Therapeutic drug monitoring of antiretroviral agents.

HIV+ patients fail antiretroviral therapy due to inadequate drug concentrations reaching the site of viral replication and/or the development of viral resistance to the antiretroviral agents. Adequate drug concentrations may not be reaching the virus due to poor compliance, poor absorption, or other pharmacokinetic factors such as metabolism, elimination, and drug interactions. The most important and most common pharmacokinetic drug interactions involve inhibition of metabolism, induction of metabolism, altered drug absorption, inhibition of renal excretion, and displacement from plasma protein binding sites. If a patient is failing antiretroviral therapy, TDM of antiretroviral agents could help in determining both adequacy of drug concentrations and patients' adherence. Ongoing studies will determine whether total drug concentration or free drug concentration of the protease inhibitors is the best predictor of response. Trough concentrations could prove to be the most important predictor of response, but additional studies are needed to compare trough, peak, and AUC concentrations with response to treatment. Finally, if some patients fail therapy due to inadequate drug concentrations, then increasing the dose could benefit patients' outcome and increase longevity. Clinical trials are needed that compare patients who receive a fixed-dosage regimen with patients who have adjusted dose regimens. Such a study is the best way to determine the true value of TDM of the antiretrovirals.

Antineutrophil cytoplasmic antibodies an update on clinical utility.

Antineutrophil cytoplasmic antibodies (ANCA) are autoantibodies directed against components of neutrophils and monocytes, and are helpful in identifying different forms of vasculitis. Until the discovery of ANCA, there was no specific laboratory method for the investigation of systemic vasculitis. Many issues regarding ANCA, such as their role in the pathogenesis of vasculitis, sensitivity, specificity, predictive value, and antigen specificity are still unclear. The differential diagnosis of the vasculitic syndromes is often difficult because of the variable presentation, but is important because of the variable outcome.Wegener's granulomatosis, microscopic polyangiitis, and Churg-Strauss syndrome are three vasculidities that commonly are associated with a positive ANCA. The "gold standard" for a precise diagnosis of vasculitis depends on both the clinical and pathologic features. ANCA testing is used to support the clinical and pathologic findings. The ANCA pattern, determined by indirect immunofluorescence, can be either cytoplasmic (C-ANCA), perinuclear (P-ANCA), or atypical perinuclear (X-ANCA). Interpretation of the ANCA pattern can be problematic, and a reactive antinuclear antibody can contribute to the confusion, requiring testing for reactivities to specific autoantigens. Proteinase-3 and myeloperoxidase are the most common autoantigens and are commonly associated with C-ANCA and P-ANCA, respectively. ANCA reactivity to other autoantigens is associated with the X-ANCA pattern.

Quantitation and morphometric analysis of tumors by image analysis.

Digital image analysis provides objective measurements of tissue and cell analytes previously interpreted subjectively. Both analyte concentration determination and morphometric analyses are possible. Calibration of the instrument and the use of standards and controls are essential for precise and reproducible quantitation of the analyte. Multi-tissue blocks ensure reproducible staining of the batch in quantitative immunohistochemical assays such as breast cancer estrogen and progesterone receptors. These multi-tissue blocks can be shared among laboratories to reduce interlaboratory variation and to objectively quantitate estrogen and progesterone receptors in clinical trials. In colon carcinoma, p53 can be quantitated objectively by image analysis. In prostate carcinoma, morphometric analysis of nuclear shape, nuclear roundness factor, and variations in nuclear size are objective measurements which constitute the pathologist's nuclear grade. Developments in instrumentation have now made it possible to combine analyte determination (such as DNA ploidy) and morphologic analysis of tumors, a diagnostic improvement over either method alone. A study employing image analysis to detect and quantitate androgen receptors and p53 in formalin-fixed, paraffin-embedded prostate cancer biopsies is underway to determine the utility of androgen receptors in predicting response to hormonal therapy. Histopathological features such as nuclear size, shape, and pleomorphism must be converted to image features such as area, shape factor, and variance of the area; this feature vector must be correlated with the pathologist's expert opinion or diagnosis. Other applications of image analysis include quantitation of immunofluorescent assays such as anti-nuclear antigen or anti-cytoplasmic nuclear antigen. Fluorescent image analysis provides more precision and greater reproducibility, as well as reduced test costs.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostate-specific antigen and prostate volume: a meta-analysis of prostate cancer screening criteria.

