1-427. (canceled)
428. An anti-CD79b antibody comprising
a) a light chain variable domain amino acid sequence SEQ ID NO: 99; andor
b) a heavy chain variable domain amino acid sequence SEQ ID NO: 100.
429. An anti-CD79b antibody comprising
a) a light chain amino acid sequence SEQ ID NO: 33; andor
b) a heavy chain amino acid sequence SEQ ID NO: 35.
430. The antibody of claim 428 or 429, wherein the antibody binds to an epitope within a region of CD79b comprising an amino acid sequence comprising amino acids 29-39 of SEQ ID NO: 4.
431. The antibody of claim 428 or 429, further comprising one or more free cysteine amino acids.
432. The cysteine-engineered antibody of claim 431, which is prepared by a process comprising replacing one or more amino acid residues of a parent antibody by a free cysteine amino acid.
433. The cysteine-engineered antibody of claim 431, wherein the one or more free cysteine amino acids are at a position in the light chain according to Kabat numbering convention selected from the group consisting of 15, 43, 110, 144, 168 and 205, andor at a position in the heavy chain according to EU numbering convention selected from the group consisting of 41, 88, 115, 118, 120, 171, 172, 282, 375, and 400.
434. The antibody of claim 431, wherein the one or more free cysteine amino acids have a thiol reactivity value in the range of 0.6 to 1.0.
435. The cysteine engineered antibody of claim 432, wherein the cysteine engineered antibody is more reactive than the parent antibody with a thio-reactive reagent.
436. The cysteine engineered antibody of claim 432, wherein the process further comprises determining the thiol reactivity of the cysteine engineered antibody by reacting the cysteine engineered antibody with a thiol-reactive reagent; wherein the cysteine engineered antibody is more reactive than the parent antibody with the thiol-reactive reagent.
437. The cysteine engineered antibody of claim 431 wherein the one or more free cysteine amino acid residues are located in a light chain.
438. The cysteine engineered antibody of claim 431 wherein the one or more free cysteine amino acid residues are located in a heavy chain.
439. The cysteine engineered antibody of claim 431, which is an immunoconjugate comprising the cysteine engineered antibody covalently attached to a cytotoxic agent.
440. The cysteine engineered antibody of claim 439, wherein the cytotoxic agent is selected from a toxin, a chemotherapeutic agent, a drug moiety, an antibiotic, a radioactive isotope, and a nucleolytic enzyme.
441. The cysteine engineered antibody of claim 431, which is covalently attached to a capture label, a detection label, or a solid support.
442. The cysteine engineered antibody of claim 441, wherein the antibody is covalently attached to a biotin capture label.
443. The cysteine engineered antibody of claim 441 wherein the antibody is covalently attached to a fluorescent dye detection label.
444. The cysteine engineered antibody of claim 443 wherein the fluorescent dye is selected from a fluorescein type, a rhodamine type, dansyl, Lissamine, a cyanine, a phycoerythrin, Texas Red, and an analog thereof.
445. The cysteine engineered antibody of claim 441 wherein the antibody is covalently attached to a radionuclide detection label selected from 3H, 11C, 14C, 18F, 32P, 35S, 64Cu, 68Ga, 86Y, 99Tc, 111In, 123I, 124I, 125I, 131I, 133Xe, 177Lu, 211At, and 213Bi.
446. The cysteine engineered antibody of claim 441 wherein the antibody is covalently attached to a detection label by a chelating ligand.
447. The cysteine engineered antibody of claim 446 wherein the chelating ligand is selected from DOTA, DOTP, DOTMA, DTPA and TETA.
