SOME ASSOCIATIONS BETWEEN BLOOD GROUPS AND DISEASE

SOME ASSOCIATIONS BETWEEN BLOOD GROUPS AND DISEASE J. A. FRASER ROBERTS MX). D.Sc. F.R.C.P. Medical Research Council Clinical Genetics Research Unit Institute of Child Health The Hosital for Sick Children Great Ormond Street, 1 The reality of some associations between blood grous and disease 2 Positive findings 3 Negative findings 4 Subdivisions, and reliminary and conflicting results 5 Meaning of the associations References It is only during the ast five years that really strong evidence has been forthcoming that ersons belonging to the different ABO blood grous differ in their suscetibility to certain diseases of adult life. Strong though that evidence now seems to be, it has met with a scetical recetion in some quarters. Various objections have been set out at length by Manuila (1958) in a recent aer. Accordingly, the first art of this aer reviews in some detail the nature, and the strength, of the evidence for a few selected diseases, doing so more articularly in the light of the criticisms that Manuila has so conveniently marshalled. 1. The Reality of some Associations between Blood Grous and The first disease for which ositive findings emerged on a scale sufficient to carry fairly general conviction was cancer of the stomach (Aird, Bentall & Frascr Roberts, 1953), but the clearest icture of a blood-grou association yet obtained is that of grou O with duodenal ulceration. The results for from ten centres are shown in Table I. It will be seen that at every centre the frequency of grou O is higher, usually markedly higher, in those suffering from duodenal ulcer than it is in the controls. Conversely, at every centre A's are less numerous than in the controls. In nine of the ten B's are also less numerous. AB's are, of course, relatively few, but are reduced at seven centres, the general trend being unmistakable. A more convenient way of exressing the results, however, is to turn each into the incidence of the disease in ersons of one grou relative to the incidence in another grou or grous. This is done by simle cross-multilication. For examle, the duodenal ulcer includes 535 O's and 311 A's, the corresonding figures for the controls being 4,578 O's and 4,219 A's. 535x4,219 311x4,578 = 1.59 We can say, therefore, that in this material the incidence of duodenal ulceration in ersons of grou O is 1.59 as comared with an incidence of one in ersons of grou A. Table n reeats the results of Table I in this alternative form, the incidences shown being those in grou O as comared with an incidence of one in the other grous taken collectively. The dearture from unity is very highly significant at six centres, the robability of getting so great a deviation by chance varying from one in 100,000 to one in 100 million or less. At two centres, and Vienna, though the are large the relative incidence is lower, so the robabilities lie between one in 100 and one in 1,000. At Manchester and in the small Iowa of negroes the deviations are not significant at the 5 % level, though the trend is in the same direction. The question has to be asked, however, whether these figures can be taken at their face value. Manuila (1958) makes the general oint that the limits of chance fluctuations are TABLE I. ASSOCIATION OF BLOOD GROUPS WITH INCIDENCE OF DUODENAL ULCERATION i. ii. Manchester iii. iv. Liverool v. vi. vii. Oslo viii. Vienna ix. Iowa (whites) x. Iowa (negroes) Number in disease 946 423 482 1,059 1,642 680 579 1,160 1,301 173 Number in control 9,370 13,572 15,377 5,898 14,304 8,292 6,313 6,722 56.6 53.2 59.8 59.6 57.7 50.3 48.7 41.0 53.7 56.1 Grou O 48.4 48.6 53.9 40.6 37.8 49.2 Percentage frequencies in disease and control 32.9 35.0 32.6 29.0 31.5 38.4 43.2 41.3 25.4 Grou A 40.3 38.8 39.1 32.3 44.0 50.0 44.2 41.6 26.5 7.8 8.7 6.2 8.1 8.8 8.8 5.2 11.2 7.8 16.8 Grou B 8.9 8.4 9.7 9.4 10.8 10.9 8.2 13.4 9.0 20.1 2.7 3.1 1.5 3.3 2.0 2.5 2.9 6.5 2.2 1.7 Grou AB 3.1 19 2.9 2.5 3.0 4.5 4.0 6.0 3.6 4.3 8,445 Sources of material: l-lx, references given by Roberts (1957); x, Buckwalter, Turner, Raterman, Tldrlck & Knowler (1957) 129 VoL 15 No. 2

TABLE II. DUODENAL ULCERATION: RELATIVE IN- CIDENCE IN PERSONS OF GROUP O COMPARED WITH INCIDENCE OF ONE IN PERSONS OF GROUPS A. B AND AB Manchester Liverool Oslo Vienna Iowa (whites) Iowa (negroes) Relative incidence.54.21.57.54.17.48.56.22.37 1.32 X 1 39.83 3.76 22.75 44.26 7.51 25.06 26.77 9.95 26.49 3.21 209.59 <io- 18 <io- 5 <io- i» <io-«<io-«< io-«greater than is sometimes suosed. These limits are, of course, calculable, and the calculations are of the utmost simlicity rovided that a single comarison is being made. Almost without excetion the writers who have rovided the recent data have used erfectly efficient methods. That these methods do look after the oerations of chance is shown by results obtained by the Nuffield Blood Grou, at