Detection of Membrane-Bound Enzymes in Cells Using Immunoassay and Raman Microspectroscopy

Hawi, Sharon R.; Rochanakij, Sunant; Adar, Fran; Campbell, William B.; Nithipatikom, Kasem

doi:10.1006/abio.1998.2661

Cited by 28 publications

(17 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Colloidal gold nanoparticles have been used widely as SERS substrates to probe components in living cells [142][143][144][145][146][147][148], especially to study the interaction of various antitumor drugs with their pharmacological targets, such as DNA, within living cancer cells [143,145,[149][150][151]. Other studies include cancer gene detection [152][153][154] and cancer protein-biomarker detection [155][156][157][158][159][160]. Recently, Huang et al demonstrated the difference in the SERS of anti-EGFR conjugated gold nanorods between cancer and normal cells (Figure 3) [161].…”

Section: Surface-enhanced Raman Scatteringmentioning

confidence: 99%

Gold Nanoparticles: Interesting Optical Properties and Recent Applications in Cancer Diagnostics and Therapy

et al. 2007

View full text Add to dashboard Cite

Recent years have seen tremendous progress in the design and study of nanomaterials geared towards biological and biomedical applications, most notable among these being the noble metal nanoparticles. In this review, we outline the surface-plasmon resonance-enhanced optical properties of colloidal gold nanoparticles directed towards recent biomedical applications with an emphasis on cancer diagnostics and therapeutics. Methods of molecular-specific diagnostics/detection of cancer, including strongly enhanced surface plasmon resonance light-scattering, surface-enhanced emission of gold nanorods and surface-enhanced Raman scattering, are described. We also discuss the plasmonic photothermal therapy of cancer achieved by using the strongly enhanced surface-plasmon resonance absorption of gold nanospheres and nanorods.

show abstract

Section: Surface-enhanced Raman Scatteringmentioning

confidence: 99%

Gold Nanoparticles: Interesting Optical Properties and Recent Applications in Cancer Diagnostics and Therapy

et al. 2007

View full text Add to dashboard Cite

show abstract

“…Although untreated rat hepatocytes possess low but significant levels of CYP2B1, 28,49,50 in the constitutive situation CYPs other than CYP2B1 or non-CYP enzymes may contribute substantially to the observed effects, potentially further extending the scope of the principle shown in this study. In this context it is interesting that e.g., prostaglandin H synthase (which has recently been found to be expressed in hepatocytes, 51 has been shown to be able to activate CPA. 52 Also, CPA-inactivating enzymes such as aldehyde dehydrogenase may be modulated by PKA-dependent phosphorylation.…”

Section: Discussionmentioning

confidence: 99%

cAMP-dependent phosphorylation of CYP2B1 as a functional switch for cyclophosphamide activation and its hormonal controlin vitro andin vivo

et al. 2001

View full text Add to dashboard Cite

An important feature of cytochrome P450 (CYP) 2B1 is its high ability to convert the prodrug cyclophosphamide (CPA) to therapeutically cytotoxic metabolites, resulting in interstrand DNA-cross-linking and cell death. We have examined whether and how the phosphorylation of CYP2B1 influences CPA metabolic activation in vitro and in vivo. We found first that only part of the total CYP2B1 pool undergoes phosphorylation. This part is fully inactivated. Second, phosphorylation of CYP2B1 in intact hepatocytes reduced by up to 75% toxification of CPA to mutagenic metabolites (totally dependent on the same preferentially CYP2B-catalyzed 4-hydroxylation of CPA as is the generation of highly cytotoxic species). Third, the phosphoacceptor-serine 128 of CYP2B1 in the consensus sequence for interaction with the protein kinase A represents an on/off switch for the activation of CPA depending on the phosphorylation conditions in the cell. Fourth, evidence is presented that the above-described events also occur in vivo. In conclusion, a successful therapy with CPA, helped by forced expression of CYP2B1 in tumor cells (as recently proposed) will, in addition, be profoundly modified by its phosphorylation status. © 2001 Wiley-Liss, Inc. Key words: cyclophosphamide; CYP2B1; phosphorylation; hormonal controlThe activity of many of the foreign compound metabolizing enzymes, e.g., CYP, is of obvious importance for both physiologic homeostasis as well as effects of drugs and environmental chemicals. 1 Much of the detoxification is performed by CYP enzymes. Many of them are inducible and have a broad and overlapping substrate specificity. 2 Interindividual differences in the expression level of CYP enzymes are known, 3 thus contributing to the variations in toxic symptoms and therapeutic response evoked by antineoplastic drugs. 4 CYP2B1 (the major phenobarbital-inducible form) has long been suggested as 1 of the most important enzymes involved in the activation of widely used antineoplastic prodrugs. It catalyzes oxidative desulfuration of thio-TEPA to TEPA as well as 1-electron reduction of Adriamycin to its semiquinone free radical [5][6][7] and, most importantly, is responsible for conversion of the oxazaphosphorines CPA and IFA to their active metabolites. 8,9 It has been shown that 3 specific rat liver CYP enzymes (CYP2B1, CYP2C6 [constitutively expressed, gender independent] and CYP2C11 [constitutively expressed, male specific]) are major catalysts of CPA activation in adult rat liver, 8,10 while a rat CYP3A enzyme additionally contributes to the activation of IFA. 9 The corresponding human enzymes CYP2B6 and CYP3A4 activate CPA and IFA in human liver, respectively. 11-13 It was demonstrated that among rat enzymes CYP2B1 (the ortholog of the human CYP2B6) is by far the most active catalyst. 8,11,12 Activation of CPA to the therapeutic active species has its initial step in the liver. 8 The CYP2B1-catalyzed 4-hydroxylation reaction forms a highly unstable metabolite, which undergoes spontaneous ␤-elimination and gives rise to 2 pro...

show abstract

“…23 The observed PGHS-l was 0.86±0.26 pg4ig of protein (n=4) in normal cells and 0.40±0. 16 pg/hg of protein (n=7) in malignant cells.…”

Section: Localization Of Pghs In Human Hepatocytesmentioning

confidence: 98%

<title>Detection of prostaglandin H synthase (cyclooxygenase) -1 and -2 expression in single cancer cells by confocal Raman microspectroscopy</title>

Nithipatikom

McCoy

Kajdacsy-Balla

et al. 1999

Biomedical Applications of Raman Spectroscopy

Self Cite

View full text Add to dashboard Cite

Prostaglandin H synthase (PGHS), commonly known as cyclooxygenase (COX), is the enzyme involved in the synthesis of biologically active prostanoids from arachidonic acid. There are two PGHS isozymes with identical functions, similar sizes and similar structures. PGHS-1 is constitutively expressed in most mammalian cells where as PGHS-2 is induced by various agents. PGHS-2 expression was increased with inflammation, mitogenesis, and some types of cancer. We have developed a method to simultaneously detect PGHS-1 and PGHS-2 in single cancer cells by using specific antibodies, surface-enhanced Raman scattering, and confocal Raman microspectroscopy. Cells were plated, cultured, incubated for immunocomplexation, and detected directly in the wells of a multiwell plate. The expression and localization of PGHS-1 and-2 in single malignant human hepatocytes was detected and compared to normal cells. Furthermore, the correlation between the expression of PGHS and the invasiveness of cancer cells was investigated in PC-3 cell sublines.

show abstract

Detection of Membrane-Bound Enzymes in Cells Using Immunoassay and Raman Microspectroscopy

Cited by 28 publications

References 30 publications

Gold Nanoparticles: Interesting Optical Properties and Recent Applications in Cancer Diagnostics and Therapy

Gold Nanoparticles: Interesting Optical Properties and Recent Applications in Cancer Diagnostics and Therapy

cAMP-dependent phosphorylation of CYP2B1 as a functional switch for cyclophosphamide activation and its hormonal controlin vitro andin vivo

<title>Detection of prostaglandin H synthase (cyclooxygenase) -1 and -2 expression in single cancer cells by confocal Raman microspectroscopy</title>

Contact Info

Product

Resources

About