Current Status of Leukocyte and Platelet Administration in Cancer Therapy

Transcription

1 ANNALS OF CLINICAL AND LABORATORY SCIENCE, Vol. 13, No. 6 Copyright 1983, Institute for Clinical Science, Inc. Current Status of Leukocyte and Platelet Administration in Cancer Therapy ERNEST M. WALKER J r, M.D., P h.d., ALBERT CANNON, M.D. and ERNESTINE N. M ITCHUM, R.N. Department of Laboratory Medicine, Medical University of South Carolina, Charleston, SC ABSTRACT Leukapheresis and plateletpheresis are rather commonly perform ed in order to obtain single donor concentrates of granulocytes and platelets. These procedures, although relatively safe, present occasional risks to donors and recipients. Some of the occasional adverse problems experienced by donors include citrate toxicity or acute hypocalcemia, hypotension, hypervolemia, venospasm or vein occlusion, chills, anaphylactoid reactions, hem orrhage, abdominal pain or complications related to equipm ent failure and related technical problems. Potential risks to donors include those related to the receiving of six percent hydroxyethyl starch (HES), dextrans, or corticosteroids, lymphocyte depletion or immunosuppression, and effects on the complement system. Prophylactic granulocyte transfusions to prevent the occurrence of infections and associated complications in neutropenic patients have not proven to be efficacious; therapeutic granulocyte transfusions appear to be more effective. Indications for therapeutic granulocyte transfusions include those patients with known infections unresponsive to appropriately aggressive antibiotic chemotherapy over a two or three day period combined with findings of a peripheral granulocyte count less than 500 mm3 and especially those with counts below 100 mm3 and/or prolonged fever greater than 38 C (100.4 F) for 24 to 48 hours. In addition, the patient should have a reasonable chance for bone marrow recovery. Hazards or complications associated with granulocyte transfusions include: (a) immediate transfusion reactions, (b) hypersensitivity reactions, (c) pulmonary infiltrates, (d) alloimmunization, (e) transmission of infections, and (f) the possibility of Graft vs. Host (GVH) disease. The current best use of apheresis platelets is to provide therapeutic doses of single donor m atched platelets for patients refractory to pooled random donor platelets. Alloimmunization represents the major complication of therapeutic platelet transfusion and is characterized clinically by the failure to achieve expected platelet count increments after transfusion. F uture developm ents which m ight greatly improve the effectiveness of therapeutic and possibly prophylactic leukapheresis and plateletpheresis include the development of effective sedimenting agents with shorter bi /83/ $02.50 Institute for Clinical Science, Inc.

2 454 WALKER, CANNON, AND MITCHUM ological half-lives, more efficient and less expensive methods of procurem ent of granulocytes and platelets, im proved m ethods of cryopreservation of granulocytes and platelets, better methods for detecting and quantitating antigranulocyte and/or antiplatelet antibodies, and more efficient evaluation of possible synergism of granulocyte transfusions with antibiotic therapy and residual host defenses. These im provem ents may be of great value in the effective utilization of granulocyte and platelet products and in determ ining which patients are most likely to receive the maximum benefit from granulocyte and platelet support. Introduction Donor Considerations Granulocyte concentrates, one of the newest of the transfused blood compon en ts, can be o b tain ed cu rre n tly by methods including: sedimentation of solutions of red blood cells by hydroxyethyl starch (HES) or o th er m acrom olecules followed by centrifugation of the cell-rich plasma to isolate the granulocytes; adhesion filtration leukapheresis using nylon fibers; apheresis techniques using discontinuous-flow centrifugation* or continuous-flow centrifu g atio n f.54,124 The transfusion of granulocytes is considered, most often, in cases of infection in patients dem onstrating n eutropenia and bone marrow aplasia as a result of chemotherapy. 28 Platelet concentrates can be obtained by the pooling of individual bags of platelets obtained from m ultiple donors or as a single donor apheresis platelet concentrate.26 These platelet concentrates are adm inistered to help p rev en t bleeding complications in patients with thrombocytopenia.26,112 C oncentrates containing appreciable numbers of platelets and granulocytes can be obtained by using the apheresis techniques of discontinuous-flow centrifugation*112 or continuous-flow centrifugation, f 47 These products can, therefore, provide th erap eu tic benefit in a single throm bocytopenic and neutropenic patien t.112 * Haemonetics-30. t IBM-2997 Celltrifuge. Only 0.27 percent of a total of 18,288 leuka- and leukaplateletphereses p e r form ed in American Red Cross regional blood centers during a 12-month period w ere discontinued due to acute donor reactions.92 None of the reactions had serious or long-term donor consequences. However, it is obvious that additional data is needed to assess the effects of repeated leukapheresis procedures on donors and to establish rational guidelines for donor recruitm ent. Common Acute Adverse Clinical Reactions Associated with Apheresis C it r a t e T o x ic it y O ne of the most common adverse reactions is citrate toxicity or the acute hypocalcemia which may result from rapid reinfusion of c itra te d donor b lo o d.92 Symptoms usually include tingling around the m outh and in the fingers and occasionally, d o n o r h y p erv en tilatio n. The reaction is usually prom ptly reversed by slowing down the reinfusion rate of citrate blood and/or by giving the patient calcium in the form of milk or calcium gluconate. H ow ever, over-adm inistration of i.v. calcium involves the risk of iatro g en ic acu te h y p ercalcem ia to th e donor.92 H y p o t e n s io n Hypotensive symptoms may be seen in situations of donor hypovolemia, especially in discontinuous-flow procedures,

3 L EU K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY 4 55 w hen as much as 550 ml or m ore whole blood may be in the extracorporeal circulation. The signs of syncope or other hypotensive reactions generally respond to rapid infusion of i.v. solutions or return of the extracorporeal blood to the donor. Occasionally, persistent hypotension, weakness or diaphoresis may occur despite adequate correction of hypovolemia; this prolonged reaction may be due to anxiety or a vasovagal reaction.92 H y p e r v o l e m ia This com plication may be associated with the infusing of six percent hydroxyethyl starch (HES) solution in norm al leukapheresis donors. Clinical symptoms include headache, swollen fingers, p e ripheral edem a, and other signs of acute plasma volume expansion. In one study, donors w ere given daily infusions of 500 ml six percent HES over a two-hour period for a total of four days. These donors dem onstrated an average increase of 37 percent in their plasma volume and an average total volume accumulation of 850 m l.87 Volume expansions of this dimension could pose a danger to donors with borderline cardiac function.92 V e n o s p a s m o r V e i n O c c l u s io n This complication is seen most often in continuous-flow p ro c e d u re s in w hich whole blood is drawn from the donor by suction through an i.v. line. Owing to the great individual variation regarding veins, the complication of venospasm may be seen at different flow rates in different individuals at a point at which the forces of suction place too m uch stress on the vein being utilized for the procedure. C h il l s Chills or shaking chills may be caused in individuals undergoing apheresis procedures by the rapid returning or infusion of cooled blood or fluids. This complication can be reduced or controlled by covering the donor w ith a blanket, increasing the donor room tem perature, or by adding in-line warming units to the return tubing.92 A n a p h y l a c t o id R e a c t io n This com plication is an acute cutaneous reaction characterized by u rticaria, p ru ritu s, flush, and occasionally angioedema. The cause is not fully understood. Attem pts have been m ade to associate the reaction with the adm inistratio n of six p e rc e n t H E S solution. H ow ever, th e reactio n has b een o b served in leukocyte donors receiving six percent HES as well as in platelet donors not receiving six percent H ES.92 In addition, antibodies to HES have not been dem onstrated convincingly in hum ans84 and recipients of HES-containing granulocytes do n o t becom e sen sitized to H E S.92 I n f e c t io n s The possibility of infections from contam inated solutions used in apheresis procedures rem ains, b u t this complication is rarely seen.92 In one case, gram negative bacterem ic shock developed in a healthy donor minutes after term ination of a discontinuous-flow leukapheresis procedure with six percent HES and trisodium citrate. It was found that an unobserved hairline crack in the HEStrisodium citrate infusion bottle allowed contamination by Enterobacter cloacae and that this solution was subsequently given to the donor.52 O t h e r O th e r ad v erse reactio n s som etim es seen in apheresis donors include occasional hem orrhage, especially in those receiving heparin or excessive six per

