IgG-recognizing shed tumor-associated antigens can promote tumor invasion and metastasis.

Tumors secreting glycoproteins that act as tumor-associated antigens have been described as highly invasive and metastatic. In this study, the consequences of the humoral immune response (HIR) against these antigens were investigated. Using an in vitro model of tumor cell invasion, results indicated that the invasiveness of tumor cells secreting antigenic secreted/shed tumor glycoproteins (STGP) increases in the presence of specific anti-STGP IgG, polymorphonuclear cells and monocytes. This in vitro model showed that the coincidental presence in the matrix of both STGP and specific anti-STGP IgG increases the local release of IL-1beta, IL-6 and vascular endothelial growth factor (VEGF) by stromal cells, but not by tumor cells. Using an in vivo model, the experiments show that immune-competent mice develop an anti-tumor HIR with anti-STGP IgG production. In this model, tumor growth was increased in parallel with the serum concentration of specific anti-STGP IgG. In athymic nude (nu/nu)-beige mice the same trend was observed, suggesting a T-cell-independent tumor-promoting effect induced by anti-STGP IgG. Tumor histology showed intense infiltration of IgG-positive plasma cells and lymphocytes. A severe combined immunodeficient-beige mouse-based in vivo model of tumors, experimentally infiltrated with monoclonal IgG plasmocytoma cells, showed that only specific anti-STGP-IgG-secreting cells could exacerbate tumor invasion, angiogenesis and metastasis. These results suggest that tumors shedding/secreting antigenic STGP can induce a host IgG immune response that can promote invasion and metastasis by inducing tumor infiltrating stromal cells to release proinflammatory cytokines and VEGF.

Tumor cells express FcgammaRI which contributes to tumor cell growth and a metastatic phenotype.

High levels of circulating immune complexes containing tumor-associated antigens are associated with a poor prognosis for individuals with cancer. The ability of B cells, previously exposed to tumor-associated antigens, to promote both in vitro and in vivo tumor growth formed the rationale to evaluate the mechanism by which immune complexes may promote tumor growth. In elucidating this mechanism, FcgammaRI expression by tumor cells was characterized by flow cytometry, polymerase chain reaction, and sequence analysis. Immune complexes containing shed tumor antigen and anti-shed tumor antigen Ab cross-linked FcgammaRI-expressing tumor cells, which resulted in an induction of tumor cell proliferation and of shed tumor antigen production. Use of selective tyrosine kinase inhibitors demonstrated that tumor cell proliferation induced by immune complex cross-linking of FcgammaRI is dependent on the tyrosine kinase signal transduction pathway. A selective inhibitor of phosphatidylinositol-3 kinase also inhibited this induction of tumor cell proliferation. These findings support a role for immune complexes and FcgammaRI expression by tumor cells in augmentation of tumor growth and a metastatic phenotype.

Direct in vivo transfection of antisense Fas-ligand reduces tumor growth and invasion.

The expression of Fas ligand (FasL) by tumor cells has been reported to have multiple, conflicting effects on tumor growth. The majority of the data support the theory that FasL expressing tumor cells evade immune surveillance by killing T cells expressing Fas. However, the role of the humoral immune-blockade by FasL expressing tumor cells has not been assessed. Using immune-competent mice, we observed that FasL expressing tumor cells reduced the antitumor antibody production together with the T and B cell content of the spleen in these mice. Further, to determine if the expression of FasL in the environment of the tumor suppresses the humoral antitumor immune response and influences tumor growth, a mouse model lacking T cells was used. To assess whether a local reduction of FasL could reduce tumor progression, a plasmid encoding antisense FasL cDNA was delivered directly into a growing tumor (SW620 colon carcinoma). Intratumoral delivery of the plasmid was able to transfect tumor cells, stromal cells, and peritumoral muscle cells. This antisense FasL tumor tissue transfection persisted for at least 25 days, produced a systemic decrease in soluble FasL, and resulted in a 50% reduction in the rate of tumor growth when compared with tumor tissue of the control groups. These results suggest that direct transfection of antisense FasL cDNA impairs FasL translation in tumor and stromal cells, and can inhibit tumor progression by impairing the FasL-mediated, stromal cell-assisted, tumor counter-attack.

B lymphocyte pathology in human colorectal cancer. Experimental and clinical therapeutic effects of partial B cell depletion.

