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Purity: ≥98%
TPI-1 (full name: Tyrosine Phosphatase Inhibitor 1) is an inhibitor of SHP-1 (Src homology region 2 domain-containing phosphatase 1) with an IC50 of 40 nM. It was identified from a library of 34,000 drug-like compounds with anticancer activities. TPI-1 selectively increased SHP-1 phospho-substrates (pLck-pY394, pZap70, and pSlp76) in Jurkat T cells but had little effects on pERK1/2 or pLck-pY505 regulated by phosphatases SHP-2 or CD45, respectively. Significantly, TPI-1 inhibited (approximately 83%) the growth of B16 melanoma tumors in mice at a tolerated oral dose in a T cell-dependent manner but had little effects on B16 cell growth in culture. TPI-1 also inhibited B16 tumor growth and prolonged tumor mice survival as a tolerated s.c. agent. TPI-1 analogs were identified with improved activities in IFN-gamma(+) cell induction and in anti-tumor actions. These results designate TPI-1 as a novel SHP-1 inhibitors with anti-tumor activity likely via an immune mechanism, supporting SHP-1 as a novel target for cancer treatment.
| Targets |
The target of TPI-1 is Src homology region 2 domain-containing phosphatase 1 (SHP-1, also known as Protein Tyrosine Phosphatase, Non-Receptor Type 6). It inhibits recombinant SHP-1 at low nM concentrations , and has little inhibitory effect on SHP-2 or MKP1 phosphatases [1]
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| ln Vitro |
One possible therapeutic target for cancer has been identified as SHP-1. As the SHP-1 phosphate substrate pLck-pY394 is increased, TPI-1 becomes effective at 10 ng/mL. TPI-1 has a slight impact on pERK1/2 and pLck-pY505, but it preferentially increases SHP-1 phosphosubstrates (pLck-pY394, pZap70, and pSlp76) in Jurkat T cells. TPI-1 stimulates IFNγ+ cells in human peripheral blood and mouse spleen [1].
1. TPI-1 selectively increased SHP-1 phospho-substrates (pLck-pY394, pZap70, pSlp76) in Jurkat T cells at low nM levels, while having minimal effects on pERK1/2 (regulated by SHP-2) or pLck-pY505 (regulated by CD45) [1] 2. In in vitro phosphatase activity assays, TPI-1 inhibited recombinant SHP-1 activity in an escalating dose-dependent manner, but showed no significant inhibition of recombinant SHP-2 or MKP1 even at higher concentrations [1] 3. TPI-1 induced IFN-γ⁺ cells in mouse splenocytes in vitro, with an efficacy approximately 58-fold higher than sodium stibogluconate (SSG), as quantified by ELISPOT assays [1] 4. The compound also induced IFN-γ⁺ cells in human peripheral blood cells in vitro when cultured for 16 hours, as detected by ELISPOT [1] 5. TPI-1 had little cytotoxic effect on Jurkat T cells in culture (assessed by MTT assay after 6 days of treatment) and showed no significant inhibition of B16 melanoma cell growth in vitro after 5 days of culture [1] |
| ln Vivo |
TPI-1 has no effect on B16 cell proliferation in culture, but at a tolerable oral dosage it reduces the formation of B16 melanoma tumors in mice in a T cell-dependent way. As a result, TPI-1 also raises IFNγ+ and pLck-pY394 in mice. As a well-tolerated SC drug, TPI-1 also prevents the formation of B16 tumors and extends the longevity of tumor mice[1].