OBJECTIVE: To establish the value of serum prostate-specific antigen (PSA) and prostate-specific antigen per unit volume of prostate gland (PSAD) in detecting prostate carcinoma (CaP) in a hypothetical screening algorithm, a meta-analysis of the sensitivities, specificities, predictive values and likelihood ratios were combined from the published data. DATA SOURCES: Journal articles identified by a MEDLINE database search from 1988 to October 1992, using prostate-specific antigen as a key word were used to calculate the distribution of PSA in healthy men, men with benign prostatic hyperplasia (BPH) and men with prostate carcinoma (CaP). STUDY SELECTION: Only studies that contained the specified serum PSA values and patient outcomes were included. DATA EXTRACTION: The distributions of the serum PSA were plotted versus serum PSA for healthy men (2567), men with BPH (798) and men with CaP (835) from the abstracted data. Prostate volume distributions were estimated from the published transrectal ultrasound (TRUS) calculations. DATA SYNTHESIS: Hypothetical cohorts of 1,000 men between the ages of 60 and 70 years were screened using three different screening decision algorithms. Using a serum PSA cutoff of 3.0 ng/ml for referral for transrectal biopsy, 59 of 80 (74%) CaP would be detected and 21 (26%) would be missed. 209 transrectal biopsies would be performed, and 150 (72%) of them would be negative for CaP. Using a serum PSA cutoff of 4.0 ng/ml, 52 of 80 (65%) CaP would be detected and 28 (35%) would be missed. 146 transrectal biopsies would be performed, and 94 (64%) of them would be unnecessary. Using a cutoff of 2.0 ng/ml for serum PSA and 0.1 ng/ml/cc for PSAD, 55 of 80 (69%) of the cancers would be detected and 25 (31%) would be missed. Only 84 transrectal biopsies would be performed, and 29 (35%) of them would be negative for cancer. CONCLUSION: This algorithm maximizes the number of cancers detected (true-positive cases) and at the same time reduces the number of false-positive cases, minimizing the number of patients who would have to receive an unnecessary transrectal biopsy, compared to using a serum PSA cutoff of 3.0 or 4.0 ng/ml.

Human papillomavirus testing in the clinical laboratory. Part I: squamous lesions of the cervix.

The aim of this study was twofold: (1) to evaluate the contribution of viral (HPV) testing for improving the sensitivity of cervical cytology and (2) to correlate HPV types with the histology of the detected cervical cancer precursors, particularly the low-grade, CIN I variant. We used the dot blot hybridization technique (ViraPap) and polymerase chain reaction (PCR) in 63 women referred to our colposcopy clinic for evaluation of an abnormal Pap test. Histopathologic samples obtained by multiple colposcope-directed punch biopsies were used for a diagnostic gold standard. Among the 53 women with histologically proven CIN, precolposcopy cytology was positive in 38 (72%) compared to 53% and 60% HPV positivity by ViraPap and PCR, respectively (p less than 0.01). When the yields of ViraPap/PCR and cytology were combined, however, the detection rate of CIN was 91%, a significant improvement over cytology alone (p less than 0.02). HPV DNA was found either by ViraPap or PCR in 45 of 63 (71%) biopsy specimens, and 37 of 38 (97%) HPV-positive CIN, including the low-grade CIN I variant, contained oncogenic HPV types. HPV type 16 was present in 22 of 38 (58%) CIN lesions and mixed with HPV 6/11, 18, or the 30s group in 6 of 38 (16%) of the cases. HPV 6/11 alone was found only in 1 case of CIN I (2.7%). HPV testing by molecular technology increases the sensitivity of cytology.(ABSTRACT TRUNCATED AT 250 WORDS)

Human papillomavirus testing in the clinical laboratory. Part II: vaginal, vulvar, perineal, and penile squamous lesions.

The accurate diagnosis of HPV-related diseases of the lower genital tract requires expertise, and sometimes even the expert may face a dilemma as to the precise nature of the biopsies submitted from colposcopically suspicious HPV-related lesions. We have evaluated the role of viral testing using dot blot hybridization (ViraType) and PCR in the diagnosis of histologically typical (42 cases) and equivocal (30 cases) squamous intraepithelial lesions of the vagina (7), vulva (30), perianal epithelium (3), penis (31), and scrotum (1). The viral kits were used according to the manufacturer's instructions in a routine laboratory setting, and the probes available were HPV 6/11, 16, 18, and 31, 33, 35 (the 30s group). HPV DNA was found in 45 of 72 (62%) of all lesions. PCR was more sensitive (58%) than ViraType (39%) for detecting HPV DNA sequences (p less than 0.02), particularly in equivocal lesions (EQHPV), 14 of 30 (47%) by PCR vs 4 of 30 (13%) by ViraType (p less than 0.004). The majority of lesions contained oncogenic type viruses irrespective of their histologic presentation, namely, type 16. Only condylomata acuminata were predominantly HPV 6 or 11 positive. Viral testing may play a role in the quality control of the diagnostic expertise of routine laboratories as well as ascertaining the HPV-relatedness of histologically equivocal lesions of the anogenital tract. In view of the relatively high false-negative rates for detecting HPV DNA by ViraType and PCR, only a positive test may be meaningful unless other HPV types are added.

