2
1055 carries the risk of obliterating the antibody combining site.22 Some groups have tried coupling the biological toxins abrin and ricin’4to antibody but their safety in vivo remains in doubt. A more attractive solution has been offered by Glennie and Stevenson. 23 They have constructed a univalent antibody by papain cleavage of one Fab fragment from each immunoglobulin molecule. The resulting antibody cannot cross-link with adjacent antigen molecules and therefore does not cause antigenic modulation. However, it does retain the ability to fix complement. To show its efficacy they have treated the guineapig prolymphocytic leukaemia L2C with a polyclonal, univalent rabbit antibody against the idiotypic determinants of the surface immunoglobulin. Univalent antibody was more effective in vitro at inducing complement- dependent cytolysis and in vivo it prolonged the life of the guineapig. In this it was three times as effective as whole IgG. This particular biochemical manoeuvre is effective only for rabbit IgG but other procedures are available to produce univalent antibody from the immunoglobulin of other species. When they are applied to murine monoclonal antibodies there are immense prospects for successful immunotherapy. MONOCLONAL ANTIBODIES IN LEUKAEMIA AND LYMPHOMA ONE of the basic tenets of the haematologist involved in the treatment of leukaemia is the belief that improved definition of the population of malignant cells that characterise the disease in a particular patient will permit a more accurate prognosis and contribute to decisions on therapy. At the International Research Symposium on Monoclonal Antibodies in Leukaemia and Lymphoma organised by the Leukaemia Research Fund in London, S. F. Schlossman described how the OKT series of monoclonal antibodies revealed normal T lymphocytes as a highly heterogeneous population, and M. F. Greaves described the use of these and other monoclonal antibodies to classify the acute lymphoblastic leukaemias (ALL) and to match the predominating leukaemic cell with its normal counterpart. For example, the common-ALL (c-ALL) antigen that characterises the surface membrane of leukaemic cells from most children and some adults with ALL and was originally recognised with antisera raised conventionally (and laboriously) in rabbits can now be defined with any one of three readily available monoclonal antibodies. 1 Cells with the same phenotypic characteristics as c-ALL cells can however be found, albeit at low concentration, in the marrow of normal individuals. Greaves showed that results with one monoclonal antibody, designated J-5, gave almost complete concordance with results obtained with conventionally raised rabbit antisera. H. Stein, from Kiel University, with an impressive series of photomicrographs, showed how monoclonal antibodies could define areas of predominant B or T cell infiltration in histological sections of non-Hodgkin’s lymphoma and could confirm the presumed monoclonality of the invading B cells; 22 Noord S, Weissman IL Radiolabelled antitumour antibodies III. Highly iodinated and highly radio-iodinated antibodies. J Natl Cancer Inst 1974, 53: 959-65. 23 Glennie MJ, Stevenson GT Univalent antibodies kill tumour cells in vitro and in vivo. Nature 1982, 295: 712-14. 