1. In C57BL/B6 mice bearing 4-day established B16 melanoma tumors (5×10⁴ cells/inoculation), oral administration of TPI-1 (3 mg/kg, daily, 5 d/wk) inhibited tumor growth by approximately 83% (p < 0.002) in a T cell-dependent manner; no significant tumor inhibition was observed in nude mice (T cell-deficient) treated with the same regimen [1] 2. Subcutaneous (s.c.) administration of TPI-1 (1 mg/kg, daily, 5 d/wk) to C57BL/B6 mice with 4-day established B16 tumors (10⁵ cells/inoculation) also inhibited tumor growth and prolonged the survival of tumor-bearing mice [1] 3. TPI-1 (1 or 3 mg/kg, s.c., daily) increased pLck-pY394 levels in mouse splenocytes (quantified by Western blotting/densitometry) and elevated the number of IFN-γ⁺ splenic cells (measured by ELISPOT) in vivo [1] 4. Sodium stibogluconate (SSG, 12 mg daily, 5 d/wk, s.c.) showed weaker anti-tumor efficacy against B16 tumors in mice compared to TPI-1 [1] 5. Interleukin-2 (IL-2, 3×10⁵ IU, bid, 5 d/wk, i.p.) had limited anti-tumor effect on B16 tumors in mice, while TPI-1 exhibited superior efficacy under the experimental conditions [1] |
| Enzyme Assay |
1. Recombinant SHP-1/2/MKP1 Phosphatase Activity Assay: Recombinant SHP-1, SHP-2, or MKP1 enzymes were incubated with escalating doses of TPI-1 in a suitable reaction buffer under optimized assay conditions. Phosphatase activity was quantified using a phosphatase substrate detection system (specific substrate not detailed). The inhibitory effect of TPI-1 on each phosphatase was determined by comparing the enzyme activity in the presence of the compound to the vehicle control (mean ± SD of triplicate determinations) [1]
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| Cell Assay |
1. Jurkat T Cell Phospho-Substrate Analysis: Jurkat cells were cultured in standard medium and treated with vehicle control or TPI-1 at various doses for 10 minutes. Total cell lysates (TCL) were prepared and analyzed by SDS-PAGE/Western blotting using antibodies against pLck-pY394, pZap70, pSlp76, pERK1/2, and pLck-pY505 to assess the selective effect of TPI-1 on SHP-1 phospho-substrates [1]
2. Jurkat Cell Viability Assay: Jurkat cells were cultured in the absence or presence of TPI-1 for 6 days. Cell growth/viability was quantified by MTT assay, with data presented as mean ± SD of triplicate samples [1] 3. IFN-γ⁺ Cell Induction Assay (Mouse Splenocytes/Human Peripheral Blood): Mouse splenocytes or human peripheral blood cells were isolated and cultured in vitro with vehicle control, TPI-1, or SSG for 16 hours. The number of IFN-γ⁺ cells was quantified by ELISPOT assays (mean ± SD of duplicate samples) [1] 4. B16 Melanoma Cell Proliferation Assay: B16 cells were cultured with TPI-1 for 5 days, and cell growth was evaluated by MTT assay (mean ± SD of triplicate samples) [1] |
| Animal Protocol |
TPI-1 (~ 1 or 3 mg/kg, s.c.) for 4 days
mice (6~8-week old, female) #### Animal Protocol 1. B16 Melanoma Tumor Model (Oral Administration): C57BL/B6 mice (n=5) were inoculated with 5×10⁴ B16 cells subcutaneously to establish tumors for 4 days. TPI-1 was administered orally at a dose of 3 mg/kg daily (5 days/week) for the experimental period. Tumor volumes were measured regularly to assess anti-tumor efficacy; body weights were monitored to evaluate tolerability [1] 2. B16 Melanoma Tumor Model (Subcutaneous Administration): C57BL/B6 mice (n=5) were inoculated with 10⁵ B16 cells subcutaneously for 4 days. TPI-1 was administered subcutaneously at 1 mg/kg daily (5 days/week). Tumor volumes and mouse survival rates were recorded; nude mice (T cell-deficient) were used as a control group to confirm T cell-dependence of the anti-tumor effect [1] 3. Splenic IFN-γ⁺ Cell and Phospho-Substrate Analysis in Mice: Mice were treated with TPI-1 (1 or 3 mg/kg, s.c., daily) for a specified period. Splenocytes were isolated to prepare total cell lysates for Western blot analysis of pLck-pY394 levels and to perform ELISPOT assays for IFN-γ⁺ cell quantification [1] 4. SSG/IL-2 Control Treatments in B16 Tumor Mice: Mice bearing B16 tumors were treated with SSG (12 mg daily, 5 d/wk, s.c.) or IL-2 (3×10⁵ IU, twice daily, 5 d/wk, i.p.) as positive/negative controls, with tumor volumes measured for comparison to TPI-1 [1] 5. Toxicity Assessment in Mice: Balb/c mice were treated with TPI-1 (~10 mg/kg, s.c., daily, 5 d/wk) for two weeks, and mouse viability/body weight was monitored to evaluate acute toxicity [1] |
