CXCR4-SDF-1 interaction potentially mediates trafficking of circulating tumor cells in primary breast cancer.

BACKGROUND: Cytokines are involved in cancer invasion and metastasis. Circulating tumor cells (CTCs) play key role in tumor dissemination and are an independent survival predictor in breast cancer patients. The aim of this study was to assess correlation between CTCs and plasma cytokines in primary breast cancer (PBC) patients. METHODS: This study included 147 chemotherapy naïve PBC patients. Peripheral blood mononuclear cells (PBMC) were depleted of hematopoetic cells using RossetteSep™ negative selection kit. RNA extracted from CD45-depleted PBMC was interrogated for expression of EMT (Twist1, Snail1, Slug, Zeb1) and epithelial (Ck19) gene transcripts by qRT-PCR. The concentrations of 51 plasma cytokines were measured using multiplex bead arrays. RESULTS: CTCs were detected in 25.2% patients. CTCs exhibiting only epithelial markers (CTC_EP) and only EMT markers (CTC_EMT) were present evenly in 11.6% patients, while CTCs co-expressing both markers were detected in 2.0% patients. Patients with presence of CTC_EP in peripheral blood had significantly elevated levels of plasma IFN-α2, IL-3, MCP-3, ß-NGF, SCF, SCGF-ß, TNF-ß and SDF-1 compared to patients without CTC_EP. CTC_EP exhibited overexpression of SDF-1 receptor and CXCR4, but not other corresponding cytokine receptor, and in multivariate analysis SDF-1 was independently associated with CTC_EP. There was an inverse correlation between CTC_EMT and plasma cytokines CTACK, ß-NGF and TRAIL, while presence of either subtype of CTCs was associated with increased level of TGF-ß2. CONCLUSION: Using cytokine profiling, we identified cytokines associated with CTCs subpopulations in peripheral blood of PBC. Our data suggest that CXCR4-SDF-1 axis is involved in mobilization and trafficking of epithelial CTCs.

Relationship between circulating tumor cells and epithelial to mesenchymal transition in early breast cancer.

BACKGROUND: Circulating tumor cells (CTCs) play a crucial role in tumor dissemination and are an independent survival predictor in breast cancer (BC) patients. Epithelial to mesenchymal transition (EMT) is involved in cancer invasion and metastasis. The aim of this study was to assess correlation between CTCs and expression of EMT transcription factors TWIST1 and SLUG in breast tumor tissue. METHODS: This study included 102 early BC patients treated by primary surgery. Peripheral blood mononuclear cells (PBMC) were depleted of hematopoietic cells using RossetteSep™ negative selection kit. RNA extracted from CD45-depleted PBMC was interrogated for expression of EMT (TWIST1, SNAIL1, SLUG, FOXC2 and ZEB1) and epithelial (KRT19) gene transcripts by qRT-PCR. Expression of TWIST1 and SLUG in surgical specimens was evaluated by immunohistochemistry and quantified by multiplicative score. RESULTS: CTCs were detected in 24.5 % patients. CTCs exhibiting only epithelial markers were present in 8.8 % patients, whereas CTCs with only EMT markers were observed in 12.8 % of pts and CTCs co-expressing both markers were detected in 2.9 % pts. We observed lack of correlation between CTCs and expression of TWIST1 and SLUG in breast cancer cells or cancer associated stroma. Lack of correlation was observed for epithelial CTCs as well as for CTCs with EMT. CONCLUSIONS: In this translational study, we showed a lack of association between CTCs and expression of EMT-inducing transcription factors, TWIST1 and SLUG, in breast tumor tissue. Despite the fact that EMT is involved in cancer invasion and metastasis our results suggest, that expression of EMT proteins in unselected tumor tissue is not surrogate marker of CTCs with either mesenchymal or epithelial features.

Long-term treatment efficacy in primary inflammatory breast cancer by hormonal receptor- and HER2-defined subtypes.