1 Greaves MF Analysis of the clinical and biological significance of lymphoid phenotypes in acute leukemia Cancer Res 1981, 41: 4752-66 addressing the thorny question of the cellular origin of the atypical histiocyte or Sternberg-Reed cell in Hodgkin’s disease, he showed that these cells lacked the specific markers associated with null cells, B cells, T cells, monocytes or macrophages, or interdigitating or dendritic reticulum cells; instead, the cytoplasm of the Hodgkin cell routinely stained with a unique monoclonal designated Ki-1, and similar results were found with atypical histiocytes from all histological subtypes of the disease. Ki-1 also identified a small population of cells present in normal lymph nodes-a population whose origin and function remains unknown. From work with monoclonal antibodies that recognise either HLA or T related determinants on thymocytes, C. Milstein, jointly responsible for initiating the whole monoclonal antibody revolution in 1975,2 reported the fascinating effects of a highly purified human interferon on the MOLT-4 cell line. Interferon at high concentration slowed the growth rate of the cell line but increased the surface expression of HLA relative to T antigen; this was apparently due to increased synthesis of HLA glycoprotein associated with increased amounts of its corresponding mRNA. This heritable effect was reversible when interferon was withdrawn. For the clinician who might ask what practical application monoclonal antibodies have in the management ofleukaemia, there were some exciting prospects. G. Janossy, from the Royal Free Hospital in London, showed how the OKT3 antibody with activity against the majority ofT lymphocytes could be used to inactivate T cells in marrow harvested from normal donors before transplantation for patients with leukaemia or aplastic anaemia; preliminary results suggested that the incidence and severity of acute graft-versus-host disease could be lessened by this approach. 3 Janossy predicted that a panel of such antibodies, some fixing complement, might be able to eliminate entirely the T cell population responsible for graft-versus-host disease. Monoclonal antibodies with T cell specificity could also be used to "decontaminate" bone marrow cells in preparation , for autografting in the management of the T acute leukaemias; and antibodies against c-ALL, such as J-5, have now been used in Boston for a similar purpose. Eventually it is possible that monoclonal antibodies coupled with cytotoxic drugs or with natural toxins, such as ricin, might play an important role in treating leukaemias as well as other neoplasms. Finally R. Levy, from Stanford University, described the use of anti-T monoclonals for treating patients with cutaneous lymphomas or Sezary syndrome; 5 of 6 patients who received murine monoclonal antibody by the intravenous route showed good partial responses and, though patients did indeed make antibodies to murine proteins, toxic manifestations were very mild or non-existent. A patient with B cell lymphoma offollicular-centre cell type was treated with an anti-idiotypic monoclonal antibody raised against his own lymphoma cells; after initial apparent failure of the experiment, his disease regressed and he achieved a complete remission that has lasted so far 11 months. There can be little 2. Kohler G, Milstein C Continuous cultures of fused cells secreting antibody of predefined specificity Nature 1975, 256: 495-97. 3 Prentice HG, Blacklock HA, Janossy G, Bradstock KF, Skeggs D, Goldstein G, Hoffbrand AV. Use of anti-T-cell monoclonal antibody OKT3 to prevent acute graft-versus-host disease in allogeneic bone marrow transplantation for acute leukaemia. Lancet 1982; i: 700-03. 4. Miller RA, Maloney DG, Warnke R, Levy R. Treatment of B-cell lymphoma with monoclonal anti-idiotype antibody. N Engl J Med 1982; 306: 517-22 5. Miller RA, Levy R Response of cutaneous T cell lymphoma to therapy with hybridoma monoclonal antibody. Lancet 1981, ii: 226-29.

