Myelopreservation with the CDK4/6 inhibitor trilaciclib in patients with small-cell lung cancer receiving first-line chemotherapy: a phase Ib/randomized phase II trial.

BACKGROUND: Chemotherapy-induced damage of hematopoietic stem and progenitor cells (HSPC) causes multi-lineage myelosuppression. Trilaciclib is an intravenous CDK4/6 inhibitor in development to proactively preserve HSPC and immune system function during chemotherapy (myelopreservation). Preclinically, trilaciclib transiently maintains HSPC in G1 arrest and protects them from chemotherapy damage, leading to faster hematopoietic recovery and enhanced antitumor immunity. PATIENTS AND METHODS: This was a phase Ib (open-label, dose-finding) and phase II (randomized, double-blind placebo-controlled) study of the safety, efficacy and PK of trilaciclib in combination with etoposide/carboplatin (E/P) therapy for treatment-naive extensive-stage small-cell lung cancer patients. Patients received trilaciclib or placebo before E/P on days 1-3 of each cycle. Select end points were prespecified to assess the effect of trilaciclib on myelosuppression and antitumor efficacy. RESULTS: A total of 122 patients were enrolled, with 19 patients in part 1 and 75 patients in part 2 receiving study drug. Improvements were seen with trilaciclib in neutrophil, RBC (red blood cell) and lymphocyte measures. Safety on trilaciclib+E/P was improved with fewer ≥G3 adverse events (AEs) in trilaciclib (50%) versus placebo (83.8%), primarily due to less hematological toxicity. No trilaciclib-related ≥G3 AEs occurred. Antitumor efficacy assessment for trilaciclib versus placebo, respectively, showed: ORR (66.7% versus 56.8%, P = 0.3831); median PFS [6.2 versus 5.0 m; hazard ratio (HR) 0.71; P = 0.1695]; and OS (10.9 versus 10.6 m; HR 0.87; P = 0.6107). CONCLUSION: Trilaciclib demonstrated an improvement in the patient's tolerability of chemotherapy as shown by myelopreservation across multiple hematopoietic lineages resulting in fewer supportive care interventions and dose reductions, improved safety profile, and no detriment to antitumor efficacy. These data demonstrate strong proof-of-concept for trilaciclib's myelopreservation benefits. CLINICAL TRAIL NUMBER: NCT02499770.

Effects of decreased specific glutathione reductase activity in a chromate-tolerant mutant of Schizosaccharomyces pombe.

A chromate-tolerant mutant chr1-663T bearing a stable one-gene mutation and its parental strain 6chr(+) were used to investigate the background of Cr(VI) tolerance in the fission yeast Schizosaccharomyces pombe. The mutant chr1-663T displayed a significantly decreased specific glutathione reductase (GR) activity coded by the pgr1 (+) gene compared with its parental strain. Transformants of the mutant chr1-663T with a nonintegrative pUR18N vector expressing the pgr1 (+) gene exhibited the same Cr(VI) sensitivity and specific GR activity as their parental strain, demonstrating the importance of the GR-NADPH system in Cr(VI) tolerance. Transformants, nevertheless, exhibited an increased intracellular peroxide concentration, a decreased Cr(VI)-reducing and HO*-producing ability, which suggested an unbalanced oxidoreduction state of cells and partial complementation of the GR function. No mutation was found in the sequences of the pgr1 (+) and the pap1 (+) (transcriptional regulatory gene of GR) genes of the Cr(VI)-tolerant mutant by sequence analysis.

The dose-dependent H2O2 stress response promotes increased survival for Schizosaccharomyces pombe cells expressing HIV-1 Vpr.

Human immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) exerts multiple effects on viral and host cellular activities during infection, including induction of the cell cycle G2 arrest, and cell death in both human cells and the fission yeast Schizosaccharomyces pombe. We show that treament of exponential-phase wild-type Vpr-expressing S. pombe cells with a low, subinhibitory concentration (0.15 mmol/L) of hydrogen peroxide and 0.1 mmol/L thiamine significantly increased both cell proliferation and survival rates and decreased the number of elongated G2-arrested cells. Short-term, H2O2-induced adaptive stress increased the survival of the cells while acute stress conditions interrupted the Vpr-mediated death of the cells; however, no changes in cell length or cell phase were detected. The results suggest the importance of the oxidative status of the cells in Vpr-mediated processes. Our findings contribute to the development of a new approach via which to investigate the contribution of Vpr to HIV pathogenesis and to reduce the Vpr-mediated effects in HIV-infected patients.

