Throughout this specification, including any 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 any 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.
Mammalian cells are normally subject to tight controls regulating replication in order to maintain organ structure and function. Conversely, the disease of cancer is characterized by uncontrolled proliferation. Compromise of any of the steps involved in cell cycle regulation could be involved in escape from regulatory mechanisms and therefore lead to neoplasia. However, even if a cell escapes proliferation suppression, there are limitations to the number of replicative cycles it can progress through before safety mechanisms cause cell cycle shutdown, and this restriction is thought to be a component of the process of organismal aging. Although aging is a complex process, a major candidate for the molecular signal for replicative senescence is that of telomere shortening.
Telomeres are nucleoprotein structures at the ends of linear chromosomes consisting of DNA sequences arranged in tandemly repeated units which extend from less than 100 to several thousands of bases. In contrast to chromosome ends created by random breakage, telomeres are stable structures not prone to degradation or fusion with other chromosome ends and are not subject to DNA repair mechanisms.
During each round of cellular replication, both strands of DNA separate and daughter strands are synthesized in a slightly different manner on the leading and lagging strand. While the lead strand replicates in a continuous fashion using conventional DNA polymerase, the lagging strand replicates in a discontinuous fashion using Okazaki fragments. The gaps between individual Okazaki fragments are filled by the regular DNA polymerase. However, this sets the stage for a potential “end replication problem.” This arises because Okazaki fragment priming will not necessarily start at the very end of the DNA and because the RNA primer, once removed, would result in a portion of unreplicated 3′-DNA (an unrepaired 3′-overhang). This can lead to a loss of 50–200 base pairs with every round of somatic cell division, with eventual shortening of telomeres to a length that coincides with the activation of an antiproliferative mechanism termed “mortality stage 1” (M1), and at this stage, senescence in somatic cells occurs. Thus, telomere shortening functions as a “mitotic clock” and limits division in somatic cells to about 50–70 times, thereby contributing to cell aging.
In some cells, due to various mechanisms, the M1 stage is bypassed and cells can continue to divide until telomeres become critically shortened (“mortality stage 2,” M2). At this M2 stage, in many immortalized cells, a specialized DNA polymerase called “telomerase” appears and utilizes its internal RNA template to synthesize the telomeric sequence and compensate for the loss of telomeric DNA due to incomplete replication. This prevents further shortening of telomeres, and the resulting stabilization of their length contributes to immortalization.
Telomerase is not expressed, or if it is, its activity is repressed, in most normal mammalian somatic cells. Exceptions to this rule include male germ line cells and some epithelial stem cells (e.g., as in the intestinal crypts, the basal layer of the epidermis, and within human hair follicles). Nonetheless, both telomerase activity and shortened but stabilized telomeres have been detected in the majority of tumours examined (and in over 90% of all human cancers examined), and consequently, telomeres and telomerase are recognized targets for anti-neoplastic (e.g., cancer) chemotherapy.
The absence of telomerase in most normal cells makes this enzyme a particularly attractive target, considering that its inhibition would probably cause minimal damage to the whole patient. The fact that tumour cells have shorter telomeres and higher proliferation rates than normal replicative cell populations suggests that a therapeutic telomerase inhibitor may cause tumour cell death well before damage to regenerative tissues occurs, thereby minimizing undesirable side-effects.
For a more detailed discussion of telomeres and telomerase, and their role as anti-proliferative targets, see, for example, Sharma et al., 1997; Urquidi et al., 1998; Perry et al., 1998c; Autexier, 1999; Neidle et al., 1999; 2001; Rezler et al., Parkinson et al., 2002; Neidle et al., 2002, Gowan et al., 2002; Herbert et al., 2002; Shay et al., 2002; Mergny et al., 2002; Corey et al., 2002; and references therein.
A range of telomerase inhibitors have been investigated. See, for example, Carrasco et al., 2002; Alberti et al., 2002; Riou et al., 2002; Kim et al., 2002; Kern et al., 2002; Gomez et al., 2002.
A number of polycyclic compounds, including polycyclic acridines, anthraquinones, and fluorenones have been shown to inhibit telomerase and/or to have anti-tumour effects in vitro. See, for example, Bostock-Smith et al., 1999; Gimenez-Arnau et al., 1998; Gimenez-Arnau et al., 1998; Hagan et al., 1997; Hagan et al., 1998; Harrison et al., 1999; Julino et al., 1998; Perry et al., 1998a, 1998b, 1999a, 1999b; Sun et al., 1997.
Harrison et al., 1999, describe certain 3,6-disubstituted acridines which are shown to inhibit telomerase, and to inhibit cell growth in certain ovarian carcinoma cell lines.

Read et al., April 2001, describe certain 3,6-disubstituted and 3,6,9-trisubstituted acridines (see compounds 1, 2, 3, and 4 in FIG. 1 therein), shown below (where n is 4 or 7, and R is —H or -Me). These compounds were shown to have potent in vitro inhibitory activity against human telomerase.

Neidle et al., 2002, describe various disubstituted acridines and trisubstituted acridines, shown below, which inhibit telomerase and/or regulate cell proliferation.

Lorente et al., 1996, describe the synthesis of certain 3,6-disubstituted acridines (which are also 9-H), shown below (wherein R′ is —H or —Boc and R is —H or —CPh3), which apparently are catalytically active and cleave t-RNA.

I.G. Farbenindustrie Akt.-Ges in Frankfurt a.M., 1930, describes one 3,6-disubstituted acridine (which is also 9-H), shown below.

Although many are known, there remains a great need for potent telomerase inhibitors, anticancer agents, antiproliferative agents, etc. particularly for such compounds which offer one or more additional pharmacological advantages, such as:                (a) improved activity.        (b) improved selectivity (e.g., against tumour cells versus normal cells).        (c) complement the activity of other treatments (e.g., chemotherapeutic agents);        (d) reduced intensity of undesired side-effects;        (e) fewer undesired side-effects;        (f) simpler methods of administration;        (g) reduction in required dosage amounts;        (h) reduction in required frequency of administration;        (i) increased ease of synthesis, purification, handling, storage, etc.;        (j) reduced cost of synthesis, purification, handling, storage, etc.        
For example, particularly preferred telomerase inhibitors are ones which are characterized by one or more of the following properties:                (a) no inhibition of Taq polymerase at 10–50 μM (in order to provide specificity and eliminate broad-spectrum polymerase inhibitors);        (b) cell free telomerase inhibition (at <1 μM) at concentrations more than 5 to 10-fold less than for concentrations for acute cytotoxicity;        (c) shortening of telomere length in tumour cells at concentrations 5 to 10-fold less than concentrations for acute cytotoxicity;        (d) telomere shortening in human tumour xenografts; and,        (e) oral bioavailability.        
Thus, one aim of the present invention is the provision of compounds which are telomerase inhibitors, anticancer agents, antiproliferative agents, etc. which offer one or more of the above advantages and properties.
USPTO Backgrounds
USPTO Backgrounds