MONOCLONAL ANTIBODIES IN LEUKAEMIA AND LYMPHOMA

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1055

carries the risk of obliterating the antibody combining site.22Some groups have tried coupling the biological toxins abrinand ricin’4to antibody but their safety in vivo remains indoubt. A more attractive solution has been offered by Glennieand Stevenson. 23 They have constructed a univalent antibodyby papain cleavage of one Fab fragment from each

immunoglobulin molecule. The resulting antibody cannotcross-link with adjacent antigen molecules and therefore doesnot cause antigenic modulation. However, it does retain theability to fix complement. To show its efficacy they havetreated the guineapig prolymphocytic leukaemia L2C with apolyclonal, univalent rabbit antibody against the idiotypicdeterminants of the surface immunoglobulin. Univalentantibody was more effective in vitro at inducing complement-dependent cytolysis and in vivo it prolonged the life of theguineapig. In this it was three times as effective as whole IgG.This particular biochemical manoeuvre is effective only forrabbit IgG but other procedures are available to produceunivalent antibody from the immunoglobulin of other

species. When they are applied to murine monoclonalantibodies there are immense prospects for successful

immunotherapy.

MONOCLONAL ANTIBODIES IN LEUKAEMIA ANDLYMPHOMA

ONE of the basic tenets of the haematologist involved in thetreatment of leukaemia is the belief that improved definitionof the population of malignant cells that characterise thedisease in a particular patient will permit a more accurateprognosis and contribute to decisions on therapy. At theInternational Research Symposium on MonoclonalAntibodies in Leukaemia and Lymphoma organised by theLeukaemia Research Fund in London, S. F. Schlossmandescribed how the OKT series of monoclonal antibodiesrevealed normal T lymphocytes as a highly heterogeneouspopulation, and M. F. Greaves described the use of these andother monoclonal antibodies to classify the acute

lymphoblastic leukaemias (ALL) and to match the

predominating leukaemic cell with its normal counterpart.For example, the common-ALL (c-ALL) antigen thatcharacterises the surface membrane of leukaemic cells frommost children and some adults with ALL and was originallyrecognised with antisera raised conventionally (andlaboriously) in rabbits can now be defined with any one ofthree readily available monoclonal antibodies. 1 Cells with thesame phenotypic characteristics as c-ALL cells can howeverbe found, albeit at low concentration, in the marrow ofnormal individuals. Greaves showed that results with onemonoclonal antibody, designated J-5, gave almost completeconcordance with results obtained with conventionally raisedrabbit antisera.

H. Stein, from Kiel University, with an impressive series ofphotomicrographs, showed how monoclonal antibodiescould define areas of predominant B or T cell infiltration inhistological sections of non-Hodgkin’s lymphoma and couldconfirm the presumed monoclonality of the invading B cells;

22 Noord S, Weissman IL Radiolabelled antitumour antibodies III. Highly iodinated andhighly radio-iodinated antibodies. J Natl Cancer Inst 1974, 53: 959-65.

23 Glennie MJ, Stevenson GT Univalent antibodies kill tumour cells in vitro and in vivo.Nature 1982, 295: 712-14.

1 Greaves MF Analysis of the clinical and biological significance of lymphoidphenotypes in acute leukemia Cancer Res 1981, 41: 4752-66

addressing the thorny question of the cellular origin of theatypical histiocyte or Sternberg-Reed cell in Hodgkin’sdisease, he showed that these cells lacked the specific markersassociated with null cells, B cells, T cells, monocytes ormacrophages, or interdigitating or dendritic reticulum cells;instead, the cytoplasm of the Hodgkin cell routinely stainedwith a unique monoclonal designated Ki-1, and similar resultswere found with atypical histiocytes from all histologicalsubtypes of the disease. Ki-1 also identified a small populationof cells present in normal lymph nodes-a population whoseorigin and function remains unknown.

From work with monoclonal antibodies that recogniseeither HLA or T related determinants on thymocytes, C.Milstein, jointly responsible for initiating the wholemonoclonal antibody revolution in 1975,2 reported the

fascinating effects of a highly purified human interferon onthe MOLT-4 cell line. Interferon at high concentrationslowed the growth rate of the cell line but increased thesurface expression of HLA relative to T antigen; this wasapparently due to increased synthesis of HLA glycoproteinassociated with increased amounts of its correspondingmRNA. This heritable effect was reversible when interferonwas withdrawn.

For the clinician who might ask what practical applicationmonoclonal antibodies have in the management ofleukaemia,there were some exciting prospects. G. Janossy, from theRoyal Free Hospital in London, showed how the OKT3antibody with activity against the majority ofT lymphocytescould be used to inactivate T cells in marrow harvested fromnormal donors before transplantation for patients withleukaemia or aplastic anaemia; preliminary results suggestedthat the incidence and severity of acute graft-versus-hostdisease could be lessened by this approach. 3 Janossypredicted that a panel of such antibodies, some fixingcomplement, might be able to eliminate entirely the T cellpopulation responsible for graft-versus-host disease.Monoclonal antibodies with T cell specificity could also beused to "decontaminate" bone marrow cells in preparation

, for autografting in the management of the T acute

leukaemias; and antibodies against c-ALL, such as J-5, havenow been used in Boston for a similar purpose. Eventually itis possible that monoclonal antibodies coupled with cytotoxicdrugs or with natural toxins, such as ricin, mightplay an important role in treating leukaemias as well as otherneoplasms. Finally R. Levy, from Stanford University,described the use of anti-T monoclonals for treating patientswith cutaneous lymphomas or Sezary syndrome; 5 of 6

patients who received murine monoclonal antibody by theintravenous route showed good partial responses and, thoughpatients did indeed make antibodies to murine proteins, toxicmanifestations were very mild or non-existent. A patientwith B cell lymphoma offollicular-centre cell type was treatedwith an anti-idiotypic monoclonal antibody raised against hisown lymphoma cells; after initial apparent failure of the

experiment, his disease regressed and he achieved a completeremission that has lasted so far 11 months. There can be little

2. Kohler G, Milstein C Continuous cultures of fused cells secreting antibody ofpredefined specificity Nature 1975, 256: 495-97.