Accumulating data are showing that the humoral immune response against tumors could favor tumor progression. However, no B lymphocyte pathology has been reported in cancer. Using anti-IgM Ab we nonspecifically depleted B cells in tumor-bearing mice, a treatment that resulted in significant reduction of tumor burden. We analyzed the B lymphocyte phenotype of abdominal lymph nodes and peripheral blood from advanced colon cancer patients by flow cytometry, and compared the B cell phenotype with that found in samples from normal donors. In both lymph nodes and peripheral blood of cancer patients, abnormal populations of B lymphocytes appeared that express an increased CD21 and/or sTn antigens on their cell surface. All patients showed a reduction of CD19+ cells. In a limited clinical test, we analyzed the effects of a partial B cell depletion with Rituximab. The treated patients did not develop any side-effects; the CD21-hyperpositive lymphocytes were reduced, but the proportion of sTn-positive lymphocytes remained unaffected. Apparent reduction of the tumor burden was reported in 50% of the patients when the treatment was ended.

Promotion of tumor invasion by cooperation of granulocytes and macrophages activated by anti-tumor antibodies.

We investigated the potential role of anti-tumor antibodies and tumor antigens in the formation of immune complexes which promote matrix degradation and angiogenesis. B-cell deficient or B-cell depleted mice showed a reduction in tumor invasion and metastasis. In vitro invasion assays and in vivo models of metastasis showed that anti-sTn antibodies and sTn tumor antigens form complexes which induce granulocytes and macrophages together to mediate tumor invasion and metastasis by processes including extracellular matrix degradation and angiogenesis. These results suggest the existence of a tumor promoting role of a B-cell immune response induced by shed tumor associated antigens of solid, nonlymphoid tumors.

Staging of colorectal cancer: biology vs. morphology.

PURPOSE: An accurate determination of the extent or staging of a disease is critical, because it provides the basis for making therapeutic decisions. Staging is a collaborative effort by the surgeon and the pathologist. Radioimmunoguided surgery has been evaluated for its ability to help surgeons determine the extent of disease during surgery, when management decisions have the most impact on patient care. This study was done to compare radioimmunoguided surgery "biostaging" with traditional pathologic staging (TNM) as predictors of survival in patients undergoing curative resections for colorectal cancer. METHODS: Ninety-seven patients with colorectal cancer were prospectively enrolled in radioimmunoguided surgery protocols. Evaluation of follow-up survival data was performed. All patients underwent exploratory laparotomy and radioimmunoguided surgery with resection of their primary colorectal tumor. Survival data were analyzed with the Kaplan-Meier method with log-rank comparisons. RESULTS: Of 97 patients enrolled in the study, 59 were evaluable and completely resectable by radioimmunoguided surgery. Mean follow-up was 62 months, with a range of 34 to 89 months. By traditional staging 13 patients were pStage I, 18 patients were pStage II, and 28 patients were pStage III. By radioimmunoguided surgery biostaging, 24 patients were radioimmunoguided surgery-negative whereas 35 patients were radioimmunoguided surgery-positive. Survival rates by pathologic stage approached a significant difference, but did not, as of the conclusion of the study period, reach it (P = 0.12). Survival rates based on radioimmunoguided surgery status demonstrated a highly significant difference (P = 0.0002). CONCLUSIONS: Radioimmunoguided surgery biostaging provides new information intraoperatively on cancer staging that has not been available before. This may lead to new strategies for therapy that can be individualized and optimized for each patient with cancer.

First results for resetting the antitumor immune response by immune corrective surgery in colon cancer.

BACKGROUND: A critical step for cancer recurrence is the failure of the cellular immune response. It is suspected that chronic humoral immune responses against some tumor-associated antigens (TAA) can contribute to that failure. METHODS: In this study, we tested the ability of an immune corrective surgical procedure to prevent recurrences of colon cancer in stages I, II, and III. Radiolabeled anti-TAG antibodies injected intravenously become concentrated on TAG-72 immune complexes presented by follicular dendritic cells, which are responsible for the persistent humoral response against TAG-72 TAA. Using a hand-held gamma probe, we can intraoperatively detect and remove lymph nodes involved in TAG-72 presentation. By removing these lymph nodes, together with the tumor tissue, presentation and source of TAG-72 are drastically reduced. RESULTS: The impact of this TAA suppression on the tumor recurrence process is analyzed in a sample of 24 patients. The immune corrective surgical procedure did not increase morbidity. Five years after surgery the following were disease free: 5 of 5 stage I, 6 of 6 stage II, and 10 of 13 stage III. The global survival of this group was 87.5%. Compared with the standard surgical treatment of colon cancer (58% survival for the same stages), this surgical immune corrective procedure introduces a statistically significant improvement of 29% (P <0.001). CONCLUSIONS: The surgical removal of lymph nodes involved in the persistent humoral immune response against TAA has an important beneficial impact on colon cancer treatment.