BACKGROUND: Subtypes defined by hormonal receptor (HR) and HER2 status have not been well studied in inflammatory breast cancer (IBC). We characterized clinical parameters and long-term outcomes, and compared pathological complete response (pCR) rates by HR/HER2 subtype in a large IBC patient population. We also compared disease-free survival (DFS) and overall survival (OS) between IBC patients who received targeted therapies (anti-hormonal, anti-HER2) and those who did not. PATIENTS AND METHODS: We retrospectively reviewed the records of patients diagnosed with IBC and treated at MD Anderson Cancer Center from January 1989 to January 2011. Of those, 527 patients had received neoadjuvant chemotherapy and had available information on estrogen receptor (ER), progesterone receptor (PR), and HER2 status. HR status was considered positive if either ER or PR status was positive. Using the Kaplan-Meier method, we estimated median DFS and OS durations from the time of definitive surgery. Using the Cox proportional hazards regression model, we determined the effect of prognostic factors on DFS and OS. Results were compared by subtype. RESULTS: The overall pCR rate in stage III IBC was 15.2%, with the HR-positive/HER2-negative subtype showing the lowest rate (7.5%) and the HR-negative/HER2-positive subtype, the highest (30.6%). The HR-negative, HER2-negative subtype (triple-negative breast cancer, TNBC) had the worst survival rate. HR-positive disease, irrespective of HER2 status, had poor prognosis that did not differ from that of the HR-negative/HER2-positive subtype with regard to OS or DFS. Achieving pCR, no evidence of vascular invasion, non-TNBC, adjuvant hormonal therapy, and radiotherapy were associated with longer DFS and OS. CONCLUSIONS: Hormone receptor and HER2 molecular subtypes had limited predictive and prognostic power in our IBC population. All molecular subtypes of IBC had a poor prognosis. HR-positive status did not necessarily confer a good prognosis. For all IBC subtypes, novel, specific treatment strategies are needed in the neoadjuvant and adjuvant settings.

Gene expression profiles of inflammatory breast cancer: correlation with response to neoadjuvant chemotherapy and metastasis-free survival.

BACKGROUND: Inflammatory breast cancer (IBC) is an aggressive disease. To date, no molecular feature reliably predicts either the response to chemotherapy (CT) or the survival. Using DNA microarrays, we searched for multigene predictors. PATIENTS AND METHODS: The World IBC Consortium generated whole-genome expression profiles of 137 IBC and 252 non-IBC (nIBC) samples. We searched for transcriptional profiles associated with pathological complete response (pCR) to neoadjuvant anthracycline-based CT and distant metastasis-free survival (DMFS) in respective subsets of 87 and 106 informative IBC samples. Correlations were investigated with predictive and prognostic gene expression signatures published in nIBC (nIBC-GES). Supervised analyses tested genes and activation signatures of 19 biological pathways and 234 transcription factors. RESULTS: Three of five tested prognostic nIBC-GES and the two tested predictive nIBC-GES discriminated between IBC with and without pCR, as well as two interferon activation signatures. We identified a 107-gene signature enriched for immunity-related genes that distinguished between responders and nonresponders in IBC. Its robustness was demonstrated by external validation in three independent sets including two IBC sets and one nIBC set, with independent significant predictive value in IBC and nIBC validation sets in multivariate analysis. We found no robust signature associated with DMFS in patients with IBC, and neither of the tested prognostic GES, nor the molecular subtypes were informative, whereas they were in our nIBC series (220 stage I-III informative samples). CONCLUSION: Despite the relatively small sample size, we show that response to neoadjuvant CT in IBC is, as in nIBC, associated with immunity-related processes, suggesting that similar mechanisms responsible for pCR exist. Analysis of a larger IBC series is warranted regarding the correlation of gene expression profiles and DMFS.

Statin use in primary inflammatory breast cancer: a cohort study.

BACKGROUND: Some studies have suggested that statins, which have cholesterol-lowering and anti-inflammatory properties, may have antitumor effects. Effects of statins on inflammatory breast cancer (IBC) have never been studied. METHODS: We reviewed 723 patients diagnosed with primary IBC in 1995-2011 and treated at The University of Texas MD Anderson Cancer Center. Statin users were defined as being on statins at the initial evaluation. Based on Ahern et al's statin classification (JNCI, 2011), clinical outcomes were compared by statin use and type (weakly lipophilic to hydrophilic (H-statin) vs lipophilic statins (L-statin)). We used the Kaplan-Meier method to estimate the median progression-free survival (PFS), overall survival (OS) and disease-specific survival (DSS), and a Cox proportional hazards regression model to test the statistical significance of potential prognostic factors. RESULTS: In the multivariable Cox model, H-statins were associated with significantly improved PFS compared with no statin (hazard ratio=0.49; 95% confidence interval=0.28-0.84; P<0.01); OS and DSS P-values were 0.80 and 0.85, respectively. For L-statins vs no statin, P-values for PFS, DSS, and OS were 0.81, 0.4, and 0.74, respectively. CONCLUSION: H-statins were associated with significantly improved PFS. A prospective randomised study evaluating the survival benefits of statins in primary IBC is warranted.

