DƯỢC LÍ Goodman & Gilman's The Pharmacological Basis of Therapeutics 12th, 2010

new agents have greater capacity for transport into tumor cells
(pralatrexate) and exert their primary inhibitory effect on TS
(raltitrexed, TOMUDEX), early steps in purine biosynthesis (lometrexol),
or both (the multitargeted antifolate, pemetrexed, ALIMTA)
(Vogelzang et al., 2003; O’Connor et al., 2007).
Aside from its antineoplastic activity, methotrexate also has
been used with benefit in the therapy of psoriasis (see Chapter 65).
Additionally, methotrexate inhibits cell-mediated immune reactions
and is employed to suppress graft-versus-host disease in allogenic
bone marrow and organ transplantation and for the treatment of dermatomyositis,
rheumatoid arthritis, Wegener’s granulomatosis, and
Crohn’s disease (see Chapters 35 and 47).
Structure-Activity Relationship. The primary target of
methotrexate is the enzyme DHFR (Figure 61–5).
Inhibition of DHFR leads to partial depletion of the FH 4
cofactors (5-10 methylene tetrahydrofolic acid and N-10
formyl tetrahydrofolic acid) required for the respective
synthesis of thymidylate and purines. In addition,
methotrexate, like its physiological counterparts (the
folates), undergoes conversion to a series of polyglutamates
(MTX-PGs) in both normal and tumor cells. These
MTX-PGs constitute an intracellular storage form of
folates and folate analogs and dramatically increase
inhibitory potency of the analog for additional sites,
including TS and two early enzymes in the purine biosynthetic
pathway (Figure 61–5). Finally, the dihydrofolic
acid polyglutamates that accumulate in cells behind the
blocked DHFR reaction also act as inhibitors of TS and
other enzymes (Figure 61–5) (Allegra et al., 1987b).
Because folic acid and many of its analogs are polar, they
cross the blood-brain barrier poorly and require specific transport
mechanisms to enter mammalian cells. Three inward folate transport
systems are found on mammalian cells: 1) a folate receptor,
which has high affinity for folic acid but much reduced ability to
transport methotrexate and other analogs; 2) the reduced folate transporter,
the major transit protein for methotrexate, raltitrexed, pemetrexed,
and most analogs (Westerhof et al., 1995); and 3) a
transporter that is active at low pH. The importance of transport in
determining drug sensitivity is illustrated by the finding that the
reduced folate transporter is highly expressed in the hyperdiploid
subtype of acute lymphoblastic leukemia, due to the presence of
multiple copies of chromosome 21, on which its gene resides; these
cells have extreme sensitivity to methotrexate (Pui et al., 2004). Once
in the cell, additional glutamyl residues are added to the molecule by
the enzyme folylpolyglutamate synthetase. Intracellular methotrexate
polyglutamates have been identified with up to six glutamyl
residues. Because these higher polyglutamates are strongly charged
and cross cellular membranes poorly, if at all, polyglutamation
serves as a mechanism of entrapment and may account for the prolonged
retention of methotrexate in chorionic epithelium (where it is
a potent abortifacient); in tumors derived from this tissue, such as
choriocarcinoma cells; and in normal tissues subject to cumulative
drug toxicity, such as liver. Polyglutamylated folates and analogs
have substantially greater affinity than the monoglutamate form for
folate-dependent enzymes that are required for purine and thymidylate
synthesis and have at least equal affinity for DHFR.
New folate antagonists that are better substrates for the
reduced folate carrier have been identified. In efforts to bypass the
obligatory membrane transport system and to facilitate penetration
of the blood-brain barrier, lipid-soluble folate antagonists also have
been synthesized. Trimetrexate (NEUTREXIN), a lipid-soluble analog
that lacks a terminal glutamate, was one of the first to be tested for
clinical activity. The analog has modest antitumor activity, primarily
in combination with leucovorin rescue. However, it is beneficial
in the treatment of P. jiroveci (Pneumocystis carinii) pneumonia
(Allegra et al., 1987a), where leucovorin provides differential rescue
of the host but not the parasite.
The most important new folate analog, MTA or pemetrexed
(ALIMTA) (Figure 61–6), is a pyrrole–pyrimidine structure. It is avidly
transported into cells via the reduced folate carrier. It readily is converted
to polyglutamates that inhibit TS and glycine amide ribonucleotide
transformylase, as well as DHFR. It has shown activity
against ovarian cancer, mesothelioma, and non–small cell adenocarcinomas
of the lung (Vogelzang et al., 2003).
Inhibitors of DHFR differ in their relative potency for blocking
enzymes from different species. Agents have been identified that
have little effect on the human enzyme but have strong activity
against bacterial and parasitic infections (see discussions of
trimethoprim, Chapter 52, and pyrimethamine, Chapter 49). By contrast,
methotrexate effectively inhibits DHFR in all species investigated.
Crystallographic studies have revealed the structural basis for
the high affinity of methotrexate for DHFR (Blakley and Sorrentino,
1998) and the species specificity of the various DHFR inhibitors.
Mechanism of Action. To function as a cofactor in onecarbon
transfer reactions, folate must first be reduced by
DHFR to FH 4
. Single-carbon fragments are added enzymatically
to FH 4
in various configurations and then may
be transferred in specific synthetic reactions. In a key
metabolic event catalyzed by TS (Figure 61–5), deoxyuridine
monophosphate (dUMP) is converted to thymidine
monophosphate (TMP), an essential component of DNA.
In this reaction, a one-carbon group is transferred to
dUMP from 5,10-methylene FH 4
, and the reduced folate
cofactor is oxidized to dihydrofolate (FH 2
). To function
again as a cofactor, FH 2
must be reduced to FH 4
by
DHFR. Inhibitors such as methotrexate, with a high affinity
for DHFR (K i
~0.01-0.2 nM), prevent the formation of
FH 4
and allow an accumulation of the toxic inhibitory
substrate, FH 2
polyglutamate, behind the blocked reaction.
The absence of reduced folates shuts down the onecarbon
transfer reactions crucial for the de novo synthesis
of purine nucleotides and thymidylate and interrupts the
synthesis of DNA and RNA. The toxic effects of
methotrexate may be terminated by administering leucovorin,
a fully reduced folate coenzyme, which repletes the
intracellular pool of FH 4
cofactors.
As with most antimetabolites, methotrexate is
only partially selective for tumor cells and kills rapidly
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CHAPTER 61
CYTOTOXIC AGENTS