[Lymphadenectomy in colorectal carcinoma]. / Lymphadenektomie beim colorectalen Carcinom.

Recurrence of colorectal carcinomas occurs in about 50% of the cases with localized neoplasia. It is understood that the tumor recurrence is due to residual micrometastases not found during surgery or extraregional (peripheral blood or bone marrow). We developed a procedure to detect non-visible, abdominal metastases using a radiolabeled anti-tumor cell antibody injected before the operation (radioimmunoguided surgery RIGS). However, even with the best technique, it is not possible to remove all micrometastasis if a hematogenic dissemination happens. Based on the knowledge of disturbing humoral immune reaction is mounted against shed tumor associated antigens (TAA), we developed a new method to reduce and correct the B cell response and B cell recruitment due to chronic TAA immun complex presentation on follicular dendritic cells (immune corrective surgery, ICS). This method is based on a selective lymphadenectomy. The target lymph nodes were those loaded with TAA-immune complex. The detection method used was the injection of radiolabeled antibody able to recognize the immune complex. From 20 patients (stage I, II and III) treated with ICS, 17 survived more than 5 years 'showing a statistically significant increase of survival compared to patients treated with standard procedures. In conclusion, these data show that surgery of colorectal cancer should be selectively extended to specific anatomical regions in order to remove hidden micrometastases, and more importantly, correct postoperative immune processes that could suppress the T cell response against residual tumor cells.

Extramedullary hematopoiesis in the adult mouse liver is associated with specific hepatic sinusoidal endothelial cells.

In previous work, two anatomically distinct-liver sinusoid endothelial cells (LEC): LEC-1 and LEC-2, have been described. We also reported that extramedullary hepatic hematopoiesis occurs only in close contact with LEC-1, suggesting that these cells may provide the microenvironment necessary for the maintenance and growth of hematopoietic cells. In the present work, we studied the capacity of LEC-1 and LEC-2 to maintain in vitro hematopoiesis. LEC-1 and LEC-2 were isolated and cloned from livers of adult mice. Bone marrow cells (BM) enriched with primitive hematopoietic progenitors were isolated from day-2, post-5-FU-treated mice (5-FUBMC). LEC-1 supported the maintenance and differentiation of hematopoietic progenitors for more than 6 weeks in vitro. In contrast, LEC-2 cells poorly supported the proliferation of hematopoietic cells for only two weeks of the co-culture. LEC-1 and 5-FUBMC cocultures showed cobblestone-area formation and the presence of hematopoietic progenitors that are able to form colonies (CFC) in the adhering fraction after six weeks of coculture. LEC-1 co-cultures treated with a cocktail of cytokines (stem cell factor, interleukin [IL]1alpha, IL-3, and Epo) showed that megakaryocyte (CFU-Mk) and erythrocyte progenitors (BFU-e) were present during the entire period of the culture. Granulocyte-macrophage progenitors (CFU-GM) were present only during the first three weeks of the culture. These results suggest that LEC-1, but not LEC-2, provide an appropriate hematopoietic microenvironment for supporting the proliferation and differentiation of primitive hematopoietic cells. This could explain the anatomical restriction of hematopoietic cells for growing in LEC-1 domains during liver extramedullary hematopoiesis.

Radioimmunoguided Surgery for Gastrointestinal Malignancies: An Analysis of 14 Years of Clinical Experience.

BACKGROUND: The identification of all sites of intra-abdominal adenocarcinoma is key to optimal surgical resection and tumor staging. Conventional imaging methods and direct visualization and palpation have limited sensitivity and specificity. Radioimmunoguided surgery (RIGS) has a potential to improve these parameters. METHODS: The development of the RIGS concept is presented, and the studies of tumor localization, detection of disseminated disease, staging, and survival are correlated with the tumor biopsy of gastrointestinal carcinoma, particularly colorectal carcinoma. RESULTS: RIGS can detect clinically and histologically occult neoplasm. Also, by providing immediate intraoperative information, the RIGS approach improves surgical staging, impacts on surgical and medical care, and affects patient prognosis. CONCLUSIONS: RIGS may become the standard of care for the surgical staging and treatment of colorectal cancer and other gastrointestinal malignancies.

Tumor-host interaction in non-random metastatic pattern distribution.