Clinical relevance of cancer stem cells in bone marrow of early breast cancer patients.

BACKGROUND: Cancer stem cells (CSCs) are epithelial tumor cells that express CD44(+)CD24(-/lo). CSCs can be further divided into those that have aldehyde dehydrogenase (ALDH) activity (Aldefluor(+)) and those that do not. We hypothesized that if CSCs are responsible for tumor dissemination, their presence in bone marrow (BM) would be prognostic in early stages of breast cancer (EBC) patients. PATIENTS AND METHODS: BM aspirates were collected at the time of surgery from 108 patients with EBC. BM was analyzed for CSCs and ALDH activity by flow cytometry. Overall survival and disease-free survival (DFS) were calculated from the date of diagnosis and analyzed with Kaplan-Meier survival plots. Cox multivariate proportional hazards model was also carried out. RESULTS: Patients with CSCs in BM had a hazard ratio (HR) of 8.8 for DFS (P = 0.002); patients with Aldefluor(+) CSCs had a HR of 5.9 (P = 0.052) for DFS. All deceased patients (n = 7) had CSCs in BM. In multivariate analysis, the presence of CSCs in BM was a prognostic factor of DFS (HR = 15.8, P = 0.017). CONCLUSIONS: The presence of BM metastasis is correlated with CSCs and these CSCs irrespective of ALDH activity are an independent adverse prognostic factor in EBC patients.

Comparison of assay methods for detection of circulating tumor cells in metastatic breast cancer: AdnaGen AdnaTest BreastCancer Select/Detect™ versus Veridex CellSearch™ system.

The detection of CTCs prior to and during therapy is an independent and strong prognostic marker, and it is predictive of poor treatment outcome. A major challenge is that different technologies are available for isolation and characterization of CTCs in peripheral blood (PB). We compare the CellSearch system and AdnaTest BreastCancer Select/Detect, to evaluate the extent that these assays differ in their ability to detect CTCs in the PB of MBC patients. CTCs in 7.5 ml of PB were isolated and enumerated using the CellSearch, before new treatment. Two cutoff values of ≥2 and ≥5 CTCs/7.5 ml were used. AdnaTest requires 5 ml of PB to detect gene transcripts of tumor markers (GA733-2, MUC-1, and HER2) by RT-PCR. AdnaTest was scored positive if ≥1 of the transcript PCR products for the 3 markers were detected at a concentration ≥0.15 ng/µl. A total of 55 MBC patients were enrolled. 26 (47%) patients were positive for CTCs by the CellSearch (≥2 cutoff), while 20 (36%) were positive (≥5 cutoff). AdnaTest was positive in 29 (53%) with the individual markers being positive in 18% (GA733-2), 44% (MUC-1), and 35% (HER2). Overall positive agreement was 73% for CTC≥2 and 69% for CTC≥5. These preliminary data suggest that the AdnaTest has equivalent sensitivity to that of the CellSearch system in detecting 2 or more CTCs. While there is concordance between these 2 methods, the AdnaTest complements the CellSearch system by improving the overall CTC detection rate and permitting the assessment of genomic markers in CTCs.

Circulating tumor cells in immunohistochemical subtypes of metastatic breast cancer: lack of prediction in HER2-positive disease treated with targeted therapy.

BACKGROUND: Circulating tumor cells (CTCs) are associated with inferior prognosis in metastatic breast cancer (MBC). We hypothesized that the relationship between CTCs and disease subtype would provide a better understanding of the clinical and biologic behavior of MBC. PATIENTS AND METHODS: We retrospectively analyzed 517 MBC patients treated at a single institution. Subtypes of primary tumors were analyzed by immunohistochemical (IHC) or fluorescent in situ hybridization analyses and CTCs were enumerated by CellSearch(®) at starting a new therapy. Overall survival (OS) and progression-free survival durations for each IHC subtype were determined. RESULTS: At a median follow-up of 24.6 months, 276 of 517 (53%) patients had died. The median OS for patients with <5 and ≥ 5 CTCs were 32.4 and 18.3 months, respectively (P < 0.001). Except in HER2+ patients, the prognostic value of CTCs was independent of disease subtype and disease site. CONCLUSIONS: In this large retrospective study, CTCs were strongly predictive of survival in all MBC subtypes except HER2+ patients who had been treated with targeted therapy. Our results clearly demonstrate the value of enumerating CTCs in MBC and strongly suggest an interesting biological implication in the HER2+ subset of patients that need to be further explored.

