Journal of Clinical Apheresis 7:129-131 (1992)

Peripheral Blood Stem Cell Mobilization by Cytoki nes

Tauseef Ahmed, David Wuest, and David Ciavarella

New York Medical College (T.A., D. W., D.C.) and Hudson Valley Blood Services, Valhalla, New York (D. W., D.C.)

Hematopoietic stem cells circulate in the peripheral blood. These cells can be collected by apheresis techniques either in the unperturbed state, after mobilization following the administration of cytokines like G-CSF o r GM-CSF, or during the phase of early blood count recovery following chemotherapy-induced myelosuppression. The number of cells collected following mobilization is greater than that obtained after apheresis in the unperturbed state. There are, however, qualitative differences between unperturbed and mobilized cells. Chemotherapy related mobilization can be potentially dangerous in that severe myelosuppression necessary to achieve mobilization can have serious consequences. There are no controlled studies that evaluate the relative merits of each method of collection. Regardless of the techniques employed peripheral blood stem cells can reliably accelerate hematologic recovery after potentially myeloblative therapy and provide an alternative to bone marrow support. 0 1992 Wiley-Liss, Inc.

Key words: peripheral stem cells, apheresis techniques, marrow transplantation

INTRODUCTION

Hematopoietic stem cells circulate in the peripheral blood [ 1,2]. They can be damaged or destroyed during the course of aggressive chemotherapeutic regimens. The storage and subsequent reinfusion of these cells allows for dose intensification with chemotherapeutic agents 14-61, In recent years methods of collection and storage of stem cells have been improved 17-91. Hema- topoietic stem cells harvested from peripheral blood via cytapheresis are now used frequently during the setting of autologous bone marrow transplantation. Since pe- ripheral stem cells have a higher proportion of committed progenitors, they are often used to further accelerate he- matologic recovery after potentially myeloablative che- motherapeutic regimens and autologous marrow trans- plantation [ 10,l I] . Peripheral stem cells are also used when marrow contamination with tumor cells is likely or marrow collection is difficult, e .g . , in cases where radiation has been given to the pelvis [4].

The number of stem cells in the peripheral blood is minuscule compared to other cells [ 121. Unperturbed stem cells can be collected by using apheresis tech- niques. The number of collections and the amount of blood processed during each procedure may have to be quite large [ 12,131, and a fair amount of time may be expended to achieve an adequate collection. Hence, dif- ferent techniques that result in enrichment of the periph- eral blood with progenitors, and thus faster collection, are often employed. Prominent among these are stem cell collections during blood count recovery from che- motherapy-induced myelosuppression [ 51 and collections after stimulation of hematopoiesis by cytokines [ 1 11.

POSTCHEMOTHERAPY MOBILIZATION

Stem cell collections following recovery from chemo- therapy have been extensively studied by several groups. The mechanism of enhanced collection during blood count recovery is probably dependent on endogenous cy- tokine production. The rate of progenitor cell production is regulated by endogenous cytokines. When the hemato- poiesis compartment is depleted by chemotherapy, or when stimuli such as infection, stress, and hemorrhage exist, endogenous cytokine production increses and re- sults in greater progenitor and, subsequently, greater dif- ferential cell production. Stem cell collection following chemotherapy induced myelosuppression must be timed precisely. There is a narrow window, during early blood count recovery, when effective collections of large num- bers of CD-34 + cells can be performed [ 3,131. Logisti- cally this can be difficult, and apheresis procedures may have to be performed during weekends when trained per- sonnel may not be readily available. Moreover, the tech- nique is not without risks. The degree of myelosup- pression that has to be induced to ensure adequate mobilization can result in a significant increase in the risk of “nadir” septic episodes.

It should be noted that stem cells are collected to prevent sepsis following severely myelosuppressive ther- apies. The degree of mobilization can be enhanced fur- ther by using cytokines in conjunction with chemother-

Address reprint rcquests t o Tauseef Ahmed, M.D. . Professor of Medi- cine and Director. Bone Marrow Transplantation. Ncw York Medical College. Valhalla. NY 10595.

0 1992 Wiley-Liss, Inc.

