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Biomedical subjects

David N Krag

Publications and source records attributed to David N Krag.

At least 19 recordsLinked to original sources
PubMed2006

Detection of occult sentinel lymph node micrometastases by immunohistochemistry in breast cancer. An NSABP protocol B-32 quality assurance study.

BACKGROUND: Occult metastases, by definition, are not detected on initial examination. They may be present on slides but missed during screening or may be present in paraffin embedded tissue blocks and undetected without additional levels. Anticytokeratin immunohistochemistry (CK IHC) enhances detection of occult metastases, particularly micrometastases (> 0.2 mm but not larger than 2.0 mm) or isolated tumor cell clusters (< or = 0.2 mm). This study defines the rate at which pathologists miss metastases on CK IHC of sentinel lymph nodes (SLN). METHODS: CK IHC sections 0.5 and 1.0 mm from the original surface of SLN tissue blocks were screened by pathologists using standard bright field light microscopes (LM) and by supervised computer assisted cell detection (CACD). All blocks were from breast cancer patients, initially classified 'node negative' on review of routinely stained sections from the surface of each block. Cases missed by LM screening but detected by CACD defined false negative screens. RESULTS: Of 236 cases screened, LM detected 34 (14.4%; 95% CI: 9.6-20.2) cases and, in the 202 cases negative by LM, CACD detected an additional 30 (14.9%; 95% CI: 9.6-21.2%) cases with occult metastases. Occult metastases missed by LM screening ranged from 0.01 to 0.1 mm in greatest dimension. The probability of missing an occult metastasis < or = 0.02 mm; < or = 0.05 mm, and < or = 0.10 mm was 75%, 69.2%, and 61.2%, respectively. No occult metastases larger than 0.10 mm were missed by LM screening. CONCLUSIONS: Pathologists screening the CK IHC stained slides may frequently miss detecting metastases < 0.10 mm.

Axilla↗
PubMed2006

Selection of tumor-binding ligands in cancer patients with phage display libraries.

Phage display has been used extensively in vitro and in animal models to generate ligands and to identify cancer-relevant targets. We report here the use of phage-display libraries in cancer patients to identify tumor-targeting ligands. Eight patients with stage IV cancer, including breast, melanoma, and pancreas, had phage-displayed random peptide or scFv library (1.6 x 10(8)-1 x 10(11) transducing units/kg) administered i.v.; tumors were excised after 30 minutes; and tumor-homing phage were recovered. In three patients, repeat panning was possible using phage recovered and amplified from that same patient's tumor. No serious side effects, including allergic reactions, were observed with up to three infusions. Patients developed antiphage antibodies that reached a submaximal level within the 10-day protocol window for serial phage administration. Tumor phage were recoverable from all the patients. Using a filter-based ELISA, several clones from a subset of the patients were identified that bound to a tumor from the same patient in which clones were recovered. The clone-binding to tumor was confirmed by immunostaining, bioassay, and real-time PCR-based methods. Binding studies with noncancer and cancer cell lines of the same histology showed specificity of the tumor-binding clones. Analysis of insert sequences of tumor-homing peptide clones showed several motifs, indicating nonrandom accumulation of clones in human tumors. This is the first reported series of cancer patients to receive phage library for serial panning of tumor targeting ligands. The lack of toxicity and the ability to recover clones with favorable characteristics are a first step for further research with this technology in cancer patients.

Amino Acid Sequence↗
PubMed2006

Specific peptide ligand for Grb7 signal transduction protein and pancreatic cancer metastasis.

