Automated Preparation of [68Ga]Ga-3BP-3940 on a Synthesis Module for PET Imaging of the Tumor Microenvironment

This research focuses on the development of an automated radio labeling protocol for the preparation of Gallium 68 3BP3940. This compound is an experimental radiopharmaceutical for petaimaging Of the tumor microenvironment. Targeting the tumor microenvironment has gained significant interest in recent years. In the field of radiopharmaceuticals a key challenge is to develop efficient and safe automated radio labeling protocols for their preparation. The main experimental challenge in producing gallium labeled investigational radiopharmaceuticals is to implement a radio labeling protocol that is finally adapted to both the molecule to be radio labeled and the synthesizer used. In the field of gallium 68 labeled experimental radiopharmaceuticals. We highlighted that a careful optimization of the automated reaction conditions can significantly improve the radio labeling process for a given compound. This protocol addresses the lack of automated Gallium 68 radio labeling of three BP 3940, and offers a turnkey method for its preparation using a given synthesizer, enabling synthesis in just 20 minutes. Its GMP compliance enables radio labeling of pharmaceutical grade three BP3940, supporting the clinical applications of this innovative radiotracer.

[Narrator] To begin unwrap the gallium 68 cassette envelope. After checking for any damage, tighten each lurer connection on the cassette using a five milliliter lurer lock syringe fitted with a 21 gauge needle. Withdraw five milliliters of absolute ethanol from the reagent kit and pass it very slowly over the C18 cartridge. Then using the same syringe, withdraw five milliliters of water for injection and pass it very slowly over the same cartridge. Position Ramp A of the tubing set on the synthesis module and turn the two latches to hold it in place. Connect the free end of the vertical A one tubing to a 19 gauge needle and insert it into the waste vial. Insert a venting needle into the waste vial and orient it backward behind the shielded container that will receive the evacuation vial places 0.22 micrometer filter in position a four. Connect the horizontal A one tubing to the pressure sensor located at the bottom left of the module front panel. Using a male to male adapter connect a 30 centimeter extension line in the horizontal A five position ending in a 0.22 micrometer terminal filter and an 80 millimeter 20 gauge needle. Insert the 20 gauge needle into a sealed sterile evacuation vial. Then add an aeration needle and place the vial in its shielded container. Place the tubing line connecting vertical A one to vertical C one behind the retaining hooks above Ramp B. Connect horizontal manifolds A two and b one using a short extension line. After removing the adapter mounted in position A two, position ramp B on the synthesis module and secure it by turning the two latches. Connect the precondition C 18 cartridge to the horizontal C two position, making sure the adapter linking horizontal valve B five to C two is kept on the left side. Position Ramp C on the synthesis module before securing it by locking both latches. Using a male to male adapter, connect a 50 centimeter extension line from horizontal C five to the gallium 68 generator. Place the stained glass reaction vial of the tubing set in the module oven. Then carefully place the tubing from vertical A five to vertical C five in the peristaltic pump. Close the pump and after verifying that the tubing is properly positioned, pass the tubing through the activity sensor on the left hand side of the pump. Connect the 250 milliliter water for injection bag to c four tubing using the spike adapter and hang the bag on the designated hook located on the right side of the module. Using a three milliliter, three piece lurer lock syringe with a 20 gauge needle withdraw 1.5 milliliters of 10 milligrams per milliliter L methionine solution. Connect the syringe to horizontal C one tubing and hang it in the designated slot on the right hand side of the module. Leave approximately two milliliters of air between the liquid surface and the syringe plunger seal to ensure complete transfer. Using a one milliliter syringe with a 20 gauge needle, withdraw 750 microliters of the 10 milligrams per milliliter L methionine solution and inject it into the 0.9% sodium chloride vial after disinfecting the septum. Using a 10 milliliter three piece lurer lock syringe with a 20 gauge needle, withdraw the contents of the 0.9% sodium chloride and L methionine vial. Adjust the volume to 8.6 milliliters. Remove the spike in position B four and connect the syringe in B four instead. After disinfecting the septum, withdraw the contents of the 60% ethanol vial using a three milliliter three piece lurer lock syringe with a 20 gauge needle. After confirming that the volume is at least 1.5 milliliters, remove the spike from position B five and connect the syringe in its place. Using a low dead volume one milliliter syringe fitted with a 20 gauge needle withdraw 0.25 milliliters of 0.8 molar sodium acetate buffer solution, and inject it into the vial containing 30 micrograms of three BP3940. Solubilize it by successive injection and aspiration cycles. Withdraw the 0.25 milliliter solution into the same syringe. Remove the needle and place the syringe into position B three. Click run synthesis when all reagents are placed on the ramps and all required information is correctly recorded in the software. Transfer the terminal vial to an appropriate shielded cell. To prepare it for radioactivity measurement and patient dose preparation. Measure the radioactivity of the terminal vial using a properly calibrated dos calibrator and record the preparation on the computer. After labeling the vial correctly, place it inside an appropriate shielded container. Using aseptic and radiation protection techniques withdraw approximately 0.5 milliliters of the sample from the terminal vial for quality control testing. Examine the sample visually to assess clarity and color. Assess the pH by applying a drop of the product onto a pH paper strip. Deposit a drop of the product solution on each of the two ITLCSG plates to measure the radio chemical purity by radio TLC. Allow the plates to develop in the appropriate mobile phases and read them using the radio chromatograph. Integrate the resulting radio chromatogram by measuring the area under the curve of the product signal and the impurity signal. Then calculate the radio chemical purity according to the formula shown on the screen to measure radio chemical purity. Using radio HPLC inject approximately 50 microliters of the sample into the HPLC vial. Load the vial into the HPLC autos sampler at the correct position and start the analysis sequence. After the sample has been injected, remove the vial from the autos sampler before returning it to the shielded container to minimize exposure. At the end of the run integrate the radio chromatogram by measuring the area under the curve for the product and impurity signals as shown previously, and calculate the radio chemical purity using the given formula. A comparison of the effectiveness of different anti radiologists agents in preserving radio chemical purity during crude gallium three BP 3940 synthesis is shown in this figure. Without terminal purification, Methionine at 10 milligrams per milliliter achieved the highest radio chemical purity of 94.7% compared to gentisic acid and ascorbic acid. The final product collected in the terminal vial retained an average of above 75.1% of total radioactivity confirming high yield and efficient collection. Total synthesis related losses averaged only about 24.9% distributed among the waste vial, SPE cartridge, and reaction vial indicating low waste generation. The average final activity of the radio labeled compound across the three test batches was above 737 megabecquerel indicating consistent synthesis performance.