In the clinical evolution of malign tumors, prognosis depends on whether metastasis develops or not. Biologically speaking, the formation of metastasis implies the existence of tumor cells capable of successfully performing all the steps in the metastatic process: local invasion, lymphatic or hematogenous dissemination, arrest in the microvascular bed of an organ, extravasation and growth of a secondary colony. Clinical observations have demonstrated that for each primary tumor there is a colonization pattern determined by the characteristics of the microvascular endothelium and the functional environment of the target organ. Moreover, the formation of metastasis depends on at least two additional factors: a) tumor cell-tumor cell and tumor cell-host cell relations modulated by intercellular contact and/or soluble paracrine or autocrine growth factors; b) the antitumor efficiency of the immune system, mediated primarily by the action of NK/LAK cells, macrophages and cytolytic T-lymphocytes, whose activity is in turn regulated by a complex of cytokines, including interferons, tumor necrosis factors and interleukins. In this work, we first review certain aspects of tumor biology that are specifically involved in tumor cell-host cell interactions determining non-random metastatic pattern distribution, and then review the implication of certain cytokines in the regulation of tumor proliferation.

Automated imaging and quantitation of tumor cells and CFU-GM colonies in microcapillary cultures: toward therapeutic index-based drug screening.

Human colony forming units (CFUs) from both malignant and hematopoietic tissues can be assayed in vitro in microcapillary cultures, an alternative cloning system to the Petri dish methodology. For technical reasons, microcapillary culture may be ideally suited for new drug screening by therapeutic index. To achieve the high output required by screening programs, automated quantitation of CFUs is required. Toward this end, this paper reports the development of a prototype CapScan, an image analysis system that uses a novel axial laser illumination system to detect tumor cell colonies and, with technical modifications, CFU-granulocyte-macrophage (CFU-GM) colonies in microcapillary cultures. As currently configured, the CapScan can quantify colonies grown in a rack of 18 microcapillary cultures in 30 minutes or less. The sensitivity and detection specificity of tumor cell colonies is >90% with a coefficient of variance of 5-40%, dependent upon colony number. Over a range of colony numbers, CapScan and manual colony counts showed a linear correlation > -0.9, and yielded identical results in assays of doxorubicin inhibition of clonogenic P388 cells. As an additional advantage, the growth kinetics of individual colonies can also be monitored with the CapScan, making distinctions between cytotoxic and cytostatic drugs possible; colonies of freshly isolated human tumor cells can also be quantified. Thus, a microcapillary-based human tumor cloning assay that tests for resistance and/or sensitivity to chemotherapeutic agents may be useful in drug development programs and may also facilitate the development of chemotherapy for individual patient tumors, especially when tumor availability is limited.

The addition of interleukin-2 to cyclophosphamide therapy can facilitate tumor growth of B16 melanoma.

The role of interleukin-2 (IL-2) on tumor growth of B16F10 melanoma cells was assessed in two sets of mice with different immune status: normal (immunocompetent) mice and immunodeficient mice. The two sets of animals were treated with cyclophosphamide (CY) or IL-2 alone or with a combined therapy of CY+IL-2. On days 6 and 10 after tumor cell injection, we evaluated the incidence of hepatic B16 melanoma metastases and the percentage of hepatic volume occupied by metastatic tissue. We observed that the CY alone (300 mg/kg, days 3 and 8 post-tumoral inoculation) significantly reduced tumor growth in all treated mice; however, CY proved more effective in normal recipients than in immunodeficient hosts. On the other hand, whereas administration of IL-2 alone (10(5) IU daily, from day 3 to day 7) in immunocompetent mice significantly reduced tumor growth on days 6 and 10, in immunodeficient mice, no significant differences were observed in tumor growth either on the 6th or on the 10th day, in comparison to control groups. Finally, when the combined CY+IL-2 therapy was administered, an antisynergistic effect between these therapeutic agents was achieved both in normal and in immunodeficient mice. Thus, the addition of low-dose IL-2 (25 x 10(3) IU daily, from day 4 to day 7) to high-dose CY (300 mg/kg, days 3 and 8) significantly increased tumor growth in both the early and later periods, compared to the effect of CY alone. It is concluded that exogenous IL-2 can facilitate tumor growth of B16 melanoma cells in vivo.

A simple cell labeling technique by means of lectins linked to fluorochromes for the detection of cells on tissue sections.