Quantitation of estrogen and progesterone receptors by immunocytochemical and image analyses.

Compared to other techniques, the ability to detect estrogen and progesterone receptors by immunocytochemical analysis in formalin-fixed, paraffin-embedded sections has clear advantages, including the ability to assay small biopsy specimens, fine-needle aspirate samples, and archival material. Twenty-two cases of breast carcinoma were evaluated for estrogen and progesterone receptors by immunocytochemical analysis and enzyme immunoassay. Using a true color-based image analysis system, histograms of area versus the optical density of the positive staining nuclei were generated. A binary decision algorithm was derived from these histogram parameters by the CART computer program. Estimates generated by the algorithm for image analysis/immunocytochemical analysis had a 90% concordance with the enzyme immunoassay values. It is concluded that quantitative immunocytochemical results for estrogen and progesterone receptor content in formalin-fixed, paraffin-embedded tissue can be generated using image analysis.

DNA ploidy and cell-cycle analysis: tools for assessment of cancer prognosis.

DNA ploidy and cell cycle analysis as measured by flow cytometry (FC) and image analysis (IA) have moved out of the realm of the research laboratory to become valid clinical tests used in the assessment of prognosis of the cancer patient. Although much information on the relationship of DNA ploidy/%S-phase analysis to patient prognosis is available in the literature, the data are not presented in such a way as to be helpful in clinical decision making. Because predictive values and confidence intervals, which measure the likelihood that a given clinical test will rule in or rule out a clinical outcome, were not calculated in previous reviews, conclusions about the clinical utility of these analyses were not possible. Using the available raw data on DNA ploidy and %S-phase analysis from previously published papers, predictive values and confidence limits were calculated for specific clinical presentations. In several such clinical situations (tumor type, stage, etc.), predictive value of greater than 90% was derived. We conclude that in these situations DNA ploidy and %S-phase analysis can be used to predict clinical outcome, to design treatment, and to guide patient management. The evaluation of the clinical utility of these tests must ultimately rest on prospective trials which show that randomized arms respond to treatment regimens dependent upon the DNA ploidy and %S-phase status.

Severe immunodeficiency disease induced by a defective murine leukaemia virus.

Different classes of retroviruses have been shown to induce immunodeficiency diseases in various animal species. These animal models may provide an insight into our understanding of AIDS but, with the exception of one strain of feline leukaemia virus, the determinants of pathogenicity have not yet been mapped to these viral genomes. The immunodeficiency-inducing feline leukaemia virus is replication-defective, harbouring the determinant of pathogenicity within its env sequences. We have studied the Duplan strain of murine leukaemia virus which induces, in C57BL/6 mice, a severe immunodeficiency disease with striking similarities to human AIDS. We have identified the aetiological agent of this murine immunodeficiency disease as another defective retrovirus, with a genome of 4.8 kilobases. Molecular cloning and sequencing of this DNA showed that the pol and env genes have been deleted, but that the complete gag region has been conserved and has a novel sequence encoding the p12 protein. As with the feline leukaemia virus, these results provide evidence for the role of defective retroviruses in inducing immunodeficiency and facilitate the study of the mechanisms underlying the pathogenesis of retrovirus-induced immunodeficiency syndromes, including AIDS.

Contribution of immunohistochemistry to the diagnosis of breast cancer in men.

In men with advanced carcinoma of the prostate who have a breast tumour, it is often difficult to distinguish a primary from a secondary breast lesion. The authors describe the case of a 72-year-old man who presented with a poorly differentiated carcinoma in one breast after receiving estrogen therapy for disseminated prostatic cancer. Application of the unlabelled antibody peroxidase-antiperoxidase immunohistochemical method demonstrated prostate-specific antigen in the tumour cells, thus establishing the secondary origin of the lesion. Five controls--men with primary breast cancer--when tested by the same method did not have this marker. The authors conclude that in this clinical context, prostate-specific antigen is a useful marker of breast cancer in men.