HPCs can be cryopreserved at -80 degrees C with 5-percent DMSO and stored at -80 degrees C no longer than 6 months. A 5-percent DMSO concentration is comparable to a with 10-percent concentration in terms of recovery and MNC viability.

We report the long-term evaluation over 12 years of a simplified technique for stemcell cryopreservation at -80ºC without rate-controlled freezing and with 5% (n=251) or 10% (n=47) DMSO as the sole cryoprotectant. Platelet recovery was greater in the 5% DMSO group while long-term hematologic recovery did not differ. Factors influencing a faster hematologic recovery were infusion of more than 2.7×10 6 /kg of CD34 + cells, 10% DMSO cryopreservation and G-CSF. We confirm that the procedure is feasible with a reduction in infusion-related toxicity from 60% using 5% DMSO. Differences in hematologic reconstitution were not clinically significant if a minimum of 1. 1 The toxic effects related to DMSO infusion are generally dose-related and while they are usually mild, they can become severe.2-4 HES is a relatively non-toxic drug but it is related with long-lasting pruritus 5 and osmotic nephrotoxicity. Cryopreservation protocols usually involve ratecontrolled freezing followed by the storage of the HSC in either the liquid or vapor phase of liquid nitrogen. These procedures are time consuming and require expensive computerassisted devices.

The clinical significance of resistance/intolerance to hydroxycarbamide (HC) was assessed in a series of 890 patients with polycythaemia vera (PV). Resistance/intolerance to HC was recorded in 137 patients (15·4%), consisting of: need for phlebotomies (3·3%), uncontrolled myeloproliferation (1·6%), failure to reduce massive splenomegaly (0·8%), development of cytopenia at the lowest dose of HC to achieve a response (1·7%) and extra-haematological toxicity (9%). With a median follow-up of 4·6 years, 99 patients died, resulting in a median survival of 19 years. Fulfilling any of the resistance/intolerance criteria had no impact on survival but when the different criteria were individually assessed, an increased risk of death was observed in patients developing cytopenia [Hazard ratio (HR): 3·5, 95% confidence interval (CI): 1·5-8·3, P = 0·003]. Resistance/intolerance had no impact in the rate of thrombosis or bleeding. Risk of myelofibrotic transformation was significantly higher in those patients developing cytopenia (HR: 5·1, 95% CI: 1·9-13·7, P = 0·001) and massive splenomegaly (HR: 9·1, 95% CI: 2·3-35·9, P = 0·002). Cytopenia at the lowest dose required to achieve a response was also an independent risk factor for transformation to acute leukaemia (HR: 20·3, 95% CI: 5·4-76·5, P < 0·001). In conclusion, the unified definition of resistance/intolerance to HC delineates a heterogeneous group of PV patients, with those developing cytopenia being associated with an adverse outcome.

This simplified cryopreservation technique will be useful for institutions without rate-controlled freezing facilities. Moreover, this method diminishes the amount of DMSO infused to patients, as well as its toxicity.

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