BACKGROUND: Pancreatic cancer is one of the most aggressive malignancies, with high rates of invasion and metastasis and with generally poor prognosis. We previously found that metastasis was strongly associated with the expression of growth factor receptor-bound protein 7 (Grb7), which contains a Src homology 2 (SH2) domain. In this study, we evaluated Grb7 protein as a molecular target of therapy for metastatic pancreatic cancer. METHODS: Grb7 protein expression was measured by immunohistochemistry in 36 human pancreatic cancer specimens and adjacent normal pancreatic tissue. We synthesized a nonphosphorylated peptide inhibitor that binds specifically to the SH2 domain of Grb7. Intracellular signaling was assessed by immunoprecipitation and immunoblot assays in cultured human pancreatic cancer cells. Cell migration was measured with a modified Boyden chamber method. Peritoneal metastasis of the pancreatic cancer cells was measured with a mouse model. All statistical tests were two-sided. RESULTS: We found that 22 (61%) of 36 pancreatic cancer specimens had higher levels of Grb7 protein than their corresponding normal pancreatic tissue specimens. Grb7 expression was statistically significantly different between specimens from patients without lymph node metastasis (stage N0; two of the 10 patients) and patients with lymph node metastasis (stages N1 + N2; 20 of the 26 patients) (P = .006). The Grb7 peptide inhibitor selectively blocked the interaction between Grb7 and focal adhesion kinase and blocked the phosphorylation of Grb7 protein. In vivo Grb7 peptide inhibitor statistically significantly attenuated cell migration (for control peptide, 87.5 cells migrated, 95% confidence interval [CI] = 82.6 to 92.4 cells; for Grb7 peptide, 5.7 cells migrated, 95% CI = 2.3 to 9.0 cells; P < .001) and peritoneal metastasis of the pancreatic cancer cells in a mouse model, as assessed by the number of nodules (control = 72.6 nodules, 95% CI = 55.8 to 89.4 nodules; and for Grb7 peptide = 3.2 nodules, 95% CI = 1.6 to 4.8 nodules; P < .001, t test) and their weight (control = 4.13 g, 95% CI = 3.40 to 4.86 g; Grb7 peptide = 0.19 g, 95% CI = 0.06 to 0.32 g; P < .001, t test). CONCLUSIONS: The Grb7 peptide inhibitor appears to be a promising molecularly targeted therapeutic agent against metastatic pancreatic cancer.

Adenocarcinoma↗
PubMed2006

Selective delivery of therapeutic agents for the diagnosis and treatment of cancer.

Research activity aimed towards achieving specific and targeted delivery of cancer therapeutics has expanded tremendously in the last decade, resulting in new ways of directing drugs to tumours, as well as new types of drugs. The available strategies exploit differences in the nature of normal and cancer cells and their microenvironment. The discovery and validation of cancer-associated markers, as well as corresponding ligands, is pivotal for developing selective delivery technology for cancer. Although most current clinical trials are either monoclonal antibody- or gene-based, methodological advances in combinatorial libraries of peptides, single chain variable fragments and small organic molecules are expected to change this scenario in the near future. Nanotechnology platforms today allow systematic and modular combinations of therapeutic agents and tumour-binding moieties that may generate novel, personalised agents for selective delivery in cancer. This paper discusses recent developments and future prospects of targeted delivery technologies in the management of cancer.

Animals↗
PubMed2005

The detection of isolated tumor cells in bone marrow comparing bright-field immunocytochemistry and multicolor immunofluorescence.

BACKGROUND: The detection of isolated tumor cells in bone marrow by immunocytochemistry (ICC) has been reported to predict progression of early-stage breast cancer. The most common staining procedure uses bright-field ICC with cytokeratin (CK) antibodies to label isolated tumor cells. However, this method can result in false-positive staining events. We used multicolor immunofluorescence (IF) to develop a more specific assay for detecting isolated tumor cells in marrow samples from breast cancer patients. METHODS: We compared ICC and IF side by side for detection of cancer cells and false-positive staining events on bone marrow aspirates from breast cancer patients, bone marrow from healthy donors, and healthy donor blood spiked with cancer cells. The primary target for isolated tumor cell detection was CK for both methods. IF used an additional set of antibodies to label hematopoietic cells (HCs). RESULTS: The detection rate of CK+ events in breast cancer patient bone marrow aspirates was 18 (58%) of 31 for ICC and 21 (68%) of 31 for IF. However, with IF, 17 of 21 CK+ cases were stained with HC markers and thus were identified as false-positive events. A surprisingly high CK+ event rate was observed in healthy donor blood and marrow. In all healthy donor samples, CK+ events were readily identified as HCs by IF. Detection sensitivity of spiked cancer cells in donor blood was similar for both methods. CONCLUSIONS: There is a high frequency of CK+ events in blood and marrow, and it is important to note that this is observed both in patients with and those without cancer. IF with multiple HC markers allows straightforward discrimination between CK+ cells of hematopoietic and nonhematopoietic origin.

Bone Marrow Examination↗
PubMed2005

Phage display selection for cell-specific ligands: development of a screening procedure suitable for small tumor specimens.

