So, to start off with, the 5% “unclassified cells” were myeloid blasts without Auer rods.  In addition, the peripheral smear shows mostly granulocytes in varying stages of maturation.  The mature bands and neutrophils do not show toxic granulation or vacuolation or Dohle bodies.  Only 2% of the cells were basophils, but this still qualifies as an absolute basophilia.

Given these finidings, this is very suspicious for a leukemia, especially given the high serum LDH and the extreme leukocytosis. Leukemoid reaction is certainly in the differential – by pure virtue of its name, “leukemoid”, it mimics a leukemia.  However, leukemoid reaction rarely attains leukocyte counts of >100,000/ul and usually shows toxic changes in the granulocytes (vacuolation, granulation, cytoplasmic Dohle bodies).  Without any other history of infection and the man’s mild symptoms, a leukemia must be considered.  The 5% blast level would also be unlikely in a leukemoid reaction.  The low hemoglobin and platelet counts are also in favor of a process in the bone marrow that is “bullying” the other cell lineages.

These were the “unclassified cells” we counted as blasts.

The differential diagnosis could also include chronic myelomonocytic leukemia (more monocytes) a myeloproliferative neoplasm with PDGFRalpha/beta rearrangements or FGFR1 abnormalities (unlikely), or a myeloproliferative neoplasm, unclassified (or atypical CML).

I spoke with the Heme/Onc fellow who also reviewed the slide and my hematopathology attending and everyone was on the same page that this was likely CML.  A bone marrow had been done already.  The treating team started hydroxyurea, allopurinol, and hydration immediately and Gleevec (imatinib) was started the following day.

The bone marrow showed a typical picture of CML: a very hypercellular marrow with an immature shift of granulocytes, small and hypolobated megakaryocytes, and decreased erythroid precursors.  FISH for the t(9;22) BCR-ABL fusion gene showed the fusion in 98% of cells analyzed and the conventional karyotype showed the Philadelphia chromosome in all 20 cells.

The patient is doing well despite persistent mild fatigue and is due for his 3 month follow-up marrow in September.  He will also get bone marrows at 6 months and 12 months.  The most sensitive methods of follow-up, though, are by cytogenetic and molecular testing for BCR-ABL.  CML is one of the success stories in medicine, as a once fatal disease now sees 85% of patients alive and disease free at 7 years while on Gleevec.  Alternative tyrosine kinase inhibitors (e.g. dasatinib) are available should patients fail this regimen.

It should be pointed out that the plantiffs include women with breast and ovarian cancer, physicians, cancer and genetics researchers as well as the Association for Molecular Pathology, American College of Medical Genetics, American Society of Clinical Pathologists, Breast Cancer Action & the College of American Pathologists among others.

1) Molecular profiling may help determine patient’s response to cancer therapies, research suggests.

A pilot study of molecular profiling of tumors, helped to identify therapies that ultimately had an impact on the disease.The University of Texas MD Anderson Cancer Center and the Memorial Sloan-Kettering Cancer Center are “striving to profile individual tumors so that therapy can be personalized, which means that it has a better chance of working because it targets specific mutations found in that tumor. This also prevents patients from being exposed to drugs that have a limited chance of success, eliminating toxicity and improving quality of life.

2 )Scientists developing new techniques for detecting CTCs in cancer patients’ blood (Medscape Report)

Measuring “circulating tumor cells (CTCs) in the blood of cancer patients” gives “an indication of whether or not the patient is responding” to treatment.” Presently, “there is only one commercially available product to measure CTCs — CellSearch (Veridex LLC).” Now, one device in development “promises to be cheaper and faster,” say University of California-Los Angeles researchers. The “new technique is based on a microfilter device.”

3) Bladder cancer cells may have two distinct genetic patterns, research suggests.

Bladder cancer cells have two distinct genetic patterns, depending on whether they are invasive or not, say University of Southern California-Los Angeles scientists. The “discovery opens the possibility of monitoring the disease by a simple urine test,” which would enable clinicians to sidestep “invasive procedures.”

4) Immune cells could be reprogrammed to attack prostate cancer, scientists say.

Reprogrammed immune cells could become targeted ‘killing machines’ against prostate cancer.” In fact, “these reprogrammed T cells sharply reduced the levels of prostate specific antigen (PSA) in two patients with metastatic prostate cancer,” scientists at the Roger Williams Medical Center said. But first, the team had to “isolate a patient’s T cells from a blood sample and use genetic engineering techniques to make them sensitive to a molecule that only occurs in prostate cancer — prostate specific membrane antigen, or PSMA.”

