22 replies to this topic

[hannibellemo]

Poor Response to Second-Line Kinase Inhibitors in CML Patients With Multiple Low-Level Mutations, Irrespective of Their Resistance Profile.

This was originally published in the Dec. 30 issue of Blood. I know some publications are more respected than others and I don't know where this one falls. Unfortunately, all I can access is the abstract, wonder if anyone knows more about this Australian study?  This was the "take home message" from the article:

In this study of patients with imatinib-resistant CML and no identifiable BCR-ABL1 kinase domain mutations by direct sequencing, Australian investigators used a multiplex mass spectrometry assay to identify a poor-risk subgroup with multiple low-level mutations who did not respond to second-line tyrosine kinase inhibitors.

[Trey]

This is very interesting.  The Journal "Blood" is one of the most respected hematology publications.  But it is always about the researchers and author(s), who in this case are first class. 

This article says there are many kinase sub-mutations that are not detected by current kinase mutation testing.  It has been known for quite a while that there are over 100 verified kinase mutations, most of which do not affect TKI drug performance much, if at all.  The big ones are T315i, F317L, Y253H, E255, F359 and a few others.  But there are also many other kinase mutations that have not been verified, and some may affect TKI performance.  And some of these are very small sub-mutations that may not, by themselves, affect TKI drug performance; but if they are present in large numbers, these sub-mutations can aggregate to affect TKI performance.

By example, TKI drugs are like complex puzzle pieces.  A puzzle piece only fits in one spot.  If that puzzle spot has been "damaged" (surrounding pieces chewed on by the dog, etc) then our "TKI puzzle piece" will not fit, or maybe will not fit securely (may pop out).  So if the dog just chomped once lightly on a neighboring puzzle piece, the TKI puzzle piece may still fit well enough, although not perfectly.  If the dog really chewed on the surrounding pieces (multiple sub-mutations -- teeth marks) then the aggregated damage will prevent the TKI piece from fitting at all.  So if the leukemic cell's bcr-abl kinase has mutated in numerous small ways (chomp, chomp, chomp), then the aggregated sub-mutations can prevent the TKI drug from fitting into the slot where it does its work.  So this is one reason, although not necessarily the only explanation, for why TKI drugs can fail to work even when no kinase mutation is identified during a kinase mutation test. 

I explained the issue above as one of "damage", although it may rather be that the non-mutated leukemic cells are killed off first, and then the remaining mutated ones are those which can multiply without being affected by the TKI drug.  This likely explains why most TKI drug failures occur within the first 2 years after diagnosis, and that after approximately 2 years there are very few TKI drug failures for those who have done very well (MMR or better). 

http://bloodjournal....375535.abstract

[hannibellemo]

That was my assumption from the abstract but your analogy, Trey, made things crystal clear. Thanks, as always.

One thing, in your second scenario you mention the mutated cells gaining strength after the regular leukemic cells are killed off by the TKI drug. Wouldn't the mutated cells at that point be visible through current kinase mutation testing and, therefore, verifiable?

This sensitive mass spectometry; how available is it for general use and how expensive is it? Does this translate into early testing for those who burn through the current treatments and yet don't show any mutations through current kinase mutation tests?

This was one of the most interesting abstracts I've read in a long time but for now it's back to work...

Pat

(This was supposed to be a response to Trey but obviously those two "answer windows" threw me and I clicked on the wrong one.)

[Trey]

You asked "Wouldn't the mutated cells at that point be visible through current kinase mutation testing...?"

Kinase mutation testing is an inexact science.  It is not very accurate, from what I can tell.  Mainly, it can only detect the worst types of kinase mutations, not all 100+ of them.

"This sensitive mass spectometry; how available is it for general use and how expensive is it?"

It is not available to the public, and would require you to re-mortgage your house. 

[GerryL]

Hi Trey, possibly impossible question to answer at this time, but have they got any answers yet as to why people can suddenly become resistent to Gleevec at around 7 years?

Is it because they haven't reached CyCR or MMR?

Is it because they weren't taking their TKI regularly?

Or is it simply because the body/stem cells work out how to get round the TKI?

[Trey]

It is a rare event for those MMR or better, and no real answer. 

[matt92711]

I would suspect that in such cases there is a stem cell (or cells) that have some mutations that the TKI is having difficulty eradicating 100% of the time and as a result it persists and the longer it persists it gives it more time to develop a more serious mutation which once it shows up will send things quickly out of control. To contrast if the CML cells do not have mutations that give the TKI any difficulty then it is less likely that a line of mutated stem cells would persist for long giving rise to problematic mutations.

