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Test to test, I will survive

Preetha K

19 Jan 2023

‘Diagnosed, frightened, tested, treating, waiting, hoping’.

A six-word narrative about living with blood cancer from patients in Leukemia and Lymphoma society. 

Cancer is a condition in which the cells of the body exhibit abnormal uncontrolled growth leading to the formation of tumors. The reason why cancer is alarming is the fact that cells in any part of the body can become cancerous and start spreading. Also, when cancer cells crowd normal cells, they fight for the same resources with an increasing ferocity which leads to the death of normal cells, leading to a host of other problems. 

The GLOBOCAN 2020, reported 19.2 million new cancer cases and 9.9 million deaths globally. By 2040, cancer incidence and mortality are expected to rise to 29.5 million and 16.3 million, respectively.

Metastasis is the single most dangerous hallmark of cancer. Metastasis happens when cancerous cells break away from the primary tumor and spread to other parts of the body. These metastasized cells attach themselves to healthy tissues and form secondary tumors. 

Over 200 types of cancer are known, and they can be classified based on how they originate in the body. Leukemia is one of the most common cancers.

Leukemia, a type of malignant cancer, starts in the blood-forming cells in the bone marrow. Bone marrow is the sponge-like tissue in the center of most bones that produces red blood cells (RBCs), white blood cells (WBCs), and Platelets.

In leukemia, cancerous blood cells form and crowd out healthy blood cells in the bone marrow.  Leukemia is classified as either “acute” or “chronic.” These two terms illustrate how quickly the disease progresses without treatment. 

Acute forms of leukemia progress rapidly and produce cells that are underdeveloped. These immature cells cannot perform their normal functions. Chronic forms of leukemia usually progress slowly, and patients have greater numbers of mature cells. Chronic myeloid leukemia (CML) is a type of leukemia that progresses slowly and involves the myeloid WBCs in the bone marrow. 

The World Health Organization (WHO) classifies CML as a “myeloproliferative neoplasm,” a type of disease in which the bone marrow makes too many white blood cells. CML usually gets worse over time, as the extra cells build up in the blood and/or bone marrow. The accumulation of WBCs may eventually cause fatigue, bleeding, and other problems. If left untreated, CML shows  the possibility of turning into acute leukemia, which is challenging to treat. 

Chronic myeloid leukemia and Abnormal gene expression

Cells in the human body make new copies of themselves to replace worn-out cells by a  mechanism called “cell division”. In order to make a new copy of itself, a cell duplicates its contents, including its chromosomes, and then splits to form two cells. Errors can  occasionally occur during this process. 

One such error is translocation. Translocation occurs when a piece of chromosome breaks off and attaches to another chromosome. During this process, the genes affected by translocation become abnormal. This abnormal gene is called the fusion gene, which is made by joining different parts of the genes.

For Instance,  a portion of the ABL1 gene from chromosome 9 translocates and fuses with the remaining portion of the BCR gene on chromosome 22. The translocated piece of chromosome 9 results in an oncogene called BCR-ABL1 t(9;22)(q34;q11) (BCR-ABL1: BCR - Breakpoint cluster gene; ABL1 - Abelson Kinase 1), also known as the Philadelphia chromosome”,- discovered at the University of Pennsylvania School of Medicine,  in Philadelphia in 1960.

Image 1: Formation of BCR-ABL1 fusion gene by the translocation of chromosome 22 and chromosome 9

Genes provide instructions for a cell to make protein. The ABL1 gene helps the cell to make a protein called Tyrosine Kinase required for cell proliferation. The abnormal BCR-ABL1 protein produces BCR-ABL1 tyrosine kinase, which causes a  spike  in  levels of tyrosine kinase in the cell, and in turn, results in cell proliferation; Thereby  more WBCs are  produced. These WBCs with the  BCR-ABL1 oncogene cause CML. CML cells divide faster than normal WBCs. 

The breakpoint in the BCR gene influences the size of  the fusion protein encoded by BCR-ABL. To date, Three breakpoint cluster regions have been characterized: Major (M-bcr), Minor (m-bcr), and Micro (µ-bcr).

Signs and Symptoms:

CML is a slow, progressive  disease. majority of the signs and symptoms occur because the CML cells crowd out the healthy normal cells (red blood cells, white blood cells, and platelets). Many individuals with CML remain asymptomatic when diagnosed. 

