, cyclopentenone, cyclopentadienone;

C₆: cyclohexanone, cyclohexenone, cyclohexadienone;

O₁: furanone (C₅), pyrone (C₆);

N₁: pyrrolidone (pyrrolidinone) (C₅), piperidinone (piperidone) (C₆), piperidinedione (C₆);

N₂: imidazolidone (imidazolidinone) (C₅), pyrazolone (pyrazolinone) (C₅), piperazinone (C₆), piperazinedione (C₆), pyridazinone (C₆), pyrimidinone (C₆) (e.g., cytosine), pyrimidinedione (C₆) (e.g., thymine, uracil), barbituric acid (C₆);

N₁S₁: thiazolone (C₅), isothiazolone (C₅); N₁O₁: oxazolinone (C₅).

Polycyclic examples of such groups include, but are not limited to, those derived from:

C₉: indenedione; C₁₀: tetralone, decalone;

C₁₄: anthrone, phenanthrone;

N₁: oxindole (C₉);

O₁: benzopyrone (e.g., coumarin, isocoumarin, chromone) (C₁₀);

N₁O₁: benzoxazolinone (C₉), benzoxazolinone (C₁₀); N₂: quinazolinedione (C₁₀); benzodiazepinone (C₁₁); benzodiazepinedione (C₁₁);

N₄: purinone (C₉) (e.g., guanine). Still more examples of cyclic groups which bear one or more oxo (=0) groups on ring carbon atoms include, but are not limited to, those derived from: cyclic anhydrides (-C(=O)-O-C(=O)- in a ring), including but not limited to maleic anhydride (C₅), succinic anhydride (C₅), and glutaric anhydride (C₆); cyclic carbonates (-O-C(=O)-O- in a ring), such as ethylene carbonate (C₅) and 1 ,2-propylene carbonate (C₅); imides (-C(=O)-NR-C(=O)- in a ring), including but not limited to, succinimide (C₅), maleimide (C₅), phthalimide, and glutarimide (C₆); lactones (cyclic esters, -O-C(=O)- in a ring), including, but not limited to,

/?-propiolactone, γ-butyrolactone, <J-valerolactone (2-piperidone), and e-caprolactone; lactams (cyclic amides, -NR-C(=O)- in a ring), including, but not limited to, /?-propiolactam (C₄), γ-butyrolactam (2-pyrrolidone) (C₅), cJ-valerolactam (C₆), and e-caprolactam (C₇); cyclic carbamates (-O-C(=O)-NR- in a ring), such as 2-oxazolidone (C₅); cyclic ureas (-NR-C(=O)-NR- in a ring), such as 2-imidazolidone (C₅) and pyrimidine-2,4-dione (e.g., thymine, uracil) (C₆).

The above groups, whether alone or part of another substituent, may themselves optionally be substituted with one or more groups selected from themselves and the additional substituents listed below.

Hydrogen: -H. Note that if the substituent at a particular position is hydrogen, it may be convenient to refer to the compound or group as being "unsubstituted" at that position.

Halo: -F, -Cl, -Br, and -I.

Hydroxy: -OH.

Ether: -OR, wherein R is an ether substituent, for example, a C_1-7alkyl group (also referred to as a C_1-7alkoxy group, discussed below), a C3-₂oheterocyclyl group (also referred to as a C₃.₂oheterocyclyloxy group), or a C_5-2oaryl group (also referred to as a C₅.₂₀aryloxy group), preferably a C_1-7alkyl group. Alkoxy: -OR, wherein R is an alkyl group, for example, a C_1-7alkyl group. Examples of C_1-7alkoxy groups include, but are not limited to, -OMe (methoxy), -OEt (ethoxy), - O(nPr) (n-propoxy), -O(iPr) (isopropoxy), -O(nBu) (n-butoxy), -O(sBu) (sec-butoxy), -O(iBu) (isobutoxy), and -O(tBu) (tert-butoxy).

Acetal: -CH(OR¹)(OR²), wherein R¹ and R² are independently acetal substituents, for example, a C_1-7alkyl group, a C₃-₂oheterocyclyl group, or a C_5-2oaryl group, preferably a Ci_-7alkyl group, or, in the case of a "cyclic" acetal group, R¹ and R², taken together with the two oxygen atoms to which they are attached, and the carbon atoms to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms.

Examples of acetal groups include, but are not limited to, -CH(OMe)₂, -CH(OEt)₂, and -CH(OMe)(OEt).

Hemiacetal: -CH(OH)(OR¹), wherein R¹ is a hemiacetal substituent, for example, a C_1-7alkyl group, a C₃.₂₀heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples of hemiacetal groups include, but are not limited to, -CH(OH)(OMe) and -CH(OH)(OEt).

Ketal: -CR(0R¹)(0R²), where R¹ and R² are as defined for acetals, and R is a ketal substituent other than hydrogen, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples ketal groups include, but are not limited to, -C(Me)(OMe)₂, -C(Me)(OEt)₂, -C(Me)(OMe)(OEt), - C(Et)(OMe)₂, -C(Et)(OEt)₂, and -C(Et)(OMe)(OEt).

Hemiketal: -CR(OH)(OR¹), where R¹ is as defined for hemiacetals, and R is a hemiketal substituent other than hydrogen, for example, a Ci_-7alkyl group, a C_3-2oheterocyclyl group, or a C_5-20aryl group, preferably a Ci_-7alkyl group. Examples of hemiacetal groups include, but are not limited to, -C(Me)(OH)(OMe), - C(Et)(OH)(OMe), -C(Me)(OH)(OEt), and -C(Et)(OH)(OEt).

Oxo (keto, -one): =O.

Thione (thioketone): =S.

lmino (imine): =NR, wherein R is an imino substituent, for example, hydrogen,

C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably hydrogen or a C_1-7alkyl group. Examples of ester groups include, but are not limited to, =NH, =NMe, =NEt, and =NPh.

Formyl (carbaldehyde, carboxaldehyde): -C(=O)H.

Acyl (keto): -C(=O)R, wherein R is an acyl substituent, for example, a C_1-7alkyl group (also referred to as Ci_-7alkylacyl or C_1-7alkanoyl), a C_3-2oheterocyclyl group (also referred to as C_3-2oheterocyclylacyl), or a C_5-2oaryl group (also referred to as C_5-2oarylacyl), preferably a C_1-7alkyl group. Examples of acyl groups include, but are not limited to, -C(=O)CH₃ (acetyl), -C(=O)CH₂CH₃ (propionyl), -C(=O)C(CH₃)₃ (t-butyryl), and -C(=O)Ph (benzoyl, phenone).

Carboxy (carboxylic acid): -C(=O)OH.

Thiocarboxy (thiocarboxylic acid): -C(=S)SH.

Thiolocarboxy (thiolocarboxylic acid): -C(=O)SH.

Thionocarboxy (thionocarboxylic acid): -C(=S)OH.

lmidic acid: -C(=NH)OH.

Hydroxamic acid: -C(=NOH)OH.

Ester (carboxylate, carboxylic acid ester, oxycarbonyl): -C(=O)OR, wherein R is an ester substituent, for example, a C_1-7alkyl group, a C_3-2oheterocyclyl group, or a C₅.₂₀aryl group, preferably a C_1-7alkyl group. Examples of ester groups include, but are not limited to, -C(=O)OCH₃, -C(=O)OCH₂CH₃, -C(=O)OC(CH₃)₃, and -C(=O)OPh.

Acyloxy (reverse ester): -OC(=O)R, wherein R is an acyloxy substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a Ci_-7alkyl group. Examples of acyloxy groups include, but are not limited to, -OC(=O)CH₃ (acetoxy), -OC(=O)CH₂CH₃, -OC(=O)C(CH₃)₃, -OC(=O)Ph, and -OC(=O)CH₂Ph.

Oxycarboyloxy: -OC(=O)OR, wherein R is an ester substituent, for example, a

C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples of ester groups include, but are not limited to, -0C(=0)0CH_3l -OC(=O)OCH₂CH₃, -OC(=O)OC(CH₃)₃, and -OC(=O)OPh.

Amino: -NR¹R², wherein R¹ and R² are independently amino substituents, for example, hydrogen, a C_1-7alkyl group (also referred to as C_1-7alkylamino or di- C_1-7alkylamino), a C_3-2oheterocyclyl group, or a C₅.₂oaryl group, preferably H or a C_1-7alkyl group, or, in the case of a "cyclic" amino group, R¹ and R², taken together with the nitrogen atom to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms. Amino groups may be primary (-NH₂), secondary (-NHR¹), or tertiary (-NHR¹R²), and in cationic form, may be quaternary (-⁺NR¹R²R³). Examples of amino groups include, but are not limited to, -NH₂, -NHCH₃, -NHC(CH₃)₂, -N(CH₃)₂, -N(CH₂CH₃)₂, and -NHPh. Examples of cyclic amino groups include, but are not limited to, aziridino, azetidino, pyrrolidino, piperidino, piperazino, morpholino, and thiomorpholino.

Amido (carbamoyl, carbamyl, aminocarbonyl, carboxamide): -C(=O)NR¹R², wherein R¹ and R² are independently amino substituents, as defined for amino groups. Examples of amido groups include, but are not limited to, -C(=O)NH₂, -C(=O)NHCH₃, -C(=O)N(CH₃)₂, -C(=O)NHCH₂CH₃, and -C(=O)N(CH₂CH₃)₂, as well as amido groups in which R¹ and R², together with the nitrogen atom to which they are attached, form a heterocyclic structure as in, for example, piperidinocarbonyl, morpholinocarbonyl, thiomorpholinocarbonyl, and piperazinocarbonyl.

Thioamido (thiocarbamyl): -C(=S)NR¹R², wherein R¹ and R² are independently amino substituents, as defined for amino groups. Examples of amido groups include, but are not limited to, -C(=S)NH₂, -C(=S)NHCH₃, -C(=S)N(CH₃)₂, and -C(=S)NHCH₂CH₃.

Acylamido (acylamino): -NR¹C(=O)R², wherein R¹ is an amide substituent, for example, hydrogen, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably hydrogen or a C_1-7alkyl group, and R² is an acyl substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably hydrogen or a C_1-7alkyl group. Examples of acylamide groups include, but are not limited to, -NHC(=O)CH₃ , -NHC(=O)CH₂CH₃, and -NHC(=O)Ph. R¹ and R² may together form a cyclic structure, as in, for example, succinimidyl, maleimidyl, and phthalimidyl:

succinimidyl maleimidyl phthalimidyl

Aminocarbonyloxy: -OC(=O)NR¹R², wherein R¹ and R² are independently amino substituents, as defined for amino groups. Examples of aminocarbonyloxy groups include, but are not limited to, -OC(=O)NH₂, -OC(=O)NHMe, -OC(=O)NMe₂, and -OC(=O)NEt₂.

Ureido: -N(R¹)CONR²R³ wherein R² and R³ are independently amino substituents, as defined for amino groups, and R1 is a ureido substituent, for example, hydrogen, a C_1-7alkyl group, a C₃.₂oheterocyclyl group, or a C_5-20aryl group, preferably hydrogen or a C_1-7alkyl group. Examples of ureido groups include, but are not limited to, - NHCONH₂, -NHCONHMe, -NHCONHEt, -NHCONMe₂, -NHCONEt₂, -NMeCONH₂, - NMeCONHMe, -NMeCONHEt, -NMeCONMe₂, and -NMeCONEt₂.

Guanidino: -NH-C(=NH)NH₂.

Tetrazolyl: a five membered aromatic ring having four nitrogen atoms and one carbon atom,

Imino: =NR, wherein R is an imino substituent, for example, for example, hydrogen, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably H or a C_1-7alkyl group. Examples of imino groups include, but are not limited to, =NH, =NMe, and =NEt.

Amidine (amidino): -C(=NR)NR_2l wherein each R is an amidine substituent, for example, hydrogen, a C_1-7alkyl group, a C₃.₂₀heterocyclyl group, or a C_5-20aryl group, preferably H or a Ci_-7alkyl group. Examples of amidine groups include, but are not limited to, -C(=NH)NH₂, -C(=NH)NMe₂, and -C(=NMe)NMe₂. Nitro: -NO₂.

Nitroso: -NO.

Azido: -N₃.

Cyano (nitrile, carbonitrile): -CN.

Isocyano: -NC.

Cyanato: -OCN.

Isocyanato: -NCO.

Thiocyano (thiocyanato): -SCN.

lsothiocyano (isothiocyanato): -NCS.

Sulfhydryl (thiol, mercapto): -SH.

Thioether (sulfide): -SR, wherein R is a thioether substituent, for example, a C_1-7a!kyl group (also referred to as a C_1-7alkylthio group), a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples of C_1-7alkylthio groups include, but are not limited to, -SCH₃ and -SCH₂CH₃.

Disulfide: -SS-R, wherein R is a disulfide substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group (also referred to herein as C_1-7alkyl disulfide). Examples of C_1-7alkyl disulfide groups include, but are not limited to, -SSCH₃ and -SSCH₂CH₃.

Sulfine (sulfinyl, sulfoxide): -S(=O)R, wherein R is a sulfine substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples of sulfine groups include, but are not limited to, -S(=O)CH₃ and -S(=O)CH₂CH₃. Sulfone (sulfonyl): -S(=O)₂R, wherein R is a sulfone substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-2oaryl group, preferably a C_1-7alkyl group, including, for example, a fluorinated or perfluorinated C_1-7alkyl group. Examples of sulfone groups include, but are not limited to, -S(=O)₂CH₃ (methanesulfonyl, mesyl), -S(=O)₂CF₃ (triflyl), -S(=O)₂CH₂CH₃ (esyl), -S(=O)₂C₄F₉ (nonaflyl), -S(=O)₂CH₂CF₃ (tresyl), -SC=O)₂CH₂CH₂NH₂ (tauryl), -S(=O)₂Ph (phenylsulfonyl, besyl), 4-methylphenylsulfonyl (tosyl), 4-chlorophenylsulfonyl (closyl), 4-bromophenylsulfonyl (brosyl), 4-nitrophenyl (nosyl), 2-naphthalenesulfonate (napsyl), and 5-dimethylamino-naphthalen-1-ylsulfonate (dansyl).

Sulfinic acid (sulfino): -S(=O)OH, -SO₂H.

Sulfonic acid (sulfo): -S(=O)₂OH, -SO₃H.

Sulfinate (sulfinic acid ester): -S(=O)OR; wherein R is a sulfinate substituent, for example, a

group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a d-jalkyl group. Examples of sulfinate groups include, but are not limited to, -S(=O)OCH₃ (methoxysulfinyl; methyl sulfinate) and -S(=O)OCH₂CH₃ (ethoxysulfinyl; ethyl sulfinate).

Sulfonate (sulfonic acid ester): -S(=O)₂OR, wherein R is a sulfonate substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group. Examples of sulfonate groups include, but are not limited to, -S(=O)₂OCH₃ (methoxysulfonyl; methyl sulfonate) and -S(=O)₂OCH₂CH₃ (ethoxysulfonyl; ethyl sulfonate).

Sulfinyloxy: -OS(=O)R, wherein R is a sulfinyloxy substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably a d.₇alkyl group. Examples of sulfinyloxy groups include, but are not limited to, -OS(=O)CH₃ and -OS(=O)CH₂CH₃.

Sulfonyloxy: -OS(=O)₂R, wherein R is a sulfonyloxy substituent, for example, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-2Oaryl group, preferably a C_1-7alkyl group. Examples of sulfonyloxy groups include, but are not limited to, -OS(=O)₂CH₃ (mesylate) and -OS(=O)₂CH₂CH₃ (esylate). Sulfate: -OS(=O)₂OR; wherein R is a sulfate substituent, for example, a C_1-7alkyl group, a C₃-₂oheterocyclyl group, or a C_5-2oaryl group, preferably a C_1-7alkyl group. Examples of sulfate groups include, but are not limited to, -OSC=O)₂OCH₃ and -SO(=O)₂OCH₂CH₃.

Sulfamyl (sulfaimoyl; sulfinic acid amide; sulfinamide): -S(=O)NR¹R², wherein R¹ and R² are independently amino substituents, as defined for amino groups. Examples of sulfamyl groups include, but are not limited to, -S(=O)NH₂, -S(=O)NH(CH₃), -S(=O)N(CH₃)₂, -S(=O)NH(CH₂CH₃), -S(=O)N(CH₂CH₃)₂, and -S(=O)NHPh.

Sulfonamido (sulfinamoyl; sulfonic acid amide; sulfonamide): -S(=O)₂NR¹R², wherein R¹ and R² are independently amino substituents, as defined for amino groups. Examples of sulfonamido groups include, but are not limited to, -S(=O)₂NH₂, -SC=O)₂NH(CH₃), -S(=O)₂N(CH₃)₂, -S(=O)₂NH(CH₂CH₃), -S(=O)₂N(CH₂CH₃)_2l and -S(=O)₂NHPh.

Sulfamino: -NR¹S(=O)₂OH, wherein R¹ is an amino substituent, as defined for amino groups. Examples of sulfamino groups include, but are not limited to, -NHS(=O)₂OH and -N(CH₃)S(=O)₂OH.

Sulfonamino: -NR¹S(=O)₂R, wherein R¹ is an amino substituent, as defined for amino groups, and R is a sulfonamino substituent, for example, a C_1-7alkyl group, a C_3- ₂oheterocyclyl group, or a C₅.₂₀aryl group, preferably a Ci_-7alkyl group. Examples of sulfonamino groups include, but are not limited to, -NHS(=O)₂CH₃ and -N(CH₃)SC=O)₂C₆H₅.

Sulfinamino: -NR¹S(=O)R, wherein R¹ is an amino substituent, as defined for amino groups, and R is a sulfinamino substituent, for example, a C_1-7alkyl group, a C_3- ₂₀heterocyclyl group, or a C_5-2oaryl group, preferably a C_1-7alkyl group. Examples of sulfinamino groups include, but are not limited to, -NHS(=O)CH₃ and -N(CH₃)S(=O)C₆H₅.

Phosphino (phosphine): -PR₂, wherein R is a phosphino substituent, for example, -H, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C₅.₂₀aryl group, preferably -H, a Ci-₇alkyl group, or a C_5-2oaryl group. Examples of phosphino groups include, but are not limited to, -PH₂, -P(CHa)₂, -P(CH₂CH₃)₂, -P(t-Bu)₂, and -P(Ph)₂.

Phospho: -P(=O)₂.

Phosphinyl (phosphine oxide): -P(=O)R₂, wherein R is a phosphinyl substituent, for example, a Ci_-7alkyl group, a C₃-₂₀heterocyclyl group, or a C_5-20aryl group, preferably a C_1-7alkyl group or a C_5-20aryl group. Examples of phosphinyl groups include, but are not limited to, -P(=O)(CH₃)₂, -P(=θχCH₂CH₃)₂, -P(=θχt-Bu)₂, and -P(=O)(Ph)₂.

Phosphonic acid (phosphono): -P(=O)(OH)₂.

Phosphonate (phosphono ester): -P(=O)(OR)₂, where R is a phosphonate substituent, for example, -H, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-2oaryl group, preferably -H, a C_1-7alkyl group, or a C₅-₂₀aryl group. Examples of phosphonate groups include, but are not limited to, -P(=O)(OCH₃)₂, -P(=O)(OCH₂CH₃)₂, -P(=O)(O-t-Bu)₂, and -P(=O)(OPh)₂.

Phosphoric acid (phosphonooxy): -OP(=O)(OH)₂.

Phosphate (phosphonooxy ester): -OP(=O)(OR)₂, where R is a phosphate substituent, for example, -H, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably -H, a C_1-7alkyl group, or a C_5-20aryl group. Examples of phosphate groups include, but are not limited to, -OP(=O)(OCH₃)₂, -OP(=O)(OCH₂CH₃)₂, -OP(=O)(O-t-Bu)₂, and -OP(=O)(OPh)₂.

Phosphorous acid: -OP(OH)₂.

Phosphite: -OP(OR)₂, where R is a phosphite substituent, for example, -H, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably -H, a C_1-7alkyl group, or a C_5-20aryl group. Examples of phosphite groups include, but are not limited to, -OP(OCH₃)₂, -OP(OCH₂CHa)₂, -OP(CM-Bu)₂₁ and -OP(OPh)₂.

Phosphoramidite: -OP(OR¹ )-NR² ₂, where R¹ and R² are phosphoramidite substituents, for example, -H, a (optionally substituted) C_1-7alkyl group, a

C₃.₂₀heterocyclyl group, or a C_5-20aryl group, preferably -H, a C_1-7alkyl group, or a C_5-2oaryl group. Examples of phosphoramidite groups include, but are not limited to, -OP(OCH₂CH₃)-N(CH₃)₂, -OP(OCH₂CH₃)-N(i-Pr)₂, and -OP(OCH₂CH₂CN)-N(I-Pr)₂.

Phosphoramidate: -OP(=O)(OR¹)-NR² ₂, where R¹ and R² are phosphoramidate substituents, for example, -H, a (optionally substituted) C_1-7alkyl group, a

C_3-20heterocyclyl group, or a C₅-₂₀aryl group, preferably -H, a C^alkyl group, or a C_5-2oaryl group. Examples of phosphoramidate groups include, but are not limited to, -OP(=OXOCH₂CH₃)-N(CH₃)₂, -OP(=O)(OCH₂CH₃)-N(i-Pr)₂, and -OP(=O)(OCH₂CH₂CN)-N(i-Pr)₂.

SiIyI: -SiR₃, where R is a silyl substituent, for example, -H, a C_1-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably -H, a C_1-7alkyl group, or a C_5-2oaryl group. Examples of silyl groups include, but are not limited to, -SiH₃, -SiH₂(CH₃), -SiH(CH₃)₂, -Si(CH₃)₃ , -Si(Et)₃, -Si(JPr)₃, -Si(tBu)(CH₃)₂, and -Si(tBu)₃.

Oxysilyl: -Si(OR)₃, where R is an oxysilyl substituent, for example, -H, a Ci_-7alkyl group, a C_3-20heterocyclyl group, or a C_5-20aryl group, preferably -H, a C_1-7alkyl group, or a C₅.₂₀aryl group. Examples of oxysilyl groups include, but are not limited to, - Si(OH)₃, -Si(OMe)₃ , -Si(OEt)₃, and -Si(OtBu)₃.

Siloxy (silyl ether): -OSiR₃, where SiR₃ is a silyl group, as discussed above.

Oxysiloxy: -OSi(OR)₃, wherein OSi(OR)₃ is an oxysilyl group, as discussed above. -

In many cases, substituents are themselves substituted.

Eor example? a d_-7alkyl group may be substituted with, for example: hydroxy (also referred to as a hydroxy-C_1-7alkyl group); halo (also referred to as a halo-C₁.₇alkyl group); amino (also referred to as a amino-C_1-7alkyl group); carboxy (also referred to as a carboxy-Ci.₇alkyl group); C_1-7alkoxy (also referred to as a C_1-7alkoxy-C_1-7alkyl group); C_5-20aryl (also referred to as a C_5-2OaIyI-C₁.₇alkyl group).

Similarly, a C_5-20aryl group may be substituted with, for example: hydroxy (also referred to as a hydroxy-C₅.₂oaryl group); halo (also referred to as a halo-C_5-2oaryl group); amino (also referred to as an amino-C_5-2oaryl group, e.g., as in aniline); carboxy (also referred to as an carboxy-C₅.₂oaryl group, e.g., as in benzoic acid); C_1-7alkyl (also referred to as a C^alkyl-Cs^oaryl group, e.g., as in toluene); C_1-7alkoxy (also referred to as a C_1-7alkoxy-C_5-2oaryl group, e.g., as in anisole); C_5-20aryl (also referred to as a C_5-2Qa ryl-C_5-2oary I, e.g., as in biphenyl).

These and other specific examples of such substituted-substituents are described below.

Hydroxy-Ci_-7alkyl: The term " hydroxy-C_1-7alkyl," as used herein, pertains to a C_1-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with a hydroxy group. Examples of such groups include, but are not limited to, -CH₂OH, -CH₂CH₂OH, and -CH(OH)CH₂OH.

