American Association for Clinical Chemistry
Better health through laboratory medicine
May 2009 Clinical Laboratory News: Diagnostic Profiles

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May 2009: Volume 35, Number 5

Colorectal Cancer Recurrence

New research indicates that the bio-marker guanylyl cyclase 2C (GUCY2C) in regional lymph nodes is an independent predictor of disease recurrence in patients with colorectal cancer. The study also points to the presence of occult metastases that escape current methods of histopathological detection. If confirmed in further studies, the findings could better inform the use of adjuvant chemotherapy in colorectal cancer and lead to more precise risk stratification of patients with pN0 colorectal cancer, meaning their lymph nodes are free of tumor cells based on histopathology (JAMA, 2009;301:745–742).

The prospective study involved 2,570 fresh lymph nodes obtained from 257 patients with pN0 colorectal cancer. Half of each lymph node was formalin fixed and paraffin embedded for histopathological examination. Specimens that expressed GUCY2C mRNA above background levels in disease-free lymph nodes and for which at least one lymph node yielded RNA of sufficient integrity for analysis were analyzed via quantitative PCR. Patients were followed for a median of 24 months for disease recurrence or death.

The researchers found that 12.5% of patients had lymph nodes negative for GUCY2C and their overall rate of recurrence was 6.3% with a 95% CI. In contrast, 87.5% of patients had lymph nodes positive for GUCY2C and their overall rate of recurrence was 20.9% with a 95% CI. Multivariate analysis demonstrated that GUCY2C in lymph nodes was an independent predictor of prognosis, and that patients with histologically negative but GUCY2C positive lymph nodes exhibited earlier time to recurrence (adjusted hazard ratio, 4.66; CI, 95%) and reduced disease-free survival (adjusted hazard ratio, 3.27; CI 95%).

Consensus qPCR Guidelines Published 

First-ever consensus guidelines on quantitative PCR aim to improve the quality and transparency of studies involving qPCR (Clin Chem, 2009;55:611-622). The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines outline the minimum information necessary to evaluate qPCR studies, including all relevant experimental conditions and assay characteristics, and full disclosure of all reagents, sequences, and analysis methods. The guidelines include an 85-item checklist of desirable and essential steps to be followed when using qPCR and information to be divulged from experiments involving qPCR. The purpose of the guideline is to encourage better experimental practice, so as to enable more reliable and unequivocal interpretation of qPCR results.

Use of qPCR has proliferated, yet studies “invariably use diverse reagents, protocols, analysis methods, and reporting methods,” the authors wrote. “This remarkable lack of consensus on how best to perform qPCR experiments has the adverse consequence of perpetuating a string of serious shortcomings that encumber its status as an independent yardstick.” If researchers follow the guidelines, they should be able to design and report qPCR experiments with greater inherent value, and fellow researchers, editors, and laboratorians should be able to evaluate the technical quality of the published data against an established standard.

Higher Blood Lead Concentrations Associated with Increased Cardiovascular Death 

A prospective cohort study of elderly women revealed the lingering effects of environmental exposure to lead, suggesting that lead and other environmental toxicants may account for a portion of the burden of cardiovascular disease (Environmental Health, doi:10.1186/1476-069X-8–15). The researchers found that women with blood lead concentrations ≥8µg/dL had a 59% increased risk of multivariate adjusted mortality (HR, 1.59; CI, 95%) and a 3-fold greater risk of coronary heart disease mortality (HR, 3.08; CI, 95%) than those with blood lead concentrations <8 µg/dL. There was no significant association between blood lead levels and stroke or non-cardiovascular deaths.

The data came from a cohort of 533 women from the Study of Osteoporotic Fractures. The ancillary lead study was conducted between 1990 and 1991 and the women were followed for more than 12 years, with >95% complete contacts. Blood lead concentrations were determined us-ing graphite furnace atomic absorption spectrometry.

Mean blood lead concentrations in the U. S. population older than age 1 have declined to 1.45 µg/dL as reported in NHANES IV. However, the women in this study were exposed to lead before concerted efforts to limit environmental lead usage began in the 1970s. Although the skeleton is the main repository of lead, bone loss and demineralization after menopause appear to release bone lead into circulation. In addition to its well-described impact on cognitive function, lead also affects the cardiovascular, renal, and nervous systems. The researchers called for more studies to further the understanding of the mechanisms for lead toxicity on multiple organ systems.