Symptoms include

  • Bleeding

  • Enlarged spleen

  • Night sweats

  • Poor appetite and weight loss

  • Weakness

  • Fatigue

  • Bone pain

  • Fever

  • Unexplained weight loss

  • Shortness of breath


The most common sign of CML is an abnormal WBC count. To diagnose CML, doctors use a wide range of tests to analyze the blood cells and the bone marrow cells. Some of the standard tests done to diagnose CML include 

  • Complete Blood Count (CBC)

  • Peripheral Blood Smear (PBS)

  • Bone marrow aspiration and Biopsy (BMA)

  • Conventional Cytogenetic Analysis (CCA)

  • Fluorescence In Situ Hybridization (FISH)

  • Quantitative Polymerase Chain Reaction (qPCR) for BCR ABL1 Detection

Treatment Options

Before initiating the treatment, the hematologist-oncologist will discuss the treatment options with the patients. CML treatment has improved significantly since using Tyrosine Kinase Inhibitors, targeted toward the tyrosine kinase of ABL1. The approach of treating each patient and their choice of treatment depends on the phase of CML during the diagnosis.

Tyrosine Kinase Inhibitor (TKI) Therapy: 

Tyrosine Kinase Inhibitor Therapy is a targeted therapy that inhibits the BCR-ABL1 gene kinase activity, inhibiting uncontrolled cell proliferation. TKI therapy causes less damage to the normal cells, compared to conventional treatment. 

The following drugs are used for standard treatment for CML:

  1. Imatinib Mesylate (Gleevec)

  2. Dasatinib (Sprycel)

  3. Nilotinib (Tasigna)

  4. Bosutinib (Bosulif)

  5. Ponatinib (Iclusig)


TKIs can control CML in most patients.

Measuring Treatment Response:

After the patients begin treatment, the hematologist-oncologist will periodically monitor the patient using blood and bone marrow tests to determine the treatment response. Monitoring the treatment response is one of the key strategies for managing CML. In general, the better the response to drug therapy, the overall survival (OS) of the patient is improved.

There are 3 types of responses

  1. Hematologic

  2. Cytogenetic

  3. Molecular

Type of Response

  1. Hematologic

  2. Complete Hematologic Response 

  3. Blood count is back to normal

  4. No blasts in peripheral blood

  5. Absence of signs and symptoms of the disease

  6. Cytogenetic

  7. Complete Cytogenetic Response

  8. No Ph chromosome detected

  9. Partial Cytogenetic Response

  10. 1%-35% of the cells have Ph chromosome

  11. Major Cytogenetic Response

  12. 35% or fewer cells have Ph chromosome

  13. Minor Cytogenetic Response

  14. More than 35% of cells have Ph chromosome

  15. Molecular

  16. Complete Molecular Response

  17. No detection of BCR-ABL1 gene

  18. Major Molecular Response (MMR)

  19. 3-log reduction in BCR-ABL1 levels or BCR-ABL1 0.1%

Major Molecular Response (MMR

Molecular response is defined by the magnitude of reduction in peripheral blood BCR-ABL1 transcript levels. A qPCR test is used to measure the number of cells in the blood that contain the BCR-ABL1 gene and is explained as a percentage. A patient’s initial molecular response to treatment is significant in predicting outcomes and determining further treatment options. The molecular response is the most sensitive method of monitoring BCR-ABL1 levels in the blood. 

Early molecular response (EMR)—The BCR-ABL1 level is 10 percent or less, 3 and 6 months after the start of treatment. This means that the leukemia cells have been reduced by 90 percent or more.

Major molecular response (MMR)—The BCR-ABL1 level has decreased to 0.1 percent. This means that the leukemia cells have been reduced by 99.9 percent or more.

Deep molecular response (DMR)—The BCR-ABL1 level has decreased to 0.01 percent or less. 

Image 2:-  Treatment Response International Scale (Source - Information provided from Leukemia and Lymphoma Society)

International Scale (IS) - This is a standardized scale for measuring qPCR test results. The qPCR test reflects the number of cells that have the BCR-ABL1 gene. It is used to determine how well treatment is working. The International Scale (IS) defines the standard baseline as BCR-ABL1 100 percent. (This baseline was developed from the IRIS clinical trial, by testing many patients’ pretreatment samples and normalizing the average patient results to create this baseline.) The term “baseline” refers to the start of treatment. The International Scale baseline is standardized and is used for all CML patients.