the Royal Anthroological Institute,. Some hundreds of thousands of grouings of donors have been analysed for Great Britain, and reeated tests made of the agreement between successive samles from the same centres. The differences turn out to be extremely close to those theoretically exected (Koed, ersonal communication, 1958). The ercentagefiguresgiven as illustrations by Manuila look curiously large. This is because he is adoting the highly unusual level of conventional significance of one in instead of the one in 20 or one in 100 commonly chosen. This underlying assumtion was stated in an earlier aer (Manuila, 1945), but is not exlained in the aer of 1958. The next oint is the ossible contribution of technical errors. It is unnecessary to exand this further. All the data used in the recent analyses come from countries with highly develoed transfusion and hosital services, and all the tests have been carried out in well-equied laboratories. Anything more than an exceedingly small roortion of wrong ABO grouings is inconceivable; such errors would have become known through trouble and accidents on a massive scale. Moreover, even if any areciable roortion of wrong grouings could be conceived these would robably be distributed at random and merely dilute any association that was resent. A much more imortant objection is the ossibility that hosital atients and donor controls might not be drawn from entirely corresonding oulations. For this reason second line controls are very desirable. These can be furnished by atients from the same hositals suffering from other diseases found not to show ABO associations. To some extent this has already been done at a number of centres. There is also the direct comarison of duodenal ulceration with cancer of the stomach in atients from the same hositals. The differences are very highly significant. So if, for examle, there were a tendency at any centre for the control samle to be, as comared with the hosital atients, rather too high in A, the association with duodenal ulcer would be somewhat increased, but the association of A and cancer of the stomach would be reduced, and vice versa if the controls were too high in O. In any event the only disease associations that can be recognized with racticable numbers are those which are relatively very large. The differences in frequencies to which they lead are of an entirely different order of magnitude from any discreancies between alternative rationally selected control samles. Such discreancies could have no areciable effect on the results. Turning to cancer of the stomach, Table m gives the findings at a number of centres. This time only grous O and A are shown, though it may be mentioned in assing that grou B once again goes with the grou in which the incidence of the TABLE III. ASSOCIATION OF BLOOD GROUPS WITH INCIDENCE OF CANCER OF THE STOMACH i. ii. Manchester iii. iv. Liverool v. Birmingham vi. Leeds vii. viii. Basel ix. x. Vienna xi. Milan xii. Iowa xiii. Sydney Number in disease 1,437 770 101 217 100 217 299 704 798 1,146 678 908 483 Number in control 9,370 13,572 15,377 9,590 6,260 5,898 4,518 14,304 2,346 6,313 30,000 Di tease 42.4 44.5 43.6 39.2 37.0 42.4 53.2 38.6 47.8 Grou O Percent jge frequencies 48.4 48.6 47.5 47.4 53.9 41.7 40.6 42.9 46.8 45.3 43.6 44.7 57.0 47.9 34.8 53.1 51.0 44.1 48.2 40.0 Grou A 40.3 38.8 39.1 42.1 41.2 32.3 45.1 44.0 44.2 41.8 41.6 38.4 7,858 Sources of material: i-x, xli, xiii, references given by Roberts (1957); for, data from Jordal (1956) are also used; two combined; xi, Beolchini, Cresseri, De Maria, Morgantl, Peruzzotti & Serra (1957) 130 Brit. med. Bull. 1959

BLOOD GROUPS AND DISEASE J. A. Fraser Roberts TABLE IV. CANCER OF THE STOMACH: RELATIVE INCIDENCE IN PERSONS OF GROUP A COM- PARED WITH INCIDENCE OF ONE IN PERSONS OF GROUP O Manchester Liverool Birmingham Leeds Basel Vienna Milan Iowa Sydney Relative incidence.20.22.25.31.60.31.09.36.30.00.28.20.06 X 1 8.89 6.39 1.12 2.75 4.46 3.52 0.45 12.16 10.86 0.00 6.85 5.61 0.41 63.47 < 0.001 < 0.001 disease is low, this time grou O. At 12 of the 13 centres, the atients show a higher frequency of grou A, and the frequency of grou O is higher in all 13 sets of controls. At one centre, Vienna, the figures are ractically identical. It will be seen, however, that the differences are much smaller on the average than in duodenal ulceration. Table IV shows the same figures in terms of relative incidences. The figure for A relative to O is higher at all centres excet Vienna, but the differences from unity are on the average only about half those shown by duodenal ulceration. The samles are also smaller on the whole, so these two factors lead to quite a different icture in terms of x*. At two centres, Basel and, the robability is just a little less than one in 1,000; at two, and Milan, it is less than one in 100; at two, Manchester and Iowa, it