4 4 5 6 W A L K E R, C A N N O N, A N D M IT C H U M cent HES during the procedure; abdominal pain;133 and complications related to equipm ent failure or related technical problem s.92 O ne recent report suggests a relationship betw een the administration of six percent HES to a donor and the subsequent development of lichen planus in the anticubital fossa near the infusion site a few days after completion of a leukapheresis procedure.5 Potential Risks to Donors P o t e n t ia l T o x ic it y o r H a z a r d s o f Six P e r c e n t H ES H ydroxyethyl starch is a branching glucose p o ly m er d eriv ed from am ylopectin w hich is treated w ith ethylene oxide to convert some of the hydroxyl groups of individual glucose units to hydroxyethyl eth er groups.92 Hydrolysis of the resulting polym er yields a m ixture of molecules with a molecular weight range of 10,000 to 2,500,000 daltons. The average M.W. of com m ercially available products are 70,000 and 450,000, respectively. 67 The hydroxyethyl groups are attached most frequently to carbon-2 of the glucose units, which retards the action of serum amylases, preventing hydrolysis of HES m olecules and p erm itting longer intravascular survival.25,26 T hese agents w ere originally developed to serve as volume expanders.92 Virtually all of an injected dose of six percent HES is recovered in urine and feces.56,67,86 Approximately 50 percent of an i.v. adm inistered dose of six percent HES is cleared from the blood and excreted via the kidneys w ith 72 hours.67 The smaller HES molecules (<50,000 daltons) are rapidly cleared by th e kidneys; the portion of larger molecules which are susceptible to hydrolysis by alpha amylase are hydrolyzed before excretion.92 One study showed that the serum HES concentration did fall to the one percent level (of the initial level after administration) after 17 weeks of observation.8 The M.W. of HES molecules rem aining 28, 56, and 108 days follow ing an in te r- m ittent-flow centrifugation p rocedure dropped to and rem ained at 95,000 ± 5,000 so that the vast majority of the remaining molecules present w ere of a size that would not pass through renal glom eruli. These same m olecules apparen tly co n tain ed sufficient n u m b ers of hydroxyethyl groups to greatly retard enzymatic hydrolysis by plasma amylases.67 C oncern has recently increased re garding m ultiple infusions of six percent H ES into donors. Serial infusions of HES may result in intravascular accumulation of H ES and H ES has been detected, transiently, in both parenchymal and reticuloendothelial system cells of several organs.45,75 Com plete elimination of HES from the circulation may require several m onths or longer after m ultiple injections.66,86 In one study, a group of eight volunteers underw ent four interm ittent-flow centrifugation leukapheresis procedures spaced four days apart and w ere given a total of 500 m l of six p e rc e n t H E S in normal saline solution over a period of about three hours per procedure. Each donor received a total of 120 grams of H E S. S erum co n cen tratio n s of H ES dropped to ten p ercen t of prepheresis levels 78 days after the first procedure.67 Forward projection of the final portion of the elimination curve (assuming that the rate of elimination remains constant) allowed for the estimation that about 78 weeks of tim e are required for com plete return of serum HES concentrations to baseline levels.67 Another concern is the effect of massive doses of HES on the donor. Hydroxyethyl starch appears to affect the coagulation system in a dose-related fashion.49,109,117,119 M oderate amounts of H E S (not exceeding 1,500 m l total volume, 20 ml per kg, or 800 ml p er m2 during a 24-hour period) may cause mild

5 LEU K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY 4 57 to m oderate effects on th e coagulation system including prolonging of p rothrom bin tim e (PT) and activated partial throm boplastin tim e (PTT), and d e creased activity of factors V and VIII and fibrinogen, and decreased tensile strength of clots These changes are statistically significant but are slight and transient and not considered to pose a real threat to effective hemostasis or to produce a te n d en cy tow ard b le e d in g or thrombosis.114 However, massive doses of HES (>25 percent of the recipient s blood volume) may be more hazardous. Most studies have been done in dogs and show th a t m assive doses of H ES (> 25 percent of blood volume) cause laboratory abnorm alities in essentially all aspects of hem ostasis, d etectab le almost im m ediately after H ES infusion.109 Although the param eters of hemostasis began to improve within 48 hours after HES infusion, as long as seven days were re q u ire d for hem ostasis to approach normal values.109 The mechanism of the effects was not determ ined. There is an extrem e paucity of information regarding the effects of massive doses of HES in hum ans.109 M oderate difficulties have been en countered due to rouleaux formation in blood samples containing large quantities of HES (>30 percent) regarding the inte rp re ta tio n of ty p in g and antibody screening. However, th e rouleaux formation is readily dispersed with saline and can easily be distinguished from true agglutination by m icroscopic exam ination. The difficulty should be easily resolvable by experienced technologists, especially if they are aware that a blood specimen contains H E S.15 T here has been a noted absence of reported complications in the tens of thousands of leukapheresis donors that have received six percent H ES, so that the potential risks to HES appear to be minimal or a b s e n t.92 S everal suggestions have been made regarding future use of HES. For example, if frequent leukapheresis of individual donors is necessary, then it may be possible to use lower doses of HES in each procedure.67 However, the effects of lower doses of HES on neutrophil collection have not been reported in clinical trials. Finally, preparations of H ES that are m ore rapidly elim inated from the bloodstream should be available for general use in the future D e x t r a n s The dextrans have been tried as sedim enting agents in place of six percent HES with varying degrees of success.40 D extran-150 appears to be th e m ost promising of these agents (dextrans and gelatin) used as sedim enting agents and in a total of 700 procedures in which it has been utilized, no significant adverse reactions have been reported.44 No anaphylactoid reactions and only one episode of m ild co n ju n ctiv itis has b een noted. G ranulocyte yields com pare favorably w ith th o se using six p e rc e n t H E S.44 D extran-150 m ay in d e e d be a safe and effective macromolecular sedim enting agent for use with the interm ittent-flow cell separators. Too few studies have been done to fully evaluate its usefulness in continuous-flow leukapheresis. C o r t ic o s t e r o id s D exam ethasone and prednisone are commonly used corticosteroids given to leukapheresis donors to demarginate leukocytes into the circulation and thus improve granulocyte yields in subsequent granulocyte collections Dexamethasone is used at most centers because it has the following advantages: (a) it is potent and long-acting, (b) it may be given orally, (c) it has a m inim um of mineralocorticoid activity, and (d) its use does not appear to im pair the accumulation of neutrophils at inflammatory sites as may occur when prednisone is used.3-58in ad

6 4 58 W A L K E R, C A N N O N, A N D M IT C H U M ditio n, dexam ethasone in single high doses (10 mg) probably does not cause any clinically im portant effect on platelet function.58 Dexamethasone is given orally at an average dose of four mg per m2 body surface area, ten to 12 hours prior to the granulocyte donation at m ost centers, while prednisone, at 1.7 mg per kg body weight, is usually given orally, eight to 12 hours prior to leukapheresis at some centers.23 Single doses of corticosteroids have been reported to rarely evoke episodes of head ach e, fever, tire d n e ss, sweating and changes in mood and performance. O ther rare complications include: exacerbation of diabetes mellitus, peptic ulcers, tuberculosis and hypertension. 92 The theoretical risk of suppression of adrenal function is considered to be im probable.92 L y m p h o c y t e D e p l e t i o n The total body pool of lymphocytes has been estim ated to be at least 1 x 1012 cells.70 The total num ber of lymphocytes rem oved during a single centrifugation leukapheresis has been estim ated to be 2 to 4 X 109 cells or about twice that taken in an ordinary whole blood donation.64 Theoretically, as judged from these figures, a single leukapheresis might remove approximately 0.1 percent to 0.4 percent of the donor s lymphocyte pool so that lymphocyte depletion and m easurable im m unosuppression would appear to be unlikely. Theoretically, 60 daily leu k ap h eresis p ro ced u res w ould be needed to remove 10 to 200 X 109 lymphocytes w hich reportedly will cause lym phopenia and im m unosuppression.70 H ow ever, several recen t studies have failed to confirm this optimism. In one study, 21 healthy donors underw ent an average of nine cytaphereses (range six to 17) and w ere com pared to a m atched group of w hole blood donors who served as controls. An average of six passes p er procedure was done using the Haemonetics-30 discontinuous-flow equipm ent.98 The following changes were observed in the donors w hen com pared to the parallel control whole blood donors and included: (a) 23 percent lower m ean absolute lym phocyte count (p < 0.01), (b) 25 percent lower m ean T-lymphocyte cell count (p < 0.01), (c) 46 p erc e n t low er m ean B -lym phocyte cell co u n t (p < 0.001), (d) seru m p ro tein changes including a 27 percent decrease in m ean gamma globulin level (p < 0.01) and a 14 percent decreased IgG level (p < 0.05). Changes in IgA and IgM levels w ere not statistically significant. The loss of im m unocom petent cells in each donation was about four to five tim es that of a stan d ard w hole blood u n it. T he alarm ing part of the study is that all but two of the apheresis donors w ere re tested eight months after the first observation with no cytapheresis procedures done on th e se donors d u rin g th e in terval. There was no significant improvem ent in the total peripheral lymphocyte count, T-lymphocyte cell count, or im munoglobulin levels. There was a slight but not statistically significant increase in the num ber of circulating B-lymphocyte cells.98 A short-term study of plateletpheresis donors50 revealed a 20 percent decrease in total lymphocyte count and a statistically significant decrease in the num ber of B cells. A nother group51 observed no differences in IgG levels on a short-term protocol bu t noted appreciable differences in a long-term study of apheresis donors. A third group noted significant alterations in serum immunoglobulins in long-term plasmapheresis donors.24 A study in which rheum atoid arthritis patients underw ent removal of about 3.5 X 109 lymphocytes, twice or three times p er w eek for five to seven weeks re vealed an absolute lym phocyte depletion in the patients com parable to that re ported for chronic thoracic duct drainage.136 The greatest changes in the peripheral lym phocyte count occurred