3 Prentice HG, Blacklock HA, Janossy G, Bradstock KF, Skeggs D, Goldstein G,Hoffbrand AV. Use of anti-T-cell monoclonal antibody OKT3 to prevent acutegraft-versus-host disease in allogeneic bone marrow transplantation for acuteleukaemia. Lancet 1982; i: 700-03.

4. Miller RA, Maloney DG, Warnke R, Levy R. Treatment of B-cell lymphoma withmonoclonal anti-idiotype antibody. N Engl J Med 1982; 306: 517-22

5. Miller RA, Levy R Response of cutaneous T cell lymphoma to therapy with

hybridoma monoclonal antibody. Lancet 1981, ii: 226-29.

1056

doubt that for patients with leukaemia and lymphoma, as forpatients with other diseases, the advent of monoclonalantibody technology offers prospects of considerable benefit.

BRANHAMELLA CATARRHALIS: PATHOGEN OROPPORTUNIST?

Branhamella catarrhalis (formerly Neisseria catarrhalis) is agram-negative coccus which occurs as a commensal in theupper respiratory tract. This organism can, however, bepathogenic, and some laboratories draw attention to it whensputum contains large numbers. If we disregard B.catarrhalis on most occasions when it is isolated, will we beunderestimating its potential as a pathogen? Isolation of aheavy growth of the organism from sputum, in the presenceof large numbers of polymorphs, is suspicious, but sputumcultures may be difficult to interpret. If the patient has latelybeen on antibiotics, the bacterial population of the sputummay be completely different from the original infectingorganism. The sample itself may be heavily contaminatedwith pharyngeal flora or the culture may be made from a non-purulent portion of the sample; the result is therefore

unrepresentative of the bacteria present in the lungs. Whatbetter evidence of pathogenicity should we seek in thesedifficult circumstances? Transtracheal aspiration is one wayof obtaining sputum samples uncontaminated by upper-respiratory-tract flora; patients do not enjoy this procedure.Another approach is to search for high or rising antibodytitres against branhamella. Isolations of the organism fromblood or other normally sterile body fluids are rare butconstitute definite evidence of invasion.There have been a few reports of pneumonia due to

branhamella, based on sputum culture and transtrachealaspiration, together with suggestive X-ray appearance andclinical signs ; 1,2 almost without exception, the patients wereimmunocompromised or had underlying lung disease. Animportant piece of work was reported from Belgium byNinane, Joly, and Kraytman;3 they did transtracheal

aspirations on 193 coalminers with pneumoconiosis whowere febrile. Branhamella was obtained in pure culture from14 of 104 bacteriologically positive aspirates. When sputumhas been secured by coughing, the meaning of branhamella isuncertain; other workers2,4 point out the frequency withwhich it is isolated in mixed culture with recognisedpathogens, and the high proportion of isolates made frompatients already treated with antibiotics. Lately branhamellahas been cultured from the throats of 55% of 40 adults with

laryngitis. This was significantly different from the isolationrate in a control group without throat infections, and also farexceeded isolations of the pneumococcus or Haemophilusinfluenzae. Virus culture was attempted in those with

laryngitis but no virus was isolated. In view of the highproportion of sore throats which are thought to have a viral

1. Srinivasan G, Raff MJ, Templeton WC, et al. Branhamella catarrhalis

pneumonia-report oftwo cases and review ofthe literature. Am Rev Resp Dis 1981;123: 553-55.

2 Johnson MA, Drew WL, Roberts M. Branhamella catarrhalis—a lower respiratorytract pathogen? J Clin Microbiol 1981; 13: 1066-69

3. Ninane G, Joly J, Kraytman M. Bronchopulmonary infection due to Branhamellacatarrhalis; 15 cases assessed by transtracheal puncture. Br Med J 1978, 1. 276-78.