Characterization of chromate-sensitive and -tolerant mutants of Schizosaccharomyces pombe.

Stable chromium(VI)-sensitive and -tolerant mutants were obtained by induced mutagenesis of Schizosaccharomyces pombe lysine and leucine auxotrophic heterothallic strains 6chr+ and 9chr+. Eleven of them were selected for further studies. Fast transport of 51CrO4(2-) was detected in a representative sensitive mutant, chr-51S, while the tolerant mutant chr1-66T and the parental strain 6chr+ exhibited significantly lower 51CrO4(2-) uptake. The segregation of tetrads of three selected CrVI-tolerant mutants, chr1-66T, chr1-14T and chr2-04T, strongly indicated that tolerance was determined by single mutations. Random spore analysis proved that the mutations of chr1-66T and chr1-14T were allelic and the mutation of mutant chr2-04T was not allelic with the mutation of chr1-66T. Recombinants carrying the ura4D18 selective marker were created for transformation experiments. Two of them (chr1-661T and chr2-046T) can be used to clone and identify the genes responsible for their CrVI tolerance phenotype.

Specificity assay of serine proteinases by reverse-phase high-performance liquid chromatography analysis of competing oligopeptide substrate library.

In this paper we present an HPLC method developed for quick activity and specificity analysis of serine proteinases. The method applies a carefully designed peptide library in which the individual components differ only at the potential cleavage site for enzymes. The library has seven members representing seven different cleavage sites and it offers substrates for both trypsin and chymotrypsin-like enzymes. The individual peptide substrates compete for the proteinase during the enzymatic reaction. The reaction is monitored by RP-HPLC separation of the components. We describe the systematic design of the competitive peptide substrate library and the test of the system with eight different serine proteinases. The specificity profiles of the investigated enzymes as determined by the new method were essentially identical to the ones reported in the literature, verifying the ability of the system to characterize substrate specificity. The tests also demonstrated that the system could detect even subtle specificity differences of two isoforms of an enzyme. In addition to recording qualitative specificity profiles, data provided by the system can be analyzed quantitatively, yielding specificity constant values. This method can be a useful tool for quick analysis of uncharacterized gene products as well as new forms of enzymes generated by protein engineering.

Amyloid beta and amylin fibrils induce increases in proinflammatory cytokine and chemokine production by THP-1 cells and murine microglia.

Activated microglia surrounding amyloid beta-containing senile plaques synthesize interleukin-1, an inflammatory cytokine that has been postulated to contribute to Alzheimer's disease pathology. Studies have demonstrated that amyloid beta treatment causes increased cytokine release in microglia and related cell cultures. The present work evaluates the specificity of this cellular response by comparing the effects of amyloid beta to that of amylin, another amyloidotic peptide. Both lipopolysaccharide-treated THP-1 monocytes and mouse microglia showed significant increases in mature interleukin-1beta release 48 h following amyloid beta or human amylin treatment, whereas nonfibrillar rat amylin had no effect on interleukin-1beta production by THP-1 cells. Lipopolysaccharide-stimulated THP-1 cells treated with amyloid beta or amylin also showed increased release of the proinflammatory cytokines tumor necrosis factor-alpha and interleukin-6, as well as the chemokines interleukin-8 and macrophage inflammatory protein-1alpha and -1beta. THP-1 cells incubated with fibrillar amyloid beta or amylin in the absence of lipopolysaccharide also showed significant increases of both interleukin-1beta and tumor necrosis factor-alpha mRNA. Furthermore, treatment of THP-1 cells with amyloid fibrils resulted in an elevated expression of the immediate-early genes c-fos and junB. These studies provide further evidence that fibrillar amyloid peptides can induce signal transduction pathways that initiate an inflammatory response that is likely to contribute to Alzheimer's disease pathology.

Proteinase inhibitors from desert locust, Schistocerca gregaria: engineering of both P(1) and P(1)' residues converts a potent chymotrypsin inhibitor to a potent trypsin inhibitor.