is less than one in 20; but at six centres it fails to attain the 5 % level of significance. Clearly the evidence from individual centres taken in isolation is far from strong and, unless it can be ooled in some way, the conclusion must remain doubtful. On this oint, as on others, Manuila aears misleading. He considers that the combination of results from different centres, with different blood-grou frequencies in their oulations, is inadmissible, being under the imression that what is being combined is frequencies of blood grous. He says (Manuila, 1958):... the methodology which enabled these authors to combine their cases, and which is said to ermit other workers to do so, is not accetable to the anthroologist, since it would tend to render the control so highly heterogeneous that, in fact, they would reresent a urely artificial, unreliable oint of reference. What in fact is being combined is not, of course, blood donors, but indeendent tests of significance, a rocedure which is a commonlace of everyday statistical work. An entirely general method has been given in all editions save the earliest of Fisher's Statistical methods (Section 21.1, Fisher, 1941). This takes account only of the robabilities, and not of the detailed comosition of the data from which they are derived, which might be of very different kinds. Alying this method to the data on which Table IV is based, it is found that the chance of obtaining the aggregate of robabilities observed at the 13 centres is about one in 50,000. This general test is, however, making use of only a art of the available information. An excellent method, articularly aroriate to the data unocr consideration, has been devised by Woolf (1955). This leads to an efficient estimate of the average relative incidence, which is 1.20. The total x 1 of 63.47 is divided into two arts. The first art, with one degree of freedom, tests the significance of the difference of the general average from 1.00. It is 48.77, or very nearly seven times its standard error, and the robability of obtaining so great a deviation by chance is infinitesimal. The second art, ax 1 of 14.70, with 12 degrees of freedom, reresents heterogeneity, that is, the deartures of the individual incidences from the general average. The robability is 0.3, so there is no evidence that the individual centres are showing anything more than chance fluctuations about the general average. It should be reeated, erhas, that this rocedure does not roduce a "heterogeneous" control, or an "artificial, unreliable oint of reference." At each centre, quite indeendently, the question is asked: Is cancer of the stomach commoner in ersons of grou A than in those of grou O, and by how much? These indeendent estimates are then efficiently combined to give, first, a general estimate, and, second, a measure of the deartures of the individual centres from the general average. A final examle illustrates evidence which, though far from overwhelming as in duodenal ulcer and cancer of the stomachi is nevertheless strong. The ublished for ernicious anaemia amount to only 1,498 atients altogether. The results are summarized in Table V. At two centres, and San Francisco, the excess of grou A relative to grou O is marked, giving quite low robabilities. At Iowa the robability just attains the 5% level of significance. Sheffield and give a moderate, non-significant excess. At Oxford, Cambridge and the excess is negligible, while at there is a small excess of grou O. The general test mentioned above gives this collection of robabilities a combined robability of about one in 500. Woolf's method gives a combined weighted relative incidence of 1.26, and the robability of so great a deviation from unity TABLE V. PERNICIOUS ANAEMIA: RELATIVE IN- CIDENCE IN PERSONS OF GROUP A COMPARED WITH INCIDENCE OF ONE IN PERSONS OF GROUPO Oxford Cambridge Sheffield Iowa San Francisco Number tn disease 244 258 110 123 109 270 III 158 115 1,498 Relative incidence.63.09.01.36.03.19 c).98.42.70 X* 12.51 0.41 0.00 2.35 0.02 I.SS 0.01 3.97 6.96 27.78 0.0004 O.S 0.1 0.9 0.2 0.9 0.05 0.008 Sources of material: references given by Roberts (1957). Samles combined for and for. 131 Vol. 15 No. 2

