Blood, 1 March 2002, Vol. 99, No. 5, pp. 1503-1503
Sugar in erythropoietin: clinical and forensic implications
Most proteins secreted into the plasma are heavily
glycosylated. Complex branched polysaccharides are N-linked
to certain asparagine residues and O-linked to certain
serine or threonine residues. The carbohydrate content and structure of
plasma proteins are important determinants of their half-lives in the
circulation. The cleavage of terminal sialic residues generally results
in the rapid clearance of the protein by "asialo" receptors in
the liver.
Erythropoietin (EPO) contains 40% carbohydrate, most of which is
attached to 3 Asn sites. In the December 15, 2001 issue of Blood, Skibeli and colleagues (Blood. 2001;98:3626-3634) report on
rather striking differences in the carbohydrate structure of endogenous
plasma EPO versus that of recombinant human erythropoietin (rhEPO)
produced by high-level expression of the human EPO
gene in hamster cells. They show that 3 different commercial
preparations of rhEPO (alfa, beta, and omega) have a higher molecular
weight than endogenous plasma EPO and a high content of fully
sialylated tetra-antennary glycans versus none in plasma EPO.
The analysis of endogenous EPO was done on protein purified from the
plasma of 2 patients with aplastic anemia. The differences in
carbohydrate structure between endogenous and rhEPO are likely due to
the lack of certain monosaccharide transferases in the hamster rather
than modification of rhEPO during circulation in vivo.
These differences in carbohydrate structure are likely to have
important clinical consequences. The development of rhEPO is arguably
the most successful therapeutic application of recombinant DNA
technology. Few pharmaceutical agents can match rhEPO's
combination of efficacy and safety. However, there is growing concern
about case reports of red cell aplasia developing in patients on
chronic rhEPO therapy. The neutralizing antibody that is induced by the recombinant product cross-reacts with endogenous EPO, thereby markedly
suppressing erythropoiesis. Because the only structural difference lies
in the carbohydrate, it is possible that this is the antigenic stimulus
responsible for this rare but serious complication. In this report
Skibeli and colleagues demonstrate subtle but significant differences
in the carbohydrate structures of rhEPO alfa, beta, and omega. It will
be of interest to learn whether these correlate with antigenicity and
development of erythroid aplasia. Recently Amgen has developed a
superglycosylated rhEPO, Aranesp, that has a markedly prolonged
half-life in the circulation and therefore can be administered less
frequently. In view of the possible relationship between carbohydrate
structure and antigenicity, there will be heightened awareness
regarding the development of erythroid aplasia in patients treated with
this promising new agent.
In recent years there have been an alarming number of reports of sudden
deaths among athletes who have "doped" themselves with rhEPO in
order to improve their performance in competition. Prevention of this
illegal practice is thwarted by the difficulty in detecting
surreptitious use of rhEPO. Radioimmune and ELISA assays in current
clinical use cannot distinguish endogenous EPO from rhEPO. Moreover,
because the half-life of rhEPO is approximately 5 hours, the levels of
plasma EPO will be normal the day following self-administration.
The measurement of reticulocytes or surface markers of a cohort of
young erythrocytes offers a longer window of time in which the abuse of
rhEPO can be detected. Alternatively, the development of a sensitive
immunologic assay that can distinguish rhEPO from endogenous EPO would
also enable the detection of surreptitious drug use.
H. Franklin Bunn
Harvard Medical
School