Two peptides, SGCI and SGTI, that inhibited chymotrypsin and trypsin, respectively, were isolated from the haemolymph of Schistocerca gregaria. Their primary structures were found to be identical with SGP-2 and SGP-1, two of a series of peptides isolated from ovaries of the same species (A. Hamdaoui et al., FEBS Lett. 422 (1998) 74-78). All these peptides are composed of 35-36 amino acid residues and contain three homologous disulfide bridges. The residues imparting specificity to SGCI and SGTI were identified as Leu-30 and Arg-29, respectively. The peptides were synthesised by solid-phase peptide synthesis, and the synthetic ones displayed the same inhibition as the natural forms: SGCI is a strong inhibitor of chymotrypsin (K(i) = 6.2 x 10(-12) M), and SGTI is a rather weak inhibitor of trypsin (K(i) = 2.1 x 10(-7) M). The replacement of P(1) then P(1)' residues of SGCI with trypsin-specific residues increased affinity towards trypsin 3600- and 1100-fold, respectively, thus SGCI was converted to a strong trypsin inhibitor (K(i) = 5.0 x 10(-12) M) that retained some inhibitory affinity towards chymotrypsin (K(i) = 3.5 x 10(-8) M). The documented role of both P(1) and P(1)' highlights the importance of S(1)'P(1)' interactions in enzyme-inhibitor complexes.

Cloning and functional expression of the cytoplasmic form of rat aminopeptidase P.

A rat cytoplasmic aminopeptidase P was purified from liver cytosol with a procedure including an affinity elution step with 3 microM inositol 1,3,4-trisphosphate. Proteolytic fragments were generated, sequenced and the enzyme was cloned from a rat liver cDNA library. The structure shows high (87.8% and 95.5%, respectively) sequence identity at the nucleotide and amino acid levels with the previously described human putative cytoplasmic aminopeptidase P. The cloned rat enzyme was functionally expressed in Escherichia coli and also in COS-1 cells. Western blot analysis, using an antibody generated against the recombinant protein, and Northern blot hybridization showed ubiquitous expression of the protein in different tissues with the highest expression level in the testis.

A restriction cleavage and transfection system for introducing foreign DNA sequences into the genome of a herpesvirus.

This report describes a simple and efficient system for construction of recombinant pseudorabies (Aujeszky's disease) virus (PrV) which is based on the use of a unique restriction site inserted into the viral genome. This system enables the recovery of genetically modified viruses without screening or selection for a specific phenotype, since practically all mature viral particles obtained carry the foreign sequences. To demonstrate, we introduced the tumour suppressor protein-53 (p53) gene into two different intergenic locations of PrV: the ribonucleotide reductase (rr) gene and the promoter of a putative latency gene (PLAT), located at the inverted repeat (IR) region of the viral genome. As a first step, we engineered a unique EcoRI recognition site into the rr gene or into both copies of PLAT with the help of marker transfer using the bacterial lacZ gene. Then, in both cases viral DNAs were cut with the restriction endonuclease EcoRI followed by treatment with calf intestinal phosphatase and used for cotransfection into porcine kidney cells with a plasmid containing the p53 gene flanked by viral DNAs homologous to the target region. As a result of this process, in most of the experiments, we obtained recombinant viruses without the background of parental viruses. Here we show that this method can be used for directional insertion of exogenous sequences into either the unique or the IR region of the PrV chromosome. In principle, this system should be applicable to the construction of recombinant derivatives of any viruses having infectious DNA.

Alcohol dependence: is carbohydrate-deficient transferrin a marker for alcohol intake?

We investigated %CDT (carbohydrate-deficient transferrin) in 92 ethanol-intoxicated alcohol-dependent patients after consecutive admission to hospital and followed the for 28 days under controlled conditions. At admission, 63% (58 patients) showed elevated CDT (> 2.5%) and 34 patients (37%) had normal CDT levels (< 2.5%). No correlation of the %CDT values to alcohol-related disabilities, severity of the withdrawal syndrome, alcohol-drinking pattern before admission, or several other factors was found. The sensitivity of GGT (gamma-glutamyl transferase) was 58% for the same group of patients. Levels of %CDT decreased during the 28 days following abstinence, whereby we could separate four statistically different groups of "CDT decrease'. In two of these groups, comprising most of the cases studied, normal %CDT levels were reached after 14 days of abstinence. Those patients with %CDT levels exceeding the upper normal level after 14 days of sobriety, showed a decrease during the following 14 days to levels of 2.55-2.61%.