Levels of specific serum N-glycans identify breast cancer patients with higher circulating tumor cell counts.

BACKGROUND: Metastatic breast cancer (MBC) is currently an incurable condition that is primarily treated with palliative measures. Isolation of circulating tumor cells (CTCs) from the peripheral blood of these patients provides a predictive prognostic indicator, independent of the type of therapy, site of occurrence and biological characteristics of the primary disease. It has been well established that glycosylation processing pathways are disturbed in cancer, leading to alterations in the glycan content of glycoproteins. MATERIALS AND METHODS: The bi-, tri- and tetraantennary glycans containing sialyl Lewis x (sLe(x)) epitopes (A2F1G1, A3F1G1, A4F1G1 and A4F2G2) were quantified using normal phase high-performance liquid chromatography in combination with exoglycosidase array digestions in the glycan pools released from sera of 27 patients with advanced breast cancer (16 with CTCs <5/7.5 ml and 11 with CTCs ≥5/7.5 ml) and 13 healthy women. RESULTS: The levels of all these glycans were significantly higher in patients with CTCs ≥5/7.5 ml compared with patients with CTCs <5/7.5 ml. CONCLUSIONS: As high levels of glycans containing sLe(x) epitopes were associated with CTCs, their measurement may provide a new noninvasive approach for determining prognosis in women with MBC.

Circulating tumor cells and bone metastases as detected by FDG-PET/CT in patients with metastatic breast cancer.

BACKGROUND: We evaluated the relationship between the detection and prognostic significance of circulating tumor cells (CTCs) and sites of metastases detected by 2-[fluorine-18]fluoro-2-deoxy-D-glucose-positron emission tomography/computed tomography (FDG-PET/CT) in patients with metastatic breast cancer (MBC). PATIENTS AND METHODS: From May 2004 to January 2008, 195 patients with relapsed/progressive MBC underwent whole-body FDG-PET/CT and provided blood samples for assessment of CTC count. RESULTS: Higher CTC numbers were detected in patients with bone metastases relative to those with no bone lesions (mean 65.7 versus 3.3, P = 0.0122) and in patients with multiple bone metastases relative to those with one or two bone lesions (mean 77.7 versus 2.6, P < 0.001). CTCs predicted overall survival (OS) in 108 patients with multiple sites of metastases including bone (P = 0.0008) but not in 58 without bone metastases (P = 0.4111) and in 29 with bone involvement only (P = 0.3552). All 15 patients but one with human epidermal growth factor receptor 2 (HER-2) positive tumors who were treated with trastuzumab-based regimens had <5 CTCs at progression. In multivariate analysis, CTCs, but not bone metastases, remained a significant predictor of OS. CONCLUSION: Presence of extensive bone metastases as detected by FDG-PET/CT is associated with increased CTC numbers in MBC.

Circulating tumour cells are associated with increased risk of venous thromboembolism in metastatic breast cancer patients.

BACKGROUND: Cancer is a risk factor for venous thromboembolism (VTE). Circulating tumour cells (CTCs) are an independent predictor of survival in metastatic breast cancer (MBC) patients. The aim of this study was to test the hypothesis that CTCs are associated with the risk of VTE in MBC patients. METHODS: This retrospective study included 290 MBC patients treated in the MD Anderson Cancer Center from January 2004 to December 2007. Circulating tumour cells were detected and enumerated using the CellSearch system before starting new lines of therapy. RESULTS: At a median follow-up of 12.5 months, 25 patients experienced VTE and 53 patients died without experiencing thrombosis. Cumulative incidence of thrombosis at 12 months was 8.5% (95% confidence interval (CI)=5.5%, 12.4%). Patients with CTCs > or = 1 and > or = 5 had a higher incidence of VTE compared with patients with 0 and <5 CTCs (12-month estimate, 11.7 and 11.6% vs 3 and 6.6%; P=0.006 and P=0.076, respectively). In the multivariate model, patients with CTCs > or = 1 had a hazard ratio of VTE of 5.29 (95% CI=1.58, 17.7, P=0.007) compared with patients with no CTCs. CONCLUSION: These results suggest that CTCs in MBC patients are associated with increased risk of VTE. These patients should be followed up more closely for the risk of VTE.