7 L EU K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY after only three or four leukaphereses. T here was a selective depletion of T-lymphocytes. In addition, there was a loss of PHA mitogenic responsiveness, and decreases in im munoglobulin (IgG, IgA and IgM) levels. In follow-up evaluations of th ese patients, th e lym phocyte counts returned to at least 50 percent of normal in some in a period of days to weeks, but rem ained less than 50 percent in some patients for over a year.136 Lymphocytopenia and selective depletion of B-lymphocytes have been noted in plateletpheresis donors undergoing frequent donations.50 Several questions have developed from these studies.98 First, do frequent cytaphereses offer long-term hazards to the im m une status of donors? Second, could HES cause reticuloendothelial blockage which might interfere with antigen processing by macrophages? Third, does the failure of donors to dem onstrate a return of peripheral lymphocyte count and imm unoglobulin levels to norm al after a series of cytopheresis procedures rep resent a need for concern? Fourth, does long-term cytapheresis produce an im balance in helper vs. suppressor T- lymphocytes? F urther studies are definitely needed to determ ine the effects of cytapheresis on the ratios of helper T- cells to suppressor T-cells in these individuals. Fifth, what is the nature of the observed decreases in num bers of B-lymphocytes and im m unoglobulin (IgG) levels? Is the decrease in IgG due to the decreased num bers of B-lymphocytes or due to a decrease in T-cell helper function?98 E f f e c t s o n t h e C o m p l e m e n t S y s t e m Profound changes in the com plem ent system appear to occur d u rin g in term ittent-flow centrifugal leukapheresis (IFCL) and may be due to the adsorption of complement proteins onto the surfaces of IFCL software.116 There is some evidence that there is com plem ent activation involving both the classic and alternate pathways at the filter site as blood flows through the filters and returns to th e donor in continuous-flow filtration leukapheresis (CFFL) procedures.116 In addition, changes in donor venous blood are a new finding that may indicate in vivo activation of the alternative pathway.115 Certainly, additional studies are needed to firmly establish the safety or hazards of cytapheresis to individuals regarding the complement system. G r a n u l o c y t e P r o g e n i t o r C e l l s (CFU-C) D uring leukapheresis, the donor undergoes cellular depletion to yield a cellular suspension containing a certain proportion of granulocyte progenitors that form colonies in agar (CFU-C).68 The effects of cytapheresis on the num ber of CFU -C cells was investigated in a total of 25 donors.68 Blood samples and evaluation w ere done on the day of donation and at further fixed dates for a period of six m onths after donation. C orticosteroids w ere given to granulocyte donors b u t not to platelet donors. In most cases th ere was no postdonation effect, and in those donors d em o n stratin g m inor changes, all values returned to normal (0-250 CFU -C per ml) w ithin three months after donations. Several observations and conclusions w ere apparent in the study. First, the CFU-C count is very low in normal individuals. There is much individual variation, but the count remains relativ ely constan t in a given in d i vidual.91 Second, th e cell suspensions obtained by cytapheresis contained CFU- C num bers proportional to the CFU-C count of th e d o nor p e rip h e ra l blood. Third, the findings appeared to apply in cases of collection of granulocytes and immunocompetent cells.73,128

8 4 6 0 W A LK ER, C A N N O N, A N D M IT C H U M S u g g e s t e d G u i d e l i n e s f o r A l l o w a b l e F r e q u e n c y o f C y ta- a n d / o r P l a t e l e t p h e r e s is Suggested lim itations have recently been m ade regarding donors undergoing cytapheresis procedures.2,76 Some of these suggestions are: (a) no m ore than eight leukocytapheresis procedures within a 12- m onth period of tim e should be p e r form ed on the same donor.76 (b) a donor should not generally undergo a com bined total of more than 12 plateletpheresis and/or leukacytapheresis procedures during a calendar year, (c) a donor should usually not undergo more than two procedures w ithin a w eek and six procedures w ithin a m onth w ith at least 48 hours betw een p ro ced u res.2 The same general criteria should be used to select donors for apheresis procedures as is used to select donors for whole blood donation. In addition, and especially in cases of multiple frequent donations, the donors should have satisfactory white blood cell counts in clu d in g d ifferential and platelet counts prior to each procedure.2 Most centers require the platelet count to be greater than 150,000 p er (uli. and the white blood cell count to be greater than 4,000 p er xl. O ther centers impose ad d itio n al c rite ria for d o nor selection other than those already mentioned. D o n o r S e l e c t i o n o f D o n o r s f o r P l a t e l e t p h e r e s is ( L e u k a p h e r e s is ) T w o main systems have been considered in donors to help prevent alloimm unization in recipients of p latelets.79 These are: (a) ABH and (b) HLA, which is an extremely immunogenic and polym orphic system of antigens carried by platelets.79 Antibodies to platelet-specific antigens (PLA, Ko, PLE, and BAKa system s) are less com m only e n c o u n te re d (only one case of anti-b A K a) b u t can cause serious problem s w henever they occur.16,30 Anti-PLAi is probably the most frequently encountered platelet-specific antibody.16 Platelet transfusions m atched for HLA antigens (A and B loci) are usually effective in preventing hem orrhage in some throm bocytopenic patients refractory to random -donor platelet transfusions.59,77,120,122,138 However, recent observations suggest that HLA m atching does not reliably predict platelet transfusion responses.11,19,31,121,122 For example, poor posttransfusion increm ents are occasionally o b serv ed follow ing H LA -m atched p latelet transfusion and alternatively, excellent posttransfusion increm ents have resulted after HLA-mism atched p latelet transfusions.122 It has been suggested that the platelet migration in h ib itio n assay is p re d ic tiv e of p latelet response in both HLA-com patible and H LA -incom patible d o nor-recipient pairs.122 These same authors suggest th a t lym phocytotoxicity te sts are p re d ic tiv e only in H L A -incom patible donor-recipient pairs, and that granulocytotoxicity, m icroleukoagglutination, and capillary leukoagglutination are of alm ost no value in p re d ic tin g p la te le t transfusion increm ents, either in HLAcompatible or HLA-incompatible donorrecipient pairs.122 The presence of HLA-A2 antigen in alloimmunized recipients may be of significance in locating therapeutically suitable p la telets.18 R ecent data18 suggests that adequate platelet support of alloimmunized recipients may be m ore difficult in those who possess the HLA-A2 antigen. Patient or Recipient Considerations I n d ic a t io n s f o r G r a n u l o c y t e T r a n s f u s io n s The indications for granulocyte transfusion to granulocytopenic patients are usually based on the following criteria: (a) reasonable chance for bone marrow re