130 Ahmed et al.

apy induced myelosuppression and subsequent recovery. Schwartzberg and co-workers have shown that adequate stem cell collections (defined as 2.5 x lo6 CD-34+ cellsikg body weight) can be performed with 1-3 cy- topheresis procedures if patients are preconditioned with cyclophosphamide 4 g/m2, etoposide 600 mg/m2, and G-CSF I50 mg/m2 [ 141. However, while this methodol- ogy does reduce the number of apheresis producers, there is an enhanced greater risk of nadir septic episodes and consequent morbidity and even mortality. Interestingly, maximal effects are seen when cytokines are added to the chemotherapeutic regimen.

CYTOKl NE MOBILIZATION

Cytokines like G-CSF or GM-CSF can be used to enhance stem cell recovery without inducing myelo- suppression with chemotherapy. The commercially available cytokines are, in general, well tolerated. Both G-CSF and GM-CSF can be administered subcutane- ously and can be self-injected by the patient after a mini- mum of training. Occasional myeloid pain and fevers can be well controlled with nonsteroidal antiinflamma- tory drugs or TNF blockers such as isopentoxiphylline. As new cytokines become available, it is likely that im- proved stem cell mobilization will result.

At New York Medical College and Hudson Valley Blood Services, we generally use GM-CSF at a dose of 500 p,g, given subcutaneously daily starting 3 days be- fore the first collection and continuing through the entire period of collection. Cytapheresis procedures are per- formed on a daily basis. However, if this is not possible logistically, cytokine administration is continued until such time that cytapheresis is possible. Ten to 14 I of blood are processed at each session. Each collection yields of 0.5-1.5 X lo8 mononuclear cellsikg patient body weight. Thus, if no bone marrow is used 5-10 apheresis procedures are necessary to have a total collec- tion with 8-10 x 10' cellsikg. If peripheral stem cells are used in conjunction with bone marrow, then 4.0 x 10' mononuclear cells/kg are collected. The addition of peripheral stem cells to bone marrow progenitors in the setting of autologous transplantation hastens blood count recovery significantly compared to using marrow alone. In fact, when peripheral stem cells are used, blood count recovery is not accelerated any further by the concurrent use of currently available cytokines (T. Ahmed, unpub- lished data). The use of newer apheresis machines, such as the Code Spectra and Fenwal CS 3000, provide effi- cient mononuclear cell recovery and reduced product volumes with minimal red blood cell or granulocyte con- tamination. This is important when multiple collections are required.

STEM CELL STORAGE

Methods of stem cell storage are likewise important. In our laboratory we freeze marrow and stem cells in a mechanical freezer without controlled rate freezing. These techniques are important fiscally, since the added expense of controlled rate freezing and liquid nitrogen can be avoided. When large volumes are stored and later transplanted, DMSO toxicity may be problematic. A mixture of 6% Hetastarch, 5% DMSO, and 4% albumin used as the cryprotectant reduces the amount of DMSO necessary. Methods of volume reduction and thus de- creased DMSO content can be used upon thawing as well. Several commercial systems that reportedly posi- tively select CD-34 + cells and reduce the volume of the stem cell product have been described. These are still investigational and the impact of such technology on non-CD-34 + populations and subsequent hematologic recovery has not been fully explored.

EX VlVO EXPANSION

Incubation of stem cells with cytokines in vitro may allow for expansion of the hematopoietic stem cell pool. In preliminary work done with this technique at New York Medical College/Hudson Valley Blood Services, we have seen a tendency toward an acceleration in blood count recovery postmarrow transplant. Interestingly, the concentrations of GM-CSF seen in the stem cell collec- tion bag after a subcutaneous injection of commercially available GM-CSF (LeukineR) are virtually identical to the concentration of GM-CSF shown to be effective in enhancing CFU-GM production in vitro (L. Helson, per- sonal communication).

FUTURE DIRECTIONS

As yet there are no randomized controlled studies that compare the use of unperturbed peripheral stem cells, cytokine stimulated cells, chemotherapy recovery cells, or cells collected at the time of chemotherapy recovery along with cytokine stimulation. The differences, quali- tatively, appear to be few. Fewer apheresis procedures are necessary when cells are collected during blood count recovery and cytokines are given. This procedure is risk- ier, however. Cytokine stimulation offers a safe and ef- fective method of mobilizing peripheral stem cells.

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Cytokine Mobilization of Stem Cells 131

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