arising by chance is about one in 16,000. x* for heterogeneity is 11 for eight degrees of freedom, corresonding to a robability of 0.2. With scries of this size, therefore, the wide range of variation shown by the centres is only what is to be exected owing to the oerations of chance. 2. Positive Findings Two associations for which the evidence is overwhelming have been mentioned at some length in the receding section. Duodenal ulceration is commoner in ersons of grou O than in those of grous A, B or AB. Even for the AB's the difference is very highly significant. The incidences in grous A, B and AB are closely similar. Cancer of the stomach is commoner in ersons of grou A than in those of grous O and B, for which the incidences are again closely similar. Numbers are as yet insufficient for any conclusion about grou AB. The evidence is also overwhelming for an increased incidence of gastric ulceration in ersons of grou O, but the relative increase is definitely less than for duodenal ulcer, the figure for O as comared with the other grous being about 1.20. The data are summarized by Fraser Roberts (1957). Some qualification is required, however. The classification of gastric ulceration is neither clear cut nor generally agreed. It may be that there are gastric ulcers, as ordinarily classified, which should be considered together with duodenal ulcer, while others arc different and might show no excess of grou O. Some interesting attemts have been made at further analyses, but all that can be said with confidence at the moment is that, taking gastric ulcer as classified by those who have ublished the, there is a highly significant excess of O. Moreover, the are erfectly homogeneous on the numbers available. Some have been concerned because the figures at different centres range from no excess to an excess as great as in duodenal ulcer, but this is no more than would be exected by chance, and there is no evidence that the are not showing random fluctuations about a general average figure of 1.2. For ernicious anaemia, as mentioned in the receding section, the evidence for an excess of A is very strong. The evidence for an excess of A in diabetes mellitus is fairly strong (Fraser Roberts, 1957). A articularly interesting association was discovered by Clarke, Edwards, Haddock, Howel-Evans, McConnell & Sheard (1956). Duodenal ulceration is commoner in nonsecretors of grou secific substances than in secretors. This association is a good deal stronger than the association with grou O. Very recently Helmbold (1958) has roduced evidence showing that uterine cancers are commoner in women of grou A than in those of grou O. The samle consisted of 7,115 atients from a number of centres in western Germany. The incidence of uterine cancers in grou A relative to that in grou O was 1.15; x* being 21 for one degree of freedom, it differs from unity with high significance. A samle of 205 atients from Milan (Beolchini, Cresseri, De Maria, Morganti, Peruzzotti & Serra, 1957) also shows an excess of A. 3. Negative Findings Positive findings of the kind reviously described demand for their sure recognition numbers to be counted in thousands rather than hundreds. The same is true for negative findings. Of course, it can never be asserted that there is no association but, as numbers increase, the limits of ossible excess of one grou or another, using any level of robability we choose to select, become increasingly narrow. A number of negative findings in the sense of fairly narrow limits have been established. Cancers of the lung, breast, and colon and rectum fall into this category. If one cares to combine the results for these three grous of cancer, the numbers amount to 7,600. Incidence in grou A is 1.03 relative to grou O, x' being no more than 1.24 for one degree of freedom (Fraser Roberts, 1957). Maxwell & Maxwell (1955) found no significant association in 2,147 atients suffering from hyertension. A number of other results, though on rather smaller numbers, show no evidence of ABO associations. Reasonably large samles of Rhesus grouings (ositive and negative only) are available for some diseases for examle, etic ulcer and cancers of the lung, breast, and colon and rectum (Aird, Bentall, Mehigan & Fraser Roberts, 1954) but so far no clear indication of any disease association has emerged. 4. Subdivisions, and Preliminary and Conflicting Results When associations have been roved to exist, it is clearly of much interest to see whether differences are revealed on subdivision by sex, age and, more articularly, tye, course and site of the disease. The difficulty so far has lain in securing adequate numbers, and little has emerged u to the resent that can be regarded as more than suggestive and as an indication for further work. There are some interesting findings on the site of the tumour in relation to cancer of the stomach (Balme & Jennings, 1957; Billington, 1956). There robably is variation, but numbers are small as yet and the results at different centres are in at least artial disagreement. One finding, however, is outstanding. Brown, Melrose & Wallace (1956) found that stomal or anastomotic ulcers are articularly common in ersons of grou O, the incidence being almost double that in grou A. Doll (ersonal communication, 1958) has now obtained just the same result on a considerably larger. Stomal ulcers reresent the most extreme form of the ulcer diathesis, so this seems to be a ointer to a true causal connexion. Some ositive results may be emerging for a few other diseases. For examle, Aird, Lee & Fraser Roberts (1959) have found a moderately significant excess of grou A in cancer of the ancreas in a of 600 atients. This cannot be regarded as more than an indication for further work, but is interesting because it relates to yet another disease associated with the uer art of the gastrointestinal tract. Some results are definitely conflicting. For examle, Struthers (1951) found at a very large and highly significant excess of grou A in infants with bronchoneumonia coming to ost-mortem. Carter & Heslo (1957), on the other hand, found no excess in a of similar size. Mayr, Diamond, Levine & Mayr (1956) found a very large excess of grou O in atients suffering from chromohobe adenoma of the ituitary. A subsequent (Damon, 1957) showed a much smaller excess, while Aird et al. (1959) found no excess at all in a larger from Great Britain. The discreancies for both diseases are too large to be accounted for by chance. It would be unfortunate, however, if negative results were to be referred simly 132 Brit. med. Bull. 1959

BLOOD GROUPS AND DISEASE J. A. Fraser Roberts because they are negative to those which are ositive. It may well be there is still a henomenon to investigate. It is difficult to think of reasons for a large and highly significant association at one centre and no association at all at another or others, the differences being beyond the limits which could reasonably be ascribed to chance. Nevertheless, if the work is ursued it may be found that the differences are real, and if so a roblem of much interest would arise. It is because of such discreancies as these that one articularly strikingfindinghas been relegated for the time being, though erhas unfairly, to this section. Cameron (1958) found 206 who were grou A and 100 who were grou O amongst atients with salivary gland tumours against 1,906 who were grou A and 3,177 who were grou O in the corresonding controls. Highly significant though this difference is, it is desirable to await results from other centres just in case there should once again be a uzzling discreancy. A final oint should be emhasized. It is doing a oor service to the subject to mix u results for different diseases based resectively on large and very small numbers, to mix u REFERENCES Aird, I., Bentall, H. H., Mehigan, J. A & Roberts, J. A. Fraser (1954) Brit. med.j. 2, 315 Aird, I., Bentall, H. H. & Roberts, J. A. Fraser (1953) Brit. med. J. 1, 799 Aird, I., Lee, J. A. H. & Roberts, J. A. Fraser (1959) (In rearation) Balme, R. H. & Jennings, D. (1957) Lancet, 1, 1219 Beolchini, P. E., Cresseri, A., De Maria, B., Morganti, G., Peruzzotti, R. & Serra, A. (1957) Monogr. Analecta genet. 6, 109 Billington, B. P. (1956) Lancet, 2, 859 Brown, D. A. P., Melrose, A. G. & Wallace, J. (1956) Brit. med. J. 2, 135 Buckwalter, J. A., Turner, J. H., Raterman, L., Tidrick, R. J. & Knowler, L. A. (1957) /. Amer. med. Ass. 165, 327 Cameron, J. M. (1958) Lancet, 1, 239 Carter, C. O. & Heslo, B. (1957) Brit. J. rev. soc. Med. 11, 214 infinitesimal robabilities with those that are border-line, and conflicting results with those that are established. A few blood-grou and disease associations certainly exist. It is, therefore, a ossibility to be exlored in a hilosohical sirit with other diseases; time and numbers will bring enlightenment. 5. Meaning of the Associations Sace does not ermit an elaboration of this theme; but in fact there is little to say at resent. It seems highly unlikely that the roved associations are merely secondary to racial or other stratifications in the oulation. They are too large and too wide sread. The uer art of the gastrointestinal tract is too definitely imlicated. For one association nonsecretion and duodenal ulcer stratification has been disroved, for the subjects who are non-secretors are more suscetible than their sibs who are secretors (Clarke et al. 1956). If then the associations are real, in the sense of being rimary, interesting and imortant ossibilities resent themselves; but, as yet, their investigation has hardly begun. Clarke, C. A., Edwards, J. W., Haddock, D. R. W., Howel- Evans, A. W., McConneU, R. B. & Shcard, P. M. (1956) Brit. med. J. 2, 725 Damon, A. (1957) Science, 126, 452 Fisher, R. A. (1941) Statistical methods for research workers, 8th ed. Oliver & Boyd, Edinburgh Helmbold, W. (1958) Seventh Congress of the International Society of Blood Transfusion, Rome, 3-6 Setember, 1958,. 34 [Abstract] Jordal, K. E. (1956) Acta Med. leg. soc. 9, 195 Manuia, A. (1945) Arch, sidsses Anthro. gin. 11, Annexe Manuila, A. (1958) /. Amer. med. Ass. 167, 2047 Maxwell, R. D. H. & Maxwell, K. N. (1955) Brit. med. J. 2, 179 Mayr, E., Diamond, L. K., Levine, R. P. & Mayr, M. (1956) Science, 124, 932 Roberts, J. A. Fraser (1957) Brit. J. rev. soc. Med. 11, 107 Struthers, D. (1951) Brit. J. soc. Med. 5, 223 Woolf, B. (1955) Ann. hum. Genet. 19, 251 133 VoL 15 No. 2