9 L EU K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY 461 covery, (b) peripheral granulocyte count below 500 p e r cubic m illim eter, and p ro b ab ly m ore critically a p e rip h e ra l granulocyte count below 100 per cubic m illim eter, (c) fever g reater than 38 C (100.4 F) for 24 to 48 hours, and (d) known infection unresponsive to an adeq uate broad spectrum antibiotic reg im en over a p erio d of tw o or th re e days.26,38,41,78,113once granulocyte th erapy has been initiated, it should be contin u e d aggressively w ith gran u lo cy te transfusions on a daily basis in surviving patients until the infection is under control and the bone m arrow has recovered.26,41 Discontinuing the granulocyte transfusions after apparent infection control b u t before bone m arrow recovery may lead to recurrence of the infection, or the appearance of new infections.41 O th er influences on the decision to co n tin u e or d iscontin u e granulocyte transfusions include: (a) inevitable or impending patient death in spite of granulocyte transfusions, (b) failure of bone marrow response after long-term m aintenance with granulocytes and poor prospects of functional bone marrow recovery should granulocyte transfusions be continued, (c) developm ent of antibodies against donor granulocytes, and (d) occurrence of complications associated with granulocyte transfusion such as severe transfusion reactions, fluid overload, sequestration of granulocytes in pulmonary lesions, in travascular aggregation of granulocytes, and possible reactions of transfused granulocytes with circulating endotoxins.36,41 Failure of bone marrow recovery leads eventually to ineffectiveness of subsequent granulocyte transfusions due to sensitization or to infection with resistant organisms.36 It is reasonable to provide therapeutic granulocyte support for at least seven days after initiation before reassessing or changing the therapeutic program.36 Lithium carbonate is a granulopoietic agent90,107 that can lim it the degree of n eutropenia in patients receiving chemotherapy ordinarily associated with mild to m oderate m yelotoxicity.32,106 In one study, patients receiving a standard cytosine arabinoside and daunorubicin regim en as in d u ctio n or rein d u ctio n therapy of acute myelogenous leukemia (AML) w ere random ly assigned to re ceive lithium carbonate, 300 mg, three times p er day, or no lithium. Lithium did not prevent the onset of severe neutropenia (0.1 x 109 granulocytes p er liter) but did appear to shorten the duration of gran u lo cy to p en ia (tim e d u rin g w hich granulocyte count was 1.0 x 109 per liter) from a m edian duration of 24.6 days for controls to 16.0 days for those receiving lithium (p = 0.013). The shortened duration of neutropenia may reflect an earlier granulocyte recovery in patients receiving lithium. However, the possibility remains that lithium m ight stimulate the recovery of leukemic cells as well as the recovery of normal granulocytes.105 Sim ilar m y elo p ro tectiv e effects of lithium during chem otherapy have been reported in cases of malignancies other than leukemia, in which lithium would not be expected to increase recovery of the tum or cells.32,63,106 Protracted severe neutro p en ia may necessitate extensive broad spectrum antibiotic therapy with a risk of superinfection with fungi. Thus, shortening the duration of severe neutropenia during induction therapy of AML should decrease m orbidity. U nfortunately, in this study lithium failed to prevent the granulocyte count from d e creasing to values 0.1 x 109 per liter, did not appear to affect the rate and duration of remissions, and did not affect the incidence of infections so that the potential benefits of lithium appear to be lim ited. 105 P r o p h y l a c t ic G r a n u l o c y t e T r a n s f u s io n s A num ber of studies have been conducted to evaluate the ability of prophy

10 4 6 2 W A L K E R, C A N N O N, A N D M IT C H U M lactic granulocyte transfusions to prevent the occurrence or complications of infections in neutropenic leukem ic patie n ts O ne study involved p ro p h y lactic g ranulocyte transfusions during initial induction chemotherapy for A M L.111 O f 102 uninfected random ized patients with AML, 54 w ere given daily granulocyte transfusions for a total of 28 days or until one of the following occurred: bone marrow recovery (granulocyte count > 0.5 X 109 per liter for 48 hours), death, severe transfusion reactions, gram-negative septicemia or withdraw al of p atient consent and the re maining 48 patients served as controls and were not given granulocytes. G ranulocyte transfusions decreased the proportion of patients with bacterial septicem ia (nine percent vs. 27 percent in controls; p = 0.01). However, granulocyte transfusions did not reduce the incidence of other infections or improve bone marrow recovery, remission rate and duration of survival. A total of 72 percent of the patients given granulocytes experienced transfusion reactions. Pulm onary infiltrates w ere m ore com m on in granulocyte-treated than in control patients (57 percent vs. 27 percent in controls; p = 0.002). A total of 35 percent of the patients with pulm onary infiltrates died, as com pared w ith five p e rc e n t of those without infiltrates. T he in v estig ato rs in this study conclu d ed th a t p ro p h y lactic granulocyte transfusions should not be used during rem issio n -in d u ctio n ch em o th erap y in AM L because th e risks appear to o u t weigh the benefits.111 In a similar study, 13 of 24 p a tie n ts w ith AM L receiv ed granulocyte transfusions on alternate days during periods of marrow aplasia caused by initial induction chem otherapy with transfusions started w henever peripheral granulocyte counts becam e less than 500 per xl. Recipients received from one to 13 granulocyte transfusions with a m e dian num ber of 1.45 x 1010 granulocytes p er transfusion (range 0.28 to 3.45 x 1010). C onclusions re su ltin g from th e study are that prophylactic granulocyte transfusions offer no significant advantage regarding: (a) deaths owing to infection, (b) reduction in the frequency of febrile episodes, (c) delay in the onset of fever, (d) reduction in the length of febrile episodes, or (e) reduction in the frequency of proven infection.22 Of 65 noninfected patients with acute nonlym phocytic leukem ia (ANLL), 29 w ere chosen to receive daily granulocyte transfusions com m encing w hen the peripheral granulocyte count fell below 0.5 X 109 p er liter and continued daily as long as the count rem ained below 0.5 x 109 per liter.118 Conclusions of the study are that prophylactic granulocyte transfusions from random donors, given once daily to patients with ANLL during chem otherapy-induced neutropenia: (a) do not decrease the num ber of septicemic episodes, (b) do not increase the chance of remission-induction, (c) do not prolong th e p atien t s survival, and (d) have no practical value in the routine care of patients with ANLL.118 A total of 38 uninfected patients undergoing bone m arrow transplantation were assigned at random to receive prophylactic granulocyte transfusions and oral nonabsorbable antibiotics (group 1) or oral nonabsorbable antibiotics alone (group 2) when their neutrophil count fell below 0.5 X 109 p er liter. Both groups contained 19 individuals comparable in term s of age, sex, underlying disease, im m unosuppressive therapy, and days of neutropenia. Conclusions made from results of the study of prophylactic granulocyte transfusions include: (a) they may prevent or decrease the num ber of episodes of septicemia, (b) they have no effect on o th e r infections o r survival in patients undergoing bone marrow transplantation, (c) they are associated with a h ig h e r in c id e n c e of cyto m egalo v iral (CMV) infections and (d) oral nonabsorb

11 L E U K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY 463 able antibiotics alone are equally effective in preventing serious infections in bone marrow transplant recipients.134 Finally, a study has been made recently of th e cost-effectiveness of therapeutic and prophylactic leukocyte transfusions. Findings of the study lead to the estim ates th a t p ro p h y lactic granulocyte transfusions would add an average of 35.2 percent to the recipient s hospital bill and cost $57.8 million annually, while therapeutic granulocytic transfusions (in those patients at particular risk of death from infection) would add an average of 10.9 percent to the recipient s hospital bill and cost $17.7 million annually nationw ide.89 Leukocyte transfusion is an extrem ely expensive technological procedure and should be made as cost-effective as possible. O ne possibility of improving costeffectiveness would be granulocyte transfusions to patients at particular risk of death from infection rather than transfusion of all patients w ith neutropenia or neutropenia and infection, but data on this possibility is not yet available. W hen such data becom e available, cost-effectiveness analysis may be helpful in determ ining which individuals should receive this expensive treatm ent.89 An evaluation of 67 episodes of gramnegative bacterem ia was m ade.61 Patients had a m edian absolute granulocyte count of 0.1 X 109 per liter at the time of bacterem ia. Em piric antibiotic regimens w ere begun at the first evidence of suspected infection. Granulocyte transfusions w ere em ployed only as clinically indicated by inadequate patient response to antibiotic therapy. Among the 29 patients who dem onstrated an increase in their granulocyte count of >100 per xl over the 14 days subsequent to the initiation of therapy, 27 (93 percent) re covered w hile am ong th e 38 p atien ts ex hibiting no appreciable increase in granulocyte count 21 (55 percent) im proved (p = 0.006). In the latter group (no appreciable increase in granulocyte count) susceptibility of the pathogen to the initial empiric antibiotic regimen (two different antibiotics) was of major im portance. In four cases the pathogenic organism was resistant to both antibiotics and 0/4 of the patients responded. In 14 cases the organism was susceptible to one of the two antibiotics and 6/14 (44 percent) resp o n d ed. In 20 cases th e o r ganism was susceptible to both an tibiotics and 15/20 (75 percent) responded (p = 0.025). A num ber of conclusions w ere made from results of the study.61 First, patien ts w ith gram -negative b acterem ia and persistent granulocytopenia will often respond to antim icrobial therapy alone provided the initial choice of empiric antibiotics is appropriate and that their use is instituted promptly. Second, regarding granulocyte transfusions: (a) they are expensive and tim e-consum ing to obtain, (b) there are potential complications for both donor and recipient, (c) it is im perative to carefully identify the patients for whom granulocytes can be most beneficial and advisable in some patients to obtain a bone marrow aspirate in order to assess the likelihood of prolonged aplasia before reaching a decision regarding granulocyte transfusion,97 (d) granulocyte transfusions can be beneficial in patients with prolonged aplasia, gram -negative bacterem ias, and inadequate initial clinical response, and (e) granulocyte transfusions are not necessary for the majority of patients who are anticipated to have evidence of granulocyte or bone marrow recovery within a short tim e period.61 Knowledge of donor and recipient ABO-Rh blood types and, if possible, HLA types and using donor granulocytes from donors of th e same ABO-Rh ty p e and sim ilar or id en tical HLA type can usually increase survival times of granulocytes (and platelets) and probably delay alloimmunization to the granulocytes and platelets.61 Finally, in making decisions about the