Phage display technology has been widely used for developing tumor-targeting agents. Most of the efforts were directed towards identifying phage-displayed ligands against cancer-relevant purified targets and cancer cell lines. Whole cell screening procedures typically use a relatively large sample size and are not ideally suited for complex tumor tissues. We describe here a screening protocol that is suitable for non-adherent tumor cells from biopsy specimens. It requires only approximately 20,000 cells/round for biopanning and approximately 10,000 cells/well for subsequent clone binding assessment by ELISA. We standardized the newly developed protocol using erbB2-overexpressing SKBR3 breast cancer cells and compared the results with conventional protocols employing about 10-times more plate-adhered fixed or live cells. The selection rate of SKBR3-binding clones from biopanning approximately 20,000 non-adherent SKBR3 cells by our filter cup protocol was comparable to that obtained from using approximately 200,000 plate-adhered cells. Assessment of clones selected from different phage libraries showed that clones from fixed or live cells, adherent or non-adherent cells, either biopanned in filter cup or plate share specific motifs and binding properties. Some of the clones from each biopanning protocol bound to purified erbB2 and shared motifs with erbB family of receptors and their known ligands. These results demonstrated that the protocol developed in this study was capable of selecting cell-specific ligands using relatively small numbers of cells. Screening cells from a fresh human breast cancer specimen using our protocol showed enrichment of tumor binding clones at successive rounds of selection and some of the selected clones were tumor-specific in comparison to normal breast cells. These protocols have direct application to screen for tumor-binding ligands with small tumor tissue specimens.

Amino Acid Sequence↗
PubMed2005

A sensitive and rapid chemiluminescence ELISA for filamentous bacteriophages.

Filamentous bacteriophage (Ff) displayed random peptide and antibody libraries are widely used to identify specific, high affinity, binding ligands. A critical element in the identification of target-specific phages is to determine phage titers, not only at every round of selection, but also for normalizing phage titers of a set of individual clones for their comparative binding analysis. The conventional ELISA-based Ff titration methods require a minimum of 4-5 hr assay time and their lowest reported detection limit is approximately 50,000 particles/well. In this report, we present a sandwich ELISA that allows detection of approximately 1000 Ff particles/well in less than 2.5 hr. The values of correlation of coefficient (r2) for the curves at low phage concentrations (up to 106 TU/well) were about 0.999 in our ELISA. Experiments conducted at different temperatures suggest using 40 degrees C incubations when titering low phage concentration samples. Experiments were also conducted with conventional ELISA for comparison. Our ELISA method derives an advantage from using a chemiluminescence substrate that gives much larger signals and wide linear range of measurement, thus allowing discrimination between background and low Ff phage concentrations. In conclusion, the Ff titration method presented here is highly sensitive, rapid, and amenable to high throughput analysis.

Antibody Affinity↗
PubMed2005

Prerandomization Surgical Training for the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-32 trial: a randomized phase III clinical trial to compare sentinel node resection to conventional axillary dissection in clinically node-negative breast cancer.

OBJECTIVE: To train surgeons in a standardized technique of sentinel lymph node biopsy and to prepare them for the requirements of a prospective randomized surgical trial. SUMMARY BACKGROUND DATA: The NSABP B32 trial opened to accrual in May 1999. A significant component of this trial was a prerandomization training phase of surgeons performed by a group of core surgical trainers. The goals of this training phase were to expeditiously instruct surgeons in a standardized technique of sentinel lymph node biopsy and to educate those same surgeons in complete and accurate data collection and source documentation for the trial. METHODS: This study is a description of the training data collected in a prospective fashion for the training component for surgeon entry into the B32 trial, evaluating the effectiveness of the training program in regards to surgical outcomes and protocol compliance. RESULTS: Two hundred twenty-six registered surgeons underwent site visit training by a core surgical trainer and 187 completed training and were approved to randomize patients on the trial. The results of 815 training (nontrial) cases demonstrated a technical success rate for identifying sentinel nodes at 96.2% with a false negative rate of 6.7%. A protocol compliance analysis, which included the evaluation of 94 separate fields, showed mean protocol compliance of 98.6% for procedural fields, 95.5% for source documentation fields and 95.0% for data entry fields. CONCLUSIONS: This training and quality control program has resulted in a large number of surgeons capable of performing sentinel lymph node biopsy in a standardized fashion with a high degree of protocol compliance and pathologic accuracy. This will ensure optimal results for procedures performed on the randomized phase of the trial.