448. The antibody of any one of claims 428, 429, and 431 comprising an albumin binding peptide.
449. The antibody of claim 448, wherein the albumin binding peptide is selected from SEQ ID NOs: 52-56.
450. The antibody of claim 433, wherein a cysteine is at position 205 of the light chain.
451. The antibody of claim 433, wherein a cysteine is at position 118 of the heavy chain.
452. The antibody of claim 433, wherein a cysteine is at position 400 of the heavy chain.
453. The antibody of any one of claims 428, 429 and 431 which is selected from the group consisting of a monoclonal antibody, a bispecific antibody, a chimeric antibody, a human antibody, and a humanized antibody.
454. The antibody of claim 431 which is an antibody fragment.
455. The antibody of claim 454, wherein the antibody fragment is a Fab fragment.
456. The antibody of claim 431 which is a humanized antibody.
457. The antibody of claim 431 which is produced in bacteria.
458. The antibody of claim 431 which is produced in CHO cells.
459. A pharmaceutical formulation comprising the anti-CD79b antibody of claim 431, and a pharmaceutically acceptable diluent, carrier or excipient.
460. The antibody of claim 431, wherein the antibody is covalently attached to an auristatin drug moiety whereby an antibody drug conjugate is formed.
461. The antibody-drug conjugate of claim 460 comprising an antibody (Ab), and an auristatin drug moiety (D) wherein the cysteine engineered antibody is attached through one or more free cysteine amino acids by a linker moiety (L) to D; the compound having Formula I:
Ab-(L-D)p \u2003\u2003I
where p is 1, 2, 3, or 4.
462. The antibody-drug conjugate compound of claim 461 wherein p is 2.
463. The antibody-drug conjugate compound of claim 461 wherein L has the formula:
-Aa-Ww\u2014Yy\u2014
where:
A is a Stretcher unit covalently attached to a cysteine thiol of the cysteine engineered antibody (Ab);
a is 0 or 1;
each W is independently an Amino Acid unit;
w is an integer ranging from 0 to 12;
Y is a Spacer unit covalently attached to the drug moiety; and
y is 0, 1 or 2.
464. The antibody-drug conjugate compound of claim 463 having the formula:
where PAB is para-aminobenzylcarbamoyl, and R17 is a divalent radical selected from (CH2)r, C3-C8 carbocyclyl, O\u2014(CH2)r, arylene, (CH2)r-arylene, -arylene-(CH2)r\u2014, (CH2)r\u2014(C3-C8 carbocyclyl), (C3-C8 carbocyclyl)-(CH2)r, C3-C8 heterocyclyl, (CH2)r\u2014(C3-C8 heterocyclyl), \u2014(C3-C8 heterocyclyl)-(CH2)r\u2014, \u2014(CH2)r(C(O)NRb(CH2)r\u2014, \u2014(CH2CH2O)r, \u2014(CH2CH2O)r\u2014CH2\u2014, \u2014(CH2)C(O)NRb(CH2CH2O)r\u2014, \u2014(CH2)rC(O)NRb(CH2CH2O)r\u2014CH2\u2014, \u2014(CH2CH2O)rC(O)NRb(CH2CH2O)r\u2014, \u2014(CH2CH2O)rC(O)NRb(CH2CH2O)r\u2014CH2\u2014, and \u2014(CH2CH2O)rC(O)NRb(CH2)r\u2014; where Rb is H, C1-C6 alkyl, phenyl, or benzyl; and r is independently an integer ranging from 1 to 10.