We designed a protocol for cell labeling with the lectin wheat germ agglutinin (WGA) linked to the fluorochrome tetramethyl-rhodamine isothiocyanate (TRITC) for effective detection of the B16F10 melanoma and Lewis lung carcinoma (LLc) cells on pulmonary histological sections from C57BL/6 mice. We have also determined a suitable concentration of WGA-TRITC (10 micrograms/ml), which leads to a very intense and homogeneous labeling of the cells, as it avoids cell clumping due to the presence of the lectin WGA. In order to determine to what extent the method affects these tumor cells, we have studied some important aspects related to their metastatic behavior, taking into account three parameters: a) viability and rate of proliferation of the cells cultured in vitro; b) percentage of animals (C57BL/6 mice) bearing metastasis 15 days after intravenous inoculation with 10(5) B16F10 or LLc cells; and c) pattern of distribution of tumor foci in lung. There were no significant differences in these three parameters between the WGA-TRITC labeled-cells compared to the cultures of non-labeled cells in either of the cell lines (B16F10, LLc). Thus, we conclude that B16F10 and LLc tumor cells can be labeled following the protocol set-up in our study, as it allows these cells to be neatly identified on tissue sections and it causes no important physiological changes in the cells, with regard to metastatic behavior. These points make this technique very suitable for the detection of B16F10 and LLc cells on histological sections in studying their behavior during the first stages of the metastatic process.

Tissue distribution and therapeutic effect of intravenous free or encapsulated liposomal doxorubicin on human prostate carcinoma xenografts.

BACKGROUND: The authors compared the therapeutic effects of doxorubicin in two formulations: free in saline suspension and encapsulated in sterically stabilized liposomes composed of hydrogenated soy phosphatidylcholine/2cholesterol/polyethylene glycol-distearoyl-phosphatidyl-ethanolamine (Doxil, Liposome Technology, Inc., Menlo Park, CA). METHOD: The drug formulations were injected intravenously to treat human prostate carcinoma PC-3, implanted subcutaneously into nude Swiss mice. Confocal laser scan microscopy and microfluorometry were used to determine tissue distribution and to quantitate drug uptake. RESULTS: Laser scan microscope and microfluorometer studies showed that the liposome-encapsulated drug entered the liver, the kidneys, and the tumor in greater quantity and remained in the liver and in the tumor longer than the free drug. The liposome formulation produced a 25-fold increase in doxorubicin at the disease site. Doxil was significantly more effective than the free drug in inhibiting growth and in effecting cures and had only minor and temporary systemic toxic effects. CONCLUSIONS: The current study demonstrated the therapeutic efficacy of doxorubicin, encapsulated in sterically stabilized liposomes, against prostate carcinoma. Decreased systemic elimination, increased penetration into the tumor, and long liposome presence with slow drug release into the tumor probably accounted for the enhanced therapeutic effect of doxorubicin in sterically stabilized liposomes.

Enhancement of Ia antigen expression and nonproliferating cells correlates with metastatic capacity.

Maintaining B16F10 tumor cells in stirring culture for 48 h leads to an increase in lung and liver colonizing capacity in comparison with cells in adherent culture. Parallel to the increased metastatic capacity, we have observed a decrease in the proliferative rate of tumor cells (as the percentage of proliferating cell nuclear antigen-positive cells) and an increase in the population of tumor cells expressing Ia antigen. These results are not exclusive to B16F10 cells, since the same results were obtained when we analyzed 3LL cells maintained in identical culture conditions. In all the tumor lines tested, we found an association between the nonproliferating and the Ia-positive cell populations. We induced Ia expression by treating B16F10 cells in adherent culture with the lectin concanavalin A and again, coincident with an increase in metastatic capacity, we found the same association between the two parameters analyzed--nonproliferating state and Ia antigen expression. In addition, it was found that B16F10 cells induce lymphocytic proliferation, and a direct relationship was established between the number of Ia+ cells and lymphocytic proliferation.

Isolation and enrichment of two sublobular compartment-specific endothelial cell subpopulations from liver sinusoids.