12 4 64 W A L K E R, C A N N O N, A N D M IT C H U M adm inistration of granulocyte transfusions to granulocytopenic individuals, one should be aware of the hazards and contraindications concerning these transfusions. Some of the hazards and complications of g ran u lo cy te transfusion in c lu d e :110 (a) im m ed iate transfusion reactions, (b) hypersensitivity reactions, (c) pulm onary infiltrates, (d) alloimmunization, (e) transm ission of infections, especially viral (such as h ep atitis or CMV), and (f) the possibility of inducing graft vs. host (GVH) disease. Situations such as extracorporeal perfusion, trauma, sepsis, or acute pancreatitis may chaotically activate a p a tie n t s com p lem en t system resulting in production of complement component C5a which may cause granulocyte aggregation, increased granulocyte adherence to endothelium, and leukoembolization.46 In addition, C5a may induce granulocytes to produce toxic oxygen com pounds such as superoxide and hydrogen peroxide which can damage endothelium and, in cases of pulm onary involvement, c o n trib u te to th e d e v elo p m en t of th e ad u lt re sp irato ry d istress syndrom e (ARDS). This is c h aracterized by th e plugging of the pulmonary microvascula tu re w ith g ranulocytes w hich may cause pulm o n ary en d o th elial dam age leading to protein-aceous interstitial and, ultimately, alveolar edema. Granulocyte transfusions may be clinically dangerous u n d er these circum stances and w orsen the recipient s prognosis since the lungs might be the principal organ of granulocyte sequestration.46 Corticosteroids, in high doses, inhibit granulocyte aggregation, adherence to endothelium, and superoxide production, and thus may play an important role in preventing or lessening pulm onary complications such as ARDS in sep tic in dividuals receiving granulocytes.46 In addition, am photericin-b is used in the treatm ent of fungal infections in can cer and im m unosuppressed patients. T here have been re cent reports claiming serious interactions betw een am photericin-b and transfused granulocytes in the recipients.137 I n d ic a t io n s f o r P l a t e l e t T r a n s f u s io n s Approximately 3.5 x 1010 platelets are produced daily by the bone marrow, and production rates up to six to eight times normal have been observed." About onethird of these are normally pooled in the spleen. U nder norm al conditions, the p latelet has an age-related life span of approxim ately ten days. H ow ever, in throm bocytopenic patients with platelet counts of less than 70 X 109 per liter, p la te le ts survive for only about five days,102 possibly related to an ongoing p la te le t-e n d o th e lia l cell in teractio n. Bleeding times remain normal at four and one half ± one and one half minutes, with platelet counts of 100 x 109 p er liter or greater, and vary inversely with the platelet count in patients with depressed platelet production.35 At levels between 10 and 20 X 109 platelets per liter, the bleeding tim e may exceed 30 m inutes.102 Stool blood loss m easurem ent in patients w ith production-related throm bocytopenia has indicated that bleeding does not exceed the five ml. per day found in normal individuals until platelet counts are less than 10 x 109 per liter. Substantial increases in stool blood loss are recorded only in patients with less than 5 X 109 platelets per liter.102 U nder optim um conditions, transfused platelets should maintain hemostasis for approximately five days following transfu sio n.30 A o n e-h o u r posttransfusion platelet count is helpful in determ ining if an adequate level of platelets has been obtained to achieve im proved hem o stasis. Subsequent platelet counts at four to six hours and 24 hours postinfusion allow a rough calculation of survival and thus an indication of the necessary transfusion frequency.99 Platelet counts much higher than 10 X 109 per liter are needed

13 L E U K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A P Y 4 65 in situations in which the vascular system is not intact, prior to surgical procedures, and in many cases in which individuals are receiving therapy for cancer or other disorders.34,35,81 In addition, much higher platelet counts are needed in situations producing significant p latelet dysfunction. Common mechanisms for induction of platelet dysfunction are drugs such as aspirin129 or sem isynthetic penicillins,9,10 urem ia,127 fibrinogen-fibrin degradation p ro d u cts,74 th e events associated w ith certain types of surgery81 such as cardiopulm onary bypass surgery,34 and certain clinical conditions associated w ith the production of abnormal platelets.12,125 Absolute platelet counts cannot be used as th e only m easure for determ ining the need to transfuse platelets in these situations,99 and the need for transfusion has to be evaluated on an individual basis. Platelet transfusions have a role in preventing hem orrhage and thus reducing m ortality in throm bocytopenic cancer p atien ts.33 P latelet transfusions w ould appear to be indicated, therefore, in any patient with excessive bleeding who has a documented deficiency of either platelet num ber or function.99 Effective platelet transfusions should show im provem ents in hemostasis that directly reflect the increm ent in functional platelets achieved. Among the problems associated with freq u en t p latelet transfusions are alloim m unization and the transmission of viral infections such as hepatitis and CMV.99 T he c u rre n t b e st use of ap heresis platelets is to provide therapeutic doses of single donor matched platelets for patients refractory to pooled random donor platelets.100 It is interesting that in some series as many as half of the chronically transfused recipients never becom e immunized to platelet transfusions, whereas exposure to only a few donors may imm unize susceptible recipients to these donors as well as a large num ber of other donors. Fortunately, those individuals who do becom e im m unized to random platelet donations do not show im paired responses, in most cases, to subsequent transfusions obtained from m atched donors.100 A retrospective analysis of 352 donors who underw ent plateletpheresis at least four times* resulted in the following observations: (a) restricting plateletpheresis to donors w ith a p re p h e re sis p la te le t count of 150,000 per (jli would have allowed postpheresis platelet counts of less than 100,000 per (jli in only 1.3 percent of the procedures and would have eliminated only 3.7 percent of donations,56 and (b) plateletpheresis yields w ere related to the prepheresis platelet count, num ber of cycles, sex, type of procedure (plateletpheresis or leukoplateletpheresis), and yield recorded during previous collections.56 Various instrum ents are available to obtain single donor platelets (and granulocytes) and include one discontinuousflow* and two continuous-flow m odels.f $ T he advantages of restricting p latelet transfusions to platelets obtained from a single d o nor include: (a) selectio n for ABO, Rh, and HLA compatibility with regards to the donor-recipient pair, (b) reduced possibility of posttransfusion viral infections, and (c) streamlining of the logistics of donor processing.88 C h a r a c t e r is t ic s o f P l a t e l e t s C o l l e c t e d by P l a t e l e t p h e r e s is A num ber of studies have been made regarding th e function and survival of platelets collected by plateletpheresis or leukoplateletpheresis and subsequently adm inistered to recip ien ts.65,66,112,123 In one study, the effects of intermittent-flow centrifu g atio n p h e re sis (IFC P) w ere evaluated by com paring platelets collected from ten donors by com bined * Haemonetics Model 30 Blood Processor, t IBM $ Fenwall CS3000.

14 4 6 6 W A L K E R, C A N N O N, A N D M IT C H U M plateletleukapheresis (LP) with HES with those collected from ten donors using p la te le tp h e resis w ith o u t H E S (P P ).66 These investigators concluded that combined LP collected greater num bers of platelets than PP, and that HES, at doses currently used, does not adversely affect platelet function. C om bined LP can be perform ed by IF C P using H ES to in crease neutrophil yields and obtain large n u m b ers of p la te le ts as a b y -p ro d u ct during the procedure.66 Investigators in another study agreed that HES does not induce platelet dysfunction after one exposure in previously unpheresed donors65 and pointed out that the results are similar to those seen by using dextran at low doses.130 E f f e c t s o f ABO C o m p a t ib il it y o n P l a t e l e t T r a n s f u s io n s Studies have suggested that ABO incompatibility significantly reduces the effectiveness and survival of transfused platelets. The m ean 24-hour recovery of platelets from histocompatible donors and from donors selectively m ism atched for crossreactive HLA antigens was d e creased by approximately 23 percent of the donor types for blood group A and/ or B not found in the recipient. However, the reduction in platelet recovery associated w ith ABO incom patibility is not of a magnitude that would contraindicate transfusion of A BO-m ism atched platelets.17 A l l o im m u n iz a t io n Alloimmunization represents the major c o m p lic a tio n o f p la te le t tra n s fu s io n therapy and is characterized clinically by the failure to achieve expected platelet c o u n t in crem en ts after tran sfu sio n.96 However, recent data from a num ber of centers indicate that in cancer patients receiving platelet transfusions as well as cytotoxic and perhaps im m unosuppressive therapy, alloimmunization develops in only a minority of recipients.55,96 In recent studies of patients treated with standard, intensive induction chemotherapy, only about 40 to 50 percent of patients w ith acute leukem ia20,21,95 and an even smaller num ber (ten percent) of patients w ith solid tum ors becam e alloim m u- nized.39 A nother approach to the prevention or management of alloimmunization is to use autologous frozen platelets. Patients with leukemia undergo plateletpheresis during remission, with the platelets frozen using dim ethylsulfoxide (DM SO), and these platelets are saved for subsequent transfusion during later courses of chem o therapy. 93,94 The results using this technology have b e e n encouraging w ith transfusion recovery averaging approximately 60 to 65 percent of that expected using fresh platelets. For many patients, autologous frozen platelets have represented the only source of platelet transfusion making adm inistration of further maintenance therapy possible.96 This approach is presently not practical except at large cancer centers. S t o r a g e o f G r a n u l o c y t e s a n d P l a t e l e t s G ranulocytes retain som e functional capacity for about 24 hours when preserv ed in w hole b lo o d 62 or in concen trates obtained from leukopheresis procedures.27,83 Agitation of the granulocytes during this period is probably undesirable and may cause a reduced chemotactic response.71,72,104 Some investigators suggest storage of granulocytes at 4 C while others suggest 22 C.27,72 The use of dexamethasone to improve storage characteristics has been tried.29 Glucocorticoids have beneficial effects in the storage of c ertain tissu es and organs and are known to stabilize th e m em branes of lysosomes4,29,42,43,69,103,131 which conceivably could enhance the storage of neutrophils by lessening autolysis as well as

