George P. Stricklin

Long-term follow-up of patients with cutaneous T-cell lymphoma treated with extracorporeal photochemotherapy

BACKGROUND Few studies have assessed the long-term outcome of patients with cutaneous T-cell lymphoma (CTCL) treated with extracorporeal photochemotherapy (ECP). OBJECTIVE Our objective was to assess the efficacy, safety, and survival of a cohort of patients with refractory T-cell lymphoma in various stages of cutaneous involvement who were treated with ECP. METHODS Twenty patients who had received at least 6 months of ECP between September 1988 and April 1991 were reevaluated and the data analyzed statistically to obtain outcome data through December 1995. RESULTS A complete response (disappearance of all lesions) was obtained in five patients (25%) and a partial response (disappearance of at least 50% of lesions) in five patients (25%). Of the 10 responders, seven (70%) were weaned from ECP. Two of seven patients had a relapse. Ten patients (50%) showed no response to ECP. No statistically significant differences between responders and nonresponders were found with respect to demographic, clinical, or laboratory variables. Seven patients died of causes directly related to CTCL and two patients died of unrelated causes. Median survival time for the entire cohort was 96 months (range, 16 to 152 months). An assessment of early response after 6 to 8 months of ECP had a sensitivity of 100% and a specificity of 90% for predicting long-term (> 4 years) outcome. Adverse effects were minimal. CONCLUSION ECP is a safe effective alternative therapy for CTCL that is refractory to other therapies; it can induce a long-term, disease-free remission in a minority of patients. Response in the first 6 to 8 months of treatment predicts long-term outcome.

Extracorporeal photopheresis for the treatment of cutaneous T-cell lymphoma

BACKGROUND Many regimens are used for cutaneous T-cell lymphoma (CTCL), but with advanced disease response rates and patient survival are not adequate with any current therapy. Recently extracorporeal photochemotherapy (ECP) was proposed as an alternative therapy. OBJECTIVE Our purpose is to present the results of ECP in patients with CTCL refractory to other treatments. METHODS Patients with CTCL received ECP at 3- to 5-week intervals for at least 6 months. All patients except one were in stage T2 (patch/plaque) or higher. Eight patients had extracutaneous disease involving lymph nodes (six patients), bone marrow (five), or Sézary cells (six). The interval between initial symptoms and diagnosis was 5.9 +/- 1.9 years (mean +/- standard error of the mean) and the interval between diagnosis and ECP was 2.2 +/- 0.4 years. RESULTS A complete response (disappearance of all lesions) was obtained in five patients (25%) and a partial response (disappearance of at least 50% of lesions) in six patients (30%). Four patients (20%) showed stabilization of their disease and five progressed (25%). The only variable that predicted responders versus nonresponders was the number of ECP sessions (p < 0.05 by multivariate logistic regression). In contrast, no separate beneficial effect of adjunctive chemotherapy (p > 0.5) or electron beam therapy (p > 0.1) was found. CONCLUSION Long-term ECP may be an effective alternative treatment for CTCL refractory to other therapies and is likely to be even more useful when combined with other modalities.

Expression of Matrix Metalloproteinases and Tissue Inhibitor of Metalloproteinases in Head and Neck Squamous Cell Carcinoma

Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes implicated in the invasion and metastasis of many cancers. In situ hybridization techniques were used to reveal sites of expression of collagenase (MMP-1), gelatinase 72 kd (MMP-2), gelatinase 92 kd (MMP-9), and tissue inhibitor of metalloproteinase-1 (TTMP-1) in head and neck carcinomas (N=21). Both TTMP-1 and gelatinase 72 kd were expressed in nearly all tumors, whereas the expression of collagenase and gelatinase 92 kd showed variability. Tumor-associated expression of MMPs was strongest in stromal cells near advancing margins. No differences in expression levels were detected between primary and metastatic sites. This paper reviews the literature and discusses the significance and possible implications of MMPs in head and neck squamous cell carcinoma.

Human skin collagenase in recessive dystrophic epidermolysis bullosa. Purification of a mutant enzyme from fibroblast cultures.

Recessive dystrophic epidermolysis bullosa, a genodermatosis characterized by dermolytic blister formation in response to minor trauma, is characterized by an incresaed collagenase synthesis by skin fibroblasts in culture. Since preliminary studies of partially purified recessive dystrophic epidermolysis bullosa collagenase suggested that the protein itself was aberrant, efforts were made to purify this enzyme to homogeneity, so that detailed biochemical and immunologic comparisons could be made with normal human skin fibroblast collagenase. Recessive dystrophic epidermolysis bullosa skin fibroblasts obtained from a patient documented to have increased synthesis of the enzyme were grown in large scale tissue culture and both serum-free and serum-containing medium collected as a source of collagenase. The recessive dystrophic epidermolysis bullosa collagenase was purified to electrophoretic homogeneity using a combination of salt precipitation, ion-exchange, and gel-filtration chromatography. In contrast to the normal enzyme, the recessive dystrophic epidermolysis bullosa collagenase bound to carboxymethyl-cellulose at Ca(2+) concentrations at least 10 times higher than those used with the normal enzyme. Additionally, this enzyme was significantly more labile to chromatographic manipulations, particularly when serum-free medium was used. However, rapid purification from serum-containing medium yielded a preparation enzymatically equivalent to normal human skin collagenase. Like the normal enzyme, the recessive dystrophic epidermolysis bullosa collagenase was secreted as a set of two closely related zymogens of approximately 60,000 and approximately 55,000 daltons that could be activated by trypsin to form enzymically active species of approximately 50,000 and approximately 45,000 daltons, respectively. Amino acid analysis suggested slight variations between the normal and recessive dystrophic epidermolysis bullosa collagenases. Cyanogen bromide digests demonstrated peptides unique to the enzyme from each source. The recessive dystrophic epidermolysis bullosa proenzyme was significantly more thermolabile at 60 degrees C than the normal, a finding that correlated with an approximate fourfold decrease in the affinity of the mutant enzyme for Ca(2+), a known activator and stabilizer of human skin collagenase. Aside from the altered affinity for this metal cofactor, kinetic analysis of the structurally altered recessive dystrophic epidermolysis bullosa collagenase revealed that its reaction rates and substrate specificity for human collagen types I-V were identical to those for the normal enzyme. Likewise, enzymes from both sources displayed identical energies of activation and deuterium isotope effects. Antisera were raised to the normal and putatively mutant procollagenases respectively, and, although they displayed a reaction of identity in double diffusion analysis, immunologic differences were present in enzyme inhibition and quantitative precipitation studies. These studies indicate that recessive dystrophic epidermolysis bullosa is characterized by the increased synthesis of an enzymically normal, but structurally aberrant, collagenase.

In vitro changes in non-facial human skin following CO2 laser resurfacing: A comparison study

We evaluated the physical changes in human skin following CO2 laser cutaneous resurfacing with either the Sharplan SilkTouch® handpiece or the Coherent UltraPulse® laser.

Long-term Efficacy of Topical Fluorouracil Cream, 5%, for Treating Actinic Keratosis: A Randomized Clinical Trial

IMPORTANCE Topical fluorouracil was demonstrated to be effective in reducing the number of actinic keratoses (AKs) for up to 6 months, but no randomized trials studied its long-term efficacy. OBJECTIVE To evaluate the long-term efficacy of a single course of fluorouracil cream, 5%, for AK treatment. DESIGN, SETTING, AND PARTICIPANTS The Veterans Affairs Keratinocyte Carcinoma Chemoprevention (VAKCC) trial was a randomized, double-blinded, placebo-controlled trial with patients from dermatology clinics at 12 VA medical centers recruited from 2009 to 2011 and followed up until 2013. Our study population comprised 932 veterans with 2 or more keratinocyte carcinomas in the 5 years prior to enrollment. The mean follow-up duration was 2.6 years in both treatment and control groups. INTERVENTIONS Participants applied either topical fluorouracil cream, 5% (n = 468), or vehicle control cream (n = 464) to the face and ears twice daily for up to 4 weeks. MAIN OUTCOMES AND MEASURES This study reports on AK counts and treatments, which were secondary outcomes of the VAKCC trial. Actinic keratoses on the face and ears were counted by study dermatologists at enrollment and at study visits every 6 months. The number of spot treatments for AKs on the face and ears at semiannual study visits and in between study visits was recorded. RESULTS The number of AKs on the face and ears per participant was not different between the fluorouracil and control groups at randomization (11.1 vs 10.6, P > .10). After randomization, the fluorouracil group had fewer AKs compared with the control group at 6 months (3.0 vs 8.1, P < .001) and for the overall study duration (P < .001). The fluorouracil group also had higher complete AK clearance rates (38% vs 17% at 6 months) and fewer spot treatments at 6-month intervals, at study visits, and in between study visits during the trial (P < .01 for all). The fluorouracil group took longer to require the first spot AK treatment (6.2 months) compared with the control group (6.0 months) (hazard ratio, 0.69; 95% CI, 0.60-0.79). The number of hypertrophic AKs was not different between the 2 groups overall (P = .60), although there were fewer hypertrophic AKs in the fluorouracil group at 6 months (0.23 vs 0.41) (P = .05). CONCLUSIONS AND RELEVANCE Our results indicate that a single course of fluorouracil cream, 5%, effectively reduces AK counts and the need for spot treatments for longer than 2 years. TRIAL REGISTRATION clinicaltrials.gov Identifier:NCT00847912.

Human Fibroblast Tissue Inhibitor of Metalloproteinases: Glycosylation and Function

The glycosylation of human fibroblast tissue inhibitor of metalloproteinases and procollagenase was examined in vivo using tunicamycin B2 and in vitro using Peptide: N-glycosidase F. In the presence of tunicamycin B2, unglycosylated inhibitor continues to be synthesized and secreted at normal or increased rates. The protein core of this collagenase inhibitor has an apparent Mr of 21,000 and possesses at least two oligosaccharide linkage sites as evidenced by the accumulation of a single 25,000 dalton intermediate. Collagenase inhibitor deglycosylated by Peptide: N-glycosidase also has an apparent molecular weight of 21,000 daltons; furthermore, deglycosylated inhibitor continues to block the activity of collagenase at a 1:1 molar stoichiometry and does not differ from the native glycoprotein in its resistance to tryptic degradation. Using these same two reagents, the characteristic doublet of human fibroblast procollagenase was found to result from glycosylation in the upper (60,000 dalton) form. Secretion of procollagenase was significantly inhibited in the presence of tunicamycin B2.

Comparisons of wound healing among excisional, laser-created, and standard thermal burns in porcine wounds of equal depth.

The present study was designed to characterize similarities and differences among three wounding modalities in partial‐thickness porcine wounds. We hypothesized that inherent differences, such as endogenous cytokine delivery into excisional wounds or ablation of eschar during laser vaporization, should accelerate the magnitude and sequence of reparative events above the delayed repair that is frequently observed in patients with burns. A constant mid‐dermal depth of injury was created by a Padgett dermatome, a computer‐controlled pulsed CO2 laser, or a temperature‐controlled metal template. Wounds were harvested after 5, 10, or 15 days. After 5 days, significant resurfacing differences were apparent with values of 54% in excisions, 29% in lasers, and 12% in standard thermal burns. Sequences of fibroblastic proliferation were measurably different among the three wound modalities. At day 5 the bromodeoxyuridine labeling index for fibroblasts showed laser wound levels greater than excision wound levels, which were greater than burn wound levels; but by day 10, the proliferative profiles indicated that burn wound levels were greater than excision wound levels, which were greater than laser wound levels. Capillary areas (an assessment of angiogenesis) differed among the three wound types throughout the study. Peak values were observed at day 5 in both excisional and laser injuries; however, standard thermal burns did not peak until day 10. Both the magnitude and sequence of expression of three matrix metalloproteinases (−1, −2, and −9) differed among the three types of injuries. Laser wounds showed the earliest peak in matrix metalloproteinase‐1 expression, whereas burns showed the least expression at day 5. In conclusion, although the three types of wounds undergo similar reparative processes such as reepithelialization, fibroblastic proliferation, angiogenesis, and expression of matrix metalloproteinases, the magnitude and temporal sequences are measurably altered among the three wound modalities. A greater understanding of specific differences within wound environments may lead to more insightful design of interventional wound therapies.

Immunolocalization of collagenase inhibitor in normal skin and basal cell carcinoma.

Human collagenase inhibitor is a ubiquitous glycoprotein capable of blocking the action of several connective tissue metalloproteinases, including collagenase, gelatinase, and proteoglycanase. The action of this proteinase inhibitor may constitute a pivotal step in the control of connective tissue matrix degradation. Using monospecific antibody to collagenase inhibitor as an immunocytochemical probe, we determined its in vivo localization in normal human skin and in a pathologic state, the altered connective tissue stroma surrounding basal cell carcinoma. Collagenase inhibitor was localized diffusely throughout the dermis and appeared to be associated with the extracellular matrix components, both in normal skin and in basal cell carcinoma. Intense staining was present in the stroma surrounding islands of basal cell carcinoma. The increased amounts of collagenase inhibitor may be a result of its production by stromal fibroblasts stimulated by cytokines of tumor or inflammatory cell origin. These findings are similar to those previously described for dermal collagenase. Both collagenase inhibitor and collagenase itself appear to be normal components of the extracellular matrix, and amounts of both are increased in the altered stroma surrounding neoplastic cells. Thus we suggest that the balance of degradative proteinase(s) to specific inhibitor may be an important factor in determining the composition of the extracellular matrix.

Free Electron Laser infrared wavelength specificity for cutaneous contraction

Short pulsed and scanned CO2 lasers that target water molecules are currently used for cutaneous resurfacing. These CO2 resurfacing lasers produce acute cutaneous contraction, which can be quantitated as a measure of the lasers effect. We postulated that targeting the vibrational and rotational modes of proteins with specific infrared laser wavelengths might be more effective at inducing cutaneous contraction than the CO2 resurfacing lasers.