Circulating tumor cells in metastatic inflammatory breast cancer.

BACKGROUND: Inflammatory breast cancer (IBC) is the most aggressive form of breast cancer. Circulating tumor cells (CTCs) are an independent prognostic factor in metastatic breast cancer. The aim of this study was to assess the prognostic value of baseline CTCs in metastatic IBC patients. PATIENTS AND METHODS: This retrospective study included 42 metastatic IBC and 107 metastatic non-IBC patients treated with first- or second-line chemotherapy from January 2004 to December 2007 at MD Anderson Cancer Center. CTCs were detected and enumerated before patients started chemotherapy using the CellSearch system. RESULTS: Ten (23.8%) IBC patients versus 48 (44.9%) non-IBC patients had baseline CTCs > or =5 per 7.5 ml of peripheral blood. IBC patients had a lower mean +/- SEM CTCs than non-IBC patients (7.6 +/- 2.9 versus 34.2 +/- 9.1; P = 0.02). The estimated median overall survival was 26.5 versus 18.3 months (P = 0.68) in IBC patients and 37.4 versus 18.3 months (P = 0.016) in non-IBC patients with CTCs <5 and CTCs > or =5, respectively. CONCLUSIONS: Metastatic IBC patients had a lower prevalence and fewer CTCs in comparison to metastatic non-IBC patients. Survival of metastatic IBC patients with <5 CTCs was not significantly better than that of patients with > or =5 CTCs. Further research is warranted with prospective assessment of CTCs in IBC patients and their biological characterization.

Imatinib mesylate (Gleevec) in advanced breast cancer-expressing C-Kit or PDGFR-beta: clinical activity and biological correlations.

BACKGROUND: Novel molecular therapies for metastatic breast cancer (MBC) are necessary to improve the dismal prognosis of this condition. Imatinib mesylate (Gleevec) inhibits several protein tyrosine kinases, including platelet-derived growth factor receptor (PDGFR) and c-kit, which are preferentially expressed in tumor cells. We tested the activity of imatinib mesylate in MBC with overexpression of PDGFR or c-kit. Additionally, we sought to determine the biological correlates and immunomodulatory effects. PATIENTS AND METHODS: Thirteen patients were treated with Imatinib administered orally at 400 mg p.o. b.i.d. (800 mg/day), until disease progression. All patients demonstrated PDGFR-beta overexpression and none showed c-kit expression. RESULTS: No objective responses were observed among the 13 patients treated in an intention-to-treat analysis. All patients experienced disease progression, with a median time to progression of 1.2 months. Twelve patients have died, and the median overall survival was 7.7 months. No patient had a serious adverse event. Imatinib therapy had no effect on the plasma levels of the angiogenesis-related cytokines, vascular endothelial growth factor, PDGF, b-fibroblast growth factor, and E-selectin. Immune studies showed imatinib inhibits interferon-gamma production by TCR-activated CD4(+) T cells. CONCLUSION: Imatinib as a single agent has no clinical activity in PDGFR-overexpressing MBC and has potential immunosuppressive effects.

In vitro and in vivo effect of flutamide on steroid hormone secretion in canine and human inflammatory breast cancer cell lines.

The aim was to study the effects of flutamide on cell proliferation, in vivo tumour growth and steroid production in canine and human IBC cell lines. IPC-366 and SUM149 cell cultures were exposed to flutamide concentrations for 72 hours. Additionally, IPC-366 and SUM149 xenotransplanted mice were treated subcutaneously with flutamide 3 times a week for 2 weeks. Steroid hormones determination in culture media, serum and tumour homogenates (pregnenolone, progesterone, androstenedione, testosterone, dihydrotestosterone, 17ß-oestradiol and oestrone sulphate) were assayed by EIA. in vitro cell proliferation percentages showed a decrease in all flutamide dosages in IPC-366 and SUM149. in vivo flutamide reduced tumour size by 55% to 65%, and metastasis rates decreased. In treated groups, androgen levels in culture media, serum and tumour homogenates were increased as oestrogen levels decreased. These results suggest that flutamide treatment inhibits cell proliferation and promotes tumour reduction by increasing androgen levels and also support future therapy approaches.

Canine cell line, IPC-366, as a good model for the study of inflammatory breast cancer.