Halo-C_1-7alkyl group: The term " halo-Ci_-7alkyl," as used herein, pertains to a C_1-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with a halogen atom (e.g., F, Cl, Br, I). If more than one hydrogen atom has been replaced with a halogen atom, the halogen atoms may independently be the same or different. Every hydrogen atom may be replaced with a halogen atom, in which case the group may conveniently be referred to as a C_1-7perhaloalkyl group." Examples of such groups include, but are not limited to, -CF₃, -CHF₂, -CH₂F, -CCI₃, -CBr₃, -CH₂CH₂F, -CH₂CHF₂, and -CH₂CF₃.

Amino-C_1-7alkyl: The term " amino-C_1-7alkyl," as used herein, pertains to a Ci_-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with an amino group. Examples of such groups include, but are not limited to, -CH₂NH₂, -CH₂CH₂NH₂, and -CH₂CH₂N(CH₃)₂.

Carboxy-C_1-7alkyl: The term "carboxy-C_1-7alkyl," as used herein, pertains to a

C_1-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with a carboxy group. Examples of such groups include, but are not limited to, -CH₂COOH and -CH₂CH₂COOH.

C_1-7alkoxy-C_1-7alkyl: The term "C^alkoxy-C^alkyl," as used herein, pertains to a C_1-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with a C_1-7alkoxy group. Examples of such groups include, but are not limited to, -CH₂OCH₃, -CH₂CH₂OCH₃, and ,-CH₂CH₂OCH₂CH₃

C_5-2oaryl-C_1-7alkyl: The term "C5_-2oaryl-C_1-7alkyl," as used herein, pertains to a Ci_-7alkyl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been replaced with a C₅.₂₀aryl group. Examples of such groups include, but are not limited to, benzyl (phenylmethyl, PhCH₂-), benzhydryl (Ph₂CH-), trityl (triphenylmethyl, Ph₃C-), phenethyl (phenylethyl, Ph-CH₂CH₂-), styryl (Ph-CH=CH-), cinnamyl (Ph-CH=CH-CH₂-).

Hydroxy-C_5-20aryl: The term " hydroxy-C_5-2oaryl," as used herein, pertains to a C_5-20aryl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been substituted with an hydroxy group. Examples of such groups include, but are not limited to, those derived from: phenol, naphthol, pyrocatechol, resorcinol, hydroquinone, pyrogallol, phloroglucinol.

Halo-C_5-20aryl: The term "halo-C₅.₂₀aryl," as used herein, pertains to a C_5-20aryl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been substituted with a halo (e.g., F, Cl, Br, I) group. Examples of such groups include, but are not limited to, halophenyl (e.g., fluorophenyl, chlorophenyl, bromophenyl, or iodophenyl, whether ortho-, meta-, or para-substituted), dihalophenyl, trihalophenyl, tetrahalophenyl, and pentahalophenyl.

C_1-7alkyl-C_5-2oaryl: The term "d-ralkyl-Cs^oaryl," as used herein, pertains to a C_5-20aryl group in which at least one hydrogen atom (e.g., 1 , 2, 3) has been substituted with a C_1-7alkyl group. Examples of such groups include, but are not limited to, tolyl (from toluene), xylyl (from xylene), mesityl (from mesitylene), and cumenyl (or cumyl, from cumene), and duryl (from durene).

Hydroxy-Ci_₇alkoxy: -OR, wherein R is a hydroxy-C_1-7alkyl group. Examples of hydroxy-C_1-7alkoxy groups include, but are not limited to, -OCH₂OH, -OCH₂CH₂OH₁ and -OCH₂CH₂CH₂OH.

Halo-Cv_T-alkoxy: -OR, wherein R is a halo-C_1-7alkyl group. Examples of halo-C_1-7alkoxy groups include, but are not limited to, -OCF₃, -OCHF₂, -OCH₂F, -OCCI₃, -OCBr₃, -OCH₂CH₂F, -OCH₂CHF₂, and -OCH₂CF₃. Carboxy-C_1-7alkoxy: -OR, wherein R is a carboxy-C^alkyl group. Examples of carboxy-Ci_-7alkoxy groups include, but are not limited to, -OCH₂COOH, -OCH₂CH₂COOH, and -OCH₂CH₂CH₂COOH.

C_1-7alkoxy-C_1-7alkoxy: -OR, wherein R is a Ci_-7alkoxy-C_1-7aikyl group. Examples of C^alkoxy-C^alkoxy groups include, but are not limited to, -OCH₂OCH₃, -OCH₂CH₂OCH₃, and -OCH₂CH₂OCH₂CH₃.

C₅._2Qaryl-C_1-7alkoxy: -OR, wherein R is a C_5-20aryl-C_1-7alkyl group. Examples of such groups include, but are not limited to, benzyloxy, benzhydryloxy, trityloxy, phenethoxy, styryloxy, and cimmamyloxy.

C_1-7alkyl-C_5-2oaryloxy: -OR, wherein R is a Cv₇alkyl-C_5-20aryl group. Examples of such groups include, but are not limited to, tolyloxy, xylyloxy, mesityloxy, cumenyloxy, and duryloxy.

Amino-Ci_-7alkyl-amino: The term "amino-C_1-7alkyl-amino," as used herein, pertains to an amino group, -NR¹R², in which one of the substituents, R¹ or R², is itself a amino- C_1-7alkyl group (-Ci_-7alkyl-NR³R⁴). The amino-Ci_-7alkylamino group may be represented, for example, by the formula -NR¹-Ci.₇alkyl-NR³R⁴. Examples of such groups include, but are not limited to, groups of the formula -NR¹(CH₂)_nNR¹R², where n is 1 to 6 (for example, -NHCH₂NH₂, -NH(CH₂J₂NH₂, -NH(CH₂)₃NH₂, -NH(CH₂)₄NH₂, -NH(CH₂)₅NH₂, -NH(CH₂)₆NH₂), -NHCH₂NH(Me), -NH(CH₂J₂NH(Me), -NH(CH₂)₃NH(Me), -NH(CH₂)₄NH(Me), -NH(CH₂)₅NH(Me), -NH(CH₂)₆NH(Me), -NHCH₂NH(Et), -NH(CHz)₂NH(Et), -NH(CH₂)₃NH(Et), -NH(CH₂)₄NH(Et), -NH(CH₂)₅NH(Et)rand -NH(CH₂)₆NH(Et).

-Certain Preferred Substituents In one preferred embodiment, the substituent(s), often referred to herein as R, are independently selected from: halo; hydroxy; ether (e.g., C_1-7alkoxy); formyl; acyl (e.g.,

C^alkylacyl , C_5-20arylacyl); acylhalide; carboxy; ester; acyloxy; amido; acylamido; thioamido; tetrazolyl; amino; nitro; nitroso; azido; cyano; isocyano; cyanato; isocyanato; thiocyano; isothiocyano; sulfhydryl; thioether (e.g., C_1-7alkylthio); sulfonic acid; sulfonate; sulfone; sulfonyloxy; sulfinyloxy; sulfamino; sulfonamino; sulfinamino; sulfamyl; sulfonamido; Ci_-7alkyl (including, e.g., unsubstituted C_1-7alkyl, C_1-7haloalkyl, C_1-7hydroxyalkyl, Ci_-7carboxyalkyl, C_1-7aminoalkyl, C_5-20aryl-C_1-7alkyl); C₃.₂₀heterocyclyl; or C₅-₂₀aryl (including, e.g., C_5-2ocarboaryl, C₅.₂₀heteroaryl, C_1-7alkyl- C_5-2oaryl and C_5-2ohaloaryl)).

In one preferred embodiment, the substituent(s), often referred to herein as R, are independently selected from:

-F, -Cl₁ -Br, and -I;

-OH;

-OMe, -OEt, -O(tBu), and -OCH₂Ph; -SH;

-SMe, -SEt, -S(tBu), and -SCH₂Ph;

-C(=O)H;

-C(=0)Me, -C(=O)Et, -C(=O)(tBu), and -C(=O)Ph;

-C(=O)OH; -C(=O)OMe, -C(=O)OEt, and -C(=O)O(tBu);

-C(=O)NH₂, -C(=O)NHMe, -C(=O)NMe₂, and -C(=O)NHEt;

-NHC(=O)Me, -NHC(=O)Et, -NHC(=O)Ph, succinimidyl, and maleimidyl;

-NH₂, -NHMe, -NHEt, -NH(iPr), -NH(nPr), -NMe₂, -NEt₂, -N(JPr)₂, -N(nPr)₂, -N(nBu)₂, and -N(tBu)₂; -CN;

-NO₂;

-Me, -Et, -nPr, -iPr, -nBu, -tBu;

-CF₃, -CHF₂, -CH₂F, -CCI₃, -CBr₃, -CH₂CH₂F, -CH₂CHF₂, and -CH₂CF₃;

-OCF₃, -OCHF₂, -OCH₂F, -OCCI₃, -OCBr₃, -OCH₂CH₂F, -OCH₂CHF₂, and -OCH₂CF₃; -CH₂OH, -CH₂CH₂OH, and -CH(OH)CH₂OH;

-CH₂NH₂₁-CH₂CH₂NH₂, and -CH₂CH₂NMe₂; and, optionally substituted phenyl.

In one preferred embodiment, the substituent(s), often referred to herein as R, are independently selected from: -F, -Cl, -Br, -I, -OH, -OMe, -OEt, -SH, -SMe, -SEt, -C(=0)Me, -C(=O)OH, -C(=0)0Me, -CONH₂, -CONHMe, -NH₂, -NMe₂, -NEt₂, -N(nPr)₂, -N(JPr)₂, -CN, -NO₂, -Me, -Et, -CF₃, -OCF₃, -CH₂OH, -CH₂CH₂OH, -CH₂NH₂, -CH₂CH₂NH₂, and -Ph.

In one preferred embodiment, the substituent(s), often referred to herein as R, are independently selected from: hydroxy; ether (e.g., C_1-7alkoxy); ester, amido; amino; and, Ci_-7alkyl (including, e.g., unsubstituted C_1-7alkyl, Ci_-7haloalkyl, C_1-7hydroxyalkyl, Ci_-7carboxyalkyl, Ci-₇aminoalkyl, C₅-₂oaryl-Ci_-7alkyl).

-OH;

-OMe, -OEt, -O(tBu), and -OCH₂Ph;

-C(=O)OMe, -C(=O)OEt, and -C(=O)O(tBu);

-C(=O)NH₂, -C(=0)NHMe, -C(=O)NMe₂, and -C(=O)NHEt;

-NH₂, -NHMe, -NHEt, -NH(iPr), -NH(nPr), -NMe₂, -NEt₂, -N(iPr)₂, -N(nPr)₂, -N(nBu)₂, and -N(tBu)₂;

-Me, -Et, -nPr, -iPr, -nBu, -tBu;

-CH₂OH, -CH₂CH₂OH, and -CH(OH)CH₂OH; and,

-CH₂NH₂₁-CH₂CH₂NH₂, and -CH₂CH₂NMe₂.

Combinations

Each and every compatible combination of the embodiments described above is explicitly disclosed herein, as if each and every combination was individually and explicitly recited.

Examples of Specific Embodiments

In one embodiment, the compounds are selected from compounds of the following formulae and pharmaceutically acceptable salts, hydrates, and solvates thereof:

In one embodiment the compounds are selected from: PRD05, PRD06, PRD11, PRD17, PRD18, PRD20, PRD23, PRD25, PRD26, PRD27, PRD28, PRD29, PRD30 and PRD31.

In one embodiment the compounds are selected from: PRD05, PRD06, PRD17, PRD18, PRD23, PRD29, PRD30 and PRD31.

In one embodiment the compounds are selected from: PRD06, PRD18 and PRD29.

In one embodiment the compounds are selected from: DRD02 and DRD04. Substantially Purified Forms

One aspect of the present invention pertains to PRD or DRD compounds, as described herein, in substantially purified form and/or in a form substantially free from contaminants.

In one embodiment, the substantially purified form is at least 50% by weight, e.g., at least 60% by weight, e.g., at least 70% by weight, e.g., at least 80% by weight, e.g., at least 90% by weight, e.g., at least 95% by weight, e.g., at least 97% by weight, e.g., at least 98% by weight, e.g., at least 99% by weight.

Unless specified, the substantially purified form refers to the compound in any stereoisomeric or enantiomeric form. For example, in one embodiment, the substantially purified form refers to a mixture of stereoisomers, i.e., purified with respect to other compounds. In one embodiment, the substantially purified form refers to one stereoisomer, e.g., optically pure stereoisomer. In one embodiment, the substantially purified form refers to a mixture of enantiomers. In one embodiment, the substantially purified form refers to a equimoiar mixture of enantiomers (i.e., a racemic mixture, a racemate). In one embodiment, the substantially purified form refers to one enantiomer, e.g., optically pure enantiomer.

In one embodiment, the contaminants represent no more than 50% by weight, e.g., no more than 40% by weight, e.g., no more than 30% by weight, e.g., no more than 20% by weight, e.g., no more than 10% by weight, e.g., no more than 5% by weight, e.g., no more than 3% by weight, e.g., no more than 2% by weight, e.g., no more than 1 % by weight.

Unless specified, the contaminants refer to other compounds, that is, other than stereoisomers or enantiomers. In one embodiment, the contaminants refer to other compounds and other stereoisomers. In one embodiment, the contaminants refer to other compounds and the other enantiomer.

In one embodiment, the substantially purified form is at least 60% optically pure (i.e., 60% of the compound, on a molar basis, is the desired stereoisomer or enantiomer, and 40% is the undesired stereoisomer or enantiomer), e.g., at least 70% optically pure, e.g., at least 80% optically pure, e.g., at least 90% optically pure, e.g., at least 95% optically pure, e.g., at least 97% optically pure, e.g., at least 98% optically pure, e.g., at least 99% optically pure.

Isomers Certain compounds may exist in one or more particular geometric, optical, enantiomeric, diasteriomeric, epimeric, atropic, stereoisomeric, tautomeric, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r- forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (-) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; a- and /?-forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair-forms; and combinations thereof, hereinafter collectively referred to as "isomers" (or "isomeric forms").

Note that, except as discussed below for tautomeric forms, specifically excluded from the term "isomers," as used herein, are structural (or constitutional) isomers

(i.e., isomers which differ in the connections between atoms rather than merely by the position of atoms in space). For example, a reference to a methoxy group, -OCH₃, is not to be construed as a reference to its structural isomer, a hydroxymethyl group, -CH₂OH. Similarly, a reference to ortho-chlorophenyl is not to be construed as a reference to its structural isomer, meta-chlorophenyl. However, a reference to a class of structures may well include structurally isomeric forms falling within that class (e.g., includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl; methoxyphenyl includes ortho-, meta-, and para-methoxyphenyl).

The above exclusion does not pertain to tautomeric forms, for example, keto-, enol-, and enolate-forms, as in, for example, the following tautomeric pairs: keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime, thioketone/enethiol, N-nitroso/hydroxyazo, and nitro/aci-nitro.

keto enol enolate

Note that specifically included in the term "isomer" are compounds with one or more isotopic substitutions. For example, H may be in any isotopic form, including ¹H, ²H (D), and ³H (T); C may be in any isotopic form, including ¹²C, ¹³C, and ¹⁴C; O may be in any isotopic form, including ¹⁶O and ¹⁸O; and the like.

Unless otherwise specified, a reference to a particular compound includes all such isomeric forms, including mixtures (e.g., racemic mixtures) thereof. Methods for the preparation (e.g., asymmetric synthesis) and separation (e.g., fractional crystallisation and chromatographic means) of such isomeric forms are either known in the art or are readily obtained by adapting the methods taught herein, or known methods, in a known manner.

Salts

It may be convenient or desirable to prepare, purify, and/or handle a corresponding salt of the compound, for example, a pharmaceutically-acceptable salt. Examples of pharmaceutically acceptable salts are discussed in Berge eta/., 1977, "Pharmaceutically Acceptable Salts," J. Pharm. ScL, Vol. 66, pp. 1-19.

For example, if the compound is anionic, or has a functional group which may be anionic (e.g., -COOH may be -COO ), then a salt may be formed with a suitable cation. Examples of suitable inorganic cations include, but are not limited to, alkali metal ions such as Na⁺ and K⁺, alkaline earth cations such as Ca²⁺ and Mg²⁺, and other cations such as Al⁺³. Examples of suitable organic cations include, but are not limited to, ammonium ion (i.e., NH₄ ⁺) and substituted ammonium ions (e.g., NH₃R⁺, NH₂R₂ ⁺, NHR₃ ⁺, NR₄ ⁺). Examples of some suitable substituted ammonium ions are those derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine. An example of a common quaternary ammonium ion is N(CH₃)₄ ⁺.

If the compound is cationic, or has a functional group which may be cationic (e.g., -NH₂ may be -NH₃ ⁺), then a salt may be formed with a suitable anion. Examples of suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric, nitrous, phosphoric, and phosphorous. Examples of suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetyoxybenzoic, acetic, ascorbic, aspartic, benzoic, camphorsulfonic, cinnamic, citric, edetic, ethanedisulfonic, ethanesulfonic, fumaric, glucheptonic, gluconic, glutamic, glycolic, hydroxymaleic, hydroxynaphthalene carboxylic, isethionic, lactic, lactobionic, lauric, maleic, malic, methanesulfonic, mucic, oleic, oxalic, palmitic, pamoic, pantothenic, phenylacetic, phenylsulfonic, propionic, pyruvic, salicylic, stearic, succinic, sulfanilic, tartaric, toluenesulfonic, and valeric. Examples of suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethyl cellulose.

Unless otherwise specified, a reference to a particular compound also includes salt forms thereof.

Solvates and Hydrates

It may be convenient or desirable to prepare, purify, and/or handle a corresponding solvate of the compound. The term "solvate" is used herein in the conventional sense to refer to a complex of solute (e.g., compound, salt of compound) and solvent. If the solvent is water, the solvate may be conveniently referred to as a hydrate, for example, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.

Unless otherwise specified, a reference to a particular compound also includes solvate and hydrate forms thereof.

Chemically Protected Forms

It may be convenient or desirable to prepare, purify, and/or handle the compound in a chemically protected form. The term "chemically protected form" is used herein in the conventional chemical sense and pertains to a compound in which one or more reactive functional groups are protected from undesirable chemical reactions under specified conditions (e.g., pH, temperature, radiation, solvent, and the like). In practice, well known chemical methods are employed to reversibly render unreactive a functional group, which otherwise would be reactive, under specified conditions. In a chemically protected form, one or more reactive functional groups are in the form of a protected or protecting group (also known as a masked or masking group or a blocked or blocking group). By protecting a reactive functional group, reactions involving other unprotected reactive functional groups can be performed, without affecting the protected group; the protecting group may be removed, usually in a subsequent step, without substantially affecting the remainder of the molecule. See, for example, Protective Groups in Organic Synthesis (T. Green and P. Wuts; 4th Edition; John Wiley and Sons, 2006).

A wide variety of such "protecting," "blocking," or "masking" methods are widely used and well known in organic synthesis. For example, a compound which has two nonequivalent reactive functional groups, both of which would be reactive under specified conditions, may be derivatized to render one of the functional groups "protected," and therefore unreactive, under the specified conditions; so protected, the compound may be used as a reactant which has effectively only one reactive functional group. After the desired reaction (involving the other functional group) is complete, the protected group may be "deprotected" to return it to its original functionality.

For example, a hydroxy group may be protected as an ether (-OR) or an ester (-OC(=O)R), for example, as: a t-butyl ether; a benzyl, benzhydryl (diphenylmethyl), or trityl (triphenylmethyl) ether; a trimethylsilyl or t-butyldimethylsilyl ether; or an acetyl ester (-OC(=O)CH₃, -OAc).

For example, an aldehyde or ketone group may be protected as an acetal (R-CH(OR)₂) or ketal (RaC(OR)₂), respectively, in which the carbonyl group (>C=O) is converted to a diether (>C(OR)₂), by reaction with, for example, a primary alcohol. The aldehyde or ketone group is readily regenerated by hydrolysis using a large excess of water in the presence of acid.

For example, an amine group may be protected, for example, as an amide (-NRCO- R) or a urethane (-NRCO-OR), for example, as: a methyl amide (-NHCO-CH₃); a benzyloxy amide (-NHCO-OCH₂C₆H₅, -NH-Cbz); as a t-butoxy amide (-NHCO-OC(CHa)₃, -NH-Boc); a 2-biphenyl-2-propoxy amide

(-NHCO-OC(CH₃)₂C₆H₄C₆H₅, -NH-Bpoc), as a 9-fluorenylmethoxy amide (-NH-Fmoc), as a 6-nitroveratryloxy amide (-NH-Nvoc), as a 2-trimethylsilylethyloxy amide (-NH-Teoc), as a 2,2,2-trichloroethyloxy amide (-NH-Troc), as an allyloxy amide (-NH-Alloc), as a 2(-phenylsulfonyl)ethyloxy amide (-NH-Psec); or, in suitable cases (e.g., cyclic amines), as a nitroxide radical (>N-O*). For example, a carboxylic acid group may be protected as an ester for example, as: an C_1-7alkyl ester (e.g., a methyl ester; a t-butyl ester); a Ci.₇haloalkyl ester (e.g., a C_1-7trihaloalkyl ester); a triC_1-7alkylsilyl-C₁.₇alkyl ester; or a C_5-2oaryl-Ci-₇alkyl ester (e.g., a benzyl ester; a nitrobenzyl ester); or as an amide, for example, as a methyl amide.

For example, a thiol group may be protected as a thioether (-SR), for example, as: a benzyl thioether; an acetamidomethyl ether (-S-CH₂NHC(=O)CH₃).

Prodrugs

It may be convenient or desirable to prepare, purify, and/or handle the compound in the form of a prodrug. The term "prodrug," as used herein, pertains to a compound which, when metabolised (e.g., in vivo), yields the desired active compound. Typically, the prodrug is inactive, or less active than the desired active compound, but may provide advantageous handling, administration, or metabolic properties.

For example, some prodrugs are esters of the active compound (e.g., a physiologically acceptable metabolically labile ester). During metabolism, the ester group (-C(=O)OR) is cleaved to yield the active drug. Such esters may be formed by esterification, for example, of any of the carboxylic acid groups (-C(=O)OH) in the parent compound, with, where appropriate, prior protection of any other reactive groups present in the parent compound, followed by deprotection if required.

Also, some prodrugs are activated enzymatically to yield the active compound, or a compound which, upon further chemical reaction, yields the active compound (for example, as in ADEPT, GDEPT, LIDEPT, etc.). For example, the prodrug may be a sugar derivative or other glycoside conjugate, or may be an amino acid ester derivative.

Compositions

One aspect of the present invention pertains to a composition (e.g., a pharmaceutical composition) comprising a PRD or DRD compound, as described herein, and a pharmaceutically acceptable carrier, diluent, or excipient.

Another aspect of the present invention pertains to a method of preparing a composition (e.g., a pharmaceutical composition) comprising admixing a PRD or DRD compound, as described herein, and a pharmaceutically acceptable carrier, diluent, or excipient.

Uses The compounds described herein are useful, for example, in the treatment of diseases and conditions that are ameliorated by the inhibition of Bcl-2 protein function, such as, for example, proliferative conditions, cancer, etc.

Use in Methods of Inhibiting Bcl-2 protein function One aspect of the present invention pertains to a method of inhibiting Bcl-2 protein function (especially one or both of Bcl-X_L and Mcl-1 ), in vitro or in vivo, comprising contacting Bcl-2 protein (especially one or both of Bcl-X_L and Mcl-1 ) with an effective amount of a PRD or DRD compound, as described herein.

One aspect of the present invention pertains to a method of inhibiting Bcl-2 protein function (especially one or both of Bcl-X_L and Mcl-1 ) in a cell, in vitro or in vivo, comprising contacting the cell with an effective amount of a PRD or DRD compound, as described herein.

In one embodiment, the method further comprises contacting the cell with one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; (d) a microtubule targeted agent; and (e) ionising radiation.

Suitable assays for determining Bcl-2 protein are described herein and/or are known in the art.

Use in Methods of Inhibiting Cell Proliferation, Etc.

The PRD and DRD compounds described herein, e.g., (a) regulate (e.g., inhibit) cell proliferation; (b) promote apoptosis; or (c) a combination of both.

One aspect of the present invention pertains to a method of regulating (e.g., inhibiting) cell proliferation (e.g., proliferation of a cell), promoting apoptosis, or a combination of both, in vitro or in vivo, comprising contacting a cell with an effective amount of a PRD or DRD compound, as described herein. In one embodiment, the method is a method of regulating (e.g., inhibiting) cell proliferation (e.g., proliferation of a cell), in vitro or in vivo, comprising contacting a cell with an effective amount of a PRD or DRD compound, as described herein.