4. Percival A, Corkill JE, Rowlands J, Sykes RB. Pathogenicity of and &bgr;-lactamaseproduction by Branhamella (Neisseria) catarrhalis. Lancet 1977; ii. 1175.

5. Schalen L, Christensen P, Kamme C, et al High isolation rate of Branhamellacatarrhalis from the nasopharynx in adults with acute laryngitis Scand J Infect Dis1980; 12: 277-80.

origin, this casts some doubt on the findings. There was alsono record of attempts to isolate Streptococcus pyogenes.Branhamella is said to be responsible for some cases ofotitis

media. Coffey6 found it in pure culture in 5’ 2% of samples ofmiddle-ear fluid from children with ear infections. This is

very suggestive that the organism is a true pathogen in thesecircumstances, since middle-ear fluid would normally besterile. However, viral culture was not attempted in thisinvestigation. Likewise, isolations from the cerebrospinalfluid, of which there are sporadic reports,’,8 must be taken asevidence of pathogenicity. Feigin and colleagues7 found 19reports of branhamella meningitis between 1908 and 1969.Some of these cases also had positive blood cultures, and asmall minority presented with purpura resemblingmeningococcal septicaemia. It has been suggested thatbranhamella meningitis may have been under-reported in thepast, owing to incorrect assumptions that neisseria-likecultures from cerebrospinal fluid were meningococci.So far, serological evidence of pathogenicity has been hard

to interpret. Brorson and colleagues9 examined paired serafrom patients with maxillary sinusitis of unstated

bacteriological cause. They found precipitating antibodiesagainst branhamella in almost all these patients and in somehealthy controls. Complement fixing antibodies were foundin about one-fifth of the study group, but maxillary sinusaspirates from some of these were sterile. The antibody titreswere low and titre changes were of small magnitude. The serawere not absorbed before testing, but a separate experimentshowed no cross-reactivity of antisera (raised in rabbits)against the branhamella antigen used in the tests, and againsta control antigen prepared from Neisseria gonorrhoeae.Leinonen and colleagues 10 used an enzyme immunoassay toexamine sera from children with otitis media due to

branhamella. Again, the sera were unabsorbed before testing,so cross-reactivity was not excluded. They were unable todetect IgM antibodies, possibly because the method wasunsuitable, but demonstrated the presence of IgA and IgG.About two-thirds of the children studied showed a rise in

titre; in the control group (children with otitis media due toother organisms) no IgA was detected but some did haverecognisable titres of IgG against branhamella. In addition,IgG and IgA were identified in 8 of 11 middle-ear fluidstested. No-one seems to have looked at antibody responsesagainst branhamella in patients with chest infections.Branhamella in sputum cultures can be disregarded if the

patient was previously healthy. However, in the presence ofchronic lung disease or compromised immunity, cultures ofbranhamella from purulent sputum should be taken moreseriously. It seems to be an opportunist which is more

frequently commensal than pathogen. When it is isolatedfrom cerebrospinal fluid or blood it is obviously behaving as apathogen. Branhamella otitis media, being neither rare norconfined to the compromised host, falls somewhere in themiddle. When treatment for a branhamella infection is in

view, it should be remembered that many strains produce/3-lactamase and tetracycline is probably the best first-linetherapy for infections of chest or ear.

6. Coffey JD. Otitis media in the practice of pediatrics. Pediatrics 1966; 38: 25-32.7. Feigin RD, SanJoaquin V, Middelkamp JN. Purpura fulminans associated with

Neisseria catarrhalis septicaemia and meningitis. Pediatrics 1969, 44: 120-23.8 Pfister LE, Gallagher MV, Potterfield TG, Brown DW Neisseria catarrhalis

bacteraemia associated with meningitis. JAMA 1965; 193: 399-400.9. Brorson JE, Axelsson A, Holm SE. Studies on Branhamella catarrhalis with special

reference to maxillary sinusitis Scand J Infect Dis 1976; 8: 151-55.10. Leinonen M, Luotonen J, Herva E, Valkonen K, Makela PH. Preliminary serological

evidence for a pathogenic role of Branhamella catarrhalis. J Infect Dis 1981; 144:570-74.