5) Researchers say presence of certain variants in MC1R gene may increase melanoma risk.

Researchers from the University of Pennsylvania “analyzed 779 patients with melanoma and compared them to 325 healthy people.” The investigators found that the “presence of certain variants in the MC1R gene was linked with at least a twofold increased risk of melanoma, and was largely confined to those people who would not normally be considered at increased risk.”

Posted by Dr.Prashant A.Jani M.D.FRCPC

The molecular target of warfarin is the protein product of the Vitamin K epoxide Reductase Complex, subunit 1 gene (VKORC1). This gene product is inhibited by warfarin. Polymorphisms of this gene product have been extensively studied over the past several years, and variant genotypes are associated with variable response to warfarin.¹ Compared to patients with a wild-type genotype, patients with at least one variant allele had an increased risk for elevated INR (HR = 1.4) and required more time to achieve a stable dosing (median = 95 days).¹

Warfarin is eliminated via metabolism in the liver. It is stereoselectively metabolized by hepatic microsomal enzymes (cytochrome P-450).⁴ The warfarin breakdown products have minimal anticoagulant activity.⁴ The metabolites are principally excreted into the urine, and to a lesser extent into the bile.⁴ The drug is comprised of a mix of the R and S stereoisomers. The S isomer is 3 to 5 times more potent than the R isomer, and is metabolized by the cytochrome P450 family. The cytochrome P-450 isozymes involved in the metabolism of warfarin include 2C9, 2C19, 2C8, 2C18, 1A2, and 3A4.⁴

CYP2C9 is reported to be the main liver P-450 which modulates the in vivo anticoagulant activity of warfarin.⁴ Two variants of CYP2CP have been identified.¹ These variants ( the *2 allele -R144C and the *3 allele – I359L) are reportedly associated with a decrease in enzymatic activity of 30% and 80%.¹

To summarize, identified SNPs (single nucleotide polymorphisms) are associated with altered warfarin metabolism and utilization. The CYP2CP*2 and CYP2CP*3 genetic variations are associated with a need for a lower dose of warfarin. The VKORC1 genetic variation is also associated with a greater variation in warfarin dosing to maintain a stable INR.

In August 2007, the FDA issued a release in which it recommended genetic testing for these genetic variations in warfarin metabolism, and altered the drug label for warfarin to reflect this recommendation. This recommendation is a part of the Critical Path Initiative which seeks to harness new scientific knowledge in the areas of gene expression, bioinformatics, and new analytic methods in the service of product development while also emphasizing its commitment to personalized medicine. ⁵ Some institutions are beginning to implement testing for the identified polymorphisms using new testing modalities, such as methods utilizing nanoparticles for “NAT” testing or invader chemistry.

Specific dosing recommendations are not yet determined for patients with these known genetic variations. This is currently under study, but the hope is to give doctors a dosing regimen based on the presence or absence of these variations.^6-7 There are a myriad of other influences of dosing requirements for individual patients such as age, sex, weight, and diet, calling into question the utility of these tests. However, the FDA label change to reflect the push towards personalized medicine, along with the risk of fatal bleeding with overdosing and stroke/clotting with underdosing, the additional information given by the test will be helpful according to the studies published on this topic over the past few years.

1. The Pharmacogenomics Journal (2004) 4, 40–48. doi:10.1038/sj.tpj.6500220 (Published online 16 December 2003)
2. http://www.nanosphere.us/VerigeneWarfarin
MetabolismNucleicAcidTest_4472.aspx
3.http://www.fda.gov/bbs/topics/NEWS/2007/NEW01701.html
4. http://www.rxlist.com/cgi/generic/warfarin_cp.htm
5. http://www.fda.gov/cder/drug/infopage/warfarin/qa.htm
6. J Thromb Thrombolysis. 2007 Jul 29; 7. Clin Pharmacol Ther. 2006 Oct;80(4):346-55

This is a very superficial post touching on an increasingly utilized testing methodology.

Comparative genomic hybridization (CGH) (also has been called “competitive”) is a method for scanning the entire genome for variations in DNA copy number which is not currently common in the clinical practice of medicine, but can be a useful tool in the molecular study of tumors. CGH can also be attempted in the search for chromosomal aberrations in patients with congenital anomalies of unknown etiology when other standard methods such as cytogenetics have not yielded a satisfactory result.