In one embodiment, the method is performed in vitro. In one embodiment, the method is performed in vivo.

In one embodiment, the PRD or DRD compound is provided in the form of a pharmaceutically acceptable composition.

Any type of cell may be treated, including but not limited to, lung, gastrointestinal (including, e.g., bowel, colon), breast (mammary), ovarian, prostate, liver (hepatic), kidney (renal), bladder, pancreas, brain, and skin.

One of ordinary skill in the art is readily able to determine whether or not a candidate compound regulates (e.g., inhibits) cell proliferation, etc. For example, assays which may conveniently be used to assess the activity offered by a particular compound are described herein.

For example, a sample of cells (e.g., from a tumour) may be grown in vitro and a compound brought into contact with said cells, and the effect of the compound on those cells observed. As an example of "effect," the morphological status of the cells (e.g., alive or dead, etc.) may be determined. Where the compound is found to exert an influence on the ceils, this may be used as a prognostic or diagnostic marker of the efficacy of the compound in methods of treating a patient carrying cells of the same cellular type.

Use in Methods of Therapy

Another aspect of the present invention pertains to a PRD or DRD compound, as described herein, for use in a method of treatment of the human or animal body by therapy. In one embodiment, the method of treatment comprises treatment with both (i) a PRD or DRD compound, as described herein, and (N) one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; (d) a microtubule targeted agent; and (e) ionising radiation.

Another aspect of the present invention pertains to (a) a DNA topoisomerase I or Il inhibitor, (b) a DNA damaging agent, (c) an antimetabolite or TS inhibitor, or (d) a microtubule targeted agent, as described herein, for use in a method of treatment of the human or animal body by therapy, wherein the method of treatment comprises treatment with both (i) a PRD or DRD compound, as described herein, and (a) the DNA topoisomerase I or Il inhibitor, (b) the DNA damaging agent, (c) the antimetabolite or TS inhibitor, or (d) the microtubule targeted agent.

Use in the Manufacture of Medicaments

In one embodiment, the medicament comprises the PRD or DRD compound.

In one embodiment, the treatment comprises treatment with both (i) a medicament comprising a PRD or DRD compound, as described herein, and (ii) one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; (d) a microtubule targeted agent; and (e) ionising radiation.

Another aspect of the present invention pertains to use of (a) a DNA topoisomerase I or Il inhibitor, (b) a DNA damaging agent, (c) an antimetabolite or TS inhibitor, or (d) a microtubule targeted agent, as described herein, in the manufacture of a medicament for use in a treatment, wherein the treatment comprises treatment with both (i) a PRD or DRD compound, as described herein, and (a) the DNA topoisomerase I or Il inhibitor, (b) the DNA damaging agent, (c) the antimetabolite or TS inhibitor, or (d) the microtubule targeted agent. Methods of Treatment

Another aspect of the present invention pertains to a method of treatment comprising administering to a patient in need of treatment a therapeutically effective amount of a PRD or DRD compound, as described herein, preferably in the form of a pharmaceutical composition.

In one embodiment, the method further comprises administering to the subject one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; (d) a microtubule targeted agent; and (e) ionising radiation.

Conditions Treated - Conditions Mediated by Bcl-2

In one embodiment (e.g., of use in methods of therapy, of use in the manufacture of medicaments, of methods of treatment), the treatment is treatment of a disease or condition that is mediated by Bcl-2 (especially one or both of Bcl-X_L and Mcl-1 ).

Conditions Treated - Conditions Ameliorated by the Inhibition of Bcl-2 Function In one embodiment (e.g., of use in methods of therapy, of use in the manufacture of medicaments, of methods of treatment), the treatment is treatment of: a disease or condition that is ameliorated by the inhibition of Bcl-2 function (especially one or both of BCI-X_L and Mcl-1 ).

Conditions Treated - Proliferative Conditions and Cancer

In one embodiment (e.g., of use in methods of therapy, of use in the manufacture of medicaments, of methods of treatment), the treatment is treatment of: a proliferative condition.

The term "proliferative condition," as used herein, pertains to an unwanted or uncontrolled cellular proliferation of excessive or abnormal cells which is undesired, such as, neoplastic or hyperplastic growth.

In one embodiment, the treatment is treatment of: a proliferative condition characterised by benign, pre-malignant, or malignant cellular proliferation, including but not limited to, neoplasms, hyperplasias, and tumours (e.g., histocytoma, glioma, astrocyoma, osteoma), cancers (see below), psoriasis, bone diseases, fibroproliferative disorders (e.g., of connective tissues), pulmonary fibrosis, atherosclerosis, smooth muscle cell proliferation in the blood vessels, such as stenosis or restenosis following angioplasty.

In one embodiment, the treatment is treatment of: cancer.

In one embodiment, the treatment is treatment of: lung cancer, small cell lung cancer, non-small cell lung cancer, gastrointestinal cancer, stomach cancer, bowel cancer, colon cancer, rectal cancer, colorectal cancer, thyroid cancer, breast cancer, ovarian cancer, endometrial cancer, prostate cancer, testicular cancer, liver cancer, kidney cancer, renal cell carcinoma, bladder cancer, pancreatic cancer, brain cancer, glioma, sarcoma, osteosarcoma, bone cancer, nasopharyngeal cancer (e.g., head cancer, neck cancer), skin cancer, squamous cancer, Kaposi's sarcoma, melanoma, malignant melanoma, lymphoma, or leukemia.

In one embodiment, the treatment is treatment of: a carcinoma, for example a carcinoma of the bladder, breast, colon (e.g., colorectal carcinomas such as colon adenocarcinoma and colon adenoma), kidney, epidermal, liver, lung (e.g., adenocarcinoma, small cell lung cancer and non-small cell lung carcinomas), oesophagus, gall bladder, ovary, pancreas (e.g., exocrine pancreatic carcinoma), stomach, cervix, thyroid, prostate, skin (e.g., squamous cell carcinoma); a hematopoietic tumour of lymphoid lineage, for example leukemia, acute lymphocytic leukemia, B-cell lymphoma, T-cell lymphoma, Hodgkin's lymphoma, non- Hodgkin's lymphoma, hairy cell lymphoma, or Burkett's lymphoma; a hematopoietic tumor of myeloid lineage, for example acute and chronic myelogenous leukemias, myelodysplastic syndrome, or promyelocytic leukemia; a tumour of mesenchymal origin, for example fibrosarcoma or habdomyosarcoma; a tumor of the central or peripheral nervous system, for example astrocytoma, neuroblastoma, glioma or schwannoma; melanoma; seminoma; teratocarcinoma; osteosarcoma; xenoderoma pigmentoum; keratoctanthoma; thyroid follicular cancer; or Kaposi's sarcoma.

In one embodiment, the treatment is treatment of solid tumour cancer. In one embodiment, the treatment is treatment of: lung cancer, breast cancer, ovarian cancer, CNS cancer or leukemia.

The anti-cancer effect may arise through one or more mechanisms, including but not limited to, the promotion of apoptosis (programmed cell death), the regulation of cell proliferation, the inhibition of cell cycle progression, the inhibition of angiogenesis (the formation of new blood vessels), the inhibition of metastasis (the spread of a tumour from its origin), or the inhibition of invasion (the spread of tumour cells into neighbouring normal structures). The compounds of the present invention may be used in the treatment of the cancers described herein, independent of the mechanisms discussed herein.

Treatment

The term "treatment," as used herein in the context of treating a condition, pertains generally to treatment and therapy, whether of a human or an animal (e.g., in veterinary applications), in which some desired therapeutic effect is achieved, for example, the inhibition of the progress of the condition, and includes a reduction in the rate of progress, a halt in the rate of progress, alleviatiation of symptoms of the condition, amelioration of the condition, and cure of the condition. Treatment as a prophylactic measure (i.e., prophylaxis) is also included. For example, use with patients who have not yet developed the condition, but who are at risk of developing the condition, is encompassed by the term "treatment."

For example, treatment includes the prophylaxis of cancer, reducing the incidence of cancer, alleviating the symptoms of cancer, etc.

The term "therapeutically-effective amount," as used herein, pertains to that amount of a compound, or a material, composition or dosage form comprising a compound, which is effective for producing some desired therapeutic effect, commensurate with a reasonable benefit/risk ratio, when administered in accordance with a desired treatment regimen.

Combination Therapies

The term "treatment" includes combination treatments and therapies, in which two or more treatments or therapies are combined, for example, sequentially or simultaneously. For example, the compounds described herein may also be used in combination therapies, e.g., in conjunction with other agents, for example, cytotoxic agents, anticancer agents, etc. Examples of treatments and therapies include, but are not limited to, chemotherapy (the administration of active agents, including, e.g., drugs, antibodies (e.g., as in immunotherapy), prodrugs (e.g., as in photodynamic therapy, GDEPT, ADEPT, etc.); surgery; radiation therapy; photodynamic therapy; gene therapy; and controlled diets.

For example, it may be beneficial to combine treatment with a compound as described herein with one or more other (e.g., 1, 2, 3, 4) agents or therapies that regulates cell growth or survival or differentiation via a different mechanism, thus treating several characteristic features of cancer development.

One aspect of the present invention pertains to a compound as described herein, in combination with one or more additional therapeutic agents, as described below.

The particular combination would be at the discretion of the physician who would select dosages using his common general knowledge and dosing regimens known to a skilled practitioner.

The agents (i.e., the compound described herein, plus one or more other agents) may be administered simultaneously or sequentially, and may be administered in individually varying dose schedules and via different routes. For example, when administered sequentially, the agents can be administered at closely spaced intervals (e.g., over a period of 5-10 minutes) or at longer intervals (e.g., 1 , 2, 3, 4 or more hours apart, or even longer periods apart where required), the precise dosage regimen being commensurate with the properties of the therapeutic agent(s).

The agents (i.e., the compound described here, plus one or more other agents) may be formulated together in a single dosage form, or alternatively, the individual agents may be formulated separately and presented together in the form of a kit, optionally with instructions for their use.

Combination Therapies Employing DNA Damaging Agents As discussed herein, in some embodiments, the PRD or DRD compound is employed in combination with (e.g., in conjunction with) one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; (d) a microtubule targeted agent; and (e) ionising radiation.

When both a PRD or DRD compound and one or more other agents are employed, they may be used (e.g., contacted, administered, etc.) in any order. Furthermore, they may be used (e.g., contacted, administered, etc.) together, as part of a single formulation, or separately, as separate formulations.

For example, in regard to methods of treatment employing both a PRD or DRD compound and one or more other agents, treatment with (e.g., administration of) the PRD or DRD compound may be prior to, concurrent with, or may follow, treatment with (e.g., administration of) the one or more other agents, or a combination thereof.

In one embodiment, treatment with (e.g., administration of) a PRD or DRD compound is concurrent with, or follows, treatment with (e.g., administration of) the one or more other agents.

In one embodiment, the one or more other agents is a DNA topoisomerase I or Il inhibitor; for example, Etoposide, Toptecan, Camptothecin, Irinotecan, SN-38, Doxorubicin, Daunorubicin.

In one embodiment, the one or more other agents is a DNA damaging agent; for example, alkylating agents, platinating agents, or compounds that generate free radicals; for example, Temozolomide, Cisplatin, Carboplatin, Mitomycin C, Cyclophosphamide, BCNU, CCNU, Bleomycin.

In one embodiment, the one or more other agents is an antimetabolite or TS inhibitor; for example, 5-fluorouracil, hydroxyurea, Gemcitabine, Arabinosylcytosine, Fludarabine, Tomudex, ZD9331.

In one embodiment, the one or more other agents is a microtubule targeted agent; for example, Paclitaxel, Docetaxel, Vincristine, Vinblastine.

In one embodiment, the one or more other agents is ionising radiation (e.g., as part of radiotherapy). Other Uses

The PRD and DRD compounds described herein may also be used as cell culture additives to inhibit Bcl-2 function (especially one or both of Bcl-X_L and McM ), e.g., to inhibit cell proliferation, etc.

The PRD and DRD compounds described herein may also be used as part of an in vitro assay, for example, in order to determine whether a candidate host is likely to benefit from treatment with the compound in question.

The PRD and DRD compounds described herein may also be used as a standard, for example, in an assay, in order to identify other compounds, other Bcl-2 function inhibitors (especially one or both of Bcl-X_L and McM), other antiproliferative agents, other anti-cancer agents, etc.

Kits

One aspect of the invention pertains to a kit comprising (a) a PRD or DRD compound as described herein, or a composition comprising a PRD or DRD compound as described herein, e.g., preferably provided in a suitable container and/or with suitable packaging; and (b) instructions for use, e.g., written instructions on how to administer the compound or composition.

In one embodiment, the kit further comprises one or more other agents selected from: (a) a DNA topoisomerase I or Il inhibitor; (b) a DNA damaging agent; (c) an antimetabolite or TS inhibitor; and (d) a microtubule targeted agent.

The written instructions may also include a list of indications for which the active ingredient is a suitable treatment.

Routes of Administration The PRD or DRD compound or pharmaceutical composition comprising the PRD or DRD compound may be administered to a subject by any convenient route of administration, whether systemically/peripherally or topically (i.e., at the site of desired action).

Routes of administration include, but are not limited to, oral (e.g., by ingestion); buccal; sublingual; transdermal (including, e.g., by a patch, plaster, etc.); transmucosal (including, e.g., by a patch, plaster, etc.); intranasal (e.g., by nasal spray); ocular (e.g., by eyedrops); pulmonary (e.g., by inhalation or insufflation therapy using, e.g., via an aerosol, e.g., through the mouth or nose); rectal (e.g., by suppository or enema); vaginal (e.g., by pessary); parenteral, for example, by injection, including subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcuticular, intraarticular, subarachnoid, and intrasternal; by implant of a depot or reservoir, for example, subcutaneously or intramuscularly.

The Subject/Patient

The subject/patient may be a chordate, a vertebrate, a mammal, a placental mammal, a marsupial (e.g., kangaroo, wombat), a rodent (e.g., a guinea pig, a hamster, a rat, a mouse), murine (e.g., a mouse), a lagomorph (e.g., a rabbit), avian (e.g., a bird), canine (e.g., a dog), feline (e.g., a cat), equine (e.g., a horse), porcine (e.g., a pig), ovine (e.g., a sheep), bovine (e.g., a cow), a primate, simian (e.g., a monkey or ape), a monkey (e.g., marmoset, baboon), an ape (e.g., gorilla, chimpanzee, orangutang, gibbon), or a human.

Furthermore, the subject/patient may be any of its forms of development, for example, a foetus.

In one preferred embodiment, the subject/patient is a human.

Formulations

While it is possible for the PRD or DRD compound to be administered alone, it is preferable to present it as a pharmaceutical formulation (e.g., composition, preparation, medicament) comprising at least one PRD or DRD compound, as described herein, together with one or more other pharmaceutically acceptable ingredients well known to those skilled in the art, including, but not limited to, pharmaceutically acceptable carriers, diluents, excipients, adjuvants, fillers, buffers, preservatives, anti-oxidants, lubricants, stabilisers, solubilisers, surfactants (e.g., wetting agents), masking agents, colouring agents, flavouring agents, and sweetening agents. The formulation may further comprise other active agents, for example, other therapeutic or prophylactic agents. Thus, the present invention further provides pharmaceutical compositions, as defined above, and methods of making a pharmaceutical composition comprising admixing at least one PRD or DRD compound, as described herein, together with one or more other pharmaceutically acceptable ingredients well known to those skilled in the art, e.g., carriers, diluents, excipients, etc. If formulated as discrete units (e.g., tablets, etc.), each unit contains a predetermined amount (dosage) of the compound.

The term "pharmaceutically acceptable," as used herein, pertains to compounds, ingredients, materials, compositions, dosage forms, etc., which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of the subject in question (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. Each carrier, diluent, excipient, etc. must also be "acceptable" in the sense of being compatible with the other ingredients of the formulation.

Suitable carriers, diluents, excipients, etc. can be found in standard pharmaceutical texts, for example, Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, Pa., 1990; and Handbook of Pharmaceutical Excipients. 5th edition, 2005.

The formulations may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the compound with a carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the compound with carriers (e.g., liquid carriers, finely divided solid carrier, etc.), and then shaping the product, if necessary.

The formulation may be prepared to provide for rapid or slow release; immediate, delayed, timed, or sustained release; or a combination thereof.

Formulations may suitably be in the form of liquids, solutions (e.g., aqueous, nonaqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), elixirs, syrups, electuaries, mouthwashes, drops, tablets (including, e.g., coated tablets), granules, powders, losenges, pastilles, capsules (including, e.g., hard and soft gelatin capsules), cachets, pills, ampoules, boluses, suppositories, pessaries, tinctures, gels, pastes, ointments, creams, lotions, oils, foams, sprays, mists, or aerosols.

Formulations may suitably be provided as a patch, adhesive plaster, bandage, dressing, or the like which is impregnated with one or more compounds and optionally one or more other pharmaceutically acceptable ingredients, including, for example, penetration, permeation, and absorption enhancers. Formulations may also suitably be provided in the form of a depot or reservoir.

The compound may be dissolved in, suspended in, or admixed with one or more other pharmaceutically acceptable ingredients. The compound may be presented in a liposome or other microparticulate which is designed to target the compound, for example, to blood components or one or more organs.

Formulations suitable for oral administration (e.g., by ingestion) include liquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), elixirs, syrups, electuaries, tablets, granules, powders, capsules, cachets, pills, ampoules, boluses.

Formulations suitable for buccal administration include mouthwashes, losenges, pastilles, as well as patches, adhesive plasters, depots, and reservoirs. Losenges typically comprise the compound in a flavored basis, usually sucrose and acacia or tragacanth. Pastilles typically comprise the compound in an inert matrix, such as gelatin and glycerin, or sucrose and acacia. Mouthwashes typically comprise the compound in a suitable liquid carrier.

Formulations suitable for sublingual administration include tablets, losenges, pastilles, capsules, and pills.

Formulations suitable for oral transmucosal administration include liquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), mouthwashes, losenges, pastilles, as well as patches, adhesive plasters, depots, and reservoirs.

Formulations suitable for non-oral transmucosal administration include liquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), suppositories, pessaries, gels, pastes, ointments, creams, lotions, oils, as well as patches, adhesive plasters, depots, and reservoirs.

Formulations suitable for transdermal administration include gels, pastes, ointments, creams, lotions, and oils, as well as patches, adhesive plasters, bandages, dressings, depots, and reservoirs.

Tablets may be made by conventional means, e.g., compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the compound in a free-flowing form such as a powder or granules, optionally mixed with one or more binders (e.g., povidone, gelatin, acacia, sorbitol, tragacanth, hydroxypropylmethyl cellulose); fillers or diluents (e.g., lactose, microcrystalline cellulose, calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc, silica); disintegrants (e.g., sodium starch glycolate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose); surface-active or dispersing or wetting agents (e.g., sodium lauryl sulfate); preservatives (e.g., methyl p-hydroxybenzoate, propyl p-hydroxybenzoate, sorbic acid); flavours, flavour enhancing agents, and sweeteners. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the compound therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with a coating, for example, to affect release, for example an enteric coating, to provide release in parts of the gut other than the stomach.

Ointments are typically prepared from the compound and a paraffinic or a water- miscible ointment base.

Creams are typically prepared from the compound and an oil-in-water cream base. If desired, the aqueous phase of the cream base may include, for example, at least about 30% w/w of a polyhydric alcohol, i.e., an alcohol having two or more hydroxyl groups such as propylene glycol, butane-1,3-diol, mannitol, sorbitol, glycerol and polyethylene glycol and mixtures thereof. The topical formulations may desirably include a compound which enhances absorption or penetration of the compound through the skin or other affected areas. Examples of such dermal penetration enhancers include dimethylsulfoxide and related analogues.

Emulsions are typically prepared from the compound and an oily phase, which may optionally comprise merely an emulsifier (otherwise known as an emulgent), or it may comprises a mixture of at least one emulsifier with a fat or an oil or with both a fat and an oil. Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier which acts as a stabiliser. It is also preferred to include both an oil and a fat. Together, the emulsifier(s) with or without stabiliser(s) make up the so-called emulsifying wax, and the wax together with the oil and/or fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations.

Suitable emulgents and emulsion stabilisers include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl monostearate and sodium lauryl sulfate. The choice of suitable oils or fats for the formulation is based on achieving the desired cosmetic properties, since the solubility of the compound in most oils likely to be used in pharmaceutical emulsion formulations may be very low. Thus the cream should preferably be a non-greasy, non-staining and washable product with suitable consistency to avoid leakage from tubes or other containers. Straight or branched chain, mono- or dibasic alkyl esters such as di-isoadipate, isocetyl stearate, propylene glycol diester of coconut fatty acids, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branched chain esters known as Crodamol CAP may be used, the last three being preferred esters. These may be used alone or in combination depending on the properties required. Alternatively, high melting point lipids such as white soft paraffin and/or liquid paraffin or other mineral oils can be used.

Formulations suitable for intranasal administration, where the carrier is a liquid, include, for example, nasal spray, nasal drops, or by aerosol administration by nebuliser, include aqueous or oily solutions of the compound.

Formulations suitable for intranasal administration, where the carrier is a solid, include, for example, those presented as a coarse powder having a particle size, for example, in the range of about 20 to about 500 microns which is administered in the manner in which snuff is taken, i.e., by rapid inhalation through the nasal passage from a container of the powder held close up to the nose.

Formulations suitable for pulmonary administration (e.g., by inhalation or insufflation therapy) include those presented as an aerosol spray from a pressurised pack, with the use of a suitable propellant, such as dichlorodifluoromethane, trichlorofluoromethane, dichoro-tetrafluoroethane, carbon dioxide, or other suitable gases.

Formulations suitable for ocular administration include eye drops wherein the compound is dissolved or suspended in a suitable carrier, especially an aqueous solvent for the compound.

Formulations suitable for rectal administration may be presented as a suppository with a suitable base comprising, for example, natural or hardened oils, waxes, fats, semi-liquid or liquid polyols, for example, cocoa butter or a salicylate; or as a solution or suspension for treatment by enema.

Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the compound, such carriers as are known in the art to be appropriate.

Formulations suitable for parenteral administration (e.g., by injection), include aqueous or non-aqueous, isotonic, pyrogen-free, sterile liquids (e.g., solutions, suspensions), in which the compound is dissolved, suspended, or otherwise provided (e.g., in a liposome or other microparticulate). Such liquids may additional contain other pharmaceutically acceptable ingredients, such as anti-oxidants, buffers, preservatives, stabilisers, bacteriostats, suspending agents, thickening agents, and solutes which render the formulation isotonic with the blood (or other relevant bodily fluid) of the intended recipient. Examples of excipients include, for example, water, alcohols, polyols, glycerol, vegetable oils, and the like. Examples of suitable isotonic carriers for use in such formulations include Sodium Chloride Injection, Ringer's Solution, or Lactated Ringer's Injection. Typically, the concentration of the compound in the liquid is from about 1 ng/ml to about 10 μg/ml, for example from about 10 ng/ml to about 1 μg/ml. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze- dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets.

Dosage

It will be appreciated by one of skill in the art that appropriate dosages of the PRD or DRD compounds, and compositions comprising the PRD or DRD compounds, can vary from patient to patient. Determining the optimal dosage will generally involve the balancing of the level of therapeutic benefit against any risk or deleterious side effects. The selected dosage level will depend on a variety of factors including, but not limited to, the activity of the particular PRD or DRD compound, the route of administration, the time of administration, the rate of excretion of the PRD or DRD compound, the duration of the treatment, other drugs, compounds, and/or materials used in combination, the severity of the condition, and the species, sex, age, weight, condition, general health, and prior medical history of the patient. The amount of PRD or DRD compound and route of administration will ultimately be at the discretion of the physician, veterinarian, or clinician, although generally the dosage will be selected to achieve local concentrations at the site of action which achieve the desired effect without causing substantial harmful or deleterious side-effects.

Administration can be effected in one dose, continuously or intermittently (e.g., in divided doses at appropriate intervals) throughout the course of treatment. Methods of determining the most effective means and dosage of administration are well known to those of skill in the art and will vary with the formulation used for therapy, the purpose of the therapy, the target cell(s) being treated, and the subject being treated. Single or multiple administrations can be carried out with the dose level and pattern being selected by the treating physician, veterinarian, or clinician.