Log Reduction- A log reduction indicates the BCR-ABL1 level has decreased by a certain amount from the standard baseline.

  • 1-log reduction means that the BCR-ABL1 level has decreased to 10 times below the standardized baseline. This means that 10 percent of cells (10 out of every 100 cells) have the BCR-ABL1 gene. This reduction is equivalent to an early molecular response when achieved within 3 to 6 months of starting treatment.

  • 2-log reduction means that the BCR-ABL1 level has decreased to 100 times below the standardized baseline. This means that 1 percent of cells (1 out of every 100 cells) have the BCR-ABL1 gene.

  • 3-log reduction means that the BCR-ABL1 level has decreased to 1,000 times below the standardized baseline. This means that 0.1 percent of cells (1 out of every 1,000 cells) have the BCR-ABL1 gene. It is also known as a “major molecular response (MMR).” { 4-log reduction means that 0.01% of cells (1 out of every 10,000 cells) have the BCR-ABL1 gene.

  • 4.5-log reduction is referred to as a “complete molecular response (CMR)” or a “deep molecular response (DMR).” Doctors may refer to this as “MR4.5.” A 4.5-log reduction indicates that 0.0032% of cells (1 out of every 32,000 cells) have the BCR-ABL1 gene. Achieving a deep molecular response is a sign of disease remission. Patients who achieve and then sustain a deep molecular response for a significant period of time may be considered candidates for discontinuing TKI therapy, under careful medical supervision. 

  • 5-log reduction means that 0.001% of cells (1 out of every 100,000) have the BCR-ABL1 gene. By reaching a 5-log reduction, patients have achieved undetectable BCR-ABL1.

Image 3 - Treatment response by Log reduction (Source - Information provided from Leukemia and Lymphoma Society)3

qPCR test to determine BCR-ABL level

It is recommended that patients get a qPCR test done every 3 months, after the onset of the Tyrosine Kinase Inhibitor therapy. This is to monitor and evaluate the molecular responses achieved after the treatment.  After 2 years of achieving and maintaining a BCR-ABL1 level of 0.1 percent or less, the test should be done every 3 to 6 months.

Real-Time Quantitative PCR is an important laboratory technique to monitor the efficacy of Tyrosine Kinase Inhibitor Therapy and quantitatively assess Minimal Residual Disease (MRD). This involves the isolation of RNA from the EDTA whole blood or bone marrow aspirates of the CML patients.

Qualitative TaqMan probe RT-PCR assay can detect and differentiate the three different transcripts of BCR-ABL1. The test will confirm the presence or absence of the transcript. After the confirmation of the transcript, a quantitative test is recommended for the quantification of the BCR-ABL1 transcript.

Quantitative TaqMan probe RT-PCR assay can measure the quantity of the BCR-ABL1 gene in the blood or bone marrow aspirate. The quantitative standards with the known copy numbers should be calibrated against the WHO Ist International Genetic Reference Panel for the quantitation of BCR-ABL1 translocation for assigning IS values to a measured level of BCR-ABL/ABL%

Salient Features and Benefits of NeoDx BCR-ABL Kits

  • Easy calculations to determine the IS% and NCN% which makes the reporting more accurate and convenient

  • Multiplex, single tube assay involves amplification of all three transcripts with the internal control gene

  • No additional cDNA synthesis step is required. 

  • Minimal pipetting steps during the preparation of the master mix

  • In-silico analysis was performed for all the primer sequences and probe sequences used in the BCR-ABL Qualitative RT PCR Detection Kit - Major, Minor, Micro,  primer and probe sets are specific to BCR-ABL Major, BCR-ABL Minor, and BCR-ABL Micro respectively.

  • All the primers and probes sequences' specificity and efficiency were analyzed and have 100% specificity in identifying the respective sequences.

  • Non-CML transcripts are not detected.

  • NIBSC calibrated standards for the quantification of the BCR-ABL Major transcript.

  • High-Quality Mastermix which shows Increased Inhibitor Tolerance, Thermostability, and Reproducibility

  • The Quantitative Kit- Major can evaluate the molecular response up to 4.5 log reduction. 