| Toxicity/Toxicokinetics |
1. In mice, TPI-1 was well tolerated at oral doses of 3 mg/kg (5 days a week) and subcutaneous doses of 1 mg/kg or approximately 10 mg/kg (5 days a week for two weeks), and no significant weight loss or death was observed in the treatment groups.[1]
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| References | |
| Additional Infomation |
1. TPI-1 is a novel SHP-1 inhibitor, screened from a library of 34,000 drug compounds. Its preclinical antitumor activity is mainly mediated through immune mechanisms (T cell-dependent induction of IFN-γ⁺ cells) rather than directly producing cytotoxicity on tumor cells [1]. 2. SHP-1 is a negative regulator of immune cell activation. The inhibitory effect of TPI-1 on SHP-1 can enhance T cell signaling (by increasing pLck-pY394, pZap70, pSlp76) and the production of IFN-γ, thereby leading to an antitumor immune response [1]. 3. TPI-1 analogues (such as TPI-1a4) have been developed with higher antitumor efficacy. TPI-1a4 completely inhibited the growth of mouse K1735 melanoma tumors at a tolerable oral dose and also inhibited MC-26. The efficacy of TPI-1 in treating colon cancer is superior. [1]
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| Molecular Formula |
C₁₂H₆CL₂O₂
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| Molecular Weight |
253.08
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| Exact Mass |
251.974
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| CAS # |
79756-69-7
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| Related CAS # |
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| PubChem CID |
948589
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| Appearance |
White to yellow solid powder
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| LogP |
3.084
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| Hydrogen Bond Donor Count |
0
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| Hydrogen Bond Acceptor Count |
2
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| Rotatable Bond Count |
1
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| Heavy Atom Count |
16
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| Complexity |
385
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
ZKHFYORNAYYOTM-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C12H6Cl2O2/c13-7-1-3-11(14)9(5-7)10-6-8(15)2-4-12(10)16/h1-6H
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| Chemical Name |
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| Synonyms |
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
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| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (9.88 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (9.88 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (9.88 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.9513 mL | 19.7566 mL | 39.5132 mL | |
| 5 mM | 0.7903 mL | 3.9513 mL | 7.9026 mL | |
| 10 mM | 0.3951 mL | 1.9757 mL | 3.9513 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
TPI-1 inhibits recombinant and cellular SHP-1 and had little cytotoxicity in vitro and in mice.J Immunol.2010 Jun 1;184(11):6529-36. |
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TPI-1 selectively increases SHP-1 phospho-substrates in Jurkat cells at low nM levels.J Immunol.2010 Jun 1;184(11):6529-36. |
 TPI-1 inhibits B16 tumor growth as a tolerated single agent.J Immunol.2010 Jun 1;184(11):6529-36. |
 TPI-1 induces IFNγ+cells in mouse splenocytes and human peripheral bloodin vitro.
TPI-1 increases spleen pLck-pY394 and IFNγ+cells in mice. |
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 TPI-1 analogs TPI-1a2 and TPI-1a4 have anti-tumors in mice at tolerated oral doses.J Immunol.2010 Jun 1;184(11):6529-36. |
 TPI-1 analogs inhibit rSHP-1 in correlation with activity to increase SHP-1 phospho-substrate pLck-pY394 in Jurkat cells and induce IFNγ+ cells in mouse splenocytes. |
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