In general, a suitable dose of the PRD or DRD compound is in the range of about 10 μg to about 250 mg (more typically about 100 μg to about 25 mg) per kilogram body weight of the subject per day. Where the compound is a salt, an ester, an amide, a prodrug, or the like, the amount administered is calculated on the basis of the parent compound and so the actual weight to be used is increased proportionately. BRIEF DESCRIPTION OF THE DRAWINGS

Embodiments of the invention and experiments illustrating the advantages and/or implementation of the invention are described below, by way of example only, with respect to the accompanying drawings, in which:

Figure 1 shows a simplified diagram of apoptosis signalling in a cell; Figures 2A, 2B and 2C show the changes in the heat of binding of BH3I-1 (Figure 2A), compound PRD06 (Figure 2B) and compound PRD18 (Figure 2C) to Bcl-X_L at pH 7.0, 298 K as monitored by VP-ITC (Microcal, USA). The top panel of each data set represents the heat effects recorded as a function of time during 29 successive 10 μl injections (except the first injection of 4 μl which was not included in the data analysis) of 1mM ligand solution into the sample cell containing 50 μM BcI- X_L. The bottom panel of each data set represents the heat released after appropriate blank corrections as a function of ligand to protein ratio; and Figures 3A and 3B show assay results for PRD compounds (and BH3I-1 comparison) against A549 human alveolar basal epithelial cells.

EXAMPLES

The following examples are provided solely to illustrate the present invention and are not intended to limit the scope of the invention, as described herein.

Synthetic methodology is described below in section (A) for acid substituted pyridylrhodanines, in section (B) for ester substituted pyridylrhodanines, in section (C) for other pyridylrhodanines, and in section (D) for biphenyl rhodanines.

Characterization

¹H NMR and ¹³C NMR spectra were acquired on a Bruker 400 UltraShield Spectrometer operating at 400 MHz and 100 MHz respectively. All the proton spectra were referenced to the respective residual solvent peaks (MeOH-αV 3.20; DMSO-Of₆: 2.50 ppm) except for those recorded in CDCI₃ were referenced to TMS. Carbon spectra were referenced to the central peak of the respective residual solvents (MeOH-Cf₄: 49.0; DMSO-Ck 39.5; CDCI₃: 77.0 ppm). Chemical shifts (SH and SC) are expressed in parts per million (ppm), referenced to TMS. Coupling constants (J) are reported to the nearest 0.5 Hz. Low-resolution and high-resolution electron impact mass spectra (EIMS) were measured using a Finnigan MAT95XP double-focusing mass spectrometer.

Low-resolution electrospray ionization (ESI) mass spectra were recorded using Waters Quattro Micro™ API, and high resolution mass spectra were obtained using the Agilent 6210 Time-of-Flight LC/MS.

Infrared (IR) spectra were measured on a BioRad FTIR spectrophotometer with samples analyzed as KBr discs or thin films on a KBr plate.

Elemental analysis was performed using a EuroEA3000 series CHNS Analyzer. X- ray analysis data was obtained using a Rigaku Single Crystal X-ray Diffraction System with Saturn-70 CCD Detector.

Flash chromatography was performed using silica gel by a Biotage Combiflash unit. Thin-layer chromatography was performed on aluminium plates pre-coated with silica gel (0.2 mm, Merck 60 F254), which were visualized under UV fluorescence.

Melting points were determined on a Bϋchi Melting Point B-540 and reported to the nearest 0.5 °C.

(A) Chemical synthesis of acid substituted pyridylrhodanines

The synthesis of the pyridine-based rhodanine compounds is outlined in Scheme 1.

The convergent synthetic route enabled the rapid construction of a small compound library of the arylrhodanines. The rhodanines 7 were prepared from the natural amino acids 6a-f (glycine, alanine, valine, leucine, isoleucine, and phenylalanine) in reasonable yields following the literature procedure [Sing et al Bioorg. Med. Chem. Lett. 2001, 11 , (2), 91-94]. The 2-aryl-5-formylpyridines 9a-c were prepared in high yields via the Suzuki-Miyaura coupling of the commercially-available arylboronic acids and 2-bromo-5-formylpyridine (8) using catalytic Pd₂(dba)₃ and PCy₃ as the ligand using a procedure reported by Handy [Handy et al J. Org. Chem. 2007, 72, (22), 8496-8500.]. Knoevenagel condensation of the 2-aryl-3-formylpyridines 9a-c with the corresponding rhodanines 7a-f in buffered acetic acid furnished the desired pyridylrhodanines 10a-x (compounds PRD01 to PRD24) in good yields. The same general synthesis was used to make the corresponding compounds based on the 5-bromo-2-formylpyridine precursor.

Scheme 1

The following conditions were used:

O) CS₂, NaOH₁ H₂O, 16h.

(ii) CICH₂CO₂Et, 3h.

(iii) 6N HCI, reflux 16 h. 48-92% yield over 3 steps.

(Jv) ArB(OH)₂, aq K₃PO₄, cat. Pd₂(dba)₃, cat. PCy₃HBF₄, 1 ,4-dioxane, 10O⁰C, 16 h.

77-97% yields.

(v) glacial AcOH, NaOAc, reflux, 3h. 65-98% yields.

General Procedure for the Synthesis of Λ/-substituted Rhodanines (7a-f) To a solution of the requisite natural amino acid (typically 5 g, 30 mmol) in water (100 mL) was added NaOH (2.4 g, 2 eq) and CS₂ (1.81 ml_, 30 mmol). The reaction mixture was stirred for 16h at room temperature after which a solution of sodium chloroacetate (2.82 g, 30 mmol) was added. The reaction mixture was stirred for a further 3h at room temperature, then acidified with aqueous HCI (6N, 30 mL). The resulting solution was refluxed for 16h, then cooled to ambient temperature. The crude product was extracted from the aqueous layer with ethyl acetate (3 x 50 ml_), and the combined organic layer was dried with MgSO₄, filtered, and the solvent removed in vacuo.

2-(4-oxo-2-thioxothiazolidin-3-yl)acetic acid (7a).

The title compound was recrystallized from ethanol as a yellow solid in 55% yield. Mp 148 ⁰C (lit. 148 ⁰C). ¹H NMR (CDCI₃): δ 4.16 (s, 2H, NCH₂), 4.79 (s, 2H, SCH₂).

(S)-2-(4-oxo-2-thioxothiazolidin-3-yl)propanoic acid (7b).

The title compound was recrystallized from ethanol. The product was obtained as a yellow solid in 49% yield. Mp 152-153 ⁰C. (lit. 152-153 ⁰C). ¹H NMR (CDCI₃): ^ 1.61 (d, J = 7 Hz, 3H, CH₃), 4.00 (d, J = 2 Hz, 2H, CH₂), 5.68 (q, J = 7 Hz, 1 H, NCH).

(S)-3-methyl-2-(4-oxo-2-thioxothiazolidin-3-yl)butanoic acid (7c).

The product was obtained as a yellow solid in 92% yield. Mp 113-115 ⁰C. (lit. 113- 115 ⁰C). ¹H NMR (CDCI₃): δ 0.81 (d, J = 7 Hz, 3H, CH₃), 1.24 (d, J = 7 Hz, 3H, CH₃), 2.80 (m, 1 H, CH₃CH), 4.03 (s, 2H, SCH₂), 5.25 (d, J = 9 Hz, 1 H NCH), 9.60 (s, 1 H, CO₂H).

(S)-4-methyl-2-(4-oxo-2-thioxothiazolidin-3-yl)pentanoic acid (7d).

The product was obtained as a yellow solid in 88% yield. Mp 100 ⁰C. (lit. 99-101 ⁰C). ¹H NMR (CDCI₃): δ 0.91 (d, J = 7 Hz, 3H, CH₃), 0.95 (d, J = 7 Hz, 3H, CH₃), 1.51 (m, 1 H, CH₃CH), 2.09 (m, 2H, NCHCH₂), 3.98 (s, 2H, SCH₂), 5.66 (m, 1 H, NCH).

(2S, 3S)-3-methyl-2-(4-oxo-2-thioxothiazolidin-3-yl)pentanoic acid (7e).

The product was obtained as a yellow oil in 48% yield. ¹H NMR (CDCI₃): δ 0.86 (t, J = 7 Hz, 3H, CH₃), 0.97-1.06 (m, 1 H, CH₂₃CH₃), 1.19 (d, J = 7 Hz, 3H, CHCH₃), 1.22- 1.28 (m, 1 H, ChU_bCH₃), 2.55 (m, 1 H, CHCH₃), 4.02 (s, 2H, SCH₂), 5.31 (d, J = 9 Hz, 3H, NCH). ¹³C NMR (CDCI₃): δ 11.0, 17.5, 25.2, 33.4, 34.5, 61.9, 173.4, 173.6, 201.0. LR-ESI(-) m/z (%): 246 ([M-H]-, 23), 172 (100).

(S)-2-(4-oxo-2-thioxothiazolidin-3-yl)-3-phenylpropanoic acid (7i).

The product was obtained as a yellow solid in 72% yield Mp 170-172 ⁰C. (lit. 170- 173 ⁰C). ¹H NMR (CDCI₃): δ 3.52 (d, J = 7 Hz, 2H, CH₂Ph), 3.77 (s, 2H, SCH₂), 5.85 (br s, 1H, NCH), 7.13- 7.25 (m, 5H) 9.65 (s, 1 H, CO₂H). LR-ESI(-) m/z (%): 280 ([M- H]-, 75).

Preparation of 2-Bromo-5-formylpyridine (8)

The compound was prepared following the literature procedure [van den Heuvel et al J. Org. Chem. 2004, 69, (2), 250-262.]. To a suspension of 2-bromo-5-iodopyridine (3 g, 11 mmol) is dry Et₂O (10O mL) at -78 ⁰C was added /1-BuLi (2.2 M, 5.3 mL, 1.1 eq). The reaction mixture was stirred for 1 h prior to the addition of dry DMF (1 mL). After stirring for an additional 1 h, the mixture was warmed to room temperature and quenched by the addition of dilute HCI (1 M, 20 mL). The organic layer was separated, and the aqueous layer was further extracted with Et₂O (2 x 20 mL). The combined organic fraction was dried with MgSO₄, filtered, and the solvent removed in vacuo. Column chromatography on silica using a gradient (7-60% EtOAc, hexanes) yielded the product as a white solid (1.25 g, 64% yield). R_f (1 : 1 EtOAc/hexanes): 0.39. Mp 100-101 ⁰C. (lit. 100 ⁰C). ¹H NMR (CDCI₃): δ 7.69 (d, J = 8 Hz, 1H), 8.01 (dd, J = 8 Hz and 2 Hz, 1H), 8.83 (d, J = 2 Hz, 1H), 10.10 (s, 1H₁ CHO).

Preparation of 5-Bromo-2-formylpyridine

This isomer of (8) was prepared in the same way as for (8) but using 5-bromo-2÷ iodopyridine instead of 2-bromo-5-iodopyridine. Purified by flash chromatography (6 → 30% ethyl acetate-hexanes) to yield the desired compound (2.49 g, 76%) as a white solid: R_f 0.63 (25% ethyl acetate-hexanes); δ_H (CDCI₃, 400 MHz) 10.04 (1H, s, CHO), 8.86 (1H, d, J = 2.0 Hz, ArH), 8.03 (1 H, dd, J = 8.5, 2.0 Hz, ArH),7.86 (1H, d, J = 8.5 Hz, ArH).

General Procedure for the Suzuki coupling of 2-Bromo-5-formylpyridine to Boronic Acids (9a-c) To a solution of 8 (0.1 g, 0.54 mmol) in 1 ,4-dioxane (5 mL) was added the boronic acid (1.1 eq) and aqueous K₃PO₄ (0.23g in 1 mL H₂O, 2 eq). The reaction mixture was degassed for 1 min with argon prior to the addition of catalytic Pd₂(dba)₃ (10 mg, 2 mol%) and PCy₃HBF₄ (8 mg, 4 mol%). The reaction mixture was stirred at 100 ⁰C for 16 h in a sealed tube. The reaction mixture was then cooled to rt and the solvent was removed in vacuo. The residue was partitioned between EtOAc (20 mL) and water (20 mL). The organic layer was collected and the aqueous phase was further extracted with EtOAc (2 x 20 mL). Column chromatography on silica (EtOAc/hexanes, 0-50%) yielded the desired products as white or pale yellow solids.

2-(2,3-dimethoxyphenyl)-5-formylpyridine (9a). Pale yellow solid (95 % yield). Mp 84 ⁰C. ¹H NMR (CDCI₃): δ 3.72 (s, 3H, OCH₃), 3.93 (s, 3H, OCH₃), 7.04 (dd, J = 8 Hz and 2 Hz, 1 H), 7.20 (t, J = 8 Hz, 1 H), 7.44 (dd, J = 8 Hz and 2 Hz, 1 H), 8.09 (d, J = 8 Hz, 1H), 8.20 (dd, J = 8 Hz and 2 Hz, 1 H), 9.14 (d, J = 2 Hz, 1 H), 10.14 (s, 1 H).¹³C NMR (CDCI₃): 5 55.9, 61.0, 113.8, 122.6, 124.4, 125.1 , 129.5, 133.0, 135.6, 147.4, 151.8, 153.0, 161.0, 190.6. HRMS (ESI⁺): Calcd 244.0968 for C₁₄H₁₄NO₃ [M+H]⁺, found: 244.0961. Anal (C₁₄H₁₃NO₃) C, H, N.

2-(3,4-methylenedioxyphenyl)-5-formylpyridine (9b). White solid (97 % yield). R_f (1 : 3 EtOAc/hexanes): 0.33. Mp 133-138 ⁰C. ¹H NMR (CDCI₃): δ 6.06 (s, 2H, CH₂), 6.94 (d, J = 9 Hz, 1 H), 7.63 (m, 2H), 7.80 (d, J = 9 Hz, 1 H), 8.18 (dd, J = 8 Hz and 2 Hz, 1 H), 9.06 ( d, J = 1 Hz, 1 H), 10.11 (s, 1 H, CHO). ¹³C NMR (CDCI₃): δ 101.6, 107.7, 108.6, 119.8, 122.2, 129.4, 132.3, 136.4, 148.5, 149.7, 152.4, 161.5, 190.3. FTIR (cm ¹, KBr): 757 w, 818 m, 846 w, 895 m, 931 m, 1037 s, 1107 m, 1240 s, 1302 m, 1369 s, 1404 m, 1476 s, 1558 m, 1591 s, 1696 s. LR-ESI(+) m/z (%): 228 ([M+H]⁺, 100). HR-ESI(+): Calcd 228.0655 for C₁₃H₁₀ NO₃ [M+H]⁺, found 228.0674. Anal (C₁₃H₉ NO₃) C₁ H₁ N.

2-(3,4-dimethoxyphenyl)-5-formylpyridine (9c). Yellow crystals (77 % yield). Mp 107- 108 ⁰C. ¹H NMR (CDCI₃): δ 3.96 (s, 3H, OCH₃), 4.02 (s, 3H, OCH₃), 6.98 (d, J = 8 Hz, 1 H), 7.63 (dd, J = 8 Hz and 2 Hz, 1 H), 7.77 (d, J = 2 Hz, 1 H), 7.86 (d, J = 8 Hz, 1 H), 8.18 (dd, J = 8 Hz and 2 Hz, 1 H), 9.07 (d, J = 2 Hz, 1 H), 10.10 (s, 1 H). ¹³C NMR (CDCI₃): δ 56.0, 110.2, 111.0 , 119.7, 120.5, 129.3, 130.7, 136.2, 149.4, 151.2, 152.4, 161.6, 190.4. HRMS (ESI⁺): Calcd 244.0968 for C₁₄H₁₄NO₃ [M+H]⁺, found 244.0962. Anal (C₁₄H₁₃NO₃) C, H, N.

2-(3-chloro-4-isopropoxyphenyl)-5-formylpyridine (9d). Colorless crystals (yield 93%). Mp 83 ⁰C. ¹H NMR (CDCI₃): δ1.43 (d, J = 6 Hz, 6H, 2 x CH₃), 4.68 (m, 1 H, OCH), 7.05 (dd, J = 9 Hz, 1 H), 7.82 (d, J = 8 Hz, 1 H), 7.96 (dd, J = 8 Hz and 3 Hz, 1 H), 8.15 (d, J = 3 Hz, 1 H), 8.19 (dd, J = 8 Hz and 3 Hz, 1 H), 9.07 (d, J = 2 Hz, 1 H), 10.11 (s, 1H). ¹³C NMR (CDCI₃): δ 22.0, 72.0, 114.9, 119.7, 124.6, 126.9, 129.51 , 129.57, 130.9, 136.5, 152.4, 155.5, 160.5, 190.3. HRMS (ESI⁺): Calcd 276.0786 for C₁₅H₁₅CINO₂ [M+H]⁺, found 276.0785. Anal (C₁₅H₁₄CINO₂) C, H, N.

The 4-isopropoxyphenyl analogue was also prepared using the same methodology.

6-(4-isopropoxyphenyl)nicotinaldehyde. Beige powder (250 mg, 64%). R_f 0.35 (30% ethyl acetate-hexanes); Mp 68.0-69.0 ⁰C; δ_H (CDCI₃, 400 MHz) 10.10 (1 H, s, CHO), 9.07 (1 H, d, J = 2.0 Hz), 8.18 (1 H, dd, J = 8.5, 2.0 Hz), 8.05 (2H, d, J = 9.0 Hz), 7.83 (1 H, d, J = 8.5 Hz), 7.00 (2H, d, J = 9.0 Hz), 4.70-4.61 (1 H, m, OCH), 1.38 (6H, d, J = 6.0 Hz, OCH(CHa)₂); <*c (CDCI₃, 100 MHz) 190.5 (CHO), 161.9 (para ArCHO), 160.1 (ipso ArOCH), 152.6 (ortho ArCHO), 136.3 (ortho ArCHO), 130.1 (para ArOCH), 129.2 (ipso ArCHO), 129.1 (meta ArOCH), 119.6 (meta ArCHO), 116.0 (ortho ArCHO), 67.0 (OCH), 22.0 (CH(CH₃)₂); m/z (%) (ESI+) 296 (10) 288 (100), 274 (80), 276 ([M+H]⁺, 42). (Found [M+H]⁺ 242.1165. C₁₅H₁₅NO₂ requires [M+Hf 242.1181).

The 4-tert-butylphenyl analogue was also prepared using the same methodology.

Beige powder (240 mg, 63%). R_f 0.48 (30% ethyl acetate-hexanes); Mp 137.0-138.5 ⁰C; δ_H (CDCI₃, 400 MHz) 10.13 (1 H, s, CHO), 9.11 (1 H, d, J = 2.0 Hz, ArH), 8.22 (1 H, dd, J = 8.0, 2.0 Hz, ArH), 8.04 (2H, d, J = 8.5 Hz₁ ArH), 7.89 (1 H, d, J = 8.5 Hz, ArH), 7.55 (2H, d, J = 8.5 Hz, ArH), 1.38 (9H, s, C(CH₃)₃); δ_c (CDCI₃, 100 MHz) 190.5 (CHO), 162.2 (para ArCHO), 153.9 (/PSO AI^BU), 152.5 (ortho ArCHO), 136.4 (ortho ArCHO), 135.1 (para Ar¹Bu), 129.6 (ipso ArCHO), 127.3 (meta Arteu), 126.0 (ortho Ar'Bu), 120.2 (meta ArCHO), 34.8 (C(CH₃)₃), 31.2 (C(CH₃)₃); m/z(%) (ESH) 286 (100), 272 (50). (Found [M+H]⁺ 240.1389. C₁₆H₁₈NO requires [M+H]⁺ 240.1388). The following compounds based on the 5-bromo-2-formylpyridine precursor were also prepared using the same methodology.

5-(3-chloro-4-isopropoxyphenyήpicolina\<iehyde.

Beige powder (358 mg, 80%). R_f 0.38 (30% ethyl acetate-hexanes); Mp 49.0-51.0 ⁰C; δ_H (CDCI₃, 400 MHz) 10.11 (1 H, s, CHO)₁ 8.96 (1 H, dd, J = 2.0, 1.0 Hz, ArH), 8.02 (1 H₁ d, J = 1.0 Hz), ArH)₁ 8.01 (1 H₁ d, J = 2 Hz, ArH), 7.67 (1 H₁ d, J = 2.5 Hz₁ ArH)₁ 7.50 (1 H₁ dd, J = 8.5, 2.5 Hz₁ ArH)₁ 7.08 (1 H₁ d, J = 8.5 Hz₁ ArH), 4.61-4.70 (1 H₁ m, OCH)₁ 1.43 (6H₁ d, J = 6.0 Hz₁ OCH(CH₃)₂); ^c (CDCI₃, 100 MHz) 192.8 (CHO), 154.5 (ipso ArO'Pr), 151.2 (ipso ArCHO), 148.0 (meta ArCHO)₁ 139.1 (para ArCHO), 134.4 (meta ArCHO)₁ 129.4 (para ArO'Pr), 129.1 (mete ArO'Pr), 126.5 (meta ArO'Pr), 125.0 (ipso ArCI), 121.8 (ortho ArCHO)₁ 115.7 (ortho ArO'Pr), 72.1 (OCH), 21.9 (OCH(CH₃J₂); m/z (%) (ESI+) 298 ([M+Na]⁺, 100), 290 ([M+NH₄]⁺, 57), 276 ([M+H]⁺, 42). (Found [M+H]⁺ 276.0794. C₁₅H₁₄CINO₂ requires [M+Hf 276.0786).

5-(4-isopropoxyphenyl)picolinaldehyde.

Beige powder (194 mg, 75%). R_f 0.48 (30% ethyl acetate-hexanes); Mp 53.5-54.5 ⁰C; ό_H (CDCI₃, 400 MHz) 10.10 (1 H₁ s, CHO), 8.98 (1H, t, J = 1.5 Hz, ArH), 7.99 (2H, d, J = 1.5 Hz), ArH), 7.58 (2H₁ d, J = 9.0 Hz₁ ArH)₁ 7.02 (2H, 6, J = 9.0 Hz₁ ArH)₁ 4.68-4.60 (1 H₁ m, OCH)₁ 1.38 (6H₁ d, J = 6.0 Hz₁ OCH(CH₃)₂); δ_c (CDCI₃, 100 MHz) 192.9 (CHO), 158.9 (ipso ArO'Pr), 150.7 (ipso ArCHO)₁ 148.0 (meta ArCHO), 140.2 (para ArCHO), 134.2 (meta ArCHO), 128.5 (meta ArO'Pr), 128.2 (para ArO'Pr), 121.8 (ortho ArCHO)₁ 116.4 (ortho ArO'Pr), 69.9 (OCH), 21.9 (OCH(CH₃)₂); m/z (%) (ESI+) 256 ([M+NH₄]⁺, 100), 242 ([M+H]⁺, 70). (Found [M+H]⁺ 242.1181. C₁₅H₁₅NO₂ requires [M+H]⁺ 242.1176). 5-(4-tert-butylphenyl)picolinaldehyde.

Beige powder (203 mg, 80%). R_f 0.60 (30% ethyl acetate-hexanes); Mp 79.0-81.5 ⁰C; <∑_H (CDCI₃, 400 MHz) 10.13 (1 H, s, CHO), 9.02 (1 H, dd, J = 2.0, 1.0 Hz, ArH), 8.05 (1 H, d, J = 2.0 Hz, ArH), 8.04 (1 H, d, J = 1.0 Hz, ArH), 7.61 (2H, d, J = 8.5 Hz, ArH), 7.55 (2H, d, J = 8.5 Hz, ArH), 1.38 (9H, s, C(CH₃)₃); δ_c (CDCI₃, 100 MHz) 193.1 (CHO), 152.6 (ipso Arfeu), 151.3 (/pso ArCHO), 148.6 (meta ArCHO), 140.6 (para ArCHO), 134.9 (meta ArCHO), 133.6 (para Ai^Bu), 127.1 (meta Ar¹Bu), 126.3 (ortho Ar'Bu), 121.9 (ortho ArCHO), 34.8 (C(CH₃)₃), 31.2 (C(CH₃)₃); m/z (%) (ESI+) 262 ([M+Na]⁺, 100%), 240 ([M+H]⁺, 80). (Found [M+H]⁺ 240.1389. C₁₆H₁₈NO requires [M+H]⁺ 240.1388).

