Data interpretation, the failure missed


On Thursday/Friday we made the case for the importance of B cells.

Yesterday, we made the case that the data from clinical trials argued against a role for T cells.

MrT was quiet. 
Maybe MrT knows better than to say something:-). 
Maybe he/she has changed his/her world view.  

Have You?

There was not too much debate

This says you don't either understand what I am on about, 
you don't care or perhaps 
you don't know how to read the data ....................otherwise you may have called me on it, because in some cases I was talking rubbish.

What's new...I hear you say:-(

You would think, I would get called out, 
there are people who accept this thing without question. 

Sadly it is often made by someone called a referee:-(.

However, there are always two sides to a story and I only gave you one yesterday to make a point. 

So forgive me if you feel you were led astray

There are a large number of papers reporting an increase in the levels of memory T cells after alemtuzumab and fingolimod, as I showed you. 

However, this view is complete and utter non-sense of course, 

So the calls to ProfG by Pharma were wasted (white lie:-), alemtuzumab and fingolimod both deplete memory T cells.

So what did I show? 

I showed that the proportion of CD4, CD45RO goes up.

To make an example let's say CD4CD45RO increase from 10% to 40%, so it is a thirty percent increase right?

Wrong! It could be a number of things. 

The technique that people, typically immunologists, use to monitor cell numbers is a flow cytometer (see above) 

This detects markers say CD4 for T cells and a marker for CD45RO (memory cells) and it gives you a percentage as a read-out. So it can say of the CD4 population, x% also express CD45RO.  The flow cytometer counts say 10,000 cells per sample so in the population there were 40,000 CD4, CD45RO to be 40% or 10,000 to be 10%. This also means in the negative populations there are 90,000 or 60,000 cells. 

But is this increase because there are 30,000 more CD4, CDRO positive cells or because there are 30,000 less CD4, CD45RO negative cells? 

So in addition to knowing the percentage of cells, you need to know the absolute cell number in any given volume, so you can work out the absolute number of cells in the sample. 

This aspect is lacking in many of the publications.
This makes it very difficult to interpret.

So we could have 

10 positive + 90 negative = 10%,  1000 positive + 9000-ve = 10%  
 40 positive + 60 negative = 40%, 4000positive + 6000-ve = 40%

So in normal health  it could be that there are 1000 positive cells per mL 
and 9000 negative cells in 1 mL = 10%, but after depletion there are 40 cells that are positive cells per  mL and 60 that are negative so 40% = 30% increase in the proportion of cells, but if we say that within the mL the number of positive cells have gone from 1000 cells per mL going down to 40 cells/mL. 

Therefore in absolute numbers the numbers of cells have gone from 1000 down to 40 so actually a 96% depletion not a 30% increase. 

Do you get this...many, many scientists don't:-(. 

However, this has led to a problem because you are getting a misleading interpretation.

Yesterday, I showed you the figure from the paper indicating an increase in the effector memory (TEM) population, arguing it can't be T cells.

In the abstract it said "In MS patients, the percentages of central memory T (CCR7+CD45RO+) cells (TCM) and naïve T (CCR7+CD45RO-) cells decreased significantly, while those of effector memory T (CCR7-CD45RA-) ......increased in both CD4+T and CD8+T cells from 2 weeks to 12 months during fingolimod therapy". So authors implie the cells go up but....

Look in the TEM In the data set the first two columns are controls the next six are time after treatment so you can see the line = median halfway point goes up.

Tucked away in the supplementary data, not seen by many, is the truth. 

Of course effector memory T cells are depleted you just have to look at the absolute number of cells. The effector memory cells are depleted, but as a proportion not as much as naive and central memory.
Look at the TEM group. In the data set the first two columns are controls the next six are time after treatmentso you can see the line = median halfway point goes down

Shocking yes ......but this type of analysis is not uncommon and I am sorry to say I believe that this type of analysis has led to failure to appreciate the effects and side effects of MS drugs, and probably put people to un-necessary risk (posts being written to explain this). 

However, to go back to the point. So redoing the table of moderate/highly effective drugs based on absolute numbers you still can't argue for a direct action on depleting T cells of active drugs.
                    Memory T  Memory B 
Alemtuzumb    YES           YES
Fingolimod      YES           YES
CD20               NO             YES

T cellers still struggle and more bad news to follow

But the effects of the anti-CD20 can be indirect and block T cell function, leaving them to fight another day. 

Remember to be a lemming without questioning, can mean you end up over the cliff :-).