Inflammatory breast cancer (IBC) is an aggressive type of cancer with poor survival in women. Inflammatory mammary cancer (IMC) in dogs is very similar to human IBC and it has been proposed as a good surrogate model for study the human disease. The aim was to determine if IPC-366 shared characteristics with the IBC cell line SUM149. The comparison was conducted in terms of ability to grow (adherent and nonadherent conditions), stem cell markers expression using flow cytometry, protein production using western blot and tumorigenic capacity. Our results revealed that both are capable of forming long-term mammospheres with a grape-like morphology. Adherent and nonadherent cultures exhibited fast growth in vivo. Stem cell markers expressions showed that IPC-366 and SUM149 in adherent and nonadherent conditions has mesenchymal-like characteristics, E-cadherin and N-cadherin, was higher in adherent than in nonadherent cultures. Therefore, this study determines that both cell lines are similar and IPC-366 is a good model for the human and canine disease.

Imatinib mesylate suppresses cytokine synthesis by activated CD4 T cells of patients with chronic myelogenous leukemia.

Although imatinib mesylate (IM) is highly effective at inducing complete cytogenetic remission in patients with chronic myelogenous leukemia (CML), it is known to suppress T-cell proliferation in vitro. As cytokines are required for T-cell proliferation, we investigated the effects of IM on cytokine synthesis by T cells of CML patients by assessing cytokine synthesis by activated CD4+ and CD8+ T cells in vitro. The activation of T cells in the whole blood of IM-treated patients (CML-IM) with Staphylococcus enterotoxin B resulted in significantly lower percentages of CD4+ T cells that synthesized interleukin 2 (P = 0.017), interferon-gamma (P = 0.010), and tumor necrosis factor-alpha (P = 0.009) than did the activated T cells of control subjects. The addition of exogenous IM to the cultures of peripheral blood mononuclear cells of CML-IM patients reduced Th1 cytokine synthesis by the CD4+ T cells. Furthermore, IM therapy at clinical doses suppressed the tyrosine phosphorylation of ZAP70. These findings suggest that inhibition of ZAP70 signaling pathway and suppression of Th1 cytokine synthesis by CD4+ T cells required the presence of IM at the time of T-cell activation through the T-cell receptor.

Immunological characterizations of patients with acquired immune deficiency syndrome, acquired immune deficiency syndrome-related symptom complex, and a related life-style.

Immunological, hematological, and biochemical studies were done at the time of referral in 135 homosexual subjects, 28 of whom were symptom free (SF), 74 of whom had the acquired immune deficiency syndrome (AIDS)-related symptom complex (ARC), and 33 of whom had AIDS with Kaposi's sarcoma, opportunistic infection, or both. Of 38 laboratory parameters, 11 were significantly different than controls in the SF patients, 19 in the ARC patients, and 20 in the AIDS patients. In SF patients, delayed hypersensitivity was significantly suppressed for 6 of 12 recall antigens. In addition, the percentage of circulating lymphocytes, the percentage of T3+ cells, the percentage and absolute number of T4+ cells, the T4/T8 ratio, the blastogenic responses to phytohemagglutinin, pokeweed mitogen, and concanavalin A were depressed significantly in this group. In contrast, the percentage and absolute granulocyte count, the serum lysozyme, and the serum thymosin alpha 1 were significantly elevated in these patients. In patients with more advanced disease (ARC and AIDS), immunological and hematological parameters tended to worsen. Thus, in the AIDS patients the white blood cell count, percentage, and absolute T11+ cells, absolute T3+ cells, percentage of T4+ cells and absolute level of B-cells, as well as the monocyte adherence and delayed hypersensitivity responses to 12 of 12 recall antigens were depressed. Serum levels of thymosin alpha 1 were equally elevated in all three groups. Serum interferon was found in 15 of 18 opportunistic infection patients with or without Kaposi's sarcoma, in 3 of 9 Kaposi's sarcoma patients without opportunistic infection, but in none of the ARC or SF patients. This study has demonstrated that SF sexually active homosexuals have a characteristic pattern of immune deficiency and that immunodeficiency worsens as one compares SF to ARC to AIDS patients. The study has provided a data base for the development of prognostic criteria and for characterization and evaluation of immunorestorative and immunomodulatory therapy.

Association of levels of circulating C1q binding macromolecules with induction chemotherapy response in head and neck cancer patients.