15 L EU K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY by decreasing the lysis of adjacent cells by enzym es released into th e storage fluid.29 However, to date, results of this approach have been disappointing. Although storage of granulocytes should be possible, it is technically not clearly possible especially in regards to long-term storage or freezing. A great deal of technical advancem ent is desperately needed before long-term storage of granulocytes is possible and practical. The capabilities of platelet storage are well advanced over that of granulocyte storage. Platelets stored at room tem perature with gentle agitation can be transfused for up to three days after preparation and still exhibit reasonable survival and function although the numbers of recoverable platelets may gradually d e crease progressively during storage Recently, new p latelet container bags have been developed and m ade available* w hich allow th e p la te le ts to breathe or undergo limited gaseous exchange with the environm ent, thus d e laying ph changes in the platelet concentrate. Platelets can be transfused as long as five days after collection and maintenance in these new bags with reasonable platelet recovery and function. Five key factors are currently of importance in the short-term storage of platelets at am b ien t te m p e ra tu re and th e m aintenance of platelet viability. These are:80 (a) ph of the suspending plasma, (b) concentration of platelets, (c) continual gentle agitation during storage, (d) tem perature of storage,57 and (e) plastic composition of the storage container Platelets collected by plateletpheresis are generally adm inistered within 24 hours after collection, largely due to considerations of possible infectious contamination. A study was conducted in which platelet concentrates! w ere examined for evidence of bacterial contamination and * Fenwal. t Collected by Haemonetics-30 equipment. subsequent growth from three to 11 days after collection.13 Four to 126 p latelet concentrates exam ined revealed b acterial grow th, and grow th pattern s indicated contam ination during the microbiological m anipulations ra th e r th an contamination of the units during collection or preparation.13 These researchers concluded that platelet concentrates prepared$ can be safely transfused for up to three days after collection if stored at 4 to 6 C. New formulations, CPDA-1, CPDA-2, and CPDA-3 which contain 34 mg adenine p er 63 ml preservative and extra glucose, have m ade possible th e extended storage of whole blood or packed red blood cells.6 The effects of storing platelets in th ese new form ulations at 22 C for periods up to 72 hours have been studied.6 Morphology score, ph changes, platelet size, population distribution param eters and electron microscopic ultrastructural considerations did not show any adverse effects which could be ascribed to the presence of adenine or extra glucose or both.6 Adenine and extra glucose in the storage m edium do not appear to be harmful to platelets in vitro.6'135 Cryopreservation of platelets offers the advantages of stockpiling of serocompatible platelets and the preservation of autologous platelets at some of the larger m edical or blood c e n te rs.80 C u rre n t m ethodology involves th e storage of p latelets at 80 C in five p e rc e n t dimethylsulfoxide (DM SO )127or five p ercent glycerol80 or in glycerol-glucose (five p ercen t glycerol and four p e rcen t glucose) solutions in th e vapor p h ase of liq u id n itro g en (approxim ately q 53,80,85 Recovered platelets with both methods have been reported to perform adequately up to six hours postthaw.80 One group reported that platelets can be cry o p reserv ed by th e se m eth o d s for greater than three years w ith satisfactory posttransfusion increm ents.14 It is interestin g to n o te th a t p la te le ts frozen in

16 4 68 W A L K E R, C A N N O N, A N D M IT C H U M DM SO for periods up to 20 weeks appear to retain full HLA antigenic activity in spite of th e com bination of freezing, thaw ing and w ashing p ro c e d u re s, and the presence of the surface-active cryopreservation.82 In all cases, the previously frozen platelets w ere quantitatively equal or superior to the platelets stored at 4 C with regard to their capacity to specifically reduce the HLA antibody activities of selected typing sera against a panel of antigen-positive lymphocytes.82 P r o s p e c t s o r G o a l s f o r t h e F u t u r e Several prospects or goals have re cently been suggested regarding the collection of granulocytes and platelets by apheresis techniques and subsequent administration to recipients.37 First, more reliable m ethods are needed to d eterm ine which febrile neutropenic patients are likely to recover w ithout granulocyte support and which patients definitely req u ire this expensive form of therapy. Second, better, more efficient and less expensive methods of procurem ent are clearly n eed ed. T he lim iting factor is donor-tim e and maximizing yields during each procedure would be of great advantage, and probably m ake the prophylactic use of granulocytes more practical. Third, a m ore reliable system for m easu rin g an tig ran u lo cy te an tib o d ies is needed and would perm it better-donorrecipient pairing.36 Fourth, the possibilities of synergism betw een granulocyte transfusions and antibiotic therapy and residual host defenses should be investigated thoroughly.36 Fifth, successful cryopreservation of granulocytes might offer significant advantages especially at the larger medical or cancer centers and blood cen te rs w h ere storage of such products is practical.36 Finally, the dev elo p m en t of ad ditional sed im en tin g agents for use in leukapheresis is needed. These agents should have shorter biological half-lives w ithin donors and recipients and not present the lingering safety concerns seen with HES. R eferences 1. Author unknown: Study questions use of prophylactic granulocyte transfusions for patients with acute myelogenous leukemia. J. Am. Med. Assoc. 247:21, Author unknown: Guidelines for the collection of platelets, apheresis prepared by mechanical apheresis, August Plasm Ther. Transfus. Technol. 3: , Axelrod, L. : Glucocorticoid therapy. Medicine 55:39-65, Belzer, F. O.and Kountz, S. L. : Preservation and transplantation of human cadaver kidneys. A two-year experience. Ann. Surg. 172: , Bode, U. and Deisseroth, A. B. : Donor toxicity in granulocyte collections: Association of lichen planus with the use of hydroxyethyl starch leukapheresis. Transfusion 21:83-85, Bolin, R. B., Cheney, B. A., Simpliciano, O. A., and P eck, C. C.: In vitro evaluation of platelets stored in CPD-adenine formulations. Transfusion 20: , Bongiovanni, M. B., Katz, R. S., and W urzel, H. A. : Long-term plateletpheresis of a donor. Transfusion 20: , Boon, J. C., Jesch, F., Ring, J., and M essmer, K. : Intravascular persistence of hydroxyethyl starch in man. Eur. Surg. Res. 8: , Brown,C. H., Ill, B radshaw, M. W., N a- telson, E. A., Alfrey, Jr., C. P., and W illiams, T. W., Jr. : Defective platelet function following the administration of penicillin compounds. Blood 47: , Brown, C. H., Ill, Natelson, E. A., Bradshaw, M. W., W illiams, T. W., Jr., and A l frey, C. P., Jr.: The hemostatic defect produced by carbenicillin. Nww Engl. J. Med. 291: , Bucher, U., D ew eck, A., Spencler, H., Tschopp, L., and K ummer, H.: Platelet transfusions: Shortened survival of HLA identical platelets and failure of in vitro detection of antiplatelet antibodies after multiple transfusions. Vox Sang. 25: , C ardamon, J. M., E dson, R., M carthur, J., and Jacob, H.: Abnormalities of platelet function in the myeloproliferative disorders. J. Am. Med. Assoc. 221: , C ordle, D., Koepke, J. A., and Koontz, F. P.: The sterility of platelet and granulocyte concentrates collected by discontinuous flow centrifugation. Transfusion 20: , D aly, P. A., Schiffer, C. A., Aisner, J., and W iernik, P. H.: Successful transfusion of platelets cryopreserved for more than three years. Blood 54: , 1979.

