INST. FOR CELLFORSKNING OCH GENETIK
MEDICINSKA NOBELINSTITUTET
KAROLINSKA INSTITUTET
STOCKHOLM 60
Telefon 23 54 80

INST. FOR CELL RESEARCH AND GENETICS March 1,1953
KAROLINSKA INSTITUTET
STOCKHOLM 60

Dr Joshua Lederberg
Dept.of Genetics
College of Agriculture
University of Wisconsin
Madison 6, fisconsin

Dear Dr Lederberg:

Thank you very much for your letter and for the most interesting
reprints which you have kindly sent me. May I inquire whether you have any
plans of going to Italy this Summer and participate at the Congress of
Genetics and/or Microbiology? It would be certainly a very great pleasure
to be able to meet you there personally, as I shall probably attend the
genetical congress. Of course, it would be still more wonderful if you
could visit Sweden at the same time...

We are just finishing a paper on some quantitative studies on the
growth of two established ascites tumors. At various intervals after inoculation
of various cell doses we have determined a).the amount of peritoneal fluid,
b)ethe total number of free tumor cells, and c).the total number of non-
tumorous exudate cells. It appears that there is a certain amount of peritoneal
fluid present in the normal,untreated mouse} we call it Vo and its amount in
mice of constant weight,strain and sex is about 0.18 ml. This fluid contains
quite a number of non-tumorous cells (n-cells) the number of which we call
e,° When we inject a certain number of ascites tumor cells (T-cells) ,they intermix

with the available fluid and also provoke the production of some new fluid.
Surprisingly enough,we find that the amount of peritoneal fluid after inoculation,
and,as a matter of fact, at any given time during ascites tumor growth can be —
adequately described by the following simple expression:

Vou pT Tv,
and the number of n-cells at any given time is
e= k.T +e

where V is the amount of peritoneal fluid at a certain point of time,
T is the number of T-cells and e is the mmber of n-cells at the same time,
p and k are constants and Vo and €, are as defined above.

If we inject a comparatively small number of T-cells,tim which is
still above the critical level needed for the production of ascites tumors in 100
per cent of the inoculated animals, the product p.T and k.T,respectively,is

negligible compared to V5 and @, in the above equations, and one may say that
the cells inoculated are being simply diluted by the av@dlable peritoneal: fluid

and its content of n-cells. And still,the cells grow readily as free cells within
this fluid, and the amount of fluid increases appreciably only after the number

of cells has increased so mich t
value of p is about 130 + 10-6.}, t never neglicible (the
This would mean that a provocation of new exudate by the injected cells is not

a necessary condition for their growth,as they grow perfectly well in the fluid
that is available ~ but, and ties what surprises us too to a great extent,
only until the inoculum dose is not _ being decreased below the "eritical level"
which is usually of the order of 10° T-cells. If inoculum dose is being decreased
further, an increasing percentage of the animals develop only solid tumors ,usually
at the site of inoculation. This means that a much smaller number of T-cells can abreoclg
survive and mltiply in the solid tissues of the host than in the peritoneal fluid.
Another observation is that the"lag phase" (defined as the time needed before the
number of free T-cells increases to a value significantly higher than the dose of
jnoculum) is inversely related to the number of inoculated cells in the case

of inoculum doses above the critical level. In analogy with Hinshelwood’s theory
to explain bacterial lag,we have tried to speculate that there might be some

active substance which tem is being contributed by the inoculateaé cells and which
has to reach a certain critical concentration in the peritoneal fluid before the
cells can start to grows (I dont know how much Hinshelwood?’s theory is accepted

for the case of bacterial lag,I mst confess.) The peritoneal fluid is continuously
circulating, and some experiments of Prentice et al. with tritium-labeled water
have shown that as much as 40 to 80 per cent is being exchanged during one hour

in case of much larger volumes than what we are dealing with. One could imagine
that this continuous circulation necessitates the inoculation of at least 10%cells,
otherwise the hypothetical active product becomes too diluted and never reaches

the necessary critical concentration. Circulation of tissue fluid in solid tissues,
where the tumor cells inoculated are crowied together,might be slower andamieHET So oy b
enable the cells to build up the critical concentration of the active substance.
All this is pure speculation and we try to design experiments to get closer

to the problem. Your suggestions are certainly most welcome at any time.

The cultivation of ascites tumor cells in vitro,preferably in the form
of suspensions, is very desirable indeed. we have made some very preliminary expe-
riments several years ago, but have given it up,imewm realizing the rather enormous
technical difficulties. whet we did was to try to grow them in their own ascites
sgerum,diluted and supplied with embryo extract, and this was a complete failure.
I tried to convince Dr.Earle at the National Cancer Institute that this is just
the problem which he should solve.{f dont think he was very convinced,although he
vrote me some time ago that they will undertae some preliminary studies. I think
he has the greatest experiemce and best organization to solve the question. However,
if he looses interest,we are decided to try it again.

I am sure you will be more careful in the future to write me a letter
when you get such a long and tedious answer. There is a great number of problems
I should very much like to discuss with you in connection with the working hypothe-
sis on a "mutation-selection" origin of ascites tumors. I hope that the opportunity
will come in the not too far future.

With kindest regards,

very eincer«ly yours,
3 C .

P.SeDid your paper on genetic exchange in Salmonella appear yet? If reprints are
available, I should very much appreciate receiving one,