I think that there are different levels of stem cells causing the CML and I think the later mutations occur at the younger level of stem cells whereas the senior level does not get many mutations, but at the same time for whatever reason the TKI's are not very effective with the senior CML stem cells. I think that when the TKI's work they are eliminating the younger level of stem cells and therefore keeping out of circulation these stem cells which will likely develop further mutations if not eliminated. However if the TKI is stopped then the senior level of CML stem cells will produce younger stem cells thereby increasing the CML and eventually the CML will come back. Also I think the senior stem cell population does not increase relative to the healthy senior stem cell population so as long as the TKI is effective on the younger stem cells the person seems healthy. - Going back to your question in this scenario a possible explanation is a younger stem cell line with multiple mutations was not able to be completely eliminated by the TKI and at some point an additional mutation occurred which made the CML come back strong. - I wonder if once someone reaches CML or some other point if cycling TKI's would help target any such mutated cells. For a given TKI to be effective it must be taken in sufficient quantity for it to have a high chance of hitting all the CML cells, so taking multiple TKI's at once in smaller quantities would probably not work. However by cycling through TKI's I wonder if each cycle would increase the chance that a specific TKI would work against a specific low level mutation that is possibly present, thereby helping prevent these late term relapses.

Ultimately there is too much unknown. Until a more definitive understanding of the CML stem cells is known and how, why and when Blast Crisis occurs, all these possible answers are just theories.

[Trey]

Matt,

No one knows for sure, but would think the opposite, but have no research to prove it.  I believe the answer may lie in the quiescent cycles of the top level leukemic stem cells.  Those leukemic stem cells which hide in the marrow stromal layer niche go dormant for long periods (quiescence) may hold kinase mutations that can show up after they come out of hibernation and reproduce, introducing the mutated kinase leukemic cells into production.  Patient experience would tend to corroborate this since kinase mutations normally occur within a couple years of starting the TKI drugs, since statistics show that the probability of kinase mutations drops to near zero after a little more than 2 years.  So that is my own theory, which is made up on my own with no supporting research. 

[pamsouth]

Trey,  I posted a new question out there, but not sure I did it right.  So anyhow I will post it here, too, even though it is not the topic..

We have the bone marrow that make the stem cell, that makes all the blood.  Somewhere at the top of the chain, we have the Leukemia stem cell.  For CML + PH it is on the Myeloid side, that makes the red cells, platelets, white cell.  And these particular white cells are make up of granulocytes, which are the Neutrophils, Basophils, Eosinophils. Is that right so far?   So Where do the chromosomes come into play, what part of this diagram or equation do the chromosomes fit into all of this?

PamSouth

[matt92711]

Per wikipedia: Chromosome is an organized structure of DNA and protein found in cells. Basically the gene that is PH+ is located in a chromosome.

[pamsouth]

Matt, I guess a better question would be are the chromosomes (there are 23 #23 being the xx & xy) in every cell whether it be white cell, red cell, platelets, etc. ?

PamSouth

[matt92711]

The PH+ gene would obviously seem to be in a granulocyte stem cell.

At the top of the chain are the Hematopietic stem cells (HSC) which gives rise to all blood cells. There are different generations of these. The earliest can give rise to any blood cell line, whereas the younger generations are more specialized and give rise to specific blood lines. One of these blood lines is the Granulocytes and it would seem there is a common HSC or a Progenitor Cell that only produces blood cells in the Granulocyte series (Neutrophils, Eosinophils, and Basophils). This stem cell is apparently where the BCR-ABL mix-up occurs. Further down the Granulocyte line there may be younger generations of stem cells that give rise to each specific type of Granulocyte. Through cell signaling mechanisms the body is able to signal the appropriate level of stem cell to produce the needed blood supply.

One of the best examples I saw of this cell signalling is how the red blood cells are signaled. When the blood is filtered through the Kidneys the Kidneys secrete a certain protein in response to the level of oxygen being delivered by the red blood cells. When the oxygen levels go low more protein is secreted which when these proteins reach the red blood cell stem cells, they start creating more red blood cells. If I understand correctly the Stem Cells start replicating Stem Cells more than producing daughter cells which quickly increases the number of Stem cells available to produce daughter cells which ultimately increases the number of red blood cells being produced.