Axilla↗
PubMed2005

Selection of tumor-targeting agents on freshly excised human breast tumors using a phage display library.

The selective delivery of therapeutic agents to tumor site without harming rest of the body is a major challenge in clinical oncology today. Phage display approach has been used for searching ligands for cell-surface macromolecules on cancer cells so that they can be employed as drug targeting agents. Although basic protocols for biopanning cells are available, they are not as such suitable for screening highly complex and diverse target as whole tumor. Present study is an attempt to select peptide ligands specific to whole tumors. The cells from freshly collected human breast tumors were biopanned with phage displayed disulfide-constrained random heptapeptide library, following subtraction on normal human breast cells. Comparative analysis of amino acid frequencies in tumor-selected peptides and in random peptides from unselected library showed that selection was not random. The binding assessment of tumor-selected clones, using highly sensitive chemiluminescence ELISA, demonstrated that 47-75% of selected clones, depending on a tumor, bound to tumor cells they were panned on. Furthermore, several clones bound exclusively or preferentially to tumor cells in comparison to normal breast cells. It was interesting to note that insert sequences of tumor-binding clones from different tumors shared significant motifs. It shows the possibility of identifying ligands that may bind to tumor-specific targets common in certain tumors. The results of this investigation on seven human breast tumors demonstrated that, using procedures developed in the present study, whole tumors can be panned successfully with phage displayed library and tumor-binding ligands can be identified rapidly in high throughput manner. This is an important enabling step in identifying lead molecules for developing novel, specific, and effective agents that can be used for the diagnosis and treatment of cancer.

Amino Acid Motifs↗
PubMed2004

Identification of a small peptide that inhibits the phosphorylation of ErbB2 and proliferation of ErbB2 overexpressing breast cancer cells.

ErbB2 is overexpressed in approximately 30% of breast cancer patients with a correlation to poor prognosis. ErbB2 has been identified as a useful receptor for molecular targeting. A cyclic 20 amino acid phage display random peptide library was constructed using the fUSE5 gene III system. The library was panned against 2 different purified forms of the external domain of ErbB2. This resulted in the identification of several ErbB2-binding phage clones with variable binding to different ErbB2 preparations. One clone (EC-1) bound all preparations of ErbB2 including live cells and fresh frozen human breast cancer specimens. The synthetic peptide based on the deduced sequence of the EC-1 clone and its biotin-conjugated form retained binding affinity for purified ErbB2 and ErbB2 overexpressing cell lysates. EC-1 peptide was able to effectively inhibit the phosphorylation of ErbB2 on residues Y1248 and Y877 in a dose- and time-dependent manner. Furthermore, EC-1 peptide selectively inhibits the proliferation of ErbB2 overexpressing breast cancer cells. The linear portion of the cyclic EC-1 peptide was shown to be essential for binding ErbB2. In addition, 4 biased phage libraries were constructed allowing 4 different regions of the EC-1 peptide to have random sequence. Screening these EC-1 biased libraries did not result in higher affinity peptides but did demonstrate the importance of amino acids at position 1-4 on the N-terminal flanking arm and 11-15 within the cyclic ring. Interestingly, EC-1 contains homologous motifs with known ErbB receptor family ligands. We have identified a small peptide that binds to the extracellular domain of ErbB2, inhibits ErbB2 autophosphorylation and inhibits the proliferation of ErbB2 overexpressing cells. This supports the notion that small peptides can bind to targets important in cancer therapy even if a target does not have a natural ligand. Continuing research with this peptide includes increasing its affinity to ErbB2, evaluation of pharmacokinetics and evaluation of anti-proliferative effects with conjugate anti-cancer agents.

Amino Acid Sequence↗
PubMed2004

NSABP-32: Phase III, randomized trial comparing axillary resection with sentinal lymph node dissection: a description of the trial.