General Procedure for the Knoevenagel Condensation of Aldehydes with Rhodanines

To a buffered solution of the aldehydes 9a-d (typically 100 mg, 0.30 mmol) in glacial acetic acid (1 to 5 ml.) and sodium acetate (0.11 g, 4eq) was added the requisite rhodanine 7a-f (2 eq). The mixture was refluxed for 3 h, then cooled to room temperature. The solvent was removed in vacuo, and aqueous HCI (1 M, 50 mL) was added and the resulting mixture was refluxed for a further 1 h. The reaction mixture was then cooled to rt, and the precipitate was collected via vacuum filtration. Recrystallization from hot 1N HCI, or column chromatography on silica, gave the pure product (65-98% yields).

[PRD01] 10a. Mp 225-226 ⁰C. ¹H NMR (DMSO-d₆): δ 3.70 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 4.73 (s, 2H, NCH₂), 7.20 (m, 2H), 7.38 (dd, J = 7 Hz and 3 Hz, 1 H), 7.98 (s, 1 H), 8.01 (d, J = 8 Hz, 1 H), 8.09 (dd, J = 8 Hz and 2 Hz, 1 H), 9.01 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-CZ₆): δ 45.1 , 55.9, 60.6, 114.1 , 122.0, 125.5, 124.2, 124.7, 127.3, 130.4, 132.4, 136.7, 147.0, 152.1, 152.9, 156.5, 166.2, 167.2, 192.8. FTIR (cm ¹ , KBr): 536 w, 584 w, 786 w, 835 w, 847 w, 1125 m, 1222 s, 1265 m, 1342 s, 1387 m, 1405 m, 1470 m, 1491 m, 1595 m, 1780 s, 2833 w, 2944 w, 2987 w. Anal (C₁₉H₁₆N₂O₅S₂) C, H, N, S. [PRD02] 10b. Mp 208 ⁰C. ¹H NMR (DMSOd₆): δ 1.56 (d, J = 7 Hz, 3H, CHCH₃), 3.70 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 5.63 (q, J = 7 Hz, 1H, NCH), 7.20 (m, 2H), 7.38 (dd, J = 7 Hz and 3 Hz, 1H), 7.92 (s, 1H), 8.01 (d, J = 8 Hz, 1H), 8.06 (dd, J = 9 Hz and 2 Hz, 1 H), 8.99 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-^₆): δ 13.4, 52.9, 55.9, 60.6, 114.1 , 122.0, 123.1 , 124.1 , 124.6, 127.3, 130.1, 132.4, 136.6, 147.0, 152.0, 152.8, 156.4, 166.0, 169.5, 192.4. FTIR (cm^'1, KBr): 534 w, 745 m, 1003 m, 1026 m, 1114 m, 1235 s, 1270 s, 1470 m, 1591 m, 1724 s, 1911 br w, 2916 w. LR-ESI(-) m/z (%):429 ([M-H]^", 22), 386 (32), 271 (28), 126 (100). HR-ESI(-):Calcd. 429.0584 for C₂₀H₁₇N₂O₅S₂ [M-H]-, found 429.0566. Anal (C₂₀H₁₈N₂O₅S₂) C, H, N, calculated S, 14.90; found S₁ 14.13.

[PRD03] 10c. Brown-yellow solid. Mp 222-224 ⁰C.¹H NMR (DMSO-^₆): δ 0.76 (d, J = 7 Hz, 3H, CH₃), 1.21 (d, J = 7 Hz, 3H, CH₃), 3.69 (s, 3H, OCH₃), 2.73 (m, 1 H), 3.87 (s, 3H₁ OCH₃), 5.19 (d, J = 8 Hz, 1 H), 7.19 (m, 2H)₁ 7.38 (dd, J = 7 and 3 Hz, 1 H), 7.96 (s, 1 H), 8.00 (d, J = 8 Hz, 1 H), 8.07 (dd, J = 8 and 2 Hz₁ 1 H)₁ 8.99 (d, J = 2 Hz₁ 1 H). ¹³C NMR (DMSO-Cy₆): δ 18.9, 21.6, 27.1 , 55.8, 60.5, 62.1 , 114.1 , 122.0, 122.3,

124.1 , 124.6, 127.2, 130.9, 132.4, 136.7, 147.0, 152.0, 152.8, 156.5, 166.4, 168.5, 193.0. FTIR (KBr, cm^'1) 3423 (br), 2971 , 1722, 1609, 1478, 1248, 1029, 833, 796, 751. HRMS (ESI ): Calcd 457.0897 for C₂₂H₂₁N₂O₅S₂ [M-H]^" Found: 457.0873. Anal (C₂₂H₂₂N₂O₅S₂) H, N, S calculated C, 57.82; found C, 56.20.

[PRD04] 1Od. Yellow solid. Mp 167-169 ⁰C. ¹H NMR (CDCI₃): δ 0.95 (d, J = 6 Hz, 3H, CH₃), 1.00 (d, J = 6 Hz, 3H, CH₃), 1.57 (m, 1 H), 2.16 (m, 1 H), 2.30 (m, 1 H), 3.71 (s, 3H, OCH₃), 3.93 (s, 3H, OCH₃), 5.78 (m, 1 H, NCH), 7.03 (dd, J = 8 and 2 Hz, 1 H),

7.21 (t, J = 8 Hz, 1 H), 7.43 (dd, J = 8 and 2 Hz, 1 H), 7.77 (s, 1 H), 7.84 (dd, J = 8 and 3 Hz, 1 H), 8.07 (d, J = 8 Hz, 1 H)₁ 8.88 (d, J = 3 Hz, 1 H).¹³C NMR (CDCI₃): δ 22.2, 23.0, 25.3, 36.9, 56.0, 56.1, 61.2, 113.9, 122.6, 124.3, 124.6, 125.6, 127.8, 129.1,

132.2, 136.6, 147.5, 151.4, 153.1 , 156.6, 167.1 , 172.1 , 192.2. FTIR (KBr, cm^'1) 3440 (br), 2963, 1729, 1590, 1263, 1206, 1028, 828, 795, 749. HRMS (ESI ): Calcd

471.1054 for C₂₃H₂₃N₂O₅S₂ [M-H]- Found: 471.1026. Anal. (C₂₃H₂₄N₂O₅S₂) C, H, N₁ S.

[PRD05] 1Oe. Mp 129-130 ⁰C. ¹H NMR (DMSO-c/₆): δ 0.80 (t, J = 7 Hz, 3H, CH₂CH₃), 0.96 (m, 1 H, CH^CH₃), 1.17 (d, J = 6 Hz₁ 3H₁ CHCH₃), 1.24 (m, 1H, CH^CHy). 2.51 (m, 1 H, CH₃CH), 3.69 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 5.21 (d, J = 9 Hz, 1H, NCHCO₂H), 7.20 (m, 2H), 7.39 (dd, J = 8 Hz and 1 Hz, 1 H), 7.95 (s, 1 H), 8.01 (m, 1 H), 8.07 (dd, J = 9 Hz, 2H), 8.99 (d, J = 2 Hz, 1 H). ¹³C NMR (CDCI₃): (J 11.2, 17.6,

25.3, 33.8, 56.0, 61.1 , 62.0, 113.8, 122.6, 123.9, 124.6, 125.5, 127.7, 129.5, 132.4, 136.4, 147.5, 151.8, 153.1 , 156.8, 167.3, 171.2, 192.2. FTIR (crτϊ\ KBr): 548 w, 687 w, 738 w, 1001 m, 1036 m, 1094 w, 1126 m, 1236 br s, 1261 s, 1327 m, 1427 w, 1468 m, 1585 m, 1719 s, 2361 w, 2933 m, 2965 m. LR-ESI(-) m/z (%):471 ([M-H]^", 25), 428 (65), 357 (18), 168 (100). HR-ESI(-): Calcd. 471.1054 for C₂₃H₂₃N₂O₅S₂ [M- H]-, found 471.1038. Anal (C₂₃H₂₄N₂O₅S₂) C, H, N, S.

[PRD06] 1Of. Yellow solid. Mp 160-162 ⁰C. ¹H NMR (CDCI₃): δ 3.64 (d, J = 7 Hz, 2H), 3.70 (s, 3H, OCH₃), 3.92 (s, 3H, OCH₃), 6.00 (br s, 1 H, NCH), 7.03 (d, J = 8 Hz, 1 H), 7.20 (m, 6H), 7.40 (d, J = 8 Hz, 1 H), 7.74 (s, 1 H), 7.81 (d, J = 8 Hz, 1 H), 8.04 (d, J = 8 Hz, 1 H), 8.88 (s, 1 H). ¹³C NMR (CDCI₃): δ 33.9, 56.0, 58.2, 61.2, 1 13.9, 122.6, 124.2, 124.6, 125.6, 127.1 , 127.8, 128.5, 128.9, 129.2, 132.1, 135.9, 136.7, 147.4, 151.4, 153.1 , 156.5, 166.9, 170.9, 191.6. FTIR (KBr, cm^"1) 3410 (br), 2920, 1723, 1609, 1263, 1174, 1031 , 831 , 739, 670. HRMS (ESP): Calcd 505.0897 for C₂₆H₂₁N₂O₅S₂ [M-H]^" Found: 505.0873. Anal. (C₂₆H₂₂N₂O₅S₂) C, H, N, S.

[PRD07] 1Og. Mp 281-282 ⁰C. ¹H NMR (DMSO-d₆): δ 4.76 (s, 2H, NCH₂CO₂H), 6.12 (s, 2H, OCH₂O), 7.06 (d, J = 8 Hz, 1 H), 7.74 (d, J = 2 Hz, 1 H), 7.77 (dd, J = 8 Hz and 2 Hz, 1 H), 7.95 (s, 1 H), 8.02 (dd, J = 9 Hz and 2 Hz, 1 H), 8.09 (d, J = 9 Hz, 1 H), 8.92 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-Cf₆): δ 45.1 , 101.6, 106.8, 108.7, 119.9, 121.7, 122.8, 127.0, 130.5, 131.7, 137.5, 148.1 , 149.1 , 152.2, 156.6, 166.2, 167.2, 192.7. FTIR (cm^"1, KBr): 739 w, 807 w, 1037 m, 1105 m, 1196 s, 1251 m, 1270 m, 1328 s, 1393 m, 1439 m, 1472 s, 1584 s, 1712 s. LR-ESI(-) m/z (%): 399 ([M-H] ,73); 301 (91). HR-ESI(-): Calcd 399.011S fOr C₁₈H₁₁N₂O₅S₂ [M-H]^", found: 399.0104. Anal (C₁₈H₁₂N₂O₅S₂) C, H, N, S.

[PRD08] 10h. Mp 251-252 ⁰C. ¹H NMR (DMSO-</₆): δ 1.55 (d, J = 7 Hz, 3H, CH₃), 5.62 (q, J = 7Hz, 1 H, NCHCO₂H), 6.12 (s, 2H, OCH₂O), 7.07 (d, J = 8 Hz, 1 H), 7.74 (d, J = 2 Hz, 1 H), 7.77 (dd, J = 8 Hz and 2 Hz, 1 H), 7.89 (s, 1 H), 8.01 (dd, J = 9 Hz and 1 Hz, 1 H), 8.10 (d, J = 8 Hz, 1 H), 8.91 (d, J = 2 Hz, 1 H).¹³C NMR (DMSO-d₆): δ

13.4, 52.9, 101.6, 106.8, 108.7, 119.9, 121.7, 122.4, 127.0, 130.2, 131.7, 137.4, 148.1 , 149.1, 152.1 , 156.5, 166.0, 169.5, 192.3. Anal (C₁₉H₁₄N₂O₅S₂) Calcd C, 55.80; H, 4.21; N₁ 6.17; S, 14.90. Found C, 53.04; H, 4.25; N, 6.01; S, 15.16. FTIR (cm ¹, KBr): 467 w, 549 w, 597 w, 636 w, 686 w, 764 w, 808 m, 846 w, 894 m, 917 m, 935 m, 1001 w, 1036 m, 1053 m, 1108 m, 1144 w, 1246 s, 1257 s, 1305 m, 1348 s, 1391 m, 1442 m, 1476 s, 1501 m, 1591 m, 1608 m, 1700 s, 1740 m.LR-ESI(-) m/z (%): 413 ([M-H]^", 8); 249 (13); 155 (32); 123 (100); 69 (57). HRMS: Calcd 413.0271 for C₁₉H₁₃N₂O₅S₂ [M-H]^"; found 413.0286. Anal (C₁₉H₁₄N₂O₅S₂) C, H, N, S.

[PRD09] 10i. Mp 248 ⁰C. ¹H NMR (DMSO-d₆): δ 0.76 (d, J = 7 Hz, 3H, CH₃), 1.21 (d, J = 7 Hz, 3H, CH₃), 2.74 (m, 1H, CH(CH₃)₂), 5.18 (d, J = 7 Hz, 1 H, NCHCO₂H) 6.12 (s, 2H, OCH₂O), 7.07 (d, J = 8 Hz, 1 H), 7.75 (d, J = 2 Hz, 1 H), 7.78 (dd, J = 8 Hz and 2 Hz, 1 H), 7.94 (s, 1 H), 8.03 (dd, J = 9 Hz and 2 Hz₁ 1 H), 8.11 (d, J = 9 Hz, 1 H), 8.92 (d, J = 2 Hz, 1H). ¹³C NMR (DMSO-cfe): δ 18.9, 21.6, 27.1, 62.1, 101.6, 106.7, 108.6, 119.8, 121.5, 121.7, 126.9, 131.0, 131.6, 137.5, 148.1 , 149.1, 152.2, 156.6, 166.4,

168.5, 192.9. FTIR (cnT¹, KBr): 547 w, 808 w, 1037 m, 1201 m, 1237 s, 1330 m, 1347 m, 1475 s, 1591 m, 1607 m, 1702 s, 1734 s, 2973 w. LR-ESI (-) m/z (%): 441 ([M-H]^", 50); 398 (100); 342 (30). HRMS: Calcd 441.0584 for C₂₁H₁₇N₂O₅S₂ [M-H]^"; found 441.0568. Anal (C₂₁H₁₈N₂O₅S₂) C, H, N, S.

[PRD10] 10j. Mp 233 ⁰C. ¹H NMR (DMS0-d₆): δ 0.87 (d, J = 7 Hz, 3H, CH₃), 0.92 (d, J = 7 Hz, 3H, CH₃), 1.48 (m, 1 H, CH^CH), 2.03 (m, 1 H, CH(CH₃)₂), 2.21 (m, 1 H, CH₂J₂CH), 5.60 (br s, 1H, NCHCO₂H), 6.12 (s, 2H, OCH₂O), 7.07 (d, J = 8 Hz, 1 H), 7.74 (d, J = 2 Hz, 1H), 7.76 (dd, J = 8 Hz and 2 Hz, 1H), 7.91 (s, 1H), 8.01 (dd, J= 9 Hz and 2 Hz, 1 H), 8.09 (d, J = 9 Hz, 1 H), 8.91 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO- Of₆): (J 21.9, 22.8, 24.8, 36.3, 55.9, 101.6, 106.7, 108.6, 119.9, 121.7, 121.9, 127.0,

130.6, 131.6, 137.5, 148.1, 149.1, 152.2, 156.6, 166.4, 169.3, 193.0. FTIR (cm^"1, KBr): 747 w, 813 m, 1036 s, 1226 s, 1249 s, 1268 s, 1337 s, 1390 m, 1475 s, 1593 m, 1608 m, 1699 s, 1734 br m, 2364 w, 2905 w. LR-ESI(-) m/z (%): 455 ([M-H]^", 22); 339 (12); 325 (52); 297 (14); 243 (15); 192 (43). HR-ESI(-): Calcd 455.0741 for C₂₂H₁₉N₂O₅S₂ [M-H]-, found 455.0724. Anal (C₂₂H₂₀N₂O₅S₂) C, H, N, S.

[PRD11] 10k. Mp 221-222⁰C.¹H NMR (DMSO-c/₆): <J 0.80 (t, J= 7 Hz, 3H, CH₂CH₃), 0.95 (m, 1H, CHgaCHa), 1.16 (d, J= 7 Hz, 3H, CHCH₃), 1.23 (m, 1H, CH₂₆CH₃), 2.51 (m, 1H, CH₃CH), 5.22 (d, J= 9 Hz, 1H, NCHCO₂H), 6.12 (s, 2H, OCH₂O), 7.07 (d, J = 8 Hz, 1H), 7.74 (d, J = 2 Hz, 1H), 7.78 (dd, J = 8 Hz and 2 Hz, 1 H), 7.93 (s, 1 H), 8.02 (dd, J= 9 Hz and 2 Hz, 1H), 8.11 (d, J= 8 Hz, 1H), 8.92 (d, J = 2 Hz, 1H).¹³C NMR (DMSO-Of₆): δ 10.9, 17.5,24.9,33.0,61.6,79.1, 101.6, 106.8, 108.7, 119.9, 121.5, 127.0, 131.0, 131.7, 137.6, 148.2, 149.2, 152.2, 156.6, 166.5, 168.6, 193.0. FTIR (cm^"1, KBr): 547 w, 583 w, 660 w, 691 w, 740 w, 808 m, 845 w, 893 w, 936 w, 958 w, 1037 m, 1102 m, 1132 m, 1201 m, 1237 s, 1305 m, 1330 m, 1347 m, 1391 m, 1440 m, 1475 s, 1501 m, 1555 w, 1591 m, 1607 m, 1702 s, 1734 m, 2363 m, 2929 m, 2973 m. LR-ESI(-) m/z (%): 445 ([M-H]^", 33); 412 (78); 341 (22); 256 (25); 168 (75). HR-ESI(-): Calcd 455.0741 for C₂₂H₁₉N₂O₅S₂ [M-H]^", found: 455.0739. Anal (C₂₂H₂₀N₂O₅S₂) C, H, N, S.

[PRD12] 101. Mp 229 ⁰C. ¹H NMR (DMSO-Cf₆): δ 3.52 (d, J = 5 Hz, 2H, CH₂Ph), 5.90 (br s, 1 H, NCHCO₂H), 6.11 (s, 2H, OCH₂O), 7.05 (d, J = 8 Hz, 1 H), 7.13-7.23 (m, 5H), 7.73 (d, J = 2 Hz, 1 H), 7.76 (dd, J = 8 Hz and 2 Hz, 1 H), 7.86 (s, 1 H), 7.95 (dd, J = 9 Hz and 2 Hz, 1 H), 8.06 (d, J = 9 Hz, 1 H), 8.88 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-Cy₆): δ 33.0, 58.2, 101.6, 106.7, 108.6, 119.8, 121.5, 121.7, 126.7, 126.8, 128.3, 128.9, 130.5, 131.6, 136.4, 137.5, 148.1, 149.1 , 152.2, 156.6, 166.3, 168.6, 192.2. FTIR (Cm^"1, KBr): 839 w, 1035 m, 1170 m, 1235 s, 1266 s, 1338 s, 1475 s, 1592 m, 1609 m, 1706 s, 1732 br m, 2890 w. LR-ESI(-) m/z (%): 489 ([M-H]^", 100); 445 ([M-C₃H₈]^", 85); 354 (28); 311 (43); 243 (32); 219 (16). HR-ESI(-): Calcd 489.0584 for C₂₅H₁₇N₂O₅S₂ [M-H]^", found 489.0593. Anal (C₂₅H₁₈N₂O₅S₂) C, H, N, S.

[PRD13] 10m. Orange solid. Mp 177-178 ⁰C (dec). ¹H NMR (DMSO-c/₆): δ 3.84 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 4.77 (s, 2H, NCH₂), 7.10 (d, J = 8 Hz, 1 H), 7.79 (m, 2H), 7.97 (s, 1 H), 8.04 (dd, J = 8 Hz and 2 Hz, 1 H), 8.15 (d, J = 8 Hz, 1 H), 8.95 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-Cy₆): 5 45.1, 55.5, 55.6, 110.0, 111.8, 120.2, 120.3, 122.9, 127.0, 129.2, 130.3, 138.0, 149.0, 151.0, 151.5, 156.3, 166.2, 167.2, 192.6. FTIR (KBr, cm^"1) 3445, 2939, 2843, 1722, 1586, 1516, 1332, 1201 , 1058, 842, 734, 609. HRMS (ESI ): Calcd 415.0428 for C₁₉H₁₅N₂O₅S₂ [M-H]^" Found: 415.0418. Anal (C₁₉H₁₆N₂O₅S₂) Calcd C₁ 54.79; H, 3.87; N, 6.73; S, 15.40. Found C, 45.84; H, 3.72; N, 5.61 ; S, 12.99.

[PRD14] 10n. Brown-yellow solid. Mp 243-244 ⁰C. ¹H NMR (DMSO-d₆): δ 1.56 (d, J = 7 Hz, 3H, CH₃), 3.83 (s, 3H, OCH₃), 3.86 (s, 3H, OCH₃), 5.63 (q, J = 7 Hz, 1 H, NCH), 7.10 (d, J = 8 Hz, 1 H), 7.77 (m 2H), 7.90 (s, 1 H), 8.00 (dd, J = 8 and 2 Hz, 1H), 8.14 (d, J = 8 Hz, 1 H), 8.92 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-d₆): δ 13.4, 52.8, 55.5, 55.6, 109.9, 111.7, 1 19.8, 120.0, 122.2, 126.8, 129.9, 130.4, 137.4, 149.0, 150.8, 152.8, 156.9, 166.0, 169.5, 192.3. FTIR (KBr, cm^"1) 3424 (br), 2938, 2361 , 1721 , 1584, 1287, 1238, 814, 731 , 669. HRMS (ESI^"): Calcd 429.0584 for C₂₀H₁₇N₂O₅S₂ [M- H]^" Found: 429.0573. Anal (C₂₀H₁₈N₂O₅S₂) Calcd C, 55.80; H, 4.21 ; N, 6.51 ; S, 14.90. Found C, 53.04; H, 4.25; N, 6.01 ; S, 15.16. [PRD15] 10o. Brown-yellow solid. Mp 182-184 ⁰C. ¹H NMR (DMSO-Qf₆): δ 0.76 (d, J = 7 Hz, 3H, CH₃), 1.21 (d, J = 6 Hz, 3H, CH₃), 2.75 (m, 1 H, CH), 3.84 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 5.19 (d, J = 9 Hz, 1H₁ NCH), 7.11 (d, J = 8 Hz, 1H), 7.78 (m, 2H), 7.95 (s, 1 H), 8.04 (dd, J = 9 and 2 Hz, 1 H), 8.16 (d, J = 9 Hz, 1 H), 8.94 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-d₆): δ 19.0, 21.7, 27.2, 55.5, 55.6, 62.2, 109.9, 111.8, 119.8, 120.1, 121.4, 126.8, 130.0, 131.2, 137.6, 149.0, 150.9, 152.3, 157.0, 166.5, 168.6,

193.0. FTIR (KBr, cnY¹) 3550 (br), 2965, 2363, 1726, 1585, 1244, 1026, 836, 737. HRMS (ESI ): Calcd 457.0897 for C₂₂H_2IN₂O₅S₂ [M-H]^' Found: 457.0882. Anal ( C₂₂H₂₂N₂O₅S₂) Calcd C, 57.62; H, 4.84; N, 6.1 V₁S, 13.99. Found C, 52.40; H, 5.25; N, 5.04; S, 17.93.

[PRD16] 1Op. Yellow solid. Mp 162-164 ⁰C. ¹H NMR (CDCI₃): δ 0.95 (d, J = 7 Hz, 3H, CH₃), 1.00 (d, J = 7 Hz, CH₃), 1.56 (m, 1 H), 2.15 (m, 1 H), 2.30 (m, 1 H), 3.95 (s, 3H, OCH₃), 4.00 (s, 3H, OCH₃), 5.80 (m, 1 H, NCH), 6.97 (d, J = 9 Hz, 1 H), 7.56 (dd, J = 9 Hz and 1 Hz, 1 H), 7.69 (br s, 1 H), 7.73 (s, 1 H), 7.81 (br s, 2H), 8.82 (s, 1 H). ¹³C NMR (CDCI₃): δ 22.2, 23.0, 25.2, 36.9, 56.00, 56.03, 56.1 , 110.0, 11 1.1 , 120.3,

120.4, 123.3, 127.2, 129.4, 130.4, 137.1, 149.5, 151.1 , 152.0, 157.8, 167.0, 172.5,

192.1. FTIR (KBr, cm^"1) 3450 (br), 2959, 1722, 1584, 1275, 1208, 1025, 834, 770, 738. HRMS (ESI ): Calcd 471.1054 for C₂₃H₂₃N₂O₅S₂ [M-H]^" Found: 471.1033. Anal (C₂₃H₂₄N₂O₅S₂) C, H, N, S.