The chromosomes are present in all the stem cells. In the final differentiated cells (i.e. platelets, red blood cells, white blood cells) the chromosomes may or may not be present. In terms of CML the relevance of this is not much as the PH+ is only a problem in Stem Cells that will overproduce. Red blood cells have no nucleus so I think they have no chromosomes, yet if someone has the JAK2 mutation in the Red blood cell Stem Cell then there will be overproduction of Red Blood Cells (I think the JAK2 mutation causing ET is not considered cancer like CML since it does not progress to a worse stage if not treated. It can kill a person due to overproduction of RBC's but the JAK2 does not lead to worse mutations and a worse form of disease, whereas CML if untreated goes to Blast Phase eventually which becomes very difficult to treat). Platelets are just cell fragments yet if their precursor stem cell has an issue there can be too many or too little of them produced. Regarding the White Blood Cells there are chromosomes present in them (or at least some of the strands of DNA are present), which is actually how genetic testing is able to monitor the progression and levels of the CML. If the mutations where only visible at the stem cell level I think it would be much harder to monitor the progression of the disease as the stem cells are much lower in number than the final white blood cells (if the genes were not present in the final white blood cells you could only see the mutations at the stem cell level, whereas we can see it on every single cell that is affected by this mutation).

Not sure if any of this answers your questions. Just a note regarding any discussion of stem cells, there are too many unknowns as it has not yet been figured out how to find and observe specific stem cells, so no one has actually directly observed at the various stem cell levels what is going on. All these observations regarding stem cells are based on indirect observation.

[Trey]

Pam,

Much of what you said is not quite accurate.  If you really want to try to understand this (and I realize it is not easy) you should study this (and maybe also read hundreds of other articles and journals): 

http://community.lls.org/docs/DOC-1272

CML starts with a leukemic blood stem cell very high in the blood making system, and affects both the myeloid and lymphoid lines, as well as RBCs and platelets.  So the entire blood making system is affected, including the lymphoid line.  At CML diagnosis, about 25 - 30% of the lymphoid WBCs have the Ph+ Chromosome, and most of the myeloid line.  This is generally not true is the other leukemias, where the top level leukemic cells are believed to be lower in the chain, and affect only one line of the WBCs.  So CML is not merely a disease of the myeloid granulocytes (neutrophils, eosinophils, basophils), although they are the primary offenders in proliferation for reasons too detailed to discuss here. 

CML leukemia is a disease of the blood cells.  It starts with a translocation in the chromosomes of a blood making (hematopoietic) stem cell very high in the chain.  The chromosomes are located in the cell's nucleus, and are made up of numerous genes, which are the genetic blueprints for making the cells in the body.  There are 46 chromosomes (two pairs of 23) in every nucleated cell in the body except  the  egg and sperm, which have only one set of 23.  Some cells lose their nucleus at the end stage, such as RBCs and platelets.  "Worker bee" cells cannot reproduce even if they have a nucleus, such as end stage WBCs.  So every cell in the leukemic line which has a nucleus is leukemic, including the blood cells that make WBCs, RBCs, and Megakaryocytes (which become platelets).  Regarding your question about X and Y, females have two X chromosomes as numbers 23 and 46, while males have an X and a Y.  These are in every cell that has a nucleus, except the egg has one X, and the sperm will have either an X or a Y.

The "marrow" does not make the blood cells.  The blood cells are made in the marrow environment by blood stem cells (hematopoietic stem cells).  When they mature they leave the "nest" which is the marrow and ride the roller coaster called the circulatory system for several days.  It is a wild ride.  Lymphoid cells mainly ride the lymph system, which is more tame.  After that they are put out to pasture in the tissues (organs, saliva, muscle, etc), and then die.  This life span lasts only several days. 

There are endless issues and discussions possible about this subject area.  I would suggest that trying to explain this subject to others is not possible for most people, which is why I suggested previously to point people to my carefully crafted posting on "The Genetics of CML" which is linked above.  But in the final analysis, for most people the issue remains that either the TKI drugs work well or they do not.  The rest is an academic exercise.

[matt92711]

Trey,

Thanks for this info. I had thought that the CML was at the Granulocyte precursor, never realized that it was higher up and other lines carried the messed up gene as well. I am curious about this and guess I will be doing some homework.

[pamsouth]

Trey, 

I have copied the link and your post, and pasted  to my email, as well for future reference and further studies. 

I have read thru your post and skimmed thru parts of the link.  I am trying to digest it, but am going to have to meditate on this for awhile and let it nest in my brain.

I tried to post it, more as a question, to explain what my understanding is, for correction, and fill in the blanks or put the puzzle together in a broader sense. 

I will be studying it in more depth, (as time permits) as you said as an academic exercise!!

I guess one particular thing that has stumped me, which I think I understand in part, was this question in my mind;  (background info) When I went for an annual check up, (2005)  my platelets came back 1 million, (white cell 13 thousand)  two week later, the platelets were at 2 million (white cells at 21 thousand).  At this point I was not diagnosed and I was googling high platelets, and never suspecting leukemia. When I went to the oncologist to have another blood draw, I was thinking that someone didn't know how to read zero's, as my primary doctor said that she had gotten a call from the lab, that my platelets were only 1000. So my primary doctor called the pathologist (who is her husband) to check on the platelet count.  The platelets were not 1 thousand, they were over a million, and climbing, (now 2 week later at over 2 million).