The NSABP-32 trial is a randomized, phase III clinical trial to compare sentinel node (SN) resection to conventional axillary dissection in clinically node-negative breast cancer patients. The primary aims of the trial are to determine if removal of only SNs provides survival and regional control equivalent to those of axillary dissection, while diminishing the magnitude of surgically related side effects. In order to ensure consistency of the outcomes for this trial, a standardized method of SN surgery has been utilized for all cases. A secondary aim of the B32 trial is to evaluate whether patients with "occult" metastases in the SNs have worse survival. Accrual is taking place at 73 institutions in North America, and 217 surgeons are enrolling patients.

Axilla↗
PubMed2003

Grb7-based molecular therapeutics in cancer.

Traditional anti-cancer drugs preferentially kill rapidly growing tumour cells rather than normal cells. However, most of these drugs have no preferential selection towards cancer cells and are taken up by the whole body, resulting in significant adverse side effects. Therapeutic molecules that could specifically inhibit undesirable phenotypes are an attractive way of eliminating cancer cells. There is a widespread effort to develop inhibitors against signal transduction molecules that play a key role in the proliferative, migratory and invasive properties of a cancer cell. Grb7 is an adaptor-type signalling protein that is recruited via its Src-homology 2 (SH2) domain to a variety of tyrosine kinases. Grb7 is overexpressed in breast, oesophageal and gastric cancers, and may contribute to the invasive potential of cancer cells. Molecular interactions involving Grb7 therefore provide attractive targets for therapeutic intervention.

Drug Delivery Systems↗
PubMed2003

Effect of tamoxifen on venous thrombosis risk factors in women without cancer: the Breast Cancer Prevention Trial.

Tamoxifen reduces breast cancer incidence among healthy women, but is associated with an increased risk of venous thrombosis. We studied the 6 month effects of tamoxifen on venous thrombosis risk factors in women without cancer. One hundred and eleven women at one centre who were participants in a multicentre breast cancer prevention trial were randomized, in double-blind fashion, to receive 20 mg/d of tamoxifen or placebo. The activated protein C (APC) ratio and concentrations of antithrombin, protein C antigen, and total protein S were measured at baseline and 6 months of treatment. None of the factors changed over 6 months in placebo-treated women. Among tamoxifen-treated women, antithrombin and protein S, but not protein C or APC ratio were reduced. Sequential antithrombin concentrations with tamoxifen were 114% and 104% (P = 0.001 compared with placebo). Sequential protein S concentrations with tamoxifen were 18.42 and 17.30 micro g/ml (P = 0.02 compared with placebo). Reductions in antithrombin and protein S were greater in postmenopausal women, but did not differ by other risk factors for venous thrombosis, such as body mass index. Reductions of antithrombin and protein S, but not protein C or APC resistance, might relate to the increased risk of venous thrombosis associated with tamoxifen treatment.

Activated Protein C Resistance↗
PubMed2003

Comparison of pathologist-detected and automated computer-assisted image analysis detected sentinel lymph node micrometastases in breast cancer.

Sentinel lymph node biopsy has stimulated interest in identification of micrometastatic disease in lymph nodes, but identifying small clusters of tumor cells or single tumor cells in lymph nodes can be tedious and inaccurate. The optimal method of detecting micrometastases in sentinel nodes has not been established. Detection is dependent on node sectioning strategy and the ability to locate and confirm tumor cells on histologic sections. Immunohistochemical techniques have greatly enhanced detection in histologic sections; however, comparison of detection methodology has not been undertaken. Automated computer-assisted detection of candidate tumor cells may have the potential to significantly assist the pathologist. This study compares computer-assisted micrometastasis detection with routine detection by a pathologist. Cytokeratin-stained sentinel lymph node sections from 100 patients at the University of Vermont were evaluated by automated computer-assisted cell detection. Based on original routine light microscopy screening, 20 cases that were positive and 80 cases that were negative for micrometastases were selected. One-level (43 cases) or two-level (54 cases) cytokeratin-stained sections were examined per lymph node block. All 100 patients had previously been classified as node negative by using routine hematoxylin and eosin stained sections. Technical staining problems precluded computer-assisted cell detection scanning in three cases. Computer-assisted cell detection detected 19 of 20 (95.0%; 95% confidence interval, 75-100%) cases positive by routine light microscopy. Micrometastases missed by computer-assisted cell detection were caused by cells outside the instrument's scanning region. Computer-assisted cell detection detected additional micrometastases, undetected by light microscopy, in 8 of 77 (10.4%; 95% confidence interval, 5-20%) cases. The computer-assisted cell detection-positive, light microscopy-missed detection rate was similar for cases with one (3 of 30; 10.0%) or two (5 of 47; 10.6%) cytokeratin sections. Metastases detected by routine light microscopy tended to be larger (0.01-0.50 mm) than did metastases detected only by computer-assisted cell detection (0.01-0.03 mm). In a selected series of patients, automated computer-assisted cell detection identified more micrometastases than were identified by routine light microscopy screening of cytokeratin-stained sections. Computer-assisted detection of events that are limited in number or size may be more reliable than detection by a pathologist using routine light microscopy. Factors such as human fatigue, incomplete section screening, and variable staining contribute to missing metastases by routine light microscopy screening. Metastases identified exclusively by computer-assisted cell detection tend to be extremely small, and the clinical significance of their identification is currently unknown.