465. The antibody-drug conjugate compound of claim 463 wherein Ww is valine-citrulline.
466. The antibody-drug conjugate compound of claim 464 wherein R17 is (CH2)5 or (CH2)2.
467. The antibody-drug conjugate compound of claim 463 having the formula:
wherein R17 is a divalent radical selected from (CH2)r, C3-C8 carbocyclyl, O\u2014(CH2)r, arylene, (CH2)r-arylene, -arylene-(CH2)r\u2014, (CH2)r\u2014(C3-C8 carbocyclyl), (C3-C8 carbocyclyl)-(CH2)r, C3-C8 heterocyclyl, (CH2)r\u2014(C3-C8 heterocyclyl), \u2014(C3-C8 heterocyclyl)-(CH2)r\u2014, \u2014(CH2)rC(O)NRb(CH2)r\u2014, \u2014(CH2CH2O)r, \u2014(CH2CH2O)r\u2014CH2\u2014, \u2014(CH2)C(O)NRb(CH2CH2O)r\u2014, \u2014(CH2)rC(O)NRb(CH2CH2O)r\u2014CH2\u2014, \u2014(CH2CH2O)rC(O)NRb(CH2CH2O)r\u2014, \u2014(CH2CH2O)rC(O)NRb(CH2CH2O)r\u2014CH2\u2014, and \u2014(CH2CH2O)rC(O)NRb(CH2)r\u2014.
468. The antibody-drug conjugate compound of claim 467 wherein R17 is (CH2)5 or (CH2)2.
469. The antibody-drug conjugate compound of claim 463 having the formula:
470. The antibody-drug conjugate compound of claim 461 wherein L is SMCC, SPP or BMPEO.
471. The antibody-drug conjugate compound of claim 461 wherein D is MMAE, having the structure:
wherein the wavy line indicates the attachment site to the linker L.
472. The antibody-drug conjugate compound of claim 461 wherein D is MMAF, having the structure:
wherein the wavy line indicates the attachment site to the linker L.
473. The antibody-drug conjugate compound of claim 461 wherein D is DM1, having the structure:
wherein the wavy line indicates the attachment site to the linker L.
474. The antibody-drug conjugate compound of claim 460 wherein the antibody is a humanized antibody.
475. The antibody-drug conjugate compound of claim 460 wherein the antibody fragment is a Fab fragment.
476. An antibody-drug conjugate compound selected from the structures:
wherein Val is valine; Cit is citrulline; p is 1, 2, 3, or 4; and Ab is an antibody of claim 431.
477. The antibody drug conjugate of claim 460 wherein the auristatin is MMAE.
478. The antibody drug conjugate of claim 461 wherein L is MC-val-cit-PAB.
479. A pharmaceutical formulation comprising the antibody drug conjugate of claim 460, and a pharmaceutically acceptable diluent, carrier or excipient.
480. An article of manufacture comprising the pharmaceutical formulation of claim 479; a container; and a package insert or label indicating that the antibody drug conjugate can be used to treat cancer characterized by the overexpression of a CD79b polypeptide.
481. The article of manufacture of claim 480 wherein the cancer is selected from the group consisting of lymphoma, non-Hodgkins lymphoma (NHL), aggressive NHL, relapsed aggressive NHL, relapsed indolent NHL, refractory NHL, refractory indolent NHL, chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma, leukemia, hairy cell leukemia (HCL), acute lymphocytic leukemia (ALL), and mantle cell lymphoma.
482. A method for making an antibody drug conjugate compound comprising an anti-CD79b antibody (Ab) of claim 431, and an auristatin or maytansinoid drug moiety (D) wherein the antibody is attached through the one or more engineered cysteine amino acids by a linker moiety (L) to D; the compound having Formula I:
Ab-(L-D)p \u2003\u2003I
where p is 1, 2, 3, or 4; the method comprising the steps of:
(a) reacting an engineered cysteine group of the antibody with a linker reagent to form antibody-linker intermediate Ab-L; and
(b) reacting Ab-L with an activated drug moiety D; whereby the antibody-drug conjugate is formed; or comprising the steps of:
(c) reacting a nucleophilic group of a drug moiety with a linker reagent to form drug-linker intermediate D-L; and(d) reacting D-L with an engineered cysteine group of the antibody;
whereby the antibody-drug conjugate is formed.
483. The method of claim 482 further comprising the step of expressing the antibody in chinese hamster ovary (CHO) cells.
484. The method of claim 482 further comprising the step of treating the expressed antibody with a reducing agent.
485. The method of claim 484 wherein the reducing agent is selected from TCEP and DTT.
486. The method of claim 484 further comprising the step of treating the expressed antibody with an oxidizing agent, after treating with the reducing agent.