[PRD17] 10q. Orange solid. Mp 157-158 ⁰C. ¹H NMR (DMSO-d₆): δ 0.80 (t, J = 7 Hz, 3H, CH₃), 0.95 (m, 1H), 1.16 (d, J = 6 Hz, 3H, CH₃), 1.24 (m, 1 H), 2.54 (m, 1H), 3.87 (s, 3H, OCH₃), 3.84 (s, 3H, OCH₃), 5.24 (d, J = 9 Hz, 1H, NCH), 7.10 (d, J = 8 Hz, 1H)₁ 7.79 (m, 2H), 7.94 (s, 1 H), 8.03 (dd, J = 9 and 2 Hz, 1H), 8.15 (d, J = 9 Hz, 1H), 8.93 (d, J = 2 Hz, 1H), 13.29 (br s, 1H, CO₂H). ¹³C NMR (DMSO-d₆): δ 10.9, 17.5, 24.8, 33.0, 55.5, 55.6, 61.6, 109.9, 111.7, 119.8, 120.1 , 121.3, 126.8, 129.9, 131.2,

137.5, 149.0, 150.8, 152.3, 157.0, 166.5, 168.6, 193.0. FTIR (KBr, cm^"1) 3480 (br), 2963, 2361 , 1716, 1583, 1235, 1026, 836, 770, 736. HRMS (ESI ): Calcd 471.1054 for C₂₃H₂₃N₂O₅S₂ [M-H]^' Found: 471.1040. Anal. (C₂₃H₂₄N₂O₅S₂) C, H₁ N, S.

[PRD18] 1Or. Orange solid. Mp 202-203 ⁰C. ¹H NMR (DMSO-Cf₆): δ 3.53 (d, J= 4 Hz, 2H, PhCH₂), 3.83 (s, 3H, CH₃), 3.86 (s, 3H, CH₃), 5.86 (br s, 1 H, NCH), 7.09 (d, J= 8 Hz₁ 1H), 7.15-7.23 (m, 5H), 7.77 (m, 2H), 7.86 (s, 1 H), 7.98 (dd, J= 2 and 9 Hz, 1H), 8.13 (d, J= 9 Hz, 1 H), 8.90 (d, J= 2 Hz₁ 1 H). ¹³C NMR (CDCI₃ + CD₃OD): δ 33.6, 55.8, 58.2, 109.8, 111.0, 120.0, 120.1 , 123.0, 126.8, 127.0, 128.3, 128.95, 129.0, 130.5, 136.0, 137.0, 149.2, 150.8, 151.8, 157.8, 166.9, 169.5, 191.8. FTIR (KBr, cm^" ¹) 3476 (br), 2907, 2360, 1719, 1584, 1263, 1229, 1023, 836, 738, 698. HRMS (ESI ): Calcd 505.0897 for C₂₆H₂₁N₂O₅S₂ [M-H]^" Found: 505.0873. Anal (C₂₆H₂₃CIN₂O₅S₂) C, H, N, S.

[PRD19] 10s. Brown-yellow solid. Mp 236-237 ⁰C. ¹H NMR (DMSO-d₆): δ 1.34 (d, J = 6 Hz, 6H, 2 x CH₃), 4.76 (s, 2H, NCH₂), 4.81 (m, 1 H), 7.32 (d, J = 9 Hz, 1 H), 7.97 (s, 1 H), 8.06 (dd, J = 9 and 2 Hz, 1 H), 8.15 (dd, J = 9 and 2 Hz, 1 H), 8.18 (d, J = 9 Hz, 1 H), 8.26 (d, J = 2 Hz, 1 H), 8.96 (d, J = 2 Hz, 1 H). ¹³C NMR (DMSO-c/₆): δ 21.8, 45.1 , 71.3, 115.2, 119.9, 122.9, 123.0, 127.0, 127.3, 128.5, 130.5, 137.6, 152.3, 154.5, 155.5, 166.2, 167.3, 192.6. FTIR (KBr, cm^"1) 2980, 1708, 1582, 1321 , 1199, 1109, 951, 812, 743, 617. HRMS (ESI ): Calcd 447.0246 for C₂₀H₁₆CIN₂O₄S₂ [M-H]^" Found: 447.0231. Anal (C₂₀H₁₇CIN₂O₄S₂) Calcd C₁ 53.51; H, 3.82; N, 6.24; S, 14.28. Found C, 52.14; H, 3.85; N, 5.82; S, 13.30.

[PRD20] 10t. Yellow solid. Mp 182-183 ⁰C. ¹H NMR (DMSO-c/₆): 61.34 (d, J = 7 Hz, 6H, 2 x CH₃), 1.56 (d, J = 7 Hz, 3H, CH₃), 4.81 (m, 1 H), 5.63 (q, J = 7 Hz, 1 H, NCH), 7.32 (d, J = 9 Hz, 1 H), 7.91 (s, 1 H), 8.04 (dd, J = 8 and 2 Hz, 1 H), 8.13 (m, 2H)₁ 8.26 (d, J = 2 Hz, 1H), 8.94 (d, J = 2 Hz, 1 H). ¹³C NMR (CDCI₃): δ 13.7, 22.0, 53.1 , 72.1 , 115.1 , 120.2, 123.9, 124.7, 126.8, 127.5, 129.3, 129.4, 130.6, 137.4, 151.9, 155.4, 156.7, 166.6, 172.6, 191.4. FTIR (KBr, cm^"1) 2978, 2361 , 1711, 1582, 1475, 1247, 1109, 950, 811 , 736. HRMS (ESI ): Calcd 461.0402 for C₂₁H₁₈CIN₂O₄S₂ [M-H]^" Found: 461.0377. Anal (C₂₁H₁₉CIN₂O₄S₂) C, H, N, S.

[PRD21] 10u. Yellow solid. Mp 223-225 ⁰C. ¹H NMR (CDCI₃): δ 0.85 (d, J = 7 Hz, 3H, CH₃), 1.29 (d, J = 6 Hz, 3H, CH₃), 1.41 (s, 3H, CH₃), 1.43 (s, 3H, CH₃), 2.91 (m, 1 H), 4.66 (m, 1H), 5.38 (d, J = 9 Hz, 1H, NCH), 7.04 (d, J = 9 Hz₁ 1H), 7.73 (s, 1H), 7.78 (d, J = 8 Hz₁ 1 H), 7.83 (dd, J = 9 and 2 Hz, 1 H), 7.90 (dd, J = 8 and 2 Hz, 1 H), 8.06 (d, J = 2 Hz₁ 1H), 8.82 (d, J = 2 Hz, 1H). ¹³C NMR (CDCI₃): δ19.1, 21.7, 22.0, 27.7, 62.4, 72.1 , 115.1 , 120.2, 123.6, 124.7, 126.8, 127.5, 129.35, 129.37, 130.6, 137.4, 152.0, 155.4, 156.7, 167.2, 171.9, 192.1. HRMS (ESI ): Calcd: 489.0715 for C₂₃H₂₂CIN₂O₄S₂ [M-H]^" Found: 489.0738. Anal (C₂₃H₂₃CIN₂O₄S₂) C, H, N, S. [PRD22] 1Ov. Yellow solid. Mp 213-216 ⁰C. ¹H NMR (CDCI₃): δ 0.94 (d, J = 7 Hz, 3H, CH₃). 0.99 (d, J = 7 Hz, 3H, CH₃), 1.41 (s, 3H₁ CH₃), 1.43 (s, 3H, CH₃), 1.56 (m, 1 H), 2.15 (m, 1H), 2.31 (m, 1H), 4.66 (m, 1H, OCH), 5.80 (m, 1H, NCH), 7.04 (d, J = 9 Hz, 1H), 7.80 (d, J = 8 Hz, 1H), 7.72 (s, 1H), 7.82 (dd, J = 8 and 2 Hz, 1H), 7.91 (dd, J = 9 and 2 Hz, 1 H), 8.08 (d, J = 2 Hz, 1 H), 8.81 (d, J = 2.2 Hz, 1 H). ¹³C NMR (CDCI₃): δ

22.0, 22.2, 23.0, 25.2, 36.9, 56.0, 72.1 , 115.1 , 120.1 , 123.7, 124.7, 126.7, 127.5,

129.2, 129.3, 130.6, 137.4, 151.9, 155.4, 156.7, 167.0, 172.8, 192.0. FTIR (KBr, cm^" ¹) 2962, 1720, 1584, 1475, 1281, 1208, 1107, 952, 814, 736. HRMS (ESI ): Calcd 503.0872 for C₂₄H₂₄CIN₂O₄S₂ [M-H]- Found: 503.0897. Anal. (C₂₄H₂₅CIN₂O₄S₂)C, H, N, S.

[PRD23] 10w. Yellow solid. Mp 157-159 ⁰C. ¹H NMR (CDCI₃): δ 0.88 (t, J = 7Hz, 3H), 1.07 (m, 1 H), 1.25 (d, J = 7 Hz, 3H, CH₃), 1.31 (m, 1 H), 1.43 (d, J = 6 Hz, 6H, 2 * CH₃), 2.66 (m, 1 H), 4.67 (m, 1 H), 5.44 (d, J = 9 Hz, 1 H, NCH), 7.05 (d, J = 9 Hz, 1 H), 7.74 (s, 1 H), 7.79 (d, J = 9 Hz, 1 H), 7.83 (dd, J = 9 and 2 Hz, 1 H), 7.92 (dd, J = 9 and 2 Hz, 1 H), 8.07 (d, J = 2 Hz, 1 H), 8.81 (d, J = 2 Hz, 1 H). ¹³C NMR (CDCI₃): δ

11.1 , 17.6, 22.0, 25.3, 33.7, 61.9, 72.1 , 115.1 , 120.1 , 123.5, 124.7, 126.7, 127.5,

129.3, 129.4, 130.6, 137.3, 152.0, 155.4, 156.7, 167.2, 171.8, 192.1. FTIR (KBr, cm^' ¹) 2973, 2361 , 1721 , 1584, 1475, 1236, 1106, 951, 813, 734, 688. HRMS (ESI ): Calcd 503.0872 for C₂₄H₂₄CIN₂O₄S₂ [M-H]^' Found: 503.0853. Anal (C₂₄H₂₅CIN₂O₄S₂) C, H, N, S.

[PRD24] 10x. Yellow solid. Mp 181-183 ⁰C. ¹H NMR (DMSO-^₆): δ 1.33 (d, J = 6 Hz, 6H, 2 x CH₃), 3.53 (d, J = 5 Hz, 2H), 4.78 (m, 1H, OCH), 5.90 (br s, 1H, NCH), 7.18 (m, 5H), 7.29 (d, J = 9 Hz, 1 H), 7.86 (s, 1 H), 7.97 (dd, J = 9 and 2 Hz, 1H), 8.10 (dd, J = 9 and 3 Hz, 1H), 8.11 (d, J = 9 Hz, 1 H), 8.23 (d, J = 2 Hz, 1H), 8.89 (d, J = 3 Hz, 1 H). ¹³C NMR (DMSO-Cf₆): δ 21.7, 33.0, 58.2, 71.2, 115.1 , 119.8, 121.7, 122.8, 126.7, 126.9, 127.1, 128.2, 128.4, 128.9, 130.4, 136.5, 137.6, 152.3, 154.5, 155.4, 166.3, 168.6, 192.2. FTIR (KBr, cm^"1) 2978, 1720, 1583, 1475, 1279, 1 107, 950, 815, 736. HRMS (ESI ): Calcd 537.0715 for C₂₇H₂₂CIN₂O₄S₂ [M-H]^" Found: 537.0695. Anal. (C₂₇H₂₃CIN₂O₄S₂) C, H, N, S.

The following compound was also synthesised using the same methodology: [PRD34] (2S,3R)-2-((Z)-5-((6-(4-isopropoxyphenyl)pyridin-3-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-methylpentanoic acid

Yellow solid (190 mg, 97%). R_f 0.31 (5% methanol-DCM); mp 181.5-183.5 ⁰C; δ_H (CDCI₃, 400 MHz) 8.80 (1H, s), 7.98 (2H, d, J = 9.0), 7.79 (2H, s), 7.74 (1H, s, SCCH), 6.99 (2H, d, J = 9.0 Hz), 5.44 (1 H, d, J = 9.0 Hz, NCH), 4.67-4.61 (1 H, m, OCH), 2.72-2.61 (1 H, m, NCHCH), 1.38 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 1.33-1.28 (1H, m, IxCH₂CH₃), 1.25 (3H, d, J = 6.5 Hz, CHCH₃), 1.12-1.04 (1 H, m, IxCH₂CH₃), 0.88 (3H, t, J = 7.5 Hz, CH₂CH₃); δ_c (CDCI₃, 100 MHz) 192.2, 171.8 (CO₂H), 167.3 (NCO), 159.9, 157.9, 152.1, 137.2, 129.6, 129.5, 128.9, 127.0, 123.1, 120.2, 116.1, 70.0 (OCH), 62.1 (NCH), 33.8 (NCHCH), 25.3 (CH₂CH₃), 22.0 (CH(CH₃)₂), 17.6 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI-) 469 ([M-H]^", 40), 425 (100). (Found [M-H]^" 469.1313. C₂₄H₂₅CIN₂O₄S₂ requires [M-H]^" 469.1256).

The following compounds based on the 5-bromo-2-formylpyridine precursor were also prepared using the same methodology.

[PRD25] (S,Z)-2-(5-((5-(3-chloro-4-isopropoxyphenyl)pyridin-2-yl)methylene)-4-oxo- 2-thioxothiazolidin-3-yl) -4-methylpentanoic acid

Yellow solid (155 mg, 85%). R, 0.21 (5% methanol-DCM); Mp 108.5-110.5 ⁰C; δ_H (CDCI₃, 400 MHz) 8.95 (1 H, d, J = Hz), 7.89 (1 H, dd, J = 8.0, 2.5 Hz), 7.65 (1 H, d, J = 2.5 Hz), 7.62 (1 H, s, SCCH), 7.58 (1 H, d, J = 8.0 Hz), 7.47 (1 H, dd, J = 8.5, 2.5 Hz), 7.06 (1 H, d, J = 8.5 Hz), 5.84 (1 H, q, J = 4.0 Hz, NCH), 4.67-4.61 (1 H, m, OCH), 2.34-2.27 (1 H, m, IxNCHCH₂), 2.19-2.10 (1 H, m, IxNCHCH₂), 1.59-1.52 (1 H, m, NCHCH₂CH), 1.43 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 0.98 (3H, d, J = 6.5 Hz, CH₂CH(CH₃)₂), 0.93 (3H, d, J = 6.5 Hz, CH₂CH(CH₃)₂); δ_c (CDCI₃, 100 MHz) 192.4, 173.9 (CO₂H), 167.4 (NCO), 154.3, 150.0, 147.5, 134.7, 134.3, 129.8, 129.0, 127.9, 127.4, 127.0, 126.2, 125.0, 115.8, 72.2 (OCH), 55.5 (NCH), 36.9 (CH₂CH(CH₃)₂), 25.2 (CH₂CH(CHs)₂), 23.0 (OCH(CH₃)₂), 22.2 (1xCH₂CH(CH₃)₂), 22.0 (1xCH₂CH(CH₃)₂); m/z (%) (ESI-) 503 ([M-H]-, 55). (Found [M-H]^" 503.0881. C₂₄H₂₄CIN₂O₄S₂ requires [M-H]^" 503.0872).

[PRD26]

Yellow solid. Rf (5% MeOH in DCM) 0.32. Mp126.5-129.0 ⁰C. ¹H NMR (CDCI₃): δ 8.81 (1 H, s), 7.83 (1 H₁ d, J = 6.5 Hz), 7.60 (1 H, d, J = 2.0 Hz), 7.57 (2H, m), 7.42 (1 H, dd, J = 8.5, 2.0 Hz), 7.03 (1 H, d, J = 9.0 Hz), 5.38 (1 H, d, J = 8.0 Hz, NCH), 4.63 (1 H, m, OCH)₁ 2.56 (1 H, m, NCHCH), 1.42 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 1.26-1.23 (1 H, m, IxCH₂CH₃), 1.17 (3H, d, J = 6.0 Hz, CHCH₃), 1.04-0.97 (1H, m, IxCH₂CH₃), 0.79 (3H₁ t, J = 7.0 Hz, CH₂CH₃). LRMS {m/z, %): 503 ([M-H]^", 80), 459 (100).Calcd 503.0866 for C₂₄H₂₄CIN₂O₄S₂ [M-H]^" , found: 503.0882.Anal. Calcd (C₂₄H₂₅CIN₂O₄S₂) C, H₁N₁S.

[PRD27] (S,Z)-2-(5-((5-(4-isopropoxyphenyl)pyridin-2-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-4-methylpentanoic acid

Yellow solid (194 mg, 91%). R_f 0.35 (5% methanol-DCM); mp 108.5-110.0 ⁰C; δ_H (CDCI₃, 400 MHz) 8.95 (1 H, d, J = 2.5 Hz), 7.89 (1 H, dd, J = 8.0, 2.5 Hz), 7.62 (1 H, s, SCCH), 7.55 (1 H, d, J = 8.0 Hz), 7.54 (2H, d, J = 9.0 Hz)₁ 7.00 (2H₁ d, J = 9.0 Hz), 5.84 (1 H₁ q, J = 4.5 Hz, NCH), 4.66-4.57 (1 H, m, OCH), 2.33-2.26 (1 H, m, IxNCHCH₂), 2.17-2.10 (1H, m, IxNCHCH₂), 1.61-1.52 (1 H, m, NCHCH₂CH)₁ 1.38 (6H, d, J = 6.0 Hz₁ OCH(CH₃)₂), 0.98 (3H₁ d, J = 6.5 Hz, CH₂CH(CH₃)₂), 0.93 (3H, d, J = 6.5 Hz₁ CH₂CH(ChU)₂); δ_c (CDCI₃, 100 MHz) 199.7, 174.4 (CO₂H)₁ 167.4 (NCO), 158.6, 149.4, 147.5, 135.9, 134.0, 128.6, 128.3, 127.4, 126.4, 116.4, 70.0 (OCH), 55.5 (NCH), 36.9 (CH₂CH(CH₃)₂), 25.1 (CH₂CH(CH₃)₂), 23.0 (1xCH₂CH(CH₃)₂), 22.2 (1 xCH₂CH(CH₃)₂), 22.0 (OCH(CH₃)₂); m/z {%) (ESI-) 469 ([M-H]^", 100), 425 (94), 175 (36). (Found [M-H]^" 469.1249. C₂₄H₂₆N₂O₄S₂ requires [M-H]^" 469.1256). [PRD28] (2S,3R)-2-((Z)-5-((5-(4-isopropoxyphenyl)pyridin-2-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-methylpentanoic acid

Yellow solid (189 mg, 97%). R_f 0.33 (5% methanol-DCM); mp 1 15.5 ⁰C; δ_H (CDCI₃, 400 MHz) 10.11 (1H, broad s, CO₂H), 8.93 (1 H, d, J = 2.5 Hz), 7.88 (1 H, dd, J = 8.0, 2.5 Hz), 7.63 (1 H, s, SCCH), 7.55 (1 H, d, J = 8.0 Hz), 7.54 (2H, d, J = 9.0 Hz), 6.99 (2H, d, J = 9.0 Hz), 5.49 (1 H, d, J = 9.0 Hz, NCH), 4.66-4.57 (1H, m, OCH), 2.71-2.60 (1 H, m, NCHCH), 1.37 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 1.33-1.29 (1 H, m, IxCH₂CH₃), 1.23 (3H, d, J = 6.5 Hz, CHCH₃), 1.10-1.02 (1 H, m, IxCH₂CH₃), 0.85 (3H, t, J = 7.5 Hz₁ CH₂CH₃); (Jc (CDCI₃, 100 MHz) 199.7, 173.7 (CO₂H), 167.5 (NCO), 158.6, 149.3, 147.4, 135.8, 134.0, 128.5, 128.4, 128.8, 127.5, 126.1 , 116.4, 67.0 (OCH), 61.3 (NCH), 33.6 (NCHCH), 25.1 (CHCH₃), 21.9 (OCH(CH₃)₂), 17.5 (CH₂CH₃), 11.0 (CH₂CH₃); m/z (%) (ESI-) 469 ([M-H]^", 70), 425 (100). (Found [M-H]^" 469.1351. C₂₄H₂₆N₂O₄S₂ requires [M-H]^" 469.1256).

[PRD29] (S,Z)-2-(5-((5-(4-tert-butylphenyl)pyridin-2-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-4-methylpentanoic acid

Yellow solid (169 mg, 86%). R_f 0.34 (5% methanol-DCM); Mp 127.0-128.0 ⁰C; δ_H (CDCI₃, 400 MHz) 8.99 (1 H, d, J = 2.0 Hz), 7.94 (1 H, dd, J = 8.0, 2.0 Hz), 7.63 (1 H, s, SCCH), 7.58 (1 H, d, J = 8.0 Hz), 7.57 (2H, d, J = 8.5 Hz), 7.52 (2H, d, J = 8.5 Hz), 5.84 (1 H, d, J = 4.0 Hz, NCH), 2.33-2.26 (1 H₁ m, IxNCHCH), 2.17-2.09 (1H, m, IxNCHCH), 1.59-1.52 (1 H, m, CH(CH₃)₂), 1.37 (9H, s, C(CHg)₃), 0.98 (3H, d, J = 6.5 Hz, CH(CH₃)₂), 0.92 (3H, d, J = 6.5 Hz, CH(CH₃)₂); δ_c (CDCI₃, 100 MHz) 199.7, 174.5 (CO₂H), 167.4 (NCO), 152.1, 149.9, 147.8, 136.1 , 134.6, 133.7, 128.2, 127.4, 126.8, 126.3, 55.5 (NCH), 36.9 (NCHCH₂), 34.7 (C(CH₃J₃), 31.2 (C(CH₃)₃), 25.2 (CH(CH₃)₂), 23.0 (1xCH(CH₃)₂), 22.2 (1xCH(CH₃)₂); m/z (%) (ESI-) 467 ([M-H]^", 100), 423 (33). (Found [M-H]^" 467.1503. C₂₄H₂₅N₂O₃S₂ requires [M-H]^" 467.1463). (B) Chemical synthesis of ester substituted pyridylrhodanines

General Procedure for Esterification of Rhodanine Carboxylic Acids

The requisite rhodanine carboxylic acid (55-100 mg, 1.0 eq) and iodine (0.1 eq) were dissolved in anhydrous methanol (2 ml_). Hydrochloric acid (33% aqueous solution,

0.1 ml_) was added and the solution was heated to reflux for 24 h. The resulting brown solution was concentrated, dissolved in ethyl acetate (10 ml_) and washed with saturated aqueous sodium thioshulphate (10 ml_). The aqueous layer was extracted with ethyl acetate (2 x 15 ml_), dried (Na₂SO₄) and concentrated under reduced pressure. The yellow solids were purified by flash chromatography (ethyl acetate- hexanes) to yield the following methyl esters.

[PRD36] (S,Z)-Methyl 2-(5~((6-(3-chloro-4-isopropoxyphenyl)pyridin-3-yl)methylene)- 4-oxo-2-thioxothiazolidin-3-yl)-4-methylpentanoate

Yellow^'oil(77 mg, 75%): R, 0.43 (20% ethyl acetate-hexanes); δ_H (CDCI₃, 400 MHz)

8.77 (1 H, s, ArH), 8.12 (1 H, d, J = 2.5 Hz, ArH), 7.93 (1H, dd, J = 8.5, 2.5 Hz, ArH),

7.78 (2H, s, ArH), 7.69 (1 H, s, SCCH), 7.03 (1 H, d, J = 8.5 Hz, ArH), 5.72 (1 h, dd, J = 9.5, 5.0 Hz, NCH), 4.69-4.63 (1 H, m, ArOCH), 3.75 (3H, s, CO₂CH₃), 2.33-2.25 (1 H, m, IxNCHCH₂), 2.19-2.12 (1H, m, IxNCHCH₂), 1.59-1.51 (1H, m, CH(CH₃)₂), 1.42 (6H, d, J = 6.0 Hz₁ OCH(CH₃)₂), 1.00 (3H, d, J = 6.5 Hz, CH(CH₃)₂), 0.94 (3H, d, J = 6.5 Hz, CH(CH₃)₂); ό_c (CDCI₃, 100 MHz) 192.0, 168.8 (CO₂CH₃), 166.9 (NCO), 156.7, 155.2, 152.1 , 137.0 (SCCH), 130.9, 129.4, 129.1 , 127.2, 126.4, 124.5, 123.3, 119.5, 115.0, 71.9 (OCH), 56.0 (NCH), 52.8 (CO₂CH₃), 36.9 (NCHCH₂), 25.1 (CH₂CH(CHa)₂), 23.0 (1xCH₂CH(CH₃)₂), 22.1 (1xCH₂CH(CH₃)₂), 21.9 (OCH(CH₃)₂); m/z (%) (ESI+) 541 ([M+Na]⁺, 20), 344 (20), 336 (40), 322 (100), 226 (30), 167 (20). Found [M+H]⁺ 519.1183. C₂₅H₂₇CIN₂O₄S₂ requires [M+H]⁺ 519.1179. Anal (C₂₅H₂₇CIN₂O₄S₂) C, H, N, S.