I went to the oncologist (at the cancer center) he was trying to explain, he wanted to do a BMB, as he was sure he was looking at a blood cancer.  I was so confused because I knew leukemia was white cells, well I had over 2 million platelets. He kept explaining my granuloyctes were off, which makes up the white cells.  Thus I have been trying to figure out the question why high platelets, and where are these chromosomes, platelets are not nucleus? ??

(The chromosomes are located in the cell's nucleus)

Your post helped me to somewhat understand  ""It starts with a translocation in the chromosomes of a blood making (hematopoietic) stem cell very high in the chain.  The chromosomes are located in the cell's nucleus, and are made up of numerous genes, which are the genetic blueprints for making the cells in the body."

But I am still having a little bit of trouble understanding the high platelets as they are not nucleus.  I will reread the article and carefully read the link.

I am waiting for an insurance approval that they will be paying for the new Oncologist, as I have mailed a dispute into the insurance company, regarding the coding on my first visit to the new onc.  I have gotten an Ok, from the doctors billing that they have approval from my insurance to use a certain code, to pay for the visit.  It is kind of complicated, as insurance can be. But I want it in writing, so I am waiting the insurance has 60 days to answer me.

My new oncologist is with a group of leukemia experts,  at the number 1 premier of Indiana.  So when I go for my next visit, I don't want to go in asking dumb question or taking up valuable time, of things I can learn on my own.  I imagine they get tired of explaining the same thing over and over, especially like you said of an academic nature.

Also I guess I sort started an interest in learning the academic dynamic of all the leukemias.  Not sure why?

I know I was diagnosed in 2005 and just learned the basics of CML, and wanted to forget that I had cancer, except for taking that pill and of course the 3 month onc visits and labs. Then the question came up of changing drugs, which sent me back to getting myself up to date and the TKI and then that led me to thinking of other things and re educated myself. 

I guess there was a lot I didn't learn on the first go around.

Anyhow I will certainly be digging into more articles and reading these post, until I get it!

Thank You again for taking the time to help me and also to Matt.

PamSouth

[matt92711]

Regarding the platelets they are formed from megakaryocytes which are large bone marrow cells that become the platelets. The megakaryocytes and precursors (which are myeloid cells, hence if the precursor cells are affected with PH+ then they would also be affected by CML) do have nuclei and do have chromosomes/genes. Just when it forms platelets at that stage they no longer have nuclei and as a result the chromosomes that are normally found in cells would not be present.

[pamsouth]

Matt,  Got it!!  Thanks!!

Pamsouth

[Trey]

Pam,

Matt is correct about the platelet issue.  Also, high platelets are often the first sign of CML leukemia, not a high WBC.  Many docs do not understand that.  Again, this is because all lines of blood cells are affected by CML.  All blood forming cells have a nucleus, but the final stage for RBCs and platelets do not.  But the RBC and platelet ancestor cells all had a nucleus, which carries the chromosomes.  So the precursors to the platelets (Megakaryocytes) are cells with a nucleus, and the leukemic ones carry the Philadelphia Chromosome in their nucleus.  Same for the RBCs.  That is why in CML, unlike the other leukemias, all blood cell lines are screwed up.  But the part that is so serious about CML is the granulocyte WBCs, since they do the damage to the body if not controlled, since they will become more unstable and mutate further, becoming very aggressive at the end stages.  That is why CML focuses on the granulocyte WBCs.  But many docs do not even understand the involvement of the other blood cell lines, so the high platelets early on can be missed as a sign of CML.

[CallMeLucky]

Is there any understanding why some people have high platelets at dx and some don't?  My WBC was in the 60K range at dx, but my platelets were well in normal range (upper limits of normal).  Does this suggest any difference in the disease?  I ahve often wondered if there are sub-types of CML that have not been discovered yet.  Characteristics that explain why some people progress differently then others, develop mutations, etc.

[Tedsey]

That's interesting that you posted this.  I was thinking the same thing.  However, when I was dx, my WBC were 180,000.  My PLT were high, but a bit over high end of normal for the lab's scale, (maybe that is high enough).  Does this have anything to do with prognosis?  Or is it something that again, is little understood.  Eight months before dx, I had a normal CBC.  PLT and WBC were normal.  Crazy how this chronic disease can seemingly move so fast.  Always wondered about this too.