Carbohydrate-deficient transferrin as a marker of alcohol intake: a study with healthy subjects.

This paper reports the results of a 3-week drinking experiment in 51 healthy male subjects, examining the value of %CDT (carbohydrate-deficient transferrin) in the context of different levels of alcohol intake. All healthy persons were urine-tested drug-free and underwent daily breath alcohol tests for the 7 days preceding, and during the whole 3 weeks of, the experiment. Subjects were divided into five groups, consuming different amounts of alcohol daily over a 3-h period in the presence of the investigators. The five groups consisted of 10, 9, 10, 16 and 6 subjects respectively and consumed a daily dose of ethanol of 20, 40, 60, 80 and 80 g respectively for 3 weeks. No significant changes in %CDT were detected in most subjects, even in the 80 g alcohol-consuming groups. The results suggest that CDT is not sensitive for the detection of short-term heavy drinking by healthy subjects.

Surfactant suppresses NF-kappa B activation in human monocytic cells.

In addition to biophysical properties, pulmonary surfactant has immunomodulatory activity. We previously demonstrated that both synthetic (Exosurf) and modified natural surfactant (Survanta) downregulated endotoxin-stimulated inflammatory c ytokine mRNA levels and protein products (tumor necrosis factor-alpha [TNF], interleukin-1-beta [IL-1], interleukin-6 [IL-6]) in human alveolar macrophages. In this study, we report that both Exosurf and Survanta suppress TNF mRNA and secretion (85 +/- 4% mean percent inhibition +/- SEM by Exosurf; 71 +/- 6% by Survanta) by endotoxin-stimulated THP-1, a human monocytic cell line. Because surfactant downregulated inflammatory cytokine production similarly in both normal human alveolar macrophages and the THP-1 cell line, we used this cell line to investigate whether surfactant affected transcriptional mechanisms. Specifically, we examined nuclear factor-kappa B (NF-kappa B) activation because it is crucial in transcriptional regulation of many inflammatory cytokine genes including TNF, IL-1, and IL-6. Electrophoretic mobility shift assays showed that both surfactants decreased activation of NF-kappa B. The presence of both p65 and p50 NF-kappa B components in LPS-activated THP-1 cells was confirmed by specific antibody induction of supershifts in mobility assays. These results are the first to suggest that surfactant's suppressive effects on inflammatory cytokine production may involve transcriptional regulation through inhibition of NF-kappa B activation.

Surfactant downregulates synthesis of DNA and inflammatory mediators in normal human lung fibroblasts.

The initial inflammatory event in the adult respiratory distress syndrome (ARDS) is followed by fibroproliferation and a cascade of fibroblast-derived mediators. Because lung fibroblasts may be exposed to surfactant as well as inflammatory cytokines during ARDS, we hypothesized that surfactant might modulate fibroblast activity. We previously demonstrated that surfactant inhibited production of inflammatory cytokines from endotoxin-stimulated human alveolar macrophages. In the current study the effects of surfactant on normal human lung fibroblast proliferative capacity and mediator production were examined. Both synthetic (Exosurf) and natural (Survanta) surfactant inhibited fibroblast [3H]thymidine incorporation. Examination of pre-S-phase events indicated stimulation of the immediate response gene, c-fos, and no effect on the G1/S cyclin, cyclin D1, suggesting that the surfactant block occurred elsewhere before S phase. The antioxidant N-acetyl-L-cysteine (NAC), like surfactant, inhibited [3H]thymidine incorporation. Furthermore, menadione, a generator of intracellular H2O2, stimulated fibroblast [3H]thymidine incorporation, and this was inhibited by surfactant. Interleukin-1 (IL-1)-stimulated secretion of the inflammatory mediators, IL-6 and prostaglandin E2, was also inhibited by surfactant. These data suggest that surfactant may modify lung fibroblast participation in ARDS sequelae by downregulating DNA synthesis and secondary inflammatory mediator production.

Regulation of human alveolar macrophage inflammatory cytokines by tyloxapol: a component of the synthetic surfactant Exosurf.