Similar to the well-recognized phenotypical heterogeneity of hepatocytes, in situ sublobular variations have recently been detected in the cell structure, fenestration patterns, filtrating efficiency, surface glycosylation, scavenger function and pathological responses of the sinusoidal lining endothelium. However, unlike other liver cell populations, until now no endothelial cell subpopulations had been isolated or defined with clarity, much less with sublobular/acinar zone-related differential properties. On the basis of our previous studies showing that periportal segments of mouse liver sinusoids express a significantly higher number of wheat germ agglutinin-binding sites than do perivenous ones, we used this differential feature for in vitro labeling of the specific sublobular derivation of isolated sinusoidal lining endothelial cells to correlate their original lobular position with other features determined on flow cytometry, centrifugal elutriation, discontinuous arabinogalactan density gradients and electron microscopy. Our results revealed additional heterogeneous properties whose association with high or low wheat germ agglutinin-binding capacity made it possible to define in vitro two dominant endothelial cell subpopulations that appear similar to the differential features in the periportal and perivenous sinusoidal segments. Type 1 endothelial cells had low forward angle light scatter and high integrated side scatter, low cytoplasmic porosity index (12% +/- 5%) and high wheat germ agglutinin-binding efficiency (160 +/- 35 fluorescence intensity units/cell size); these findings are similar to what was observed in situ in the periportal sinusoidal endothelium. On the other hand, type 2 endothelial cells, with high forward angle light scatter and low integrated side scatter, had a high cytoplasmic porosity index (25% +/- 8%) and low wheat germ agglutinin-binding efficiency (60 +/- 15 fluorescence intensity units/cell size), findings similar to in situ observations of the perivenous sinusoidal lining endothelium. Moreover, these physical and morphological differences entail different cell sedimentation behaviors: type 1 endothelial cell sedimented at high centrifugal elutriation counterflow rates (23 to 37 ml/min) and high arabinogalactan density gradient levels (10% to 15%), whereas type 2 endothelial cell sedimented at low counterflow rates (18 to 23 ml/min) and low density levels (6% to 10%). The combination of these separation procedures made it possible to isolate a 90%-enriched type 1 endothelial cell population in the 12% to 15% interphase of the 23 and 37 ml/min elutriation flow rates and a 75%-enriched type 2 endothelial cell population in the 6% to 10% interphase of the 18 and 23 ml/min flow rates.

Cancer-cell traffic in the liver. III Lethal deformation of B16 melanoma cells in liver sinusoids.

It has previously been shown that most of the B16F10 melanoma cells delivered to the mouse liver via the portal vein are rapidly killed in periportal zone 1 of the sinusoids. Few intact viable cells reach pericentral zone 3 of the sinusoids and of these, only a very small proportion leave the liver to colonize the lungs. The hypothesis has been advanced that one non-exclusive, potential mechanism for cancer-cell destruction in the liver is a result of the deformation of cancer cells when they enter vessels of smaller diameter than themselves. Where entry is associated with change in shape from spheroid to cylindrical, a mandatory increase in cancer-cell surface area occurs, which is first apparent and utilizes surface membrane excess (rugosity). If this increase is insufficient, a real increase occurs which, if in excess of approximately 4%, results in lethal surface membrane rupture. This hypothesis predicts that, under these circumstances, cancer-cell resistance to mechanical trauma is favored by small diameter and utilizable surface membrane excess. To test this hypothesis, the traffic of melanoma cells in the liver following portal-vein injection has been observed by confocal microscopy and image reconstruction. In accordance with the hypothesis, non-disrupted cells within the sinusoids have a smaller mean diameter than that of the original inoculum, and show evidence of utilization of surface membrane excess. The results indicate that deformation-associated trauma, suffered by cancer cells on entry and residence in the microvasculature, may well be an important factor contributing to metastatic inefficiency.

The differential resistance of B16 wild-type and F10 cells to mechanical trauma in vitro.

Following intravenous injection of B16 melanoma cells into mice, more than 99.9% of the cells are killed by a combination of rapid and slow processes: however, the F10 line of B16 mouse melanoma cells produces approximately 10 times as many pulmonary colonies as wild-type cells. We have attempted to determine the role of one rapid cancer cell-killing process, namely deformation-driven, loss of surface membrane integrity of the type occurring in capillaries, by the use of an in vitro model in which cells are filtered through 8-microns pores in polycarbonate membranes. In accord with in vivo observations, more wild-type than F10 cells were killed by filtration in vitro. The hypothesis that resistance to mechanical trauma of this type is enhanced by a small cell diameter and a high degree of surface rugosity is supported by measurements of these parameters on viable cells and electron micrographs. Differential resistance in these cells is associated to a major extent with a high degree of utilizable surface membrane excess, and to a minor extent with the smaller mean diameter of the F10 cells. Calculations, which are in accord with previous in vivo observations, indicate that most of the cells delivered to the capillary beds of target organs during hematogenous metastasis can be destroyed by rapid mechanical trauma, which is therefore implicated as one of a number of major contributors to metastatic inefficiency.