[PRD37] (2S, 3R)-Methyl 2-((Z)-5-((6-(3-chloro-4-isopropoxyphenyl)pyridin-3- yl)methylene)-4-oxo-2-thioxothiazolidin-3-yl)-3-methylpentanoate

Yellow oil (28 mg, 50%): R_f 0.46 (20% ethyl acetate-hexanes); δ_H (CDCI₃, 400 MHz)

8.79 (1H, s, ArH), 8.13 (1H, d, J = 2.5 Hz, ArH), 7.94 (1H, dd, J - 8.5, 2.5 Hz, ArH),

7.80 (2H, d, J = 2.5 Hz, ArH), 7.71 (1H, s, SCCH), 7.04 (1H, d, J = 8.5 Hz, ArH), 5.34 (1 H, d, J = 9.5 Hz, NCH)₁ 4.70-4.64 (1 H, m, OCH), 3.72 (3H, s, CO₂CH₃), 2.72-2.61 (1 H, m, CHCH₃), 1.43 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 1.31-1.26 (1 H, m, 1 x CH₂CH₃), 1.23 (3H, d, J = 6.5 Hz, CHCH₃), 1.08-1.02 (1H, m, 1 x CH₂CH₃), 0.88 (3H, t, J = 7.5 Hz, CH₂CH₃); δ_c (CDCI₃, 100 MHz) 192.1 (NCS), 168.2 (CO₂CH₃), 167.2 (NCO), 156.8, 152.1, 137.0, 130.9, 129.6, 129.2, 127.2, 126.5, 124.6, 123.2, 119.6, 115.1, 72.0 (OCH), 61.9 (NCH), 52.5 (CO₂CH₃), 33.7 (CHCH₃), 25.1 (CH₂CH₃), 21.9 (OCH(CHs)₂), 17.5 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI+) 541 ([M+Na]⁺, 27), 381 (90), 353 (50), 226 (100). Found [M+H]⁺ 519.1176. C₂₅H₂₇CIN₂O₄S₂ requires [M+H]⁺ 519.1179. Anal (C₂₅H₂₇CIN₂O₄S₂) C, H, N, S.

[PRD38] (2S, 3R)-Methyl 2-((Z)-5-((6-(benzo[d][1 , 3]dioxol-5-yl)ρyridin-3- yl)methylene)-4-oxo-2-thioxothiazolidin-3-yl)-3-methylpentanoate

Yellow oil (15 mg, 69%): R_f 0.43 (20% ethyl acetate-hexanes); δ_H (CDCI₃, 400 MHz) 8.78 (1H, d, J = 2.0 Hz, ArH), 7.90-7.79 (2H, m, ArH, SCCH), 7.61 (2H, s, ArH), 6.93 (2H, d, J = 8.5 Hz, ArH), 6.05 (2H, s, OCH₂O), 5.34 (1H, d, J = 9.0 Hz, NCH), 3.72 (3H, s, CO₂CH₃), 2.72-2.57 (1H, m, CHCH₃), 1.33-1.26 (1H, m, IxCH₂CH₃), 1.23 (3H, d, J = 6.5 Hz, CHCH₃), 1.09-1.02 (1 H, m, IxCH₂CH₃), 0.88 (3H, t, J = 7.5 Hz, CH₂CH₃); δ_c (CDCI₃, 100 MHz) 192.2, 168.3 (CO₂CH₃), 167.2 (NCO), 157.8, 153.0, 148.6, 138.5, 137.0, 132.3, 129.7, 127.1 , 119.8, 108.7, 107.4, 101.6 (OCH₂O), 61.9 (NCH), 52.6 (CO₂CH₃), 33.7 (CHCH₃), 25.1 (CH₂CH₃), 17.5 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI+) 493 ([M+Na]⁺, 20), 477 ([M+H]⁺, 10), 226 (100), 185 (10). Found [M+H]⁺ 471.1056. C₂₃H₂₂N₂O₅S₂ requires [M+H]⁺ 471.1048. Anal (C₂₃H₂₂N₂O₅S₂) C, H, N, S. [PRD39] (S,Z)-Methyl 2-(5-((6-(benzo[d][1,3]dioxol-5-yl)pyridin-3-yl)methylene)-4- oxo-2-thioxothiazolidin-3-yl)-3-phenylpropanoate

Yellow oil (16 mg, 76%): R_f 0.31 (20% ethyl acetate-hexanes); ό_H (CDCI₃, 400 MHz) 8.74 (1H, s ArH), 7.74 (2H, d, J = 1.5 Hz, ArH), 7,63 (1 H, s, SCCH), 7.61-7.58 (2H, m, ArH), 7.25-7.16 (5H₁ m, CH₂Ph), 6.92 (1H, d, J = 8.5 Hz, ArH), 6.04 (2H, s, OCH₂O), 5.92 (1H, t, J = 8.0 Hz, NCH), 3.79 (3H, s, CO₂CH₃), 3.62 (2H, d, J = 8.0 Hz, NCHCH₂); δ_c (CDCI₃, 100 MHz) 191.7, 168.2 (CO₂CH₃), 166.9 (NCO), 157.8, 152.8, 152.2, 151.7, 149.5, 148.6, 137.0, 135.9, 129.5, 129.3, 129.2, 128.6, 127.1 , 121.8, 11.8, 108.7, 107.4, 101.6 (OCH₂O), 58.2 (NCH), 53.0 (CO₂CH₃), 33.9 (CH₂Ph); m/z (%) (ESI+) 527 ([M+H]⁺, 40), 352 (50), 279 (80), 260 (100). Found [M+H]⁺ 505.0908. C₂₆H₂₀N₂O₅S₂ requires [M+H]⁺ 505.0892. Anal (C₂₆H₂₀N₂O₅S₂) C, H₁ N₁ S.

[PRD40] (S,Z)-Methyl 2-(5-((6-(2,3-dimethoxyphenyl)pyridin-3-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-phenylpropanoate

Yellow oil (56 mg, 54%): R, 0.25 (20% ethyl acetate-hexanes); δ_H (CDCI₃, 400 MHz) 8.81 (1 H, d, J = 2.5 Hz, ArH), 8.05 (1 H, d, J = 8.5 Hz, ArH), 7.76 (1 H, dd, J = 8.5, 2.5 Hz, ArH), 7.65 (1H, s, SCCH), 7.46 (1H, dd, J = 8.0, 1.5 Hz, ArH), 7.23-7.16 (6H, m, CH₂Ph, ArH), 7.02 (1 H, dd, J = 8.0, 1.5 Hz, ArH), 5.92 (1 H, t, J = 8.0 Hz, NCH), 3.92 (3H, s, ArOCH₃), 3.79 (3H, s, ArOCH₃), 3.71 (3H, s, CO₂CH₃), 3.63 (2H, d, J = 8.0 Hz₁ CH₂Ph); δ_c (CDCI₃, 100 MHz) 191.7, 168.1 (CO₂CH₃), 166.8 (NCO), 157.1 , 153.0, 151.7, 147.5, 136.2, 135.8, 132.8, 129.5, 129.1 , 128.5, 127.2, 127.0, 125.0, 124.4, 122.4, 113.6, 61.0 (NCH), 58.1 (ArOCH₃), 55.9 (ArOCH₃), 52.9 (CO₂CH₃), 33.8 (CH₂Ph); m/z {%) (ESI+) 521 ([M+H]⁺, 20), 352 (100), 290 (30), 216 (70). Found [M+H]⁺ 521.1198. C₂₇H₂₄N₂O₅S₂ requires [M+Hf 521.1205. Anal (C₂₇H₂₄N₂O₅S₂) C, H, N, S.

(C) Chemical synthesis of further pyridylrhodanines

The following compounds were also synthesised using commercially available rhodanine (Lancaster Chemicals).

[PRDZQ](Z)-5-((6-(benzo[d][1,3]dioxol-6-yl)pyridin-3-yl)methylene)-2- thioxothiazolidin-4-one

Prepared from 9b and commercially-available rhodanine (Lancaster Chemicals).

Mp 308-309⁰C.¹H NMR (d_s-DMSO): ό 6.11 (s, 2H, OCH₂O)₁7.06 (d, J = 8 Hz, 1H, Ar-H), 7.70 (s, 1H, Ar-H)₁7.73 (d, J = 2 Hz, 1H₁ Ar-H)₁7.76 (dd, J = 8 Hz and 2 Hz, 1H, Ar-H)₁7.95 (dd, J = 9 Hz and 2 Hz, 1H, Ar-H), 8.08 (d, J = 8 Hz, 1H, Ar-H), 8.86 (d, J= 2 Hz, 1H, Ar-H).¹³C NMR (^₆-DMSO): δ 101.6 (OCH₂O), 106.7, 108.7, 119.8, 121.6, 126.6, 127.1, 128.1, 131.7, 137.2, 148.1, 149.0, 151.9, 156.2, 169.3 (CO), 195.1 (CS). FTIR (cm¹, KBr): 537 w, 588 m, 646 m, 678 m, 816 m, 1033 m, 1069 m, 1109 m, 1138 m, 1163 m, 1194 s, 1219 s, 1268 m, 1391 m, 1440 s, 1469 s, 1504 w, 1580 s, 1600 m, 1692 s. LR-ESI(-) m/z (%): 341 ([M-H]^", 100); 312 (40); 296 (51); 239 (49); 213 (25). Anal (C₁₆H₁₀N₂O₃S₂) C₁ H, N, S.

{PRϋZΛ\(Z)-5-((6-(3-chloro-4-isopropoxyphenyl)pyridin-3-yl)methylene)-2- thioxothiazolidin-4-one

Prepared from 9d and commercially-available rhodanine (Lancaster Chemicals). Mp 172 ⁰C. ¹H NMR (CZ₆-DMSO): δ 1.33 (d, J = 2 Hz, 6H, 2CH₃), 4.79 (m, 1H,

CH₃CHCH₃), 7.31 (d, J = 9 Hz, 1 H, Ar-H), 7.71 (s, 1 H, Ar-H), 7.97 (dd, J = 9 Hz and 2

Hz, 1 H, Ar-H), 8.12 (m, 2H, Ar-H), 8.24 (d, J = 2 Hz, 1 H, Ar-H), 8.88 (d, J = 2 Hz, 1 H, Ar-H). ¹³C NMR (J₆-DMSO): δ 22.0 (CH₃), 72.0 (OCH(CH₃)₂), 115.1 , 119.7, 124.7,

126.3, 126.5, 127.0, 129.2, 129.4, 130.9, 137.0, 152.1, 155.3, 157.0, 167.9 (CO),

191.8 (CS). FTIR (Cm^"1, KBr): 528 w, 676 w, 811 m, 951 m, 1061 m, 1107 m, 1192 s,

1227 s, 1258 s, 1282 s, 1393 m, 1433 m, 1475 s, 1582 s, 1711 s, 2978 w. LR-ESI(-) m/z (%):389 ([M-H]^", 100). HR-ESI(-): Calcd 389.0191 for C₁₈H₁₄CIN₂O₂S₂ [M-H]^", found 389.0177.Anal (C₁₈H₁₅CIN₂O₂S₂), C, H, N, S.

[PRD32] (Z)-5-((6-(2,3<limethoxyphenyl)pyridin-3-yl)methylene)-2-thioxothiazolidin- 4-one

Prepared from 9a and commercially-available rhodanine (Lancaster Chemicals).

Mp 220 ⁰C. ¹H NMR (CZ₆-DMSO): δ 3.68 (s, 3H, OCH₃), 3.87 (s, 3H, OCH₃), 7.20 (m, 2H₁ Ar-H), 7.36 (dd, J = 7 Hz and 3 Hz, 1 H, Ar-H), 7,74 (s, 1 H, Ar-H), 8.01 (m, 2H, Ar-H), 8,95 (d, J = 1 Hz, 1H, Ar-H). ¹³C NMR (cZg-DMSO): δ 55.9 (OCH₃), 60.6

(OCH₃), 114.1 , 122.0, 124.2, 124.8, 127.4, 127.6, 127.9, 132.2, 136.7, 146.9, 151.4, 152.8, 155.8, 169.2 (CO), 195.2 (CS). FTIR (cm ¹, KBr): 536 w, 1036 m, 1096 w, 1124 m, 1204 s,1267 s, 1433 m, 1476 s, 1542 w, 1575 s, 1623 m, 1712 s, 2835 w, 2970 w, 3021 w, 3403 br s. LR-ESI (-) m/z (%): 357 ([M-H]^', 100). HR-ESI(-):Calcd. 357.0373 for C₁₇H₁₃N₂O₃S₂ [M-H]^", found 357.0365. Anal (C₁₇H₁₄N₂O₃S₂) C, H, N, S.

[PRD33] (Z)-5-((6-(3A-dimethoxyphenyl)pyridin-3-yl)methylene)-2-thioxothiazolidin- 4-one

Prepared from 9c and commercially-available rhodanine (Lancaster Chemicals). Mp 260 ⁰C. ¹H NMR (c/₆-DMSO): δ 3.83 (s, 3H, OCH₃), 3.86 (s, 3H, OCH₃), 7.09 (d, J = 8 Hz, 1H₁ Ar-H), 7.70 (s, 1H, Ar-H), 7.75 (m, 2H, Ar-H), 7.97 (dd, J = 9 Hz and 2 Hz, 1H, Ar-H), 8.14 (d, J = 9 Hz, 1H, Ar-H), 8.88 (d, J = 2 Hz, 1H, Ar-H). ¹³C NMR (Cf₆-DMSO): δ 55.56 (OCH₃), 55.63 (OCH₃), 109.9, 111.8, 120.1, 120.2, 126.6, 127.1, 128.1 , 129.5, 137.5, 149.0, 150.8, 151.5, 156.2, 169.2 (CO), 195.1 (CS). FTIR (cm ¹, KBr): 541 w, 617 w, 912 w, 1011 m, 1062 m, 1161 s, 1209 s, 1237 s, 1274 s, 1420 m, 1437 m, 1515 s, 1585 s, 1715 s, 2361 m, 2565 br m, 2810 m, 3021 m, 3339 br m, 3582 m. LR-ESI(-) nVz (%): 357 ([M-H]^", 100). HR-ESI(-):Calcd. 357.0373 for C₁₇H₁₄N₂O₃S₂ [M-H]- , found 357.0371. Anal (C₁₇H₁₄N₂O₃S₂) C, H, N, S.

The following compound was also synthesised from the 2-bromo-5-formylpyridine precursor and rhodanine 7c.

[PRD35J 2-((Z)-5-((6-bromopyridin-3-yl)methylene)-4-oxo-2-thioxothiazolidin-3-yl)-3- methylbutanoic acid

Prepared from 2-bromo-5-formylpyridine and rhodanine 7c via Knoevenagel condensation.

Mp 160-161 ⁰C¹H NMR (GVDMSO): δ 0.84 (d, J = 7 Hz, 3H, CH₃), 1.29 (d, J = 6 Hz, 3H, CH₃), 2.90 (m, 1 H, CH(CH₃)₂), 5.37 (d, J = 9 Hz, 1 H, NCHCO₂H), 7.63 (br s, 3H, Ar-H), 8.53 (br s, 1 H, Ar-H). ¹³C NMR (GVDMSO): δ 19.1 (CH₃), 21.7 (CH₃), 27.6 (CH(CHs)₂), 62.2 (CHCO₂H), 128.0, 128.6, 128.8, 138.1 , 143.8, 151.8, 166.8 (CO), 172.3 (CO₂H), 191.5 (CS). FTIR (cm^"1, KBr): 685 w, 723w, 825 w, 1030 w, 1083 m, 1124 w, 1025 m, 1240 s, 1340 m, 1377 m, 1459 m, 1573 w, 1609 m, 1722 s, 1968 m, 3422 m. LR-ESK+) m/z (%):403 ([M+H]⁺, 100) and 401 ([M+H]⁺, 90) for ⁷⁹Br and ⁸¹Br isotopes; 344 (15); 283 (15); 261 (15); 239 (25); 217 (15). HR-ESI(+): Calcd 422.9443 for C₁₄H₁₃N₂O₃S₂Na, found 422.9444. Anal (C₁₄H₁₃BrN₂O₃S₂) C, H, N, S.

(D) Chemical synthesis of biphenyl rhodanines

The synthesis of the biphenyl analogues was carried out by coupling of arylboronic acids with 4-bromobenzaldehye. To a solution of 4-bromobenzaldehyde (100-150 mg), requisite boronic acid (1.1 eq) and anhydrous K₂CO₃ in 'PrOH (5 mL) degassed with Ar was added Pd(OAc)₂ (5 mol%). The reaction mixture was heated to reflux for 16 h after which time the black solution was diluted with sodium hydrogencarbonate (20 mL) and DCM (20 mL). The aqueous layer was extracted with DCM (2 x 20 mL) and the combined organic layers dried (MgSO₄) and concentrated under reduced pressure. The crude yellow oil was purified by flash chromatography (0 -> 10% ethyl acetate-hexanes) to yield the biaryls.

The following biphenyl precursors were synthesised.

3 '-chloro-4 '-isopropoxybiphenyl-4-carbaldehyde

White powder (109 mg, 61%). R_f 0.40 (11% ethyl acetate-hexanes); Mp 61-62 ⁰C; V_max (film) 1686, 1596, 1292, 1281 , 1263, 1170, 1103, 1058, 952 cm^"1; δ_H (CDCI₃, 400 MHz) 10.03 (1H, s, CHO), 7.92 (2H, dd, J = 8.0, 2.0 Hz, ArH), 7.68 (2H₁ dd, J = 8.0, 2.0 Hz, ArH), 7.66 (1H, d, J = 2 Hz, ArH), 7.47 (1H, dd, J = 6.0, 2.0 Hz, ArH), 7.03 (1H, d, J = 9.0 Hz, ArH), 4.63 (1H, m, OCH), 1.42 (6H, d, J = 6.0 Hz, CH(CH₃)₂); δ_c (CDCI₃, 100 MHz) 191.8 (CHO), 154.0 (ipso ArO'Pr), 145.5 {para ArCHO), 135.0 {ipso ArCHO), 132.8 (para ArO'Pr), 130.3 (ortho ArCHO), 129.2 (ortho ArCI), 127.1 {meta ArCHO), 126.4 (para ArCI), 124.7 {ipso ArCI), 115.7 {ortho ArO'Pr), 72.2

(OCH), 22.0 (OCH(CHa)₂); m/z {%) (ESI+) 298 ([M+Naf, 100), 290 ([M+NH₄]⁺, 57), 276 ([M+H]\ 42). (Found [M+H]⁺ 276.0794. C₁₅H₁₄CINO₂ requires [M+H]⁺ 276.0786).

4'-isopropoxybiphenyl-4-carbaldehyde

White powder (61 mg, 35%). R_f 0.30 (9% ethyl acetate-hexanes); Mp 76-77 ⁰C; v_max (film) 1701, 1600, 1252, 1187, 1176, 1102, 952, 820 cm^"1; δ_H (CDCI₃, 400 MHz) 10.03 (1H₁ s, CHO), 7.91 (2H, dd, J = 7.0, 2.0 Hz, ArH), 7.71 (2H, dd, J = 7.0, 2.0 Hz, ArH), 7.57 (2H, dd, J = 6.0, 2.0 Hz₁ ArH), 6.99 (2H, dd, J = 7.0, 2.0 Hz, ArH), 4.62 (1 H, m, OCH), 1.37 (6H, d, J = 6.0 Hz, CH(CH₃)₂); δ_c (CDCI₃, 100 MHz) 191.9 (CHO), 158.5 (ipso ArO'Pr), 146.9 (para ArCHO), 134.6 (ipso ArCHO), 131.7 (para ArO'Pr), 130.3 (ortho ArCHO), 128.5 (meta ArO'Pr), 127.0 (meta ArCHO), 116.2 (ortho ArO'Pr), 67.0 (OCH), 22.0 (OCH(CH₃)₂); m/z (%) (ESI+) 298 ([M+Na]⁺, 100), 290 ([M+NH₄]\ 57), 276 ([M+H]⁺, 42). (Found [M+H]⁺ 276.0794. Ci₅H₁₄CINO₂ requires [M+H]⁺ 276.0786).

4'-tert-butylbiphenyl-4-carbaldehyde [Yang et al J. Org. Chem. 2002, 67, 5057]

White powder (109 mg, 83%). R_f 0.50 (9% ethyl acetate-hexanes); Mp 103 ⁰C; δ_H (CDCI₃, 400 MHz) 10.05 (1H, s, CHO), 7.94 (2H, dd, J = 8.0, 2.0 Hz, ArH), 7.75 (2H, d, J = 8.0 Hz, ArH), 7.59 (2H, d, J = 8.0 Hz, ArH), 7.51 (2H, d, J = 8.0, 2.0 Hz, ArH)₁ 1.37 (9H, s, C(CHa)₃); δ_c (CDCI₃, 100 MHz) 191.9 (CHO), 151.7 (ipso A^Bu), 147.0 (para ArCHO), 136.7 (para Ar¹Bu), 135.0 (ipso ArCHO), 130.3 (ortho ArCHO), 127.4 (meta ArCHO), 127.0 (meta Ai^Bu)₁ 126.0 (ortho A^Bu)₁ 34.7 (C(CH₃)₃), 31.3 (C(CH₃)₃); m/z (%) (ESI+) 320 (96), 280 (60), 239 ([M+H]⁺, 100), 158 (70).

The biphenyl compounds were then combined with the appropriate N-substituted rhodanines in the same way as for the pyridine analogues described above. Thus, a solution of biaryl compound (100 mg), rhodanine (1.2 eq) and sodium acetate (2.4 eq) in glacial acetic acid (3 ml.) was heated to reflux for 4 h. The solution was concentrated under reduced pressure as orange solids which were redissolved in ethyl acetate (10 mL) and water (15 ml_). The aqueous phase was washed with ethyl acetate (2 x 10 mL) and the combined organic layers dried (MgSO4) and concentrated under reduced pressure. The crude yellow oil was purified by flash chromatography (0 ->• 7% methanol-DCM) to yield the following compounds: [DRD01] (S,Z)-2-(5-((3'-chloro-4'-isopropoxybiphenyl-4-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-4-methylpentanoic acid

Yellow solid (193 mg, 93%). R_f 0.29 (5% methanol-DCM); Mp 104.0-105.0 ⁰C; δ_H (CDCI₃, 400 MHz) 7.73 (1 H₁ s, SCCH), 7.65 (1 H, d, J = 2.0 Hz), 7.64 (2H, d, J = 8.5 Hz), 7.53 (2H, d, J = 8.5 Hz), 7.46 (1 H, dd, J = 8.5, 2.5 Hz), 7.02 (1 H, d, J = 8.5 Hz), 5.82 (1 H, d, J = 4.5 Hz, NCH), 4.67-4.58 (1 H, m, OCH), 2.34-2.27 (1 H, m, IxNCHCH₂), 2.18-2.12 (1H, m, IxNCHCH₂), 1.62-1.50 (1H, m, CH(CH₃)₂), 1-42 (6H, d, J = 6.0 Hz, 0(CHs)₂), 0.99 (3H, d, J = 6.5 Hz₁ CH(CH₃)₂), 0.94 (3H, d, J = 6.5 Hz, CH(CH₃)₂); *c (CDCI₃, 100 MHz) 192.9, 174.4 (CO₂H), 167.2 (NCO), 153.8, 141.8, 133.3, 132.7, 131.8, 131.3, 128.9, 127.3, 126.1 , 124.7, 121.4, 115.8, 72.2 (OCH), 55.8 (NCH), 36.8 (CH₂CH(CHs)₂), 25.2 (CH₂CH(CHs)₂), 22.9 (OCH(CH₃)₂), 22.1 (1xCH₂CH(CH₃)₂), 22.0 (1xCH₂CH(CH₃)₂); m/z (%) (ESI-) 502 ([M-H]^', 100), 458 (20). (Found [M-H]- 502.0992. C₂₅H₂₅CINO₄S₂ requires [M-H]^" 502.0914).