We previously demonstrated that the synthetic surfactant Exosurf and a modified natural surfactant, Survanta, both down-regulated endotoxin-stimulated production of inflammatory cytokines (tumor necrosis factor-alpha, interleukin-1 beta, interleukin-6) in human alveolar macrophages. To further characterize the source of the inhibitory effect of surfactant, the three individual components of Exosurf were evaluated. Dipalmitoylphosphatidylcholine had no effect on endotoxin-stimulated cytokine secretion. Cetyl alcohol (spreading agent) compromised macrophage function as measured by adherence. However, at concentrations equivalent to those found in the complete surfactant (Exosurf) preparation, tyloxapol (nonionic dispersing agent) was inhibitory in a dose-dependent manner. The viability of alveolar macrophages as assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide cleavage assay was not affected by incubation in Exosurf or any of its individual components. Cytokine secretion and mRNA levels of endotoxin-stimulated alveolar macrophages were decreased by tyloxapol. These data suggest that tyloxapol alone, like Exosurf, has an inhibitory effect on cytokine production which may be pretranslationally mediated.

Characterization of exosurf (surfactant)-mediated suppression of stimulated human alveolar macrophage cytokine responses.

Previous studies in our laboratory demonstrated that the synthetic surfactant Exosurf (Burroughs Wellcome Co.) inhibited endotoxin-stimulated cytokine secretion from human alveolar macrophages in vitro. The purpose of the present study was to further characterize the suppressive effects of Exosurf, which consists of dipalmitoylphosphatidylcholine (DPPC), cetyl alcohol (spreading agent), and tyloxapol (nonionic dispersing agent). Suppression was not stimulus specific in that Exosurf also significantly reduced cytokine production elicited by either Staphylococcus aureus or recombinant interleukin-1. Suppression was also mediated by a modified bovine surfactant (Survanta), which, in contrast to Exosurf, contains the surfactant-associated proteins B and C, and several different phospholipids, but no cetyl alcohol or tyloxapol. This suggests that suppression of macrophage cytokines is not specific to Exosurf. Both cell associated and secreted tumor necrosis factor and interleukin-1 were reduced by Exosurf, indicating that Exosurf is not simply blocking cytokine release. At 3 h, cytokine mRNA levels were not different between Exosurf-treated and untreated cells. However, at 8 and 24 h, cytokine mRNA levels were lower in Exosurf-treated cells. The observations that mRNA levels were decreased at 8 and 24 h and that cellular cytokine release was not blocked suggest that Exosurf's effect may in part be pretranslationally mediated. Collectively, these data add to previous work indicating that pulmonary surfactant may play a critical role in reducing inflammatory cytokine production associated with the adult respiratory distress syndrome and similar disorders.

Phase I trial of subcutaneous recombinant macrophage colony-stimulating factor: clinical and immunomodulatory effects.

PURPOSE: Recombinant human macrophage colony-stimulating factor (rM-CSF) has been demonstrated to control the growth, differentiation, and function of mononuclear phagocytes. Preclinical studies have indicated antitumor effects, and therefore a phase I trial of rM-CSF in patients with malignancy was initiated. The toxicity and hematologic and immunologic effects were investigated. PATIENTS AND METHODS: rM-CSF was administered as a subcutaneous injection on days 1 through 5 and 8 through 12. Cycles were repeated every 28 days. Cohorts of four to seven patients received rM-CSF at dose levels from 0.1 to 25.6 mg/m2/d. Forty-two patients received 88 cycles of rM-CSF. All patients had metastatic solid tumors refractory to standard therapy. RESULTS: The toxicity of rM-CSF was mild. Dose-limiting toxicity included thrombocytopenia (two patients) and iritis (one patient) occurring at a dose of 25.6 mg/m2/d. Hematologic studies demonstrated dose-related monocytosis occurring routinely at doses > or = 3.2 mg/m2/d, and thrombocytopenia. Immunologic studies demonstrated enhanced secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1-beta (IL-1 beta) by monocytes after in vitro stimulation with lipopolysaccharide, and increased expression of TNF-alpha mRNA at higher rM-CSF dose levels. Pharmacokinetic studies demonstrated that the systemic clearance rate of M-CSF increases during week 1 of therapy, resulting in lower blood levels of M-CSF during the second week of therapy. CONCLUSION: rM-CSF can be safely administered to patients, and has biologic activity on peripheral-blood monocytes.

Immunomodulatory effects of recombinant interleukin-3 treatment on human alveolar macrophages and monocytes.