487. The method of claim 486 wherein the oxidizing agent is selected from copper sulfate, dehydroascorbic acid, and air.
488. A polynucleotide encoding an antibody of claim 428, 429, or 431.
489. A vector comprising the polynucleotide of claim 488.
490. A host cell comprising the vector of claim 489.
491. A method of making an anti-CD79b antibody, the method comprising
(a) culturing a cell expressing an antibody comprising a heavy chain variable domain of claim 428 or 431 and a light chain variable domain of 428 or 431; and
(b) isolating the antibody from said cultured cell.
492. A method of treating a subject having cancer, said method comprising administering to the subject an effective amount of an antibody of any one of claims 428, 429, and 431.
493. The method of claim 492 wherein the cancer is selected from lymphoma, non-Hodgkins lymphoma (NHL), aggressive NHL, relapsed aggressive NHL, relapsed indolent NHL, refractory NHL, refractory indolent NHL, chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma, leukemia, hairy cell leukemia (HCL), acute lymphocytic leukemia (ALL), and mantle cell lymphoma.
494. The method of claim 492 wherein said antibody is conjugated to a cytotoxic agent.
495. The method of claim 492 wherein said antibody is conjugated to a growth inhibitory agent.
The claims below are in addition to those above.
All refrences to claim(s) which appear below refer to the numbering after this setence.
1. A method for heating a sample, said method comprising:
providing a heating apparatus which includes a waveguide, an applicator, and a deflector positioned in said waveguide to form a resonant cavity with the sample and said applicator;
inserting the sample in the applicator;
generating electromagnetic radiation at a first output power level; and
rotating the deflector for adjusting coupling factors between the waveguide and the resonant cavity, wherein rotating the deflector includes:
performing at least once one of the following:
positioning the deflector in a first position and measuring a first power of electromagnetic radiation reflected from the applicator, the reflected radiation corresponding to said first position of the deflector,
rotating the deflector to a second position that is different from the first position and measuring a second power of electromagnetic radiation reflected from the applicator, the reflected radiation corresponding to said second position of the deflector; and
determining a preferred position of the deflector based on the amount of power reflected from the waveguide applicator in at least the first and second positions.
2. The method according to claim 1, wherein the sample has a first temperature T1, the method further comprising:
heating the sample to obtain a second temperature T2, wherein T2>T1; and
rotating the deflector to adjust the coupling factor between the waveguide and the resonant cavity in response to a variation in a dielectric properties \u2208sample of the sample.
3. The method according to claim 1, further comprising:
providing a first storing means;
storing information relating to the first position in the storing means and storing a measured first power in relation thereto; and
storing information relating to the second position in the storing means and storing a measured second power in relation thereto.
4. The method according to claim 3, wherein determining the preferred position of the deflector further comprises processing the stored first and second measured powers for determining the preferred position of the deflector corresponding to a local or an absolute minimum in the measured power or to a predetermined ratio of the measured power to the first output power level.
5. The method according to claim 1, further comprising positioning the deflector in a preferred position.
6. The method according to claim 1, further comprising positioning the deflector in a preferred position and generating electromagnetic radiation at a second output power level which is larger than the first output power level.
7. The method according to claim 3, further comprising determining a relative permittivity measurement of the sample by comparing the stored measured powers with corresponding stored measured powers from a different sample.
8. The method according to claim 3, further comprising determining an indication of a chemical composition of the sample by comparing the stored measured powers with corresponding stored measured powers from a sample of known chemical composition.
9. The method according to claim 8, wherein the sample comprises at least one reactant for performing a chemical reaction.
10. The method according to claim 9, further comprising:
performing a chemical reaction with the at least one reactant; and
determining a degree of reaction for the chemical reaction using an indication of chemical composition of the sample.