What does <1 in 1,000,000 cells mean?
What does detected at below levels of detection mean?
Turns out a lot.
On January 21, 2020 I had a bone marrow biopsy (BMB) at UCSD with Dr. Kipps’s team in La Jolla, CA, specifically with the amazing nurse Becky, the master of the minimal pain BMB. Having an experienced and skilled technician do the procedure makes a huge difference.
That said, I doubt Becky shows what a great bone core she extracted from a hip bone to many of her patients, unless they had their sensitivity to the human condition and its related bloody tissues hardened by a life of seeing more body parts in disarray or out of place than perhaps anyone other than a homicide detective in Chicago.
I remember once early in my medical career the time a patient fainted after I urged her to look at a drop of her own urine under the microscope that was full of bacteria and white blood cells due to her raging bladder infection.
I never made that mistake again.
Becky knew that I have a morbid medical curiosity (and a toughened gut) and would want to see the enemy up close, having personally viewed many of my own blood smears over the years. “Keep your friends close and your enemies closer.” Besides, you can’t faint when you are already lying flat on your stomach with your pants down on the procedure table.
I had some extra “draws” of my marrow aspirate for research.
I received a kind call-back from Dr. Kipps when the results were available, as there was no time to meet in person at his clinic.
There were three ways the specimen was examined.
The first was old school.
The pathologist looked under the light microscope.
He looked at the health of the bone marrow and the distribution of the progenitor cells. All good.
He looked unsuccessfully for telltale lymphoid aggregates that would have suggested that my CLL had clumped together in a heterogeneous fashion in the marrow raising the possibility of a sampling error, that the cells were pulled from an area where there was no CLL, or that there might be a different story from a sample a millimeter away. But none of this was the case.
An experienced pathologist with a light microscope can find as few as one cancer cell in a hundred or more cells. None were seen in the 200 cells examined, and the cellularity and the blood precursors were healthy for red blood cells, white blood cells, and platelets.
That means based on the BMB alone I would be in a complete remission (CR). I have one persistent lymph node that measures 1 millimeter above normal and is unchanged in 20+ months, so this means I am still officially in a partial remission. That lymph node will probably never shrink back to normal due to scar tissue replacing the CLL, but it is extraordinarily unlikely that it will ever cause me any trouble.
- I am in complete remission in the marrow!
Flow cytometry is an automated test that looks at the immune fingerprint, the immunophenotype of the CLL cells by shooting different colored lasers at the surface markers seen in CLL. It can find as few as one in 10,000 or maybe even one in 100,000 cells. Although that later number is not generally accepted. 1/10,000 is widely accepted as the gold standard for measurable residual disease (MRD) testing.
- uMRD4 (undetectable measurable disease to the level of 10-4 or 1 in 10,000)!
The next test was the ClonoSEQ test that doesn’t look for individual cells but looks for the sequences of DNA bases through next generation sequencing (NGS). Highschool biology reminds us that the building blocks of life are made from only four bases that make up DNA: adenine (A), guanine (G) Cytosine (C) and thymine (T). Sequences are called out by their letters (such as AGTCCAG) and so on.
Beyond this the biology gets tricky, but let’s just say that all B-cells have their own very specific immunoglobulin sequences that help them recognize unique threats such as viruses or bacteria.
Normally all these sequences are different, but we know that all cancer including CLL is clonal, so in CLL they are all the same in the cancer clone.
When my cancer was rampant in my bone marrow, ClonoSEQ searched all the DNA sequences in my marrow for sequences that were found on my cancer clone. There are all kinds of complicated biological and statistical thresholds that the sequences must exceed to be sure they are truly from the CLL clone.
The number of these unique sequences that it is possible to find depends on the nature of the particular cancerous immunoglobulins being examined, and whether they are lambda or kappa, and I won’t go into the science behind that.
They look for those same sequences in later samples to see if the cancer is still present or if it has returned.
Suffice to say some sequences are more unique than others, meaning that they are most unlikely to rise from a normal B cell. Others have a statistically significant chance they might randomly rise in non-cancerous cells, therefore finding that sequence is less certain evidence that you have really found the cancer.
In my case I had 4 sequences from my cancer’s immunoglobulins that would signal if the CLL was back.
When my marrow checked for residual CLL one-month post CAR-T, they found none down to the level of one in a million. I also had no normal B cells at the time, so they would be no “background noise.”
In January, they looked at 2,700,000 cells from my marrow aspirate.
My two most unique sequences and my second most unique sequence showed no CLL to the level of one in a million. SUPER!
But wait, the 4th sequence, the least unique sequence was detected at <1 in a million. So, my question was, “Is the tiniest fragment of the cancer still holding on, or is it just a random chance that some nonclonal noncancerous B cells have the same sequence?”
While it can’t be said with absolute certainty which answer is correct, odds strongly favor the second more celebratory explanation.
All four sequences that were checked came from the same clone, so if the cancer is around all four sequences it should be seen at similar levels. In my case, the only sequence was the one that they detected way below the level (<1/1,000,000) where it could be determined that it actually came from the cancer, so it wasn’t random. While it is more than wishful thinking that the second more benign explanation is the correct one, it is by far the most logical and statistically likely answer.
Having a very little amount of measurable cancer versus no measurable cancer is not a trivial distinction, at least in my case. If the cancer is measurably there, it means it could and likely would eventually return and cause problems. But hopefully not for a long time.
The same could happen if the cancer is completely undetectable. Undetectable MRD can become measurable again and then take off.
But there is another possibility. The CLL is all gone, meaning every last chronic lymphocytic leukemia cell has been obliterated or it’s been knocked back so far that it isn’t never coming back.
So, what can I say about ClonoSEQ? Officially I have to say they found less than one in a million CLL cells. Unofficially, I feel great confidence that no CLL was found in the 2,700,000 cells that were interrogated.
- uMRD6 (undetectable minimal residual disease to the level of 1 in 1,000,000)!
I don’t want to be overly optimistic. I am urging myself to stay calm, ZEN, but I must acknowledge that there is the real possibility that I am done with my CLL journey which would be pretty great.
For me, stopping at less than one in 10,000 was not good enough. I wanted more reassurance, and so did my doctors. I needed to go two degrees deeper to be sure that if I stopped the ibrutinib treatment that saved my life the past seven years I would not be surprised by a sudden resurgence of either my CLL or my ITP (Immune Thrombocytopenic Purpura) that crashed my platelets and almost killed me over a dozen years ago. Now I will move ahead, and for the first time since I traveled to Ohio State where I took my first dose on May 7, 2012, of PCI-32675 (which later became ibrutinib), I will wake up in the morning and not be taking any medications to control my CLL.
I am so grateful for both the revolutionary treatment CAR-T therapy and the revolutionary testing that tells us that it is working. It is an amazing world. This all would be unimaginable when I was diagnosed almost 15 years ago.
Stay strong. We are all in this together.