  • Open instrument compatibility

  • BCR-ABL1 Qualitative RT-PCR Kit- Major, Minor, Micro

  • TaqMan probe-based Qualitative, 1-step RT-PCR, multiplex assay for the specific detection and differentiation of BCR-ABL major, BCR-ABL minor, and BCR-ABL micro transcripts along with the internal control ABL1

  • Targets are Major BCR-ABL1. Minor BCR-ABL1 and Micro BCR-ABL1

  • The kit supports reverse transcription. No additional cDNA steps are required.

  • Single tube assay, for the detection and differentiation of the three BCR-ABL transcripts.

  • Presence or Absence of the BCR-ABL1 transcripts can be detected.

  • BCR-ABL1 Quantitative RT-PCR Kit- Major:

  • IS quantitative, one-step RT-PCR, multiplex assay for quantifying the IS ratios of BCR-ABL  Major (M-BCR) p210 transcript in CML patient samples. 

  • Evaluates the molecular response up to 4.5 log reduction. The reporting is based on European Treatment and Outcome Study (EUTOS) guidelines and European Leukemia Net (ELN) recommendations

  • The kit supports reverse transcription. No additional cDNA steps are required.

  • The quantitative standards provided are with known copy numbers and are calibrated against WHO 1st International Genetic Reference Panel for the quantitation of BCR-ABL1 translocation (09/138)2 for assigning IS values to a measured level of BCR ABL/ABL %.

  • BCR-ABL1 Quantitative RT-PCR Kit- Major, Minor, Micro

  • IS quantitative, one-step RT PCR, multiplex assay for quantifying the IS ratios of BCR-ABL  Major (M-BCR) p210 transcript in CML patient samples. 

  • The minor BCR-ABL and micro BCR-ABL are reported in NCN (Normalized Copy number) %.

  • Single tube assay, for the quantification of three BCR-ABL transcripts.

  • The kit supports reverse transcription.No additional cDNA steps are required.

Applications of NeoDx BCR-ABL1 Kits:

  • Diagnosis of Chronic Myeloid Leukemia

  • To monitor the effectiveness of the Tyrosine Inhibitor Therapy

  • To calculate the IS (International Scale) % for the BCR-ABL Major transcript

  • To calculate the Normalized Copy Number (NCN)% for the BCR-ABL minor and BCR-ABL micro transcripts

  • To determine the Minimal Residual Disease or Measurable Residual Disease after the treatment

  • To detect the level of BCR-ABL throughout the drug treatment

  • To evaluate the treatment response by the log reduction

Difference between commercially available BCR-ABL1 qPCR kits and NeoDx BCR-ABL1 qPCR Kits

Image 4- Difference between NeoDx BCR-ABL1 kits and other commercially available kits

Due to the increased advances in understanding Chronic Myeloid Leukemia and the successful results of the Tyrosine Kinase Inhibitor Therapy, Treatment-Free Remission (TFR) is the new treatment goal for the CML patients. 

TFR is achieved when a patient discontinues the tyrosine kinase therapy and maintains a Deep Molecular Response and does not need treatment again. 

CML has been transformed from a life-threatening disease to a manageable condition because of TKI therapy. Patients with CML are living longer and experiencing fewer side effects from the treatment. Healthcare providers need to maintain a comprehensive knowledge of current treatment options to ensure each patient is aligned with a therapy that is both effective and also provides the best quality of life.

NeoDx Biotech Labs provides the most reliable qPCR kits to monitor CML patients and help them with  successful TKI treatment. The highly specific and sensitive kits detect, differentiate, and quantify the BCR-ABL transcripts.

Order Information: 

  1. BCR-ABL Qualitative RT-PCR Kit - Major, Minor & Micro

  2. NDXIVD001: 100 Rxns

  3. NDXIVD001_01: 50 Rxns

  4. NDXIVD001_02: 20 Rxns

  5. BCR-ABL Quantitative RT-PCR Kit -Major

  6. NDXIVD002: 100 Rxns

  7. NDXIVD002_01: 50 Rxns

  8. NDXIVD002_02: 20 Rxns

  9. BCR-ABL Quantitative RT-PCR Kit - Major, Minor & Micro

For more information, write to us at

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