Breast Neoplasms↗
PubMed2003

Breast cancer survival according to number of nodes removed.

BACKGROUND: Results from randomized trials indicate a 5.4% survival advantage associated with axillary dissection. To gain insight on survival outcomes when less than an axillary dissection is performed, we performed a retrospective analysis to determine survival outcome for node-negative and node-positive breast cancer patients when a variable number of nodes were excised. METHODS: The data analyzed in this paper are from the Surveillance, Epidemiology, and End Results (SEER) database, from which 72,102 patients were selected whose breast cancer had been diagnosed in 1988 or later and who were aged 40-79 years at diagnosis, had a single primary lesion, and had 0 to 3 positive lymph nodes. Cases were separated into age groups (40 to 49 and 50 to 79 years), and node-negative cases were separated from those with one to three positive nodes. RESULTS: This analysis indicates that even when all regional lymph nodes are pathologically negative, the number of nodes removed is associated with survival. In the group of breast cancer patients who had one to three pathologically positive nodes, as with the node-negative group, the higher the number of nodes removed, the greater the survival. The hazard rate for death in the node-negative group was roughly 5% less for each additional five nodes removed. For the node-positive group, the hazard rate for death was between 8% and 9% less for each additional five nodes removed. CONCLUSIONS: This retrospective study supports the notion that removal of regional nodes, even when such nodes are interpreted as pathologically negative, is important for the long-term survival of breast cancer patients.

Adult↗
PubMed2003

Lymphoscintigraphy and sentinel node biopsy accurately stage melanoma in patients presenting after wide local excision.

BACKGROUND: Patients have traditionally been considered candidates for sentinel node biopsy (SNBx) only at the time of wide local excision (WLE). We hypothesized that patients with prior WLE may also be staged accurately with SNBx. METHODS: Seventy-six patients, including 18 patients from the University of Virginia and 58 from a multicenter study of SNBx led by investigators at the University of Vermont, who had previous WLE for clinically localized melanoma underwent lymphoscintigraphy with SNBx. Median follow-up time was 38 months. RESULTS: Intraoperative identification of at least 1 sentinel node was accomplished in 75 patients (98.6%). The mean number of sentinel nodes removed per patient was 2.0. Eleven patients (15%) had positive sentinel nodes. Among the 64 patients with negative SNBx, 3 (4%) developed nodal recurrences in a sentinel node-negative basin simultaneous with systemic metastasis, and 1 (1%) developed an isolated first recurrence in a lymph node. CONCLUSIONS: This multicenter study more than doubles the published experience with SNBx after WLE and provides much-needed outcome data on recurrence after SNBx in these patients. These outcomes compare favorably with the reported literature for patients with SNBx at the time of WLE, suggesting that accurate staging of the regional lymph node bed is possible in patients after WLE.

Female↗
PubMed2003

Current status of sentinel node surgery in breast cancer.

Sentinel node surgery potentially increases the accuracy of identifying lymph nodes that contain breast cancer and decreases morbidity compared to conventional axillary lymph node resection. However, no long-term comparisons of the two modalities have been carried out, and the survival benefit associated with one protocol vs the other remains unknown. Although sentinel node surgery is not expected to increase the cure rate of breast cancer patients, a significant reduction in the incidence of permanent side effects associated with axillary node resection will be a considerable advance. The completion of clinical trials establishing that no meaningful reduction in survival is associated with the decrease in side effects is important.

Breast Neoplasms↗