Tumor or germ-line DNA can be studied using this method. Genomic DNA is isolated from both the patient (and tumor) and a reference population. Both the test and the reference metaphase genomic DNA is labeled with fluorescent probes with different fluorescent characteristics. This mix of reference DNA, test DNA, and hybridized fluorescent probes are studied using fluorescent microscopes and equipment with the ability to measure the intensity of fluorescent signals. If the test DNA has signal which is much greater than the reference DNA (increased copy number) this will be detected by a change in signal compared to the expected value. Conversely, if the test DNA has decreased copy number of a particular gene compared to the reference population, then the fluorescent signal will be altered and detected. CGH will detect extra chromosomes and missing or altered chromosomes.

Array CGH is a CGH method in which the genomic DNA is hybridized to a microarray. This has improved the detection limit on this test; however the single copy number aberration must occur over a region of about 10 Million base pairs. Methods utilizing a focused microarray for hybridization can improve this detection limit further.

Some argue this may completely replace the discipline of cytogenetics because the methodology is steadily improving and its flexible. You can look at multiple focused aspects of the genome or the entire genome. The current resolution is supposed to be about 10,000 base pairs, but currently the signal noise associated with this resolution confounds its utility. However, cytogenetics will not be replaced just yet, as this method does not confer information about mosaicism and would be expensive to implement currently.

References
Nature Genetics 37, S11 – S17 (2005)
Oncol Rep 2007; 18:917-26.

• High blood pressure
• Heart attacks
• Abnormal heart valves
• Congenital heart defects
• Stress due to pneumonia
• Sick sinus syndrome
• Etc

When atrial fibrillation occurs in patients without these risk factors, lone atrial fibrillation is in the consideration. The stroke risk for patients with atrial fibrillation increases per year and is as follows (stroke rate percent per patient year)³:

• 1.3 in those aged 50 to 59 years
• 2.2 in those aged 60 to 69 years
• 4.2 for those aged 70 to 79 years
• 5.1 for those aged 80 to 89 years

One company offers a home do it yourself DNA diagnostics kit for risk assessment for myocardial infarction, atrial fibrillation, and diabetes. The kit costs $200 and uses DNA derived from buccal swabs to detect 2 single nucleotide polymorphisms on chromosome 4q25 that the company has identified as major risk factors for atrial fibrillation⁴. The company conducted a genome wide analysis of 300,000 Icelandic patients with atrial fibrillation, and found 2 SNPs associated with increased atrial fibrillation on the PITX2 gene on chromosome 4q25⁴. The results were published in Nature⁵. The test was also validated against Swedish patients and patients from MGH and is also found to be prevalent in the Han Chinese population⁴. If identified, the SNPs are associated with a 1.8 fold increase in atrial fibrillation. The value of the test as reported by the company is identifying patients who are at risk so that diagnostic tests may be utilized more effectively, and the onus is on the physician directing care for the patient to determine how to utilize this information in the treatment plan. Several polymorphisms have been reported as being associated with atrial fibrillation^6,7,8.

Therefore, the significance of the results of this type of DIY test is baffling to me. The original studies were conducted and validated on a relatively homogeneous population. Currently, it is still believed that atrial fibrillation is a heterogenous disease with several contributing factors. Genetics is probably a major factor. So would a negative test result put the average non-Swedish, non-Icelandic, Non-Han Chinese citizen and the doctor treating that patient at ease? Would a positive test result at some point dictate insurance eligibility and premiums?

If you are “lucky” enough to read Bloomberg. com on the right day⁹, you would find out that a company can offer you a DNA test for myocardial infarction risk for a mere $200. If positive, the risk of MI is 1.6 (the risk for smokers is at least 2-fold higher). The population for this study was slightly more diverse (Ottawa, Texas, Denmark, Great Britain).

References
1. JAMA. 2001 May 9;285(18):2370-5
2. http://www.mayoclinic.com/health/ atrial-fibrillation/DS00291/DSECTION=3
3. http://patients.uptodate.com/topic.asp?file=carrhyth/50724
4. http://www.decodediagnostics.com/AF-genetics.php
5. Nature Volume 448(7151), 19 July 2007, pp 353-3576.
6. Heart Rhythm. 2007 Apr;4(4):469-75. Epub 2006 Dec 157.
7. Circulation. 2003 Jun 17;107(23):2880-3. Epub 2003 Jun 28.
8. N Engl J Med. 1997 Mar 27;336(13):905-119.
9. http://www.bloomberg.com/apps/news? pid=newsarchive&sid=aBpFp6D4Q9BE