The purpose of these studies was to examine the effects of in vivo and in vitro recombinant IL-3 treatment on alveolar macrophage and monocyte activities associated with antitumor and antimicrobial properties. Alveolar macrophages and blood monocytes from 6 patients receiving IL-3 (125-500 micrograms/m2/day) subcutaneously were isolated before therapy and at various times during the 15 days of therapy. Results indicated that tumor necrosis factor-alpha (TNF), interleukin-1 beta (IL-1), and interleukin-6 (IL-6) secretion were enhanced from monocytes of all patients and from alveolar macrophages of patients receiving 500 micrograms/m2/day IL-3. Constitutive cytokine gene expression was present before therapy, but further enhancement was not detectable during therapy, suggesting a rapid time course of cytokine gene transcription and translation. Serum neopterin levels were elevated 2-5 fold in all patient compatible with the presence of augmented monocyte/macrophage activity. Peak levels of neopterin did not coincide with peak levels of cytokine secretion. In vitro studies of IL-3-treated normal alveolar macrophage and monocyte population demonstrated that IL-3 significantly augmented TNF and IL-6 secretion in monocytes, but not in alveolar macrophages. These differences in alveolar macrophage cytokine secretion observed after in vivo and in vitro IL-3 treatment may reflect the involvement of other cell populations in IL-3 modulation of alveolar macrophages in vivo. Monocytes, in contrast were comparably activated by IL-3 whether presented in vitro or in vivo.

Synthetic surfactant (Exosurf) inhibits endotoxin-stimulated cytokine secretion by human alveolar macrophages.

Tumor necrosis factor-alpha (TNF), interleukin-1 beta (IL-1), interleukin-6 (IL-6), and interleukin-8 (IL-8) are inflammatory cytokines produced by alveolar macrophages (AMs) and implicated in sepsis-related adult respiratory distress syndrome (ARDS). Preliminary findings from clinical trials suggest that aerosolized delivery of the synthetic surfactant Exosurf (Burroughs Wellcome Co.) reduces mortality in patients with sepsis-induced ARDS. The purpose of the present study was to examine the effect of Exosurf on inflammatory cytokine secretion from AMs in vitro. AMs were obtained from normal nonsmoking adult volunteers. Secreted TNF, IL-1, IL-6, and IL-8 were measured by enzyme-linked immunoassays in 24 h culture fluids of AMs. Exosurf inhibited LPS-stimulated TNF, IL-1, and IL-6 secretion in a dose-dependent fashion. IL-8 secretion was not affected by Exosurf under these conditions. However, if AMs were preincubated for 24 h in media and then LPS-stimulated, IL-8 secretion was inhibited by Exosurf. Regulation of IL-8 production may differ from TNF, IL-1, and IL-6. Unstimulated cytokine secretion was not affected by any of the tested concentrations of Exosurf. The inhibitory effect of Exosurf on endotoxin-induced cytokine secretion by human AMs suggests that Exosurf may modulate inflammatory cytokine production in the lung.

Opsonin-independent phagocytosis of group B streptococci: role of complement receptor type three.

The role of complement receptor type 3 (CR3) in nonopsonic recognition of group B streptococci (GBS) by macrophages was investigated. Monoclonal anti-CR3 (anti-Mac-1) inhibited phagocytosis of GBS strains by as much as 50% in serum-free cultures of both mouse peritoneal macrophages and the macrophage cell line PU5-1.8. GBS uptake was unaffected by the presence of anti-C3 or salicylhydroxamate, an inhibitor of the covalent binding reaction of C3. Soluble antibodies to LFA-1 or to the common beta-chain (CD18) of the LFA-1/CR3/p150,95 family of cell adhesion molecules did not inhibit GBS uptake. Down-modulation of surface Mac-1 on macrophages following adherence to anti-Mac-1- or anti-CD18-coated surfaces also inhibited uptake of GBS. Further evidence for GBS interaction with CR3 was demonstrated by reduction of EC3bi rosette formation in macrophages adherent to GBS-coated plates. These studies suggest that GBS can interact with macrophage CR3, promoting phagocytosis in a C3-independent fashion. In the absence of specific immunity in neonates, this recognition mechanism may be a significant virulence determinant for GBS which poorly activate the alternate complement pathway.