[DRD02] (2S,3R)-2-((Z)-5-((3'-chloro-4'-isopropoxybiphenyl-4-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-methylpentanoic acid

Yellow solid (154 mg, 84%). R, 0.31 (5% methanol-DCM); Mp 95.0-96.5 ⁰C; δ_H (CDCI₃, 400 MHz) 7.73 (1 H, s, SCCH), 7.64 (1 H, d, J = 2.0 Hz)₁ 7.63 (2H₁ d, J = 8.5 Hz), 7.53 (2H, d, J = 8.5 Hz), 7.46 (1 H₁ dd, J = 8.5, 2.5 Hz)₁ 7.02 (1 H, d, J = 8.5 Hz), 5.46 (1 H, d, J = 9.0 Hz, NCH), 4.63-4.59 (1H, m, OCH), 2.71-2.61 (1H, m, NCHCH), 1.41 (6H, d, J = 6.0 Hz, OCH(CH₃)₂), 1.33-1.27 (1H, m, IxCH₂CH₃), 1.24 (3H, d, J = 6.5 Hz, CHCH₃), 1.10-1.02 (1 H, m, IxCH₂CH₃), 0.86 (3H, t, J = 7.5 Hz); δ_c (CDCI₃, 100 MHz) 193.0 (CO₂H), 173.3 (NCO), 167.4, 153.9, 141.9, 133.5, 132.7, 131.9, 131.3, 128.9, 127.3, 126.1, 124.8, 115.8, 72.2 (OCH), 61.7 (NCH), 33.6 (NCHCH), 25.2 (CH₂CH₃), 22.0 (CH(CH₃J₂). 17.5 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI-) 502 ([M-H]^", 100), 458 (55). (Found [M-H]^' 502.0937. C₂₅H₂₅CINO₄S₂ requires [M-H]^" 502.0914). [DRD03] (2S,3R)-2-((Z)-5-((4'-isopropoxybiphenyl-4-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-methylpentanoic acid

Yellow solid (167 mg, 86%). R, 0.34 (5% methanol-DCM); Mp 82.0-84.0 ⁰C; δ_H

(CDCI₃, 400 MHz) 7.74 (1 H, s, SCCH), 7.65 (2H, d, J = 8.5 Hz), 7.55 (2H, d, J = 9.0 Hz), 7.52 (2H, d, J = 8.5 Hz), 6.97 (2H, d, J = 9.0 Hz), 5.46 (1 H, d, J = 9.0 Hz, NCH), 4.63-4.57 (1H, m, OCH), 2.72-2.61 (1 H, m, NCHCH), 1.36 (6H, d, J = 6.0 Hz, OCH(CHa)₂), 1.33-1.27 (1 H, m, IxCH₂CH₃), 1.24 (3H, d, J = 6.5 Hz, CHCH₃), 1.10- 1.03 (1 H, m, IxCH₂CH₃), 0.86 (3H, U J = 7.5 Hz, CH₂CH₃); δ_c (CDCI₃, 100 MHz)

193.0, 173.2 (CO₂H), 167.5 (NCO)₁ 158.3, 143.3, 133.8, 131.5, 131.4, 131.3, 128.2, 127.2, 126.4, 116.2, 70.0 (OCH), 61.7 (NCH), 33.6 (NCHCH), 25.2 (CH₂CH₃), 22.0 (CH(CHa)₂), 17.5 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI-) 468 ([M-H]^", 90), 424 (100). (Found [M-H]^" 468.1303. C₂₅H₂₆NO₄S₂ requires [M-H]- 468.1321).

[DRD04] (2S,3R)-2-((Z)-5-((4'-tert-butylbiphenyl-4-yl)methylene)-4-oxo-2- thioxothiazolidin-3-yl)-3-methylpentanoic acid

Yellow solid (182 mg, 93%). R_f 0.30 (5% methanol-DCM); Mp 112.0-113.0 ⁰C; δ_H (CDCI₃, 400 MHz) 7.75 (1H, s, SCCH), 7.70 (2H, d, J = 8.5 Hz), 7.57 (2H, d, J = 8.5 Hz), 7.54 (2H, d, J = 8.5 Hz), 7.49 (2H, d, J = 8.5 Hz), 5.47 (1H, U₁ J = 9.0 Hz, NCH), 2.72-2.61 (1 H, m, NCHCH), 1.36 (9H, s, C(CH)₃), 1.32-1.27 (1H, m, IxCH₂CH₃), 1.24 (3H, d, J = 6.5 Hz, CHCH₃), 1.10-1.03 (1 H, m, IxCH₂CH₃), 0.86 (3H, t, J = 7.5 Hz, CH₂CH₃); δ_c (CDCI₃, 100 MHz) 193.1 , 173.3 (CO₂H), 167.5 (NCO), 151.6, 143.5, 136.5, 133.7, 131.8, 131.3, 127.7, 126.8, 126.0, 121.1 (SCC), 61.7 (NCH), 34.6 (C(CH₃)₃), 33.6 (CHCH₃), 31.3 (C(CH₃)₃), 25.2 (CH₂CH₃), 17.6 (CHCH₃), 11.1 (CH₂CH₃); m/z (%) (ESI-) 466 ([M-H]-, 30), 422 (20), 161 (100). (Found [M-H]^" 466.1559. C₂₆H₂₈NO₃S₂ requires [M-H]^" 466.1511 ). The following table summarises the compounds made.

TABLE 1 - compounds synthesised

Biological Methods Cell lines

All cell culture components were purchased from Invitrogen (Carlsbad, CA). Human cancer cell lines: Breast (MCF-7), lung (NCI-H460) and CNS (SF268) were obtained from National Cancer Institute; Prostate (PC3 and DU 145) and Leukemia (K562) were purchased from ATCC (Manassas, VA). The human ovarian carcinoma cell lines (IA9) and its drug resistant mutant (epothilone-resistant, A8) are available from NCI and ATCC.

All the cell lines were cultured in RPMI supplemented with 5% fetal bovine serum (FBS) except IA9 and A8 cells were cultured in RPMI supplemented with 10% FBS, 2 mM glutamine, 50 units/ml streptomycin and penicillin.

Cell density determination

The sulforhodamine B (SRB) assay was used for cell density determination, based on the measurement of cellular protein content. Under mild acidic conditions, it binds to basic amino acid residues and under mild basic conditions it can be extracted from cells and solubilized for measurement. Results of the SRB assay were linear with cell number and cellular protein measured at cellular densities ranging from 1 to 200% of confluency.

Drug solubilization

All the compounds were solubilized in neat DMSO (Sigma Aldrich, St. Louis, MO) at

10 mM stock concentration and kept frozen at -80 ⁰C before use.

BcI-X₁ expression and purification

A vector modified from pET-32a containing the construct for human BcI-XL (from residues 1-218 with the flexible loop from residues 45 to 84 removed) as described by Zhang et al [Zhang et al., J. MoI. Biol. 2006, 364, (3), 536-549] was used for expression of wild-type Bcl-X_L and its mutants (F105A, L108A, E129A, L130A, R139A, A142G & Y173F). The DNA sequences of all the constructs were confirmed by BigDye sequencing. The expression, purification and thrombin cleavage of His- tagged Bcl-X_L protein were performed as described previously [Zhang et al 2006].

A vector modified from pET-32a containing the construct for mouse McI- 1 (from residues 147-308) with GST-tag was cloned and used for expression of wild-type mMcl-1 and its mutants (H205A, A208G, M212A, V230A, K236A, T247A & F251A). The DNA sequences of all the constructs were confirmed by BigDye sequencing. The GST-tagged protein was expressed in BL21 (DE3) Escherichia coli cells: the cells were grown at 37⁰C to an OD of 1.0 at 600 nm and then induction was done at 20 ⁰C for 12 hrs with 0.5 mM isopropyl-β-D-thiogalactopyranoside (IPTG) in rich medium and M9 minimal medium with ¹⁵N ammonium chloride (1gm/lit, Cambridge Isotope Lab.) and glucose (4 gm/lit) as the sole nitrogen and carbon sources, respectively. The expressed protein was purified using the procedure described for the purification of Bcl-X_L in the previous section except GST-mMcl-1 fusion protein was purified with glutathione-sepharose affinity chromatography using 1X PBS (pH 7.0) as a equilibration buffer and 50 mM Tris-HCI (pH 7.9) containing 10 mM reduced glutathione as a elution buffer.

Cytotoxicity - Pre-screeninq assay Cytotoxicity studies were performed using the sulphorhodamine B assay. Cytotoxicity of each drug was evaluated by the GI50 value, representing the 50% growth inhibition compared to non-treated control and a control at the time of drug addition (TO). In brief, cells were seeded on 96-well plates (Greiner, Frickenhausen, Germany) in 100 //I of culture medium (10,000, 10,000 and 5000 cells/well for MCF- 7, SF268 and NCI-H460, respectively). Twenty four hours later, 100 μ\ of medium containing 10 μM of desired compounds was added duplicate to the respective well. The plates were incubated for 48 hr at 37°C before fixing with 50% cold trichloroacetic acid (Sigma Aldrich, St. Louis, MO) for one hour after which the plates were washed five times with distilled water. The plates were then air-dried at room temperature. The fixed cells were stained with 100 μ\ of 0.4% (w/v) SRB (Sigma Aldrich, St. Louis, MO) in 1% acetic acid for 10 min. Excess SRB was removed by washing the plates four times with 1% acetic acid. After drying, 100 μ\ of 10 mM Tris base (pH 10.5) were added to solubilize the protein bound SRB and mixed. The absorbance was measured at 515 nm using a Versamax microtitre plate reader (Molecular Devices) and GI50 was calculated from 5 dosage responses using Softmax®Pro 3.1 software based on point to point plot. Percentage of net growth was calculated as below:

If TO >T, % of net growth = ((T-TOy(C-TO)) * 100 If TO < T, % of net growth = ((T-TO)HO) * 100

T is the optical density of the test well after a 48-hour drug exposure. TO is the optical density at time zero, and C is the control optical density after 48 hours.

Positive control: Paclitaxel and Doxorubicin (Sigma Aldrich) Negative control: Cells without drug addition

Compounds exhibiting % of net growth <50% in the pre-screening assay were proceeded for Gl₅₀ determination.

Cytotoxicity - GIm determination

The protocol is the same as the pre-screening assay except that 100 μ\ of media containing five different concentrations (0.001 μM, 0.01 μM, 0.1 μM, 1 μM and 10 μM) of the desired compounds were added to the respective well. The seeding densities for the respective cell lines are as followed: PC3 (7500), DU 145 (7500),

K562 (5000), IA9 (5000) and A8 (10,000).

Competitive binding - Fluorescence Polarization Assays

The Bak-BH3 peptide labeled with fluorescein at the N terminus was synthesized by Mimotopes (Clayton, Victoria, Australia) and purified by HPLC. The peptide was dissolved in DMSO at 1 mM, and stock solutions of the test compounds (4mM in DMSO) were used for serial dilutions (250 μM to 0.65μM final concentrations). The reaction was carried out in a total volume of 100 μL/well containing 3 μg glutathione S-transferase (GST)-hBcl_XLΔC19 or 3 μg glutathione S-transferase (GST)-hMcl- 1ΔC20 and 60 nM labeled peptide in assay buffer (50 mM Tris, pH 8, 150 mM NaCI and 0.1% bovine serum albumin). To each well was added 10 μL of the test compounds, and the reaction mixture was incubated at rt for 1 h. The fluorescence polarization values were determined using Tecan GeniosPro plate reader using the excitation/emission wavelengths 485/535 nm.

Isothermal titration calorimetry (ITC) ITC experiments of the binding of ligands to proteins (Bcl-X_L and mMcl-1 ) were performed by VP-ITC (Microcal, USA) at 25 ⁰C, pH 7.0. In all ITC experiments, the BCI-X_L or mMcl-1 dissolved in 20 mM phosphate buffer at a concentration of 50 μM were taken into the sample cell and the compounds were dissolved in 20 mM phosphate buffer containing 1% DMSO (v/v) at a concentration of 1mM were taken into the syringe compartment of the calorimetry.

The reference cell was filled with 20 mM phosphate buffer. All solutions were degassed before usage. After thermal equilibration, aliquots of 10 μ\ of ligand solution were added into the sample cell containing protein solution, which was stirred constantly at 320 rpm. After each injection, the change in the heat of interaction was monitored by the VP-ITC and the data were processed and analyzed by Origin 5.0. All sample data obtained after appropriate blank corrections were fitted to various model equations to choose right binding model which presumably yields exact thermodynamic parameters of protein-ligand interactions. For blank ITC experiments, the sample and reference cells were filled with 20 mM phosphate buffer and the ligand solution was added from syringe into sample cell at the conditions which were identical to the conditions mentioned above for the protein samples.

Biological Data Cytotoxicity

All of the compounds tested exhibited activity. The results for specific compounds are set out in Tables 2 and 3 below.

TABLE 2 - cytotoxicity (prescreening) data

TABLE 3 - cytotoxicity data

FPA Data

PRD and DRD compounds were screened using FPA against the fluorescein-labeled Bak-peptide and the proteins Bcl-X_L and Mcl-1. The IC₅₀ values are summarized in Table 4. Whilst some active compounds exhibited low or no detectable binding at the concentrations tested, others exhibited very significant levels of binding. The structural variations between and within each series of compounds comprise the substitution pattern on the aryl ring system and the substitution on the Λ/-atom of the rhodanine.

TABLE 4 - FPA data

Within the series PRD01 to PRD06, where the aryl ring system is the 2,3- dimethoxyphen-1-yl group, PRD01 and PRD02 do not displace the Bak peptide from the BCI-X_L protein at the concentrations tested. The values for the valine, leucine and isoleucine derivatives PRD03 to PRD05 show comparable activity to each other, while the phenylalanine derivative PRD06 showed a lower IC₅₀ value against the BcI- X_L protein. Similar displacement studies with the Mcl-1 protein showed that all the compounds in the series PRD01 to PRD06 were active with IC₅₀ values ranging from 115μM to 18 μM, with PRD06 being the best inhibitor of both Bcl-X_L and Mcl-1.

For the series PRD07 to PRD12, where the aryl ring system is retained as the 3,4- dioxolobenzene, the inhibitory profile for the Bcl-X_L protein is similar with that observed with PRD01 to PRD06. For PRD07 no significant binding was observed at the concentrations studied but in displacement studies with Mcl-1 , rhodanine derivatives PRD08 to PRD12 have IC₅₀ values ranging from 138 to 21 μM.

The series PRD13 to PRD18 is similar to PRD01 to PRD06 with the exception of the methoxy group at the R₄ rather than R₂ position. PRD 16 and PRD 17 are moderately active against Bcl-X_L. Compound PRD18 has one of the lowest IC₅₀ values against Mcl-1 (22 μM), which indicates that it may be a selective Mcl-1 inhibitor.

The series of compounds PRD19 to PRD24 contain the 3-chloro-4-isopropylbenzene as the aryl group. Four compounds display significant activity against Bcl-X_L, with the leucine and isoleucine derivatives (PRD22 and PRD23) being the most active (IC₅₀=33 and 32 μM respectively). These compounds are all active against Mcl-1 (IC₅₀=13 to 63 //M).

The inhibition studies with Bcl-X_L and Mcl-1 show that some of the compounds synthesized show preferential selectivity for Mcl-1 over BCI-X_L. A number of the novel rhodanines are inhibitors of both Bcl-X_L and Mcl-1 (e.g. PRD03 to PRD06, PRD09 to PRD12, PRD16 to PRD17 and PRD21 to PRD24). The IC₅₀ values for some of these rhodanines, as measured against the Bak peptide, are lower than the ICs₀ values obtained for the known inhibitor BH3I-1. Some of the active pyridylrohodanines are Mcl-1 selective inhibitors with IC₅₀ values between 22 μM to 81 μM (e.g. PRD02, PRD15, PRD18, PRD19 and PRD20).

The potent Mcl-1 inhibitors may be important as drugs on their own, or as an effective co-therapeutic with Bcl-X_L specific inhibitors.

Binding Constants

Investigation of the protein-ligand interactions was carried out for some of the compounds using isothermal calorimetry (ITC) to validate the binding affinities of

BH3I-1 , PRD06 and PRD18 to the respective proteins. The binding affinities of BH3I- 1 , PRD06 and PRD18 to Bcl-X_L and Mcl-1 were investigated by monitoring the changes in the heat of binding of the complexes using VP-ITC (Microcal, USA) at pH

7.0, 25 ⁰C.

Figures 2A to 2C show the heat released upon titration of Bcl-X_L BH3I-1 , PRD06 or PRD18 as a function of the ligand to the protein ratio. The data is best fitted to a 2- site sequential binding model, but in general the first dissociation constant K₁ is ten to a hundred times smaller than the second dissociation constant K₂. Due to the large difference in magnitude between K₁ and K₂, the K₂ values can be ignored and the results can be interpreted in light of a single-site binding model where K₁= K_d. The dissociation constant for BH3I-1 was estimated at K_d = 9 μM against Bcl-X_L. This value is similar to the literature value previously reported (IC₅₀=5.86 μM against FITC-Bid).

For PRD06, the dissociation constant was measured to be K_d = 3.4 μM with BCI-X_L. Both K_d values measured by ITC are ten times lower than the IC₅₀ value obtained by FPA using Bcl-X_L and the labeled Bak peptide. The dissociation constant estimated for Bcl-X_L and PRD18 was higher than 750 μM, and an accurate value could not be obtained due to the higher fitted-error associated with the data. This observation is due to a very small change in the heat of binding of the Bcl-X_L- PRD18 complex indicating that the binding affinity is weak. Overall, the results suggest that the binding affinity of PRD06 to the protein is roughly twice that of BH3I-1 , while PRD18 does not bind so strongly to Bcl-X_L. With respect to Mcl-1 (data not shown), the ITC of BH3I-1 showed a poor thermal response. This would indicate that BH3I-1 binds weakly to Mcl-1. In contrast, PRD06 binds with a K_d= 16 μM. Compound PRD18 on the other hand has a weaker binding affinity of K_d=26 μM. The binding constants obtained via ITC measurements using Mcl-1 are in excellent agreement with the values obtained by FPA.

The results show that the binding affinities obtained by ITC closely mirror the results obtained by FPA. Both FPA and ITC show that compound BH3I-1 binds to Bcl-X_L as previously reported, but in contrast does not seem to bind to Mcl-1 as earlier reported. The FPA and ITC results also show that PRD06 binds both Bcl-X_L and McI- 1 at low μM concentrations (K_d=3.4 μM and 16 μM respectively). Furthermore, the ITC results verify that PRD18 binds to Mcl-1 (K_d=26 μM).

To verify that the protein-ligand complexes were stable under these conditions and were not uncoiled, the apparent free energy values of the unbound proteins and the protein-ligand complexes were examined by GdnHCI-induced denaturation processes using fluorescence spectrometry [data not shown]. It was noted that the differences in the free energy of the free protein and its complexes were negligible by accounting the fitted-error associated with the data, which implies that the structure of the protein Bcl-X_L and Mcl-1 as well as their respective complexes were indeed stable.

Figures 3A and 3B show the results of the WST Cytotoxicity Assay in which A549 cancer cells were treated with varying concentrations of the test compounds.

Thus, Figures 3A and 3B illustrate the cytotoxicity of the compounds against A549 human alveolar basal epithelial cancer cells as determined in the WST Assay. The library of lead compounds were also screened against cancer cells which over- express Bcl-X_L and Mcl-1 and the compounds were found to induce apoptosis in these cells. The cytotoxicity of these compounds was found to be better than that of BH3I-1.

It is to be understood that variants of the above described examples of the invention in its various aspects, such as would be readily apparent to the skilled person, may be made without departing from the scope of the invention in any of its aspects.

Claims

1. A compound selected from compounds of formula I, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula I wherein:

X¹ is independently selected from CH₂ and C=S;

one of X² and X³ is N and the other is CH;

R¹ is independently selected from H or R^N, wherein R^N is independently branched or unbranched saturated or unsaturated C_1-20alkyl and is optionally substituted;

each of R^A and R^B is independently selected from H, Ci_-2oalkyl, C_1-20alkoxy, C_3-20aryl, C^oaryl-C^alkyl, C_3-2oheterocyclyl, halo, amino, OH or R^A and R^B together with the ring atoms to which they are attached form C_3-7heterocyclyl, and is optionally substituted;

n is independently 0 to 5;

and is a single or double bond.

2. A compound according to claim 1 , wherein R¹ is independently R^N, and R^N is independently branched or unbranched C_1-20alkyl or C_3-2oaryl-C_1-20alkyl and is substituted with R^AN, wherein R^AN is an acid group or an ester group, and is optionally further substituted.

3. A compound according to claim 2, wherein R^N is:

wherein

R^AN is an acid group or an ester group, preferably selected from -C(O)OR^E, - P(O)(OR^E)₂, -SO₃R^E and -B(OR^E)₂; wherein each R^E is independently selected from H, C_1-7alkyl and C_3-12aryl; and

R^Nα is selected from H, branched or unbranched, saturated or unsaturated C₁. ioalkyl and C_5-6aryl-C_1-10alkyl, and is optionally substituted.

4. A compound according to any one of claims 1 to 3, wherein X¹ is C=S, n is 0, and is a double bond.

5. A compound according to any one of the preceding claims, wherein X² is N and X³ is CH.

6. A compound according to any one of the preceding claims, wherein R², R³ and R⁴ are selected as follows: (a) R² is C_1-2QaIkOXy, R³ is C_1-2oalkoxy and R⁴ is H; or (b) each of R³ and R⁴ is independently C_1-20alkyl, C_1-2oalkoxy, halo or together form a -0-CH₂-O- group, and R² is H; and wherein each of R⁵, R⁶, R^A and R^B is H.

7. A compound according to any one of the preceding claims, wherein the compound is selected from:

8. A compound selected from compounds of formula VIII, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula VIII wherein:

each of X¹, R¹, R^A, R^B and n is independently as defined in any one of claims

1 to 6; and

R^D is independently C_3-20aryl, preferably phenyl, and is optionally substituted.

9. A compound according to claim 8, wherein the compound is selected from compounds of formula IX, and pharmaceutically acceptable salts, hydrates, and solvates thereof:

Formula IX

wherein each of R jA , R DB , D R2 , n R3^ό, D R4 , π R5^s, R is as defined in any one of claims 1 to 7; and

R^N is independently selected from branched or unbranched saturated or unsaturated Ci_-2oalkyl and C_3-2Oaryl-Ci-₂oalkyl, and is substituted with R^AN, wherein R^AN is an acid group or an ester group, and is optionally further substituted.

10. A compound according to claim 9, wherein R^N is:

wherein

R^AN is an acid group or an ester group, preferably selected from -C(O)OR^E, - P(O)(OR^E)₂, -SO₃R^E and -B(OR^E)₂; wherein each R^E is independently selected from H, C^alkyl and C_3-i₂aryl; and

R^Nα is selected from H, branched or unbranched, saturated or unsaturated C_1- ioalkyl and C_5-6a ry 1-C_1-1Oa I kyl and is optionally substituted.

11. A compound according to any one of claims 8 to 10, wherein R², R³ and R⁴ are selected as follows: (a) R² is C_1-2QaIkOXy, R³ is C_1-20alkoxy and R⁴ is H; (b) each of R³ and R⁴ is independently C_1-2oalkyl, Ci-₂oalkoxy, halo or together form a -0-CH₂-O- group, and R² is H; and wherein each of R⁵, R⁶, R^A and R^B is H.

12. A compound according to any one of claims 8 to 11 , wherein the compound is selected from:

13. A compound according to any one of the preceding claims for use in a method of treatment of the human or animal body by therapy.

14. A compound according to any one of claims 1 to 12 for use in a method of treatment of cancer.

15. A method of treating cancer using a compound according to any one of claims 1 to 12.