17 L E U K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY D a n i e l s, M. J., S t r a u s s, R. G., and S m it h - F l o s s, A. M.: Effects of hydroxyethyl starch on erythrocyte typing and blood crossmatching. Transfusion 22: , D e c a r y, F. : Platelet antigens. Plasma Ther. Transfus. Technol. 3: , D u q u e s n o y, R. J., A n d e r s o n, A. J., T o m a s u l o, P. A., and A s t e r, R. H.: ABO compatibility and platelet transfusions of alloimmunized thrombocytopenic patients. Blood 54: , D u q u e s n o y, R. J., F i l i p, D. J., and A s t e r, R. H.: Influence of HLA-A2 on the effectiveness of plalelet transfusions in alloimmunized thrombocytopenic patients. Blood 50: , D u q u e s n o y, R. J., F i l i p D. J., R o d e y, G. E., and A s t e r, R. H.: Transfusion therapy of refractory thrombocytopenic patients with platelets from donors selectively mismatched for crossreactive HLA antigens. Transplant Proc. 9(Suppl. 1): , D u t c h e r, J. P., S c h i f f e r, C. A., A is n e r, J., and W i e r n i k, P. H.: Alloimmunization following platelet transfusion: The absence of a dose-response relationship. Blood 57: , D u t c h e r, J. P., S c h i f f e r, C. A., A is n e r, J., and W i e r n ik, P. H.: Long-term follow-up of patients with leukemia receiving platelet transfusions: Identification of a large group of patients who do not become alloimmunized. Blood 58: , F o r d, J. M., C u l l e n, M. H., R o b e r t s, M. M., B r o w n, L. M., O l iv e r, R. T. D., and L is t e r, T. A.: Prophylactic granulocyte transfusions: Results of a randomized controlled trial in patients with acute myelogenous leukemia. Transfusion 22: , F r e n c h, J. E.: Leukapheresis practice in the U.S.: Results of a survey of registered establishments. Conference on Leukapheresis Donor Safety. Washington, Department of Health and Human Services, Public Health Services, Food and Drug Administration, 1980, pp F r i e d m a n, B. A., S c h o r k, M. A., M o c n ia k, J. L., and O b e r m a n, H. A.: Short-term and long-term effects of plasmapheresis on serum proteins and immunoglobulins. Transfusion 15: , G a r z o n, A. A., C h e n g, C., L e r n e r, B., L i c h t e n s t e i n, S., and K a r l s o n, K. E.: Hydroxyethyl starch (HES) and bleeding. J. Trauma 7: , G i l c h e r, R. O.: Blood and apheresis products: Usage in modern transfusion therapy. Plasma Therapy 3:27-44, G l a s s e r, L. : Effect of storage on normal neutrophils collected by discontinuous-flow centrifugation leukapheresis. Blood 50: , G la sser, L.: Functional considerations of granulocyte concentrates used for clinical transfusions. Transfusion 19:1-6, G la sser, L. and H u e stis, D. W. : Characteristics of stored granulocytes collected from donors stimulated with dexamethasone. Transfusion 19:53-56, G lowitz, R. J. and Slichter, S. J.: Frequent multiunit plateletpheresis from single donors: Effects on donors blood and platelet yield. Transfusion 20: , G mur, J., v o n F elten, A., and F rick, P.: Platelet support in polysensitized patients: Role of HLA specificities and crossmatch testing for donor selection. Blood 51: , G reco, F. A. and Brererton, H. D.: Effect of lithium carbonate on the neutropenia caused by chemotherapy: A preliminary clinical test. Oncology 34: , H an, T., Stutzman, L., C ohen, E., and Kim, U.: An autopsy study of the effects of platelet transfusion on hemorrhage in patients with acute leukemia. Cancer 19: , H arker, L. A., Malpass, T. W., Branson, H. E., H essel, E. A., and Slichter, S. J.: M echanism of abnorm al bleeding in p atients undergoing cardiopulm onary bypass: A cquired transient dysfunction associated with selected alpha granule release. Blood 56: , H arker, L. A. and Slichter, S. J.: The bleeding time as a screening test for evaluation of platelet function. New Engl. J. Med 287: , H igby, D. J. and B urnett, D.: Granulocyte transfusions: Current status. Blood 55:2-8, H igby, D. J., Burnett, D., Ruppert, K., and H enderson, E. S.: Filtration leukapheresis: Studies on donors. Leucocytes: Separation, Collection and Transfusion. Goldman J. M. and Lowenthal R. M., eds. London, Academic Press, 1975, pp H igby, D. J., Yates, J. W., H enderson, E. S., and H olland, J. F.: Filtration leukapheresis for granulocyte transfusion therapy. New Engl. J. Med. 292: , H olohan, T. V., T erasaki, P. I., and D eisseroth, A. B.: Suppression of transfusion-related alloimmunization in intensively treated cancer patients. Blood 58: , H uestis, D. W.: Use of dextran 75 as a macromolecular agent in centrifugal leukapheresis. Transfusion 17: , H uestis, D. W., Bove, J. R., and Busch, S.: Practical Blood Transfusion, 3rd ed. Boston, Little, Brown and Co., Ignarro, L. J.: Dissimilar effects of anti-inflammatory drugs on stability of lysosomes from peritoneal and circulating leukocytes and liver. Biochem. Pharmacol. 20: , Ignarro, L. J.: Effects of anti-inflammatory drugs on the stability of rat liver lysosomes in vitro. Biochem. Pharmacol. 20: , I sbister, J. P., Scurr, R. D., C louten, V. C., and Biggs, J. C.: Use of dextran 150 as a macromolecular agent to improve granulocyte yields on the intermittent flow blood cell separator (Haemonetics Model 30). Transfusion 19: , Jesch, F., H ubner, G., Zumtobel, V., Zimmermann, M., and M essmer, K.: Hydroxyathylstarke (HAS) 450/0.7 in Plasma and Leber. Konzentrationsverlauf und Histopathologische Veränderungen Beim Menschen. Infusionstherapie 6: , 1979.

18 4 7 0 W A LK ER, C A N N O N, A N D M IT C H U M 4 6. J a c o b, H. S.: Complement induced vascular leukostasis: Its role in tissue injury. Arch. Pathol. Lab. Med. 104:88-91, K a l m in, N. D. and G r in d o n, A. J. Pheresis with the IBM Transfusion 21: , K a r l s o n, K. E., G a r z o n, A. A., S h a f t a n, G. W., and C h u, C. J. : Increased blood loss associated with administration of certain plasma expanders: Dextran 75, dextran 40, and hy- droxyethyl starch. Surgery 62: , K is k e r, C. T., St r a u s s, R. G., K o e p k e, J. A., M a g u ir e, L. C., and T h o m p s o n, J. S.: The effects of combined platelet and leukapheresis on the blood coagulation system. Transfusion 19: , K o e p k e, J. A., P ark s, W. M., G o e k e n, J. A., K l e e, G. G., and St r a u ss, R. G. : The safety of weekly plateletpheresis: Effect on the donor s lymphocyte population. Transfusion 21:59-63, K o e p k e, J. A., Wu, K. K., H o a k, J. C., and T h o m pson, J. S.: Effects of long-term plasmapheresis on plasma proteins. Transfusion 16: , K o s m in, M. : Bacteremia during leukapheresis. Transfusion 20:115, K o t e l b a-w it k o w s k a, B. and S c h i f f e r, C. A.: Cryopreservation of platelet concentrates using glycerol-glucose. Transfusion 22: , K u r t z, S. R., C a r c ie r o, R., and V a l e r i, (t. R.: Factors that influence the process of 51Chromium labeling of human granulocytes isolated!from blood by counterflow centrifugation. Transfusion 19: , K u t t i, J., Z a r o u l is, C. G., D in s m o r e, R. E., R e i c h, L., C l a r k s o n, B. D., and G o o d, R. A.: A prospective study of platelet-transfusion therapy administered to patients with acute leukemia. Transfusion 22:44-47, L a sk y, L. C., L i n, A., K a h n, R. A., and M c C u l l o u g h, J. : Donor platelet response and product quality assurance in plateletpheresis. Transfusion 21: , L a z a r u s, H. M., H e r z ig, R. H., W a r m, S. E., and F ish m a n, D. J.: Transfusion experience with platelet concentrates stored for 24 to 72 hours at 22 C: Importance of storage time. Transfusion 19: , L ic h t e n f e l d, K. M. and S c h i f f e r, C. A.: The effect of dexamethasone on platelet function. Transfusion 19: , L o h r m a n, H. P., B u l l, M. I., D e c t e r, J. A., Ya n k e e, R. A., and G ra w, R. J., J r.: Platelet transfusions for H L A compatible unrelated donors to alloimmunized patients. Ann. Intern. Med. 8 0 : 9-1 4, L o r ia u s, L. : Adrenocorticosteroids: Potential risks of repeated administration. Conference on Leukapheresis Donor Safety. Washington, Department of Health and Human Services, Public Health Service, Food and Drug Administration, 1980, pp L o v e, L. J., S c h im p f f, S. C., S c h if f e r, C. A., and W ie r n ik, P. H.: Improved prognosis for granulocytopenic patients with gram-negative bacteremia. Am. J. Med. 68: , L o w e n t h a l, R. M.: Chronic leukemias: Treatment of leucapheresis. Exp. Hemat. 5(suppl.): 73-84, L y m a n, G. H., W i l l ia m s, C. C., and P r e s t o n, D.: A prospective randomized study of the effect of lithium carbonate. Blood 52(Suppl. 1):228, M a c P h e r s o n, J. L., N u s b a c h e r, J., and B e n n e t t, J. M. : The acquisition of granulocytes by leukapheresis: A comparison of continuous flow centrifugation and filtration leukapheresis in normal and corticosteroid-stimulated donors. Transfusion 16: , M a g u ir e, L. C., H e n d r ik s e n, R. A., S t r a u s s, R. G., G o e d k e n, M. M., E c h t e r n a c h t, B., K o e p k e, J. A., and T h o m p f o n, J. S.: Platelet function in donors undergoing intermittent-flow centrifugation plateletpheresis or leukapheresis. Transfusion 20: , M a g u ir e, L. C., H e n d r ik s e n, R. A., S t r a u s s, R. G., S t e i n, M. N., G o e d k e n, M. M., E c h t e r n a c h t, B., K o e p k e, J. A., and T h o m p s o n, J. S.: Function and morphology of platelets produced by intermittent-flow centrifugation plateletpheresis or combined platelet-leukapheresis. Transfusion 21: , M a g u ir e, L. C., S t r a u s s, R. G., K o e p k e, J. A., B o w m a n, R. J., Z e l e n s k i, K. R., L a m b e r t, R. M., H u l s e, J. D., and A t n i p A. K.: The elimination of hydroxyethyl starch from the blood of donors experiencing single or multiple intermittent-flow centrifugation leukapheresis. Transfusion 21: , M a y e r, S., S c h m it t, M., and Noss, P.: Granulocyte progenitor cells (CFU-C) in peripheral blood of normal volunteers following cell collection by continuous-flow centrifugation. Transfusion 19: , M c C a b e, R. E., S t e v e n s, L. E., and F it z pa trick, H. F.: Corticosteroids in organ preservation. Trans. Proc. 7: , M c C u l l o u g h, J. : Leukapheresis and granulocyte transfusion. CRC Crit. Rev. Clin. Lab. Sci. 10: , M c C u l l o u g h, J. : Liquid preservation of granulocytes. Transfusion 20: , M c C u l l o u g h, J., W e i b l e n, B. J., P e t e r s o n, P. K., and Q u i e, P. G.: Effects of temperature of granulocyte preservation. Blood 52: , M c C r e d i e, K., H e r s c h, E. M., and F r e i r e ic h, E. J.: Cells capable of colony formation in the peripheral blood of man. Science 171: , M c K ay, D. G. : Disseminated Intravascular Coagulation. New York, Harper and Row, M e s s m e r, K. and J e s c h, F.: Volumenersatz and Hamodilution Durch Hydroxyathylstarke. Infiisionstherapie 5: , M is c h l e r IV, M. M.: Donor conditioning agents in leukocytapheresis and thrombocytapheresis: Preliminary guidelines for use. Plasma Therapy 3:5-26, M it t a l, K. K., R u d e r, E. A., and G r e e n, D.: Matching of histocompatibility (HLA) antigens for platelet transfusion. Blood 47:31-41, 1976.

19 L E U K O C Y T E A N D P L A T E L E T A D M IN IST R A T IO N IN C A N C E R T H E R A PY M o r s e, E. E., K a t z, A. J., B u c h h o l z, D. H., and Houx, J.: Clinical effectiveness of transfusion of granulocytes obtained by filtration or intermittent-flow centrifugation. Cancer 47: , M u l l e r, J. Y., C a r t r o n, J. P., T r i l l o t, P., H a l l e, L., and R o q u i n, H.: Platelet transfusion: Automated Selection of Donors. Transfusion 21: , M u r ra y, C. and T i s h k o f f, G. H.: Preparation of components and their characteristics; plasmapheresis and cytapheresis. Clinical Practice of Blood Transfusion. Petz, L. D. and Swisher, S. N., eds., New York, Churchill Livingstone, 1981, pp O B r i e n, J. R., J a m is o n, S., and E t h e r in g t o n, M.: Refractory state of platelet aggregation with major operations. Lancet 2: , P o l l a c k, M. S., Z a r o u l is, C. G:, K l a in b a r d, J., L e id e r m a n, I. Z., and D u p o n t, B.: Retention of H L A antigens on previously frozen human platelets. Transfusion 20: , P r i c e, T. H. and D a l e, D.: Neutrophil transfusion: Effect of storage and collection method on neutrophil blood kinetics. Blood 51: , R i c h t e r, A. W. and d e B e l d e r, A. N.: Antibodies against hydroxyethyl-starch produced in rabbits by immunization with a protein-hydroxyethylstarch conjugate. Int. Arch. Allergy Appl. Immunol. 52: , R id g w a y, R. L. and U sry, R. T.: Cryopreservation of platelets simplified: A modified glycerol-glucose method. Transfusion 20: , R i n g, J., S h a r k o f f, D., and R i c h t e r, W.: Intravascular persistence of hydroxyethyl starch ( H E S ) after serial granulocyte collections using H E S in man. Vox Sang. 39: , R o c k, G. and W i s e, P.: Plasma expansion during granulocyte procurement: Cumulative effects of hydroxyethyl starch. Blood 53: , R o c k, G.: Platelet Pheresis: Collection Methods, Yields, and Storage. Program Manual of the Fourth Annual Apheresis Symposium: Current Concepts and Future Trends. Chicago, Sponsored by the American Red Cross Blood Service, Carolinas Region, October 1-3, 1982, pp R o s e n s h e i n, M. S., F a r e w e l l, V. T., P r i c e, T. H., L a r s o n, E. B., and D a l e, D. C.: The cost effectiveness of therapeutic and prophylactic leukocyte transfusion. New Engl. J. Med. 302: , R o t h s t e i n, G., C l a r k s o n, D., L a r s e n, W., G r o s s e r, B., and A t h e n s, J. W.: Effect of lithium on neutrophil mass and production. New Engl. J. Med. 298: , R u b in, S. H. and C o w a n, D. H.: Assay of granulocytic progenitor cells in human peripheral blood. Exp. Hematol. 1: , S a n d l e r, S. G. and N u s b a c h e r, J.: Health risks of leukapheresis donors. Haematologia 15:57-69, S c h i f f e r, C. A., A i s n e r, J., D u t c h e r, J. R., D a l y, P. A., and W ie r n ik, P. H.: A clinical program of platelet cryopreservation. Cytopheresis and Plasma Exchange: Clinical Indications. Vogler, W. R., ed., New York, Alan R. Liss, in press. 94. Schiffer, C. A., Aisner, J., and Wiernik, P. H.: Frozen autologous platelet transfusion for patients with leukemia. New Engl. J. Med 299:7-12, Schiffer, C. A., Lichtenfeld, J. L., W iernik, P. H., Mardiney, M.R., Jr., and Joseph, J. M.: Antibody response in patients with acute nonlymphocytic leukemia. Cancer 37: , Schiffer, C. A., D utcher, J. P., Hocge, D. E., and Aisner, J.: Histocompatible platelet transfusion for patients with leukemia. Plasma Ther. Transfus. Technol. 3: , Schiffer, C. A.: Principles of granulocyte transfusion therapy. Med. Clin. North. Am. 61: , Senhauser, D. A., W estphal, R. G., Bohman, J. E., and N eff, J. C.: Immune system changes in cytopheresis donors. Transfusion 22: , Slichter, S. J.: Indications for platelet transfusions. Plasma Ther. Transfus. Technol. 3: , Slichter, S. J.: Clinical indications and experiences with apheresis platelets. Program Manual of the Fourth Annual Apheresis Symposium: Current Concepts and Future Trends. Chicago, Sponsored by American Red Cross Blood Services, Carolinas Region, October 1-3, 1982, pp Slichter, S. J. and H arker, L. A.: Preparation and storage of platelet concentrates. Transfusion 16:8-12, Slichter, S. J. and H arker, L. A.: Thrombocytopenia: Mechanisms and management of defects in platelet production. Clin Haematol. 7: , Starling, J. R., F erguson, W. W., Rudolf, L. E., and W angensteen, S. L.: Lysosomal enzym e release and vascular resistance changes in th e isolated p erfused kidney: Influence of m ethylprednisolone. Surg. Forum 23: , Steigbigel, R. T., Baum, J., MacPherson, J. L., and N usbacher, J.: Granulocyte bactericidal capacity and chemotaxis as affected by continuous-flow centrifugation and filtration leukapheresis, steroid administration, and storage. Blood 52: , Stein, R. S., F lexner, J. M., and Graber, S. 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