Ninety-five untreated patients with squamous cell carcinoma of the upper aerodigestive tract expressed significantly higher levels of C1q-binding macromolecules as compared to 45 noncancer-bearing controls. No relationship between C1q-binding macromolecules and levels of circulating IgG-immune complexes as determined by the solid-phase C1q-binding assay or the C3d-solid-phase assay could be defined suggesting that C1q-binding macromolecules were distinct from IgG-circulating immune complexes. An elevated level of C1q-binding macromolecules within these patients was predictive of subsequent response to induction chemotherapy; those with elevated levels characteristically showed no response. Using multivariate logistic regression analysis including the covariates of American Joint Committee staging parameters as well as C1q assay results, levels of the isolated macromolecules added significant prognostic information as to the probability of chemotherapeutic response. The quantitation of C1q macromolecules has clinical implication as to choice of therapeutic regimens against head and neck cancer. The nature of these substances remains to be defined.

Impaired in vitro interferon, blastogenic, and natural killer cell responses to viral stimulation in acquired immune deficiency syndrome.

The in vitro immune response to herpes simplex virus (HSV), type 1, strain 539, HSV type 2, strain 316D, and cytomegalovirus was studied in 20 patients (14 with acquired immune deficiency syndrome, four with the acquired immune deficiency syndrome-related symptom complex, and two sexually active asymptomatic homosexuals) and 18 heterosexual healthy controls. Peripheral blood mononuclear cells were cultured with 2 X 10(5) plaque-forming units of heat-inactivated viruses, their lymphocyte blastogenic responses were measured after 5 days in culture by [3H]-thymidine incorporation, their interferon production was measured after 24 hr and 5 days, and natural killer (NK) cell activation was measured after 24 hr and 5 days of culture. Blastogenic responses to viruses were significantly low for only HSV, type 1:1.75 X 10(3) cpm in patients' cells compared to 6.36 for controls. Interferon responses to all three viruses were significantly low at both 24 hr and 5 days; e.g., HSV, type 1:139 IU/ml in patients' cells compared to 777 for controls at 24 hr. NK cell responses of patients were lower than those of controls when tested fresh and after 24 hr of incubation: 6.1 versus 11.7% and 9.2 versus 16.8% target cell lysis, respectively. Exposure to viruses boosted NK cell responses of both patients' and controls' cells, but boosting was generally greater among the normal rather than the patients' cells. The abnormalities of response were present in all three patient groups. Addition of interleukin-2 in vitro increased the patient and control blastogenic and NK responses but did not augment the interferon responses. The in vitro responses to both HSV, type 1, and HSV, type 2, correlated significantly with our conventional assays of the percentage and absolute level of T4+-helper lymphocytes in the blood and the blastogenic responses to mitogens, such as phytohemagglutinin, pokeweed mitogen, and concanavalin A. This system should be useful for the study of host defense in acquired immune deficiency syndrome patients and those in high-risk groups, and also for the in vitro evaluation of immunomodulators.

Immunological characterization of homosexual males.

To help clarify the nature and pathogenesis of the syndrome of severely opportunistic infection associated with immune deficiency in young homosexual males, we investigated the immunological characteristics of a group of 33 young homosexual men. These young men all had the prodrome to the syndrome which included a history of multiple sexual partners and multiple sexually transmitted diseases. In addition, they all had a past history of mild to moderate viral, bacterial, parasitic, or fungal infections and had used recreational drugs. Within this group of patients, there were five men who had Kaposi's sarcoma. Compared to the 21 normal heterosexual individuals, the homosexual men were found to be anergic to a battery of recall antigens (52% versus 19%); to be hyporesponsive to mitogen stimulation (pokeweed, 30.7 x 10(-3) versus 65.3 x 10(-3) cpm, p less than or equal to 0.005; concanavalin A, 32.2 x 10(-3) versus 60.1 x 10(-3) cpm, p less than or equal to 0.006); and to have lower helper T-cells (18% versus 34.6%, p less than or equal to 0.01), inverted helper:suppressor T-cell ratios (0.85 versus 1.92, p less than or equal to 0.01), and an elevated serum thymosin alpha 1 level (1473 versus 524 pg/ml, p less than or equal to 0.001). These data suggest that the immunological defect precedes the syndrome. The mechanism of this phenomenon is unclear; however, the repeated viral infection combined with drug usage may be responsible. The five patients with Kaposi's sarcoma were compared as a group to the other patients without cancer and found to be more severely immunodeficient. This suggests that the immune suppression